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ED 209 119 SE 035 928

A .
TITLE' Microscopic Analysis, of Activated Sludge. Training

Manual. -
'INSTITUTION A Office of Viatr--4graik Operations (EPA)r, Cincin
L'Ohio. FatipAil Training and Operational Technology

Center. :'
'. '''
3, FEPORT NO EPA-430/1-80-007 ,

POP DE Jun 80
NOTE 250p.; Contains,occasional light and broken type.
-Pages'1-198 ',Field Key to Some Genera of Algae', and
. ',Ciliated isiotoZoa removed due.to copyright %
H AVAILABLE FROM EPA Indtructional'Reiources Center, 1200 Chambers
, Rd., 3rd,ylbor, Columbus, OH 432124$1.00 plus $0.:03
. .
per Otge). O


EDRS PRICE MF01/PC10 Plus Postage. * .

DESCRIPTORS .Biological Stiences; Data Analysis; *Laboratory

Equipment; *Laboratory Procedures; AIMicrobiology; .

. - *Microscopes:, Postsecondary Education; *Water ,

'Pollution; :water lesouices
'IDENTIFIERS Activated Sludge; Analytical Methods; *Waste Water

---/-: 4
This trIiniA4 manual pwents material on the use Of
a compound microscope to analyze microscope communities, present in
wastewater tteatient procefses, for operational control. Course
topics includei sampling techniques, sample ha moiling, lairdratory
analysis, iderftificatiot, of organisms, data interpretation,. and use
'4 heecomOcund microscope: This manual contains 26 chapters
including reading material,- 'laboratory activities, 'and selected
references. Prior expgriende ip-microscopy is not necessary. (Co)



* Reproductions suppliedrbEDRS atethe best that can be made,. *.--
* fromIthe'original document.-

t °
United States National Training EPA-430/1-80-007
Environmental Protection and Operational June 1980 ,
Agen Cy Technology Center
Cincinnati OH 45268

a EPA Microscopic
Analysis of N
r-4 Activated Sludge A

Training Manual
This document has been reproduced as
received from the person or organization
onginatino a
XMinor changes have been made to improve
reproduction guat
4. stated in this docu
Points of view of 01)16'10ns
mem do not necessarily represent official N1E
position or policy,


_IL J1_
LLI __
__ __
4 June-198Q 1.

Microicopic Anasis of Activated Sludge

This course is for anyone who needs the skills to

use a compound' microscope toanalyze microscopic
communities, present in wastewater treatment
processes, for operational control. Prior experience
in microscopy 'is not necessary:.
`After successfully.completing the course; the student
will be able to relate microscopic communities present
in the wastewater treatment process to operational
controls. The student will also be capable of instructing
treatril4nt plant personnel in the more proficient use of
. the compound mitroscope and re1ating communities
present to operational control.
4 The training, includes classroom instruction and
laboratory practice.



Office of, Water Program Operations
National Training and Operational Technology Center


., Reference to commercial products, trade names, or

manufacturers` is for_purposes of example and illustration.
Such reference's do not constitute ehdorgement by the
Officeof Water Program Operations, U. S. Environmental
4201). .
Protection Agency.



f Title or Description -Outline Number
The Aquatic Environrdent 1

Classification of Communities, Ecosystems, .and Trophic Levels 2

Limnology and Ecology'of Plankton - 3
Biology', of Zooplankton 4
, Optics and the Microscope 5
Strubture and Function. of Cells 6
Bacteria and Protozoa as Tokicological Indicators 7
iiamentous Bacteria
Fungi and the "Sewage Fungus" Community
Protozoa, Neniatodes, and Rotifers' 0
Activated sludge Protozoa 11 A
Free-Living Amoebae and Nematodes 12
Animal Plankton 13
Preparation and Enumeration of Plankton in the Laboratory 14

Attached Growths 15
Effect of WaVewater Treatment Plant Effluent on Small Streams 16
Ecology iol WaLste Stabilization Processes 17
Ecology Primer 18
The Laivs of Ecology 1

Application Orlifological Data 20

Signilicance of 9Limiting Factors" to POPulation Variation 21
Algae and Cultural Eutrophication 22
Calibration and Use of Plankton Counting Equipment 23
Laboratory: Proportional Counting. of Mixed Liquor . 24
. . t
Key to Selected' Groups of Freshwater Ant . 25
,A Key for the Initial Separation o f Sortie C o on Plankton Organisms 26.
Field Key to Some Genera of Algae
i . .-
Ciliated, Protozoa

r 5

Part 1: e Nature and Behavior of Water

I INTRODUCTION . forms, but also with living Organisms'

and infinite interactions that occur
The earth is physic y divisible into the bet hem and their environment.
lithospherekor land ma ses, and the
hydrosphereiwhich includes the oceans, C Water quality management, including
lakes, stre(ms, and subterranean waters; pollution control, thus looks to all
and the atmosphere. branches of aquatic science in efforts
to coordinate and improve man's
to Upon the hydrospere are based a number
of sciences which represent different
relationship with his aquatic environment.

approaches. Hydrology is the general

science of water itself with its various
special fields suclkas hydrography, - II SOME FACTS ABOUT WATER
hydri.tros, etc. These in turn merge
into physical chemistry and chemistry. AN.Water is the only abundant liquid on ota
planet( It has many properties most
B Limnology and Oceanography combine unustial for liquids, upon which depend
aspects of all of thegei and deal not oaly, most df the familiar aspebts of the world
with the physical liquid water and its about us as we know it. ZSee Table 1)
various naturally occurring solutions and


Pro2erty_ Significance
Highest heat capacity (specific heat) of any Stabilizes temperatures of oilanismstand
sulici or liquid (except NH3) ',geographical regions
Highest latent heat of fusion (except NH3 / Thermostatic effect at freezing point
Highest heat.of evaporation of any substance Important in heat and water transfer of
The only substance that has its maximum Fresh and brackish waters have maximum
density as a liquid (4°C) density abode freezing point. This is
- impo ;tant in- vertical circulation pattern
in lager.
Highest surface tension of any liquid Controls surface and drop phenomena.
iniportant in cellular physiology .
Dissolves more substances in greater Makes complex biological system possible.
quantity than any othei. liquid Important for transportation of materials
'in solution.
Pure water has the bfgheat di- electric Leads to high dissociation of inorganic
,natant Of any liquid . substances in solution .

Very little electrolytic dissaciat Neutral, yet contains, both ft+ and OH ions
Relatively transparent AbsOrbs much,energy In infra red and ultra
violet ranges, bta little in visible range.
Bence ."color less" `,1

21g23. 80 4

The Aquatic Envitomnent

i 4 a

B ysical-Factors of Significance TABLE 2

Water is not, simply "H20" but in
reality is a mixture of some 33 Temperature (°C) Density
different substances involving three
isotAeS each of hydrogen and oxygen Water
(ordinary hydrogen 111,, deuterium
and tritium H3; ordinary oxygen al°, -10 .99815 .9397
oxygen 17; and oxygen 18) plus 15 -8 99869 .9360
known ty }es of ions. The molecules ,-6 9991'2 .9020
of a water mass tend to associate .99945 .9277
themselves as polymers 'rather than 2 .99970 .92,29
to remain as discrete units. '0 .99987 - --)9168
(See Figure 1) .
'2 .99997
6 4 1.00000
6 .99997
206: .99823
40 .99225
60 .98324
80_ .97183
I 100 '" .195838

Tabular values for density, etc.; represent

estimates brvarious workers rather than
absolute values, due to the variability of
** Regular Ice is knoivn as "ice I. Four or
more other "forms" of ice are known.tq
_exist (ice II, ice III, etc. ), having densities °
at 1 atm pressure ranging'from 1.1595
I -,to 1.67. These are of.extremely restricted

Figure I
occurrence and. may be ignored in most
routine operations. -

2 Density
This epsures°That ice usually
forms on top of a body of water
a TeMperature and density: Ice.
Water is the only known substance
( and tends'to insulate the renfain-
ing water mass from fuither_loss
in which the solid.state will float of heat. Did ice sink, there
on the liquid state. (See Table 2) could be little or no carryover of
aquatic life from.season to season
in-the higher latitudes. Frazil or
needle ice forms colloid fly at a
few Thousandths of a- degree
-below CI° C. It is adhesiye arid
may build up on submerged objects
as "anchor ice's', but it is still
.typical-ice (ice I):
The Aquatic Enviionmentt

Seasonal increase in solar Mineral-rich water from the

radiation annually warms hypolimnion, for example,,,,
surface waters in summer is mi.Red with oxygenated
while other factors result in . water from the epilimnion.. war

winter cooling. The density This usually triggers a

differences resulting establish sudden growth or.ubloom"
two classic layer's: the epilimnion of. plankton organisms.
.or surface layer,' and the
hypolimnion or lower layer, and 6) When stratification is present,
in between.is the thermocline however, each layer belgiaves,
or shear-plane. relativelytindependently, and
significant quality 'differences.
2) While for certain theoretical may develop.
purposes d'"thermocline" is
defined as a zone in which the '7) Thermal stratification as
temperature changes one described above has no
degree centigrade for each reference to the size of the
meter of depth, in practice, water mass; it is found In
any transitional layer between oceans and puddles.
two relatively stable masses
of water of different temper- b The relative denSities of the
atures may be regarded as a various isotopes of water
thermOcline. influence its molecular com-
position. For example, the
3) Obviously the greater the lighter 016 tends to go off
temperature differences first in the process of evaporation,
between epilimnion and leading to the relative enrichment
hypolimnion and the sharper of air by, 016 acid the enrichmett
the gradient in the thermOcline, of water by 017 and 018. This
the more stable will the can lead to a measurably' higher
situation be. 018 content in warmer climates.
Also, the temperature of water
4) From information given above, in past geologic ages can be
it should be evident that While closely estimated from the ratio.,"
the temperature of the of 018 in the. carbonate of mollusc
hypolimnion rarely drops shells.
much below 4° C, the
epilimnion may range from c Dissolved and /or suspended solids
0° C upw' ard. may,,also affect the density of
natural water masses (see Table ,3)
r 5) When epilimnion and hypolimnion
achieve the same temperature, TAI3LE 3
stratification no longer exists. EFFECTS OF DISSOLVED SOLIDS
The entire body of water)3 ehayes ON D ENSITY
hydrologically asaunit, and
tends to assume uniform chemical Dissolved Solids Density
and physical characteristics. (Grams,per liter) (at 4°C)
Even a light breeze may then, 0 1.00000'
cause the entire body of water 1 1.00085
to circulate, Such events. are called
overturns, and usually result in . 2 1.00169
water quality changes Of consider- 3 1.00251
able physical, chernicar, and
biological significance. 10 . 1.00818
t. 35 (mean for sea water) . 1.02'822

1 The Aquatic Environment

d Types-of density stratification ';-____ This is important not only in situations

involving the control of flowing water
1) Dersity differences produce as in a sand filter, but also since, -
stratification which may be overcoining resistance to flow gen-
,_,permanent, transient, or erates heat, it is significant in the
seasonal. heating of water by internal friction
from wave and.current action.
, 2) 'Permanent stratification Living organisms more easily support
exists'for example where themselves in the more viscous
`there is a heavy mass Of (and also denser) cold waters of the
'brine in the deeper areas of arctic than in the less vis cous warm
a basin which does not respond waters of the tropic. (See Table 4).
to seasonal'or other changing
conditions. t' TABLE 4

3) Transient stratification may VISCOSITY OF WAT R (In millipoises at 1 attn)

occur with the recurrent
influx of tidal Water in an pissted solids in g/L
estuary,for example, or the -
occasional influx of cold Temp. 0 C 0 1 5 10
muddy water into a deep lake -10 26.0 1

or reservoir.. ..
t ,
- 5. 21.4 I
7,--- ---- - - --
4) Seasonal stratification is 0 '17, 94 18,1 18.24 18.7
typically t rynal in nature, 15.3 15.5 16.0
and thyolv the-annual 5 15/19
establisAm t of the epilimnion, 10 13 ,10 13.2 13.4 1.3.8
hypolimnion, and thertnocline :
30 8,00 8,1 8,2 8.6
as described above. .
5) Density stratification is not'
limited to two-layered systems;
three, four, or ven more 4 Surface-tension has biological as,well
layers may be encountered in as physical-significance. Organisms
larger bodies of water. whose body gurfaces cannot be wet by
'I water can either ride on the surface
e A "pltinge line' (sometimes called film or in some instances may be
"thezmal line) niay develop at "' "trapped" on the surface film andbe
the mouth of a stream. Heavier unable to re-enter the water.,
water flowing into>,a lake or
reservoir plunges below the ' 4. 5 :Heat or energy
lighter Water mass ofthe epiliminium
to flow along at a lower level. Such Incident solar radiation is;the'prime
a line is usually'marked by an Source of energy for virtually all
accumulation of floating debris._ organic and most inorganic processes
on earth. For the earth as a whole
f Stratification may be modified th'e total amount (of energy) received
or ,entirely -suppressed in some _annually must exactly balance that
cases *hen deemed expedient, by lost by reflection and radiation into
mean- s of a simple. air lift.' space if climatic and related con-
, ditions are to remain relatively
3 The viscosity of Water is greater at constant over geologic time.
lower temperOures (see Table 4).

The Aquatic Environment %

a' For a given body of water, significant change in surface.

immediate sources of energy level is detected. Shifts in
inclyle in addition to solar submerged water masses of 0 .

irradiation terrestrial, heat, this type can have severe effects

transformation of kinetic energy on the biota and also on human
(wave and current action) to heat, water uses where withdrawals
'chemical and biochemical are.confined toa given depth,
reactions, convection from the Descriptions and analyses of
atmosphePe, and condensation of many other types arid sub-types
ater vapor. of waves and wane -like movements
may be found in the literature.
b the progortion'of light reflected
depends on the angle of incidence, b Tides
the temperature, color, and other
qualities of the water; and the 1) Tides are the longest waves
presence or absence of films,. (' known, and are responsible for
of lighter liquids such as oil. the orrce or twice a day rythmic
In general, "as the ddpth increases rise'and fall of the',Ocean level
arithmetically, the light fends to on most shores, around the world.
decrease geometrically. Blues,
greens, and yellows tend to 2) Wylie part, and parcel of the
penetrate most deeply while ultra same phenomenon, it is'often
violet, violets, and orange-reds Convenient to refer to the rise
are most quickly absorbed. On and fall of the water level as . .0
,the order of 90% of the total "tide, " and to the resulting
illUmination which penetrates the A currents as "tidal currents. ".1
surface film is absorbed in the
first 10 meters of even the clearest 3) Tides are basically caused by the
water, thus tending to warm the attraction of the sun and moon on
upper layer,s. water masses, large and small;
however, it is abnly in the oceans
6 Water movements and peksibly certain of the larger
lakes that time. tidal action has
a Waves or rhythmic movement been,demonstrated. The patterns
ortidal action are enormously
4) The best known are traveling complicated by local, topography,
waves caused 'by wind. These are .interaction with seiches, ancrother
effective only against objects near fa,ctors. The literature on tides
the surface. They have little- is very large.
effect on the movement of large 'a

$ Currents (except tidal currents) -

masses of water. - c
are steady arythrni-c water movelneAs
2) Seiches t which have had maRir study only in
oceanography although they are,
Standingwaves seiches occur most often observed, in rivers and
in lairs, estuaries, and other, stream's. 'They are "primarily
e - ed bodies of water, but are concerned with the tranilocation of
ge enough to be water masses. They May bp generated,
observed. An "internal wave or internally by virtue of density changes,
seicli" is an oscillation in a, or externally by wind or terrestrial
subrhersed mass of water such topography,' Turbulence phenomena
asaa hypblimmon, accompanied or eddy currents are largely respbn-
by compenSating oscillation in the Sible. for, late rals.Mixing in a curtrent .
overlying Water so, thatdo, These are of far more importance
in the econry of a body of water than
. mere laminar flow.
14 10. S.

The Aquatic Environment

d °Coriolis force it a result of inter- depend on the depth of the water,

action between the rotation of the the velocity and duration of the
earth, and the movement of masses wind, and other factors. The net
or bodies on the earth. The net result is that adjacent cylinders
result is a slight tendency for moving tend to rotate in oppOsite directions
objects to veer to the right.in the like meshing cog wheels. Thus,
northern hemisphere, and to the\ the water between two given spinals
left in the southern hemisphere. maybe meeting and sinking, while
While the result in fresh waters is that between spirjls on either side
usually negligible, it may be con- will be meeting and rising. Water
siderable in marine waters.. For over the sinking, while that between
example, other factors permitting, spirals on either side will be meet-
ifiebre is a tendenby in estuaries for ing and rising:, Water over.the
fresh waters-to move toward the sinking areas tends to accumulate
ocean faster along the right bank, flotsam and jetsam on the surface
while salt tidal waterstend to 'in long conspicuous lines. '
intrude farther inland,along the.
left bank. Effects are even' more °a This phenomenon is of consider-
dramaticin the open oceans. able importance to those sampling
forplankton (or even chemicals)
e Langmuire spirals (or Langinuire near the surface when the. wind
circulation) are a relatively is blowing. Grab samples from
massive cylindrical motion imparte d either dance might obviously
to surface waters under_ the influence differ considerably, artkeif
of wind.' Thq axes of the cylinders ' a plankton tow is contemplated ..\\1
are parallel to the .direction of the. t it should be made,,across the
wind, and their depth and velocity ere wind in order that thesnet
may pass Through a succession
of both dances.

- b y b WATER



Figure 2. 'Langmuitilpirals
h.. Blue dna, water r.
dance., water sinking, floating o I

-swimming.,ohiects concentrated.

;- 1-6

The Aquatic Environment,

b Langmuire spirals. are not REFERENCES

sually established until the
wind has either beenk blowing Buswell, A. M. and Rodebush, W: H.
for an ,extended period, or Water. Sci, Am. April 1956.
, else,is blowing rather hard.
Their presence can be detected 2 Dorsey, N. ,Ernest. Ribperties of
by the lines of foam and Ordinary Water Substance.
other floating material which Reinhold Publ. Corpew. York.
coincide with the direction pp. 1-673. 1940.
of the wind.
3 Fowle, Frederick E. Smithsonian
6 The pH of pure water hasheen deter-, PhyMcal XabIes. -Smithsonian
Miscellaneous Collection, 71(1),
mined between 5.7 and 7.01-by various. 7th revised ed., 1929.
workers. The latter value is most
,,widely accepted at the present time. 4 Hutcheson, George:E. A Treatise on (
Natural waters of coarse vary widely Limnology. John Wiley Company.
according to circumstances. 1957.
C The elements of hydrology mentioned
above represent a selecton of some of
the more conspicuous physical factors
involved in working with water quality:
Other }hems no spebificallif Mentioned
include:. molecular structure of waters, /
° /7.

interaction of water and radiation,

internal pressur,e, acoustical charac-
1 .
teristics, presVure-volume-temperature'
. relationships, refractivity, luminescence, .4
color, dielectrical characteristics and
:phenomena, solubility, action and inter.-
actions .of gases; liquids and 'solids,
. water vapor, phenomena of hydrostatics
and liy.drodynamics in general.



Part 2: The Aquatic Environment as an Ecosystem'

) I


Part-1 introduced the lithosphere and the ORGANISMS, AND BETWEEN ORGA-
hydrosphere. Part 2 will deal with certain NISMS AND THEIR ENVIRONMENT.
general aspects of th.e biosphere, or the
sphere of life on this earth, which photp_-- A The ecosystem is the basic functional'
graphs from space have showri is a finite unit of ecology. Any area of nature that
globe in infinite spaces includ s ing organisms and nonliving
substances nteracting to produce an
This is the habitat of man and the other *exchange o material8 between the livihg
organisms. His relationships with tie and nonliv g pd.rtse,constitutes an
aquatic biosphere are our common concern. ecosystem. (Odurp, 1959) -
1 From a structural standpoint, it is
,II THE BIOLOGICA I., NATURE OF THE- convenient to recognize four
WQRLD WE LIVE IN cpnstituents as composing an
ecosystem (Figure.1).
4 A We can only imagine what'this world
musthave been like before there was life: a Abiotic NUTRIENT MINERALS
which ape the physical stuff of
B The" ircSrld as we know it is largely shaped ,which living protoilasdi will be
by the forces of life. synthe.sized.
1 Primitive forms of life created organic b utotrophic (self-nourishing) or
matte?.-and estab/i'shed soil. PRODUCER organisms. These
are largely the -green plants .
D Plants cover the lands and "enormously -(holophytes), .but other minor
influehte the forces of erosion. groups must also be included
..(See Figure 2). They assimilate
3 The nature and rate of 'erosion affect' the nutrient, rpinerals,..by the use
the redistribution of materialE . of conSiderable energy, and combine
(and mass) on the Surface of -the their into living organic, substance.
earth (topographic changes). '
C Heterotrophic (Ole r-nourishing)
4 Organismstie up vast quantities of coNsulAgas (holozoic), are chiefly
certain chemicals, such as carbon' the animals. They ingest (or eat)
and oXygen. Ss, and digest organic matter, releasing
considerablk energy in the process.
5 Respiration of plants and animals
releathes carbon dioxide to the . d Heterotrophic REDUCERS are chiefly
atmosphere in influential quantities... bUcteria and fungi that return
complex organic compounds back to
6 CO affects the heat transmisthori of the originaribiotic mineral condition,
the2atmosphere. thereby reletising the remaining
chemical energy.
C ,Organisms respond to and in turn affect
- their environment.. Man is one a the 2 From a functional standpoint, an ,
-'rhosi. influential. ecosystem 'ha.8 two parts (Figure 2)

1 -9
The Aquatic Environment



a The -autotrophic or producter, 2 These two groups can be `defined on

organisms, which utilize light the basis of relative complexity of 1
energy or the oxidation of in- structure.
organic bompounds, as their
sole energy source. a The bacteria andblue-gieen
lacking a nuclear membrane are .

b The heterotropic or consumer the Monerit.

and reducer organisms which
eutiliies organic compounds for b The single-celled algae and
its energy and carbon requirements. protozoa are Protista.
3 Unless the autotrophic and hetero- 1
trophic phases of the cycle approximate C Distrilpited throughout these groups will
a dynamic equilibrium, the ecosystem be found most of the traditional
'and the environment will change. . of classic biology.

B Each dethesegtoups includes simple,

single-celled representativei, persisting IV FUNCTIONING OF THE ECOSYSTEM
4arr at lower levels on the evolutionary stems
of the higher organisms. (Figure 2) A A food chain is the transfer of food energy
from plants through a aeries of ,organisms
1 These groups span the gaps between the With repeated bating and being eaten.
higher kingdoms with a multitude of Food chains are rrot isolated sequences but
transitional forms: They are collectively axe interconnected.
called.the PROTISTA and MONERA.

The Aquatic Environment


Energy'Flows from, eft to Right, Gal Evolutionary Sequence is Upward


.OrganicNiaerial PrOdtced, Arganic Material Ingested or Organic Material Reduced
UsuallyN PhotosTifFirt Consumed. by Extracellular Digestion
Digested Ipternally and Intracellular Metabolibth,
to Mineral Condition

Flowering Plants and Arachnids Mammals

Gymnosperps Basidiomycetes
Insects Birds
Club Mosses, Ferns Crustaceans Reptiles
Segmented Worms Amphibians
Liverworts, Mosses Fungi Imperfecti.
Molluscs Fishes
Bryoz oa Primitive
fylulticellular Green C'bordates
Algae Rotifers
Roundworms Echinoderms
Red Algae Flatworms
Coelenterates p
Higher Phycomycetes
Brown Algae Sponges

P r o 10 v 0 a
Unicellular Green Algae Lower
A nioet;oic.1
Diatoms Phycoznycetes
Flagellated, Suctoria -
Pigmented Flagellates (nnii-pigmented)' (Chytridiales, et. al. )'


Blue Green Algae Actinomycetes
1- "IpimenaMes
Phototropic Bacteria 4
Saprophyt lc
I Bacterial
Chenwtropie Bacteria es -

131. ECO. pl. 2a. 1.69

/The Aquatic EmAroliment


` .?:?311,

B A food web is the interlOcking pattern of D _Total Assimilation

food chains in an ecosystein. (Figures 3, 4)
In complex natural communities, organisms The amount of energy which flows through
whose food is obtained by the same number a trophic level is distributed between the
of steps are said to belong to the same production of biothass (living substance),
trophic (feeding) level. and the demands of respiration (internal
energy use by living organisms) in a ratio
C Trophic Levels of approximately 1:10.
E Trophic Structure of the Ecosystem
1 First - Green plants (producers)
(Figure 5) fix biochemical energy and The interaction of the food chain
wlthesine basic organic subotances. phenomena (with energy loss at each
This is primary production . transfer) results in various communities
2 Second - Plant eating animals (herbivores) having definite trophic structure or energy
depend on the producer organisms Mt levels. Trophic structure- may be
food. measured and described either in terms
of the standing crop per unit area or in
3 Third - Primary cazpivores, aninials terms of energy fixed per unit area per
which feed on herbivores. unit time at successive trophic leWis. ,
Trophic structure and function can be
4 Fourth --Secondary carnivores feed on shown graphically by means of ecological
primary carnivores. pyraMids (Figure 5).

5 Last Ultimate carnivores are the last

or. ultimate level of consumers. 1


Basic units are asifollows: 1, abfotic substancesbasic inorganic and

Figure 3. Diagram cf the pond ecosystem.vegetation;
organic compounds; IIA, producers--rooted IIB, producersphytoplankton; III-1A. primary consumers
(herbivores)zooplankton; 111.2, secondary consumers (car- .`
(berbivores)bottom forms; III.1B, primary con
decomposersbacteria and fungi of decay.
&Imes); 1114, tartluy consumers (secondary carnivores); IV,

* 1

The Aquatic Environment


-**. /
Nutrient E

\ 1

Bacterial Death and decay

action ti1/4 \ /
x.....-..,_. -.


topr ts,1

1;11101 LI,t1

771.r./1%..4 Ajo, so.

tta 4,01. 1

Figure 4. A MARINE ECOSYSTEM (After Clark, 1954 and Patten, 1966)

The Aquatic EfiVironment

Includes bacteria, algae, .protozoa and
Decomposers Carnivores (Secondary) other micnoscopic animals, and often the
Carmvores (Primary" young or embryonic stages of algae' and
other organisms that normally grow. up
fierbworek to become a part of the benthos (see below).
Producers any planktonic types will also adhere
(b) to surfaces as periphyton, and some
ty.pical periphyton may break off and
be collected as plankters:
C' Benthos are the plants and animals living
on, in, or closely associated with the
bottom. They include plants and ,,
D Nekton are the community of strong
aggressive swimmers of the open waters,
often called pellagic. Certain fishes,
whales, and invertebrates such as
'Figtire 5. HYPOTHETICAL PYRAMIDS of shrimps and squids are included here.
(a)'Numbers of individuals, (b) Bioniass, and
lc) Energy (Shading Indidatet Energy:Loss). E The marsh community is based on larger
"higher" plants,` floating and ernergent.
Both marine and freshwater marshes are
areas of enormous biological production.
. Collectively known at "wetlands", they
gap between the waters and the
.pA Plankton are the macroscopic and VI PRODUCTIVITY
microscopic animals, Plants, ba teria,
etc., floating free in the open wat r. A The biological resultant of all physical
Many clog filters, cause tastes, odors,. and chemical factors in the quantity of
and other troubles in watel-supplies. Hire that may actually be present. The
Eggs and larvae of larger forms are abilityto produce this "biomass" is
often presetit. often referred to as the "productivity"
of a body of water. This is neither good
1 Phytoplankton are ,T4..ese nor bad per se. &water cflow'pro-
are the dominarit producers of the duCtivity is a "poor" water biologically,
waters, fresh and salt, "the grass , and also a relatively "pure" or "clean"
of the seas ".' water; hence desirable as a water supply
or a bathing beach. ,A productive water
2; Zooplankton are animal-like. on the other hand may be a nuisance to
Includes many different animal types, man or highly desirable. it is a nuisance
range in size from minute protozoa if foul odors and/or weed-chocked
to gigantic marine jellyfishes. ; waterways result, it is 'desirable if
bumper crops of bass, catfish, or
tst Periphyton (orAufwuchs) - The communities oysters are prodliced. Open oceans have
of microscopic organisnis associated.with a low level of productivity in general.
submerged surfaces of any type or depth. .

- S
The Aquatic Environment

VII PERSISTENT CHEMICALS' IN THE a Oysters,Vor instance, will'con-

ENVIRONMENt centrafe DDT 70,000 times higher
*, \ in their tissues than it's concentration
Increasingly complex manufacturing processes, in surrounding water.
. They can
coupled with rising industrialization, create also partially cleanse themselves
health hazards for humans and aquatic life. in water free-of DDT.
Compounds besides being toxic (acutely or b Fish feeding on lower organisms
chrOnic) may produce 'mutagenic effects build up concentrations in their
' including Cancer, tumors, and teratogenicity visceral fat which may reach several
(embryo defects). Fortunately there are,tests, thousand parts per million and levels
such as the Aniis test, to screen chemical in their edible flesh of hundreds of
compounds for these effects. parts per million.
o Larger animals, such-as fish-eating
A Metals - current levers of cadmium, lead gulls and other birds, can-further
and other substances constitute a mount- concentrate the chemicals. .A survey
ing concern. Mercury pollution, as at on organochlorine residtes in aquatic
Minimata., Japan has been fully documented. birds in the Canadian prairie provinces
showed that California and ring-billed.
Pesticides gulls were among the most contaminated.
Since gulls breed in colonies, .breeding
1 A pesticide and its metabolites may , population changescan be detected and
move through an ecosystem in many related to levels of chemical con-
ways. Hard (pestidides which are tamination. Ecological research on
persistent, having a long half-life in colonial birds to monitor the effects
the environment includes the organo- of chemical pollution on the environ-
-chlotines,' ex., DDT) pesticides . inent is useful.
ingested or otherwise borne by the
target species will stay in the "Polychlorinated biphenyls" (PCB's).
'nvironment, possibly to be recycled C
PCB's were.used in plasticizers, asphalt,
or concentrated further through the ink, paper, and a host of other products.
hattiral action of food chains if the Action was Taken to curtail their release
species is eaten., Most of the volume to the environment. sincedtheir effects
Of pesticides do not reach their target are similarto hard pesticides. However
at all. ,
this doesn't solve thofroblems of con-
taininated sedinients and ecosystems and
2 Biological magnification final fate of the PCB's still circulating.
Initially, low levels of persistent '\: D There are numerous,other compounds
pesticides in air, soil, and water' may which are toxic and-accumulated in the
.. be concentrated at every step up the ecosystem.
food chain. Minute aquatic organisms
and scavengers, which screen water and
ottom mud having pegticide levels of a
few parts peribillion, can accumulate
levels measured in parts per milliona.
thousandfold increase. Thesediments
including fecal deposits are continuously
recycled by the bottom animals.
A,' I


r le-
The AqUa-tic Environment

FERprcEs 5 Odum, E.P. Fundamentals of Ecology.

W. B. Saunders Company,
1 Clarke;' G. L. ents of Ecology. Philadelphia and Londont 1959.
',john Wiley & Sons, New York. 1954.
6 Patten, B. C. Systems Ecology.
2 Cooke, W. B. ''rickling Filter Ecology. j_kige,Stance. 16(9). 1966.
Ecology 40(2):273-291. 1959.
7 Whittaker, R.H. New Concepts of
3 Hanson, E.D. Animal Diversity. Kingdoms. Science 163:150-160. 1969.
Prentice-Hall, Inc New Jersey. 1964.

4 Hedgpeth, J.W. Aspects of the Estuarine

Ecosystem. Amer. Fish. Soc., Spec.
Publ. No. 3. 1966.

a 4.
. part 3. The Freshwater Environment

I INTRODUCTION During periods of run-off after a

rain or snow -melt, -buch a gulley
The freshwater environment as considered -would have a flow of Water which
herein refers to those inland waters not might range from torrential 'to a
detectably diluted by ocean waters, although mere trickle. Erosion mayproceed
the lower portions of rivers are subject to rapidly as there is no permanent
certain 'tidal flow effects.. aquatic Llora or 'fauna to stabilize
streambed materials. On the other
Certain atypical inland waters such as saline hand, terrestrial grass or forest
or alkaline lakes,, springs, etc., are not- growth may retard 'erosion. When
treated, as the main objective here is tyPical.... "Ate run-off has passed, however,
inland water. - the "streambed" is dry.

All waters have certain basic biological cycles 2 Youthful streams. When the
and types of interactions most of Which have streambed is- eroded be/ow the
already beed presented, hence this outlinT ground water level, spring or
will concentrate on aspects essentially, seepage water enters, and the
peculiar to fresh inland waters. stream becomes permanent. An
aquatic cflora and fauna develops
and water flows the year round,.
II PRESENT WATER QUALITY AS A , Yout hful streams typically have a
relatively steep gradient, rocky.beds,
FRESH.WATERS with rapids, falls°, and small pools.
A The history of ,body of water determines 3 Mature streams. Maiure stred.ms
its present condition. Nattficarwaterd have have wide valleys, a developed
evolved in the course of geologic time flood plain, are 'deeper, more
into what :we know today. turbid, and usually have warmer '
B Streams,
water, sand, rilud, silt, O clay
bottoin materfalb Ng& shift with . 4
increase' in flow,- In Jheir more
In the course of their evolution, setealiffs favorable reaches,- streams in this
in general pass through four stages of oondition are at a peak of biological
. development which may be called: birth,'
productivity. Gradients' are moderate,
youth, maturity, and old age. _riffles, or rapids are often separated
by long pools. -
' These terms or conditions may be
employed or considered in two contexts: I/ 4 age, ',streams have approached
temporal, or spatial. In terms of geologic 4 geologic Vase level, usually the
time, a given point in a stream may pass ocean. During -flood siOgge they scour
through each of the stages described below their beds and deposit. materials on
or: at any given time, these various stages the flood Plain which may be very
of development can be loosely identified bread and flat. During normal flow
in successive reaches of a stream traveling the channel is refilled and:Many
from its he dwaters tb base level in ocean shifting bars are developed. *eanders
or major la e. -
and ox-bow lakes are often formed.
1 Establishment eir birth. *is
might be a :'dry runt' or headwater'.
stream-bed,. before it had eroded
down to the level of ground watery

The A uatic Environment

f,(Under the influ ce of man this became a lake. Or; heglacier may,-
Pattern may be b oken up, or actually scoop out a."'hole.. Landslides
temporarily inte upted. Thus an may dam valleys, extinct volcanoes;may
essentially "youthful" stream might... collapse, etc., etc.-
take on some of the *characteristics
of_ a "mature" stream following soil 2 Maturing or natural eutrophication of
erosion, organic enrichment, and lakets.
increased surface runoff. Correction IP
of these-conditions might likewise be a If not Airoor present shoal areas
followed by at least a partial reversion are developed through erosion
to the "original" Condition). and deposition of the shore material
by wave action and undertow.
C Lakes and Resertroirs
b. currents produce bars across bays
GeologiCal factors.iirhioh significantly and thud cut off irregular areas.
affedt the flatting of either a stream or
-lake include toe. following: c Silt brought in by tributary streams
settles out in. the quiet lake water
1 The geographical locatiOn of the
. ' drainage basin or watershe . ed to-eurfacet,
apd floating free plankton. Dead
2 The size and shape of the drainage organic matter begins to accumulate
basin. .on,the bottom.
45 The general topography, i.e.,' e Rooted aquatic plants grow on
mountainous or plains. shoals and bars, and in doing so
mit off bays and.coqribute to the
4 The character of the bedrocks and filling of the lake.
f Dissolyed carbonates and other
5 The character, amount, annual material's are precipitateld in the
distribution, and rate' of precipitation,. deeper popions of the lake in part
thfough the action o plants.'
6, the natval vegetative cover of the .1
land_ is, of course, responsive to and . g When filling is well.a anced,,
responsible for many of the above ' mats of sphagnum mods may extend
factors and is also severely subject outward from the shore. These .

to.the whims of civilization. This . mats are followed by sedies and

is one of the major factors determining grasses which finally con ter_ the
run-off versud soil absorption, etc. lake marsh.
D Lakes have a developmental history which 3 _Extinction of lakes. After lakes reach
somewhat parallels that'of streams. This matukty4, their progress tolyerd
process is often referred to as natural fillirig up is accelerated. They become
eutrophication:' extinct through:

1 The methods of formation vary greatlyf a- the. downcutting

of thec outlet.
bid all influence the character and ''7
subsequent.chibitory of the lake. b Fkllfng with detritus erode from
the shores or brought in b
In glaciated areas, :for example, a tributary streams.
huge block of ice may have been covered
with till. The glacier !retreated, the c Filling by the a ccumu,lation'Ok the .
ice melted, 'and the resulting hole remains of vegetable materials
grering.in the lake itself.
(Often two or three proceSdes may
4 act concurrently)
The Aquatic Environment
HP PRODUCTIVITY IN FRESH WATERS 2 As the stream flows toward a more
"mature condition, nutrients tend to
A Fresh waters'in general and under . accumulate, and. gradient diminishes
natural conditions by definition have a ,+ and So time of flow increases, fern;
lesser supply of dissolved substances perature tends to increase, and
than marine waters, and thus a lesser plankton floUrish. .
basic potential forthe growth.of aqUatic
organiSims By tile same token, theST. _Should a heavy load of-inert silt
maybe said to be more sensitive to tile develop on the other hand, the
addition of extraneous materials turbidity would reduce the light
(pollutants, nutrients, etc. ), The penetration and consequently the
following notes are directed toward general plankton prctduction would
natural geological and other environ-
mental faqtorS as they ffect the .

productivity`of fres% $ters. 3 As the streaniapproaches base/lev el

(old age) and the, time 'available for
B Factors Affecting Stream Productivity 'plankton growth increases, the c
(See Table 1) balance between to nutiie t
levels; and temper ture and ther
TABLE' I seasonal conditions, determines the
° overall productivity.
PRODUCTIVITY* C -Factors Affecting the Productivity of
lakes (See Tape 2)
(The productivity of sand bottoms is
taken as 1) 1 The size, shape, and 4epth of the
lake basin. Shallow water is more
productive than- deeper Water" since 1

Relative more light will.reach the bottom to

Bottom Material # Productivity stimulate rooted plant grOwth, As
a corollary, lakes,.with ThOre shore-
Sand 1
line, having more shallow water, '
Marl 6 are in general more productive.
Fine Gravel 9 -c,
/ Broad shallow lakes and reservoirs
Gravel and silt , '14 have the greateit production potential
Coarse gravel ' . . 32 (and hence should be avoided for
Moss on fine gravel 8g 'Nwater supplies). s
Fiasidens (moss) on coarse Lit
._ .----v
gravpl - TABLE 2.
Ranunculus (water buttercup) '19'4' , a
Watercress 301 EFFECT OF .SUBSERA TE
Elodea,(waterweed) 452 -; ON LAKE PRODUCTIVITY
.. -

*Selected from Tarzwell 1937

To be productive of aquatic life, a -
stream -must provide adequate nutrients,
light, a 1 suitable' temperatu,re:. and time
forgrowth to take place.

productivity of sand-bottcims is taken- s 1)
Relative Productivity


-Pebbles 1.
Flay ...
° --..\
1 -YOUthfurstreamS; especially on rock
.ore sandSUbstrates are Zoo in-essential Flat rubble .`
's-mutrients. Temperatures in moun- Block rubble ' 11
tainous regions are usually low,' and .
Shelving,r9ck. 77
A ...
due to thp steep,gradient, time for 4

.g.xlowth is rt. Although ample * Selected frolinTarzwell 1937

light is a,ilable; growth of true
plankt is thus greatly llmited. 9
Aquatic Environment
2 °Hard,waters are generally more . IV CULT RA EUTROPHICATION
'prOductite than soft waters as there 1
are more plant nutrient A. The genleral processes of natural
available.- This is often greatly in- eutroph cation, or natural enrichment .
fluenced byl.the character of the soil 4 and pro uctivity have been briefly out-
and rocks in the watershed'and the lined aboye.
quality and quantity of ground water . .
entering Ze lake. In general; pH B When the°\activities of 'man speed up
ranges'oP6.8 to =$. 2 appear to be these enr chfnent Processes by intro-
^ most 'productive. ducing atural quantities of nutrients
tc. ) the result is often called
, 3 Turbidity reduces productivity as cultural e tro hication. This term is
is reduced. o often exte ded beyond its original usage
, to include the enrichment (pollution) of
4 The presence or absence of thermal streams, /estuaries, and even oceans, as
stratification with its .semi-annual well as likes.
turnovers affects productivity by C

distributing nutrients throughout the

5 Glimate, temperature, prevalence of
ice and snow, Of course A The productivity of lakes and impoand-
important. ments is such a conspicuous feature that
it is often used as a convenient of
D Facto /ale ctini the uctivity of :44 :classification.
Reservoirs .

1 Oligotrozhic lakes are the younger,

1 The productivity of reservoirs is less productive lakes, which are deep,
governed by much the same principles have clear water, and usually support
as that of lakes, withoike difference Saimonoid fishes in their deeper waters.
that the water level'is much more
under the control of man. Fluctuations 2 Eutroyhic lakes' are more mature,
in water level can be used tb de- more turbid, and richer. They are
liberately increase o'rdecrease usually shallower. They are richer
productivity. This can ,be aemOnstated in dissolved solids; N, P, and Ca are
by a comparison.of the TVA reservoirs abundant.. Plankton is abundant and
- which practice a summaidrawdown ther'e is often a rich bottom fauna.
with some of those in, the 'west where
,a winter drawdown is the rule. 4 Dy strophic lakes, such as bog lakes,
are low in Ph, water yellow to brown,
2 The level at which watif is removed ;;
dispolved solids, N, 1', and Ca scanty
from a reservoir is important to the. but hun4c materials abundant, bottom
productivity of the stream below. , fauna and plankton poor, and fish
'The hypolimnion may. be anaerobic species 13.re limited.
while the epilimnion is aerobic, .for
...example, or...the epilimnion is poor in R eservoirs lay also be class ified as
nutrients. While the hypolitnnion is orage, an run of the river.
orage reserNirs have a large
3 Reservoir discharges also Profoundly volume in relation to their inflow.
affect the DO, temperature, and
turbidity in the.stream below a'dam. 2 Run'of the river reservoirs have a
Too much fluctuationin flow may'° iarge.flow-thrOZh in relation to their
permit sections of he stream, to dry, ° storage value:
or proVide inadequate dilution for
..toxic waste.
, The Aquatic Environment

C According to 'location, lakes and

reservoirs may be classified as polar,
temperate, or tropical. Differences in .

climatic .and geographic conditions

xesult in,sdifferences in theiebiology.

A A body of water such as a lake, stream, l,
or estuary represents an intricately
balanced-system in a state of dynamic
eqnilibrium. Modification imposed at
one point in the system automatically
results in compensatory adjustments at
B The mca4 th rough our knowledge of the 40.
entire system the better we can judge
where to ivnpo e control, measures to
achieve a desir d result.

1 Chamberlin, Thomas C. and Salisburg,
Rollin1). Geological Processes and
Their ResUlts. Geology 1: pp
and 1-654. Henry Holt and Company.
New York. 1904.

2 Hutcheson, George E. A-Treatise, on

Limnology Vol. I \Geography, Physics
and Chemistry. 197. Vol. II.
Introduction to Lake Biology and the
Limnoplankton. 11'l5 pp. 1967.
John Wiley CO.
3 Hynes, H. B. N. The Ecology of Running
Waters. Univ. Toronto Press.
555 pp. 1970,

4 De Santo, Robert S. Concepts of

Applied EpolOgy. Spinger-
Verlag. 1978. A


.1 -2
MO 0 .4.

Part 4. The Marine Environment and its Mile in the Total Aquatic EhAronment
. I
TABLE 1 e .. '1 k. l
-' (Data from Cthrk, F.W., 1924, '"The CoinpOsition of ffiverx
A The marine emlir onmerit is arbitrarily and Lake Waters of the United Stales", U.S. . Geal. Surv,
, Prof. Paper'No. 135; Harvey, H. W. 1957. "The Chemistry 4
defined as the water massextending ..,
and Fertility of Sea Waters", Cambridge ljniversitrPretx, ?. ,
beyond the continental land masse1/4., Cambridge) ..., V'''' " 4
including the plants and animals harbored'. -,.
1 '1 r
therein. This water`mass is large and
'i'deep, coveleing 'about 70 percent of the
earth's surface and being as deep as

Ioik:,1,-. -.rat
DeIaware ,River


' ire-
..4-)Alo Grande
. ", .
,-, at

'' -
" , - i.e.!
***-.' -4 -
7 miles. The salt content averages. ..
about 35 parts per.thousand. Life extends
"-81.46 1. ti: :85i-4- !\.1-,,, 1r
Ca - , . '17.49 - -. 13.I2 41- 1:16
.to all depths'. 1

5 03 / .
:B The general nature of the water cycle on Cl. , h44.23 e .
2t. 65 1. 55.2
earth is well knOwn. Because the largest' SO4 17.49 30.10 7.7
. . -...
portion of the; surface area of the earth CO
32.99 ., 11.55 +HCO3 0.35
is covered-with water, roughly 70 percent .
of the earth's rainfall is on the seas. C 'or this presentation, the marine
(Figure 1) .environment will be (1) described using
an ecological, approach, (2) characterized
ecolOgically by comparing it with fresh-,
water and estuarine environments, and
(3) considered as a functional: eqological
"systerh (ecosystem). .




Distinct differences are found in phgsicare
.--.--7\chemtical, dr kci biotic factors in going from
a fr,enhwater to an oceanic environment.
Since roughly one third of the .
In general, environmental factors are more
. rain which falls on the land is again constant in freswater iriVers) and oceanic'
recycled through the atmospiiere.
1414,' - ',.-environments than in the highly variable
(see Figure 1 again), the total amount and harsh environments of estuarine an
of water washing.Over-Ithei.p.rth's surface, coastal waters. (Figure 2) ,
is significantly greaterethan one third c
the total world rainfall. It is thusnot physical and Ch7nical Factors
surprising to note that the rivers which
finally empty into the sell carry'a RiVers, estfaaries, and oceans are
disproportionate burden of dissolved and ..14dompared.in Figure 2 with referenee to
suspended`golids picked up from the --, the relative instability (or variation) of
The chemical.composition of this burden several important parameters: ',In the
depends on the 'cOmpoditibn of the rocks oceans, J will be noted, very little change
and soils through vi,./hIchihe river flMvs,' occurs in any,parametei.. In fivers, while '
the'Pr'oximity of an ocean; the direction "Salinity" (usually, referred to as udissalbed -
of prevailing winds, and otherfacars. -:salids") and teMperature,(accepting normal
This latho substance of geological eroSiOn. seasokelvariation,$) changtlittle'3,the other
(Table_ 1) 4f four pars: ters'vary c nsiderphly. In
estuattes, they.all chan e.
1 AK
-The Aquatic Environment -

Degree of instability .
Type of environment - .Vertical Avail-
ageneral direction Water Strati- -
Salinity Temperature elevation -ability Turbidity
arwater movement fication
, .

Riverine -
. . .

scene .


Oceanic ,


B Biotid Factors C Zones of the Sea
A complex of physical and ch(mical The nearshore environment is often
factors determine the biotic composi- classified -in relation to tide level and
tion of an environment. In general, water depth. The nearshore and offshore
the number of species in a rigorous, oceanic regions isagether, are often
highly variable environment tends to Ike classified with reference to light penetra-
less than the number in a More stable tion and water depth. (Figure 3)
environment (Hedgpeth, 1966).
1 Neritic - Relatively shallow-water
The dominant animal species (in zone which extends froin the high- \
terms.of total biomass) which occur 4* tide mark to the edge of the -7
in estuaries ke often transient, continental shelf.
.pending only N. part of their lives in
estuaries. This results in better
utilization of a rich environment.


The Aquatic Environment'


Swm 1 T/C
Suomi 0 C EA N I C
opf P opic

L it !pot
(Intertidal) '70
Sittilittorol $0.

wecop logic. 1),

EPNI0Pe °we
.54.1ltyptlave .r.z_r_r_rx://xxx e7-7
8ENTHIC (Bottom)
Littoral (IntHIJol) ACV Mocaor CA) Kee
SOldtatel 14a NV
tl scret civotle of P.*
Inns. hVIC.
6. Xt.
e ' 4/././C'2
Jhe, txe

FIGURE 3Classikition of marine environments

..sow .
a Stability of physical factors is .1) Physical factors fluctuate
intermediate between estuarine less than in the neritic zone.
00 and oceanic environments.
2) Producers are the phyto-
b Phytoplankters are the dominant plankton and consumers are
producers but in some locations the zooplankton and nekton.
attached algae are also important
As producers. b Bathyal zone - From the bottom
of the euptiotic zone to about
c The animal consumers are 2000 meters.
zooplankton, nektoil; and benthic
forms. 1) _Physical factors relatively
constant but light is absent.
2 Oceanic - The region of the ocean .
beyond the continental shelf. Divided 2.) Producers are abSent and
into three parts, all relatively consumers are scarce.
poorly populated compared to the
neritic zone. , c Abyssal zone - All the sea below
the bathyal zone,
a EuEihotic zone - Waters into which
sunlight penetrates (often to the 1) Physical factors'more con-
bottom, in the neritic zone). The stant than in bathyal zone:.
zone of primary productivity often
extend's to 600 feet below the surface. 2) Producers absent and consumers
ven less abundant than in the
bathyal 'zone .
1-25 .
The A uatic Environment

A Sea water is a remarkably suitable ORGANISMS
environment for living cells,, as it
contains 11 of the chemical elements A Salinity. Salinity is the single most
essential to thesgrowthand maintenance. . constant and controlling facto Atn the '
of plant and animalS. The ratio and ' marine environment, probabl bllowed
often the concentration of the major by temperature. It ranges around
salts of sea water are strikingly similar 35, 000 mg. per liter, or "35 parts per . .

in the cytoplasm and body fluids of thousand" (symbol: 35 %) in the language

marine organisms. This similarity is of the oceanographer. While variations
also evident; although modified somewhat in the open ocean= are relatively small,
in the body fluids of fresh water and salinity decreases rapidly as one
terrestrial animals. For example, approaches shore'and prpceeds through
sterile sea water may be usedin the estuary and up into fresh water with
emergencies as a substitute for blood a salinity of "0 Too (see -Figure 2)
plasma in man.
B Salinity and temperature as limiting
B Since marine organisms have an internal . factors in ecological distribution.
salt content similar to that of their `*,r)
surrounding medium (isotonic condition) 1 Organisms differ in the salinities
osmoregulation poses no problem. On the and temperatures in which they
other hand, fresh water organisms are prefer to live, and in the variabilities
hypertonic (osmotic pressure of body ofthese parameters which they can
ands is higher than that of the surround- t olerate. These preferences and
ing water). Hence, fresh water animals tolerances often change with successive
must constantly expend more energy to life history stages, and in turn often
keep water out (i. e., high osmotic dictate where the organisms live:.
pressure fluids contain more salts, the 1

their "distribution.' ,
action being then to dilute this concen-
tration with more water). 2 These requirements or preferences
often lead to extensive migrations
1 Generally, marine invertebrates are of various Species for breeding,
narrowly 2oildlosmotic, i.e., the salt feeding, mild growing stages. One
concentration of the body fluids chatnges very important-resnkof this is that
with that of the external medium. This an estuarine environment is an
has 'special significance in estuarine absolute necessity for over half of
situations where salt concentrations all coastal commercial and sport
of the water often vary considerably related species- of fishes and invertebrates,' e.

in short periods of time. for eitherall or certain portions of their

life histories. (Part V; figure 8)
2 Marine bony fish (teldosts4ave loWer
salt content internally than aite external 3' The Greek word roots "eury"
environment (hypotonic).' In order to (meaning wide) and "steno '-(ineaning
prevent dehydration, water is ingested narrow) are customarily combined,
and salts are excreted through special with such Words as "haline" for salt,
cells in the gills. and "thermal" for temperature, to
give us "euryhaline" as an adjective
to characterize an organism able to
tolerate a wide range of salinity, for
example; or "stenothermal" meaning
one which cahnot stand -much change
in temperature. "Meso-" is a prefix
indicating an intermediate capacity.

The Aquatic Environment

4 Some will knoWn and interesting

examples of migratory species which
C Marine, estuarine, and fresh water change their environmental preferemes
organisms. (See-Figure 4) with the-life history stage include the
shrimp (mentioned above), striped bass,,
many herrings and relatives, the
salmons, and many others. -None are
more dramatics than the salmon hordes
which hatch in headwater streams;
migrate far out to feed and grow,
then return to the mountain stream.
where they hatched 'to lay their own
eggs before dying.
5 Among euryhaline animals landlocked
(trapped), populations,living in lowered
Nlinitieskoften have a smaller maximum
0 Salinity ",; ca. 35 size than individuals of the same species
living in more saline waters. For
Figure 4. Salinity Tolefance of Organisms example, the lamprey (Petromyzon
marinus) attains a length of 30 - 56"
1 Offshore marine organisms are, in in the sea, while in the Great Lakes
general, both stenohaline and the length is 18 -.24".
stenothermal unless, as noted above,
they have certain life history require- Usually the larvae of aquatic organisms
ments for estuarine conditions. are more sensitive to changes in
salinity than are thesodults. This
Fresh water organisms are also characteristic both limits and dictates
stenohaline, and (except for seasonal the distribution and size of populations.
adaptation) meso- or steno ,ermal.
(Figure 2) D The effects of tides on organisms.
3 Indi us or native estuarine species 1 Tidal fluctuations probably subject
at normally spend their entire lives the benthic or intertidal populations
in the estuary are relatively few in to the most extreme and rapid variations
number. (See Figure 5). Tpey are of environmental stress encountered
generally meso- or euryhaline and in any aquatic habitat. Highly specialized
meso- or eurythermal, communities have developed in this
zone, some adapted to the rocky surf
zones of the open coast, others to the
muddy inlets of protected estuaries.
Tidal reaches of fresh water rivers,,
sandy beaches, coral reefs and
mangrove swamps in the tropics; all
have their own floras and faunas. All
must emerge and flourish when whatever
water there is rises and povers.or
tears at them, all must collapse or
10 15 20
25 30 35 retract to endure drying, blazing
tropical sun, or freezing arctic ice
Figure 5. DISTRIBUTION OF during the low tide interval. Such a
ORGANISMS IN AN ESTUARY community is depicted in Figure 6.
tEuryhalirie, freshwater
b Indigenous, estuarine, (mesohaline)
c Euryhaline, marine

1-27 ,
the Aquatic Environment

Littorina neritoides
L. rudis
I.. obtusata
O. L. littorea
0!) Chthamalus stellatus
0 Balanus balanoides
13. pe Hoist uS

Figure 6
Zonation of plants, snails, and barnacles on a rocky shore, While
this diagram is based on the situation on the southwest coast of
England, the general idea of zonation may be applied to any temper- -
att. rocky ocean shore, though the species will differ, The gray
zone consists largely of lichens. At the left is the zonation of rocks
with exposure too extreme to pport algae; at the right, on a less
exposed situation, the animals re mostly obscured by the algae.
Figures at the right hand margin refer to the percent of time that
the zone is exposed.to the air, i.e., the time that the tide is out.
'Three major zones can be recognized: the L4ttorina zone (above the
gray zone); the Balan.oid zone (between the gray zone and the
laminarias); and the Laininaria zone, a. Pelvetia cankliculata;
b. Fucus spiralis; c. Ascophyllum nodosum; d. Fucus serratus;
e.Laminaria digitata. (Based on Stephenson)


The Auatic Environment


ENVIRONMENT 1 Harvey, H. W. The Chemistry and
Fertility of Sea Water (2nd Ed. ).,
A The sea is in continuous circulation. ,With- Cambridge Univ. Press, New York.
'on circulation, nutrients' of the ocea.n would 234 pp. 1957.
eventually become a part of the bottom and
biological Production would cease. Generally, 2 Wickstead,, John . Marine Zooplankton
in all oceans there.exists a warm surface Studies in Bio gy,no. 62. The Institute
layer which overlies the dolder'water and of Biology. 78.
forms a two-layer system of persistent
stability. Nutrient concentration is-usually
greatest in the lower zone. Wherever a
mixing or disturbance of these two layers
occurs biological production is greatest.
B The estuaries are also a mixing zone of
enormous importance. Here the fertility
washed off the lanclis mingled with the
nutrient capacity of seawater, and many
of the would's most productive waters
C When man adds his cultural contributions
of sewage, fertilizer, silt or toxic waste,
it is no wonder that the dynamic. equilibriu
of the ages is rudely upset, and the
environmentalist. cries; "See what man
hath wrought "!


This outline contains selected material

from other outlines prepared by C. M.
Tarzwell, Charles L. Brown, Jr.$
C. G. Gunnerson, W. Lee Trent, W. 13.
Cooke, 13. H. Ketekum, J. McNulty,'
J. L. Taylor, R. M. Sinclair,. and others.

Part 5: Wetlands

I INTRODUCTION B Estuarine pollution studies are usually

. .., devoted to the dynamicsul.the'circulattng .

A Broadly defined, wetlands are areas water, its chemical, physical, and
which are "to wetto plough but too biological parameters, bottom, deposits, etc.
thick to flow." The soil tends to be
saturated with water, salt of fresh, C It is easy to overlook the intimate relation-
and numerous channels or ponds of ships which exist between the bordering
shaliow or open water are common. marshland, the moving waters, the tidal
Due to ecological features too numerous flats, subtidal deposition, and seston
,. lid -,variable to list here, they comprise whether of locale oceanic, or riverine
general ta rigorous (highly stressed) origin.
habitat, occupied by a small relatively
specialized indigenous (native) flora D The tidal marsh (some inland areas also
and fauna. ,i have salt marshes) is generally considered
tb be the marginal areas or estuaries and
B They are prodigiously productive coasts in the intertidal zone, 4hich are
however, and many constitute an dominated by emergent vegetation. They
absolutely essential habitat for some generally extend inland to the farthest
portion of the life history of animal point reached by the spring tides, where
forms generally recognized as residents they merge into freshwater swamps and
of other habitats (Figure ti). This is marshes 4-Figure 1). They may range in
particularly true of tidal marshes as width from nonexistent on rocky coasts to
mentioned below. many kilometers.
C Wetlands intoto comprise a remarkably
large proportion of the earth's surface,
and the total organic carbon bound in
. their mass constitutes an enormous
1 sink of energy.
D Since, our main concern here is with Channel LMud Flat Tidal Marsh Marsh

the "aquatic" eironment, primary tuin Mara

emphasis will be directed toward a
descr t ption of*vetlands as the transitional
zone' etween the waters and the land, and
how their desecration by human culture
- - :::
spreads degradation in both direction,. 47 pat 0
0 - Blue Clay

Zonation in e positive New England estuary.

1. Sprtng tide level. 2. Mean higlyide.

S. Mean low tide, 4. Dog hole. S. leo eleavap pool, 4. Chunk of Spartina turf deposited by lee.
T. Organic oose with associated community. O. pipes* (Zostei ra). 2; Ribbed mussels froodiolusl-
A "There is no other case in nature, save clam QM); mud snail Meal community. 10. lettur vai
in the coral reefs, where the adjustment
of or anie'relations to physicardttlition
a see in such a. beautiful way as the
balan e between the growing marshep
and the tidal streams by which thePare
at once nourished and worn awe*:
(Shale , ).886):
The Aquatic, Envir6timent

III MARSH ORIGINS AND 'STRUCTURES Such banks are likely to be cliff-like,
and are often undercut. Chunks of
A In general, marsh substrates are high in ,peat are oen found lying about on
organic content, relatively low in minerals harder sViattrate below high tide line.
and trace elements. The upper layers If face of cliff is well above high water,
bound together with living roots called overlying vegetation is likely to be
turf, underlaid by more compacted peat typically terrestrial Of the area.
type material. Marsh type vegetation is probably'.
1 Rising or eroding coastlines may
expose peat from ancient marsh 2 Low lying deltaic, or sinking coast=
growth to wave action which cuts lines, or those with low energy wave
into the soft peat rapidly (Figure 2). action are likely to have active marsh
formation in progress. Sand dunes
.. are also common in such areas
A1)(11,1( h (Figure 34. General coastal
Terrestrial turf configuration is a factor.

Salt marsh eat


7-:;: .

Figure 2. gran.--13ta matic se ion of.erodlng prat cliff

Figure 3

Development pi a Massachusetts Marsh since .1300 BC. involving an

18 foot rise in water level. Shaded area indicates sand dunes. Note
i meandering marsh tidal drainage. A: 1300 BC, Br 1950 AD.

The Acpiatic EnVironment

a Rugged or precipitous coasts or

slowly rising coasts, typically
eghibit narrow shelves; sea/cliffs,
fjords, massive beaches, and .
relatiyely less marsh area (Figure 4).
An Alaskan fjord subject to recent
I catastrophic subsidoce and rapid -
deposition of glaciflour Shows
eVidencelif the recent encToachment
of saline waters in the presence of -
`recently buried trees and other
terrestrial vegetation, 'exposure .
of layers of salt marsh peat 4ong
the edges of channels, .and.a poorly
compacted younglmarsh turf developing
at the. new high water level (Figure 5).

Figure 4 A River Mouth on a Slowly Rising Coast. Note abseime

of deltaic development' and relatively marshland
although mud flats stippled are extensive,

Shifting flats Tidal marsh I 'Terre stria

0 Et',

8 .4
: it


. .


. ..

Figure 5 Some genial relationships in 4 northern fjord witla rising water level. 1. ,mean low
'water,' 2, maximum high tide, 3, Bedrock, 4. Glacial flour to depths in excess or-
b() meters, 5. Shifting flats and channels, 6. Channel against bedrock, 7. Buried
terrestrial vegetation, 8. Outcroppings of salt marsh peat.-
F (V
b Lowslying coastal plains tend to be cs
Deep tidal' channels fan but through
fringed by barrier islands, broad c. °
irmumerablebranching and often
estuaries and deltas, and broad Infer-connecting rivulets: The
assqciated marshlands (Figure 3). izitervening. grassy plains are _
essentially, at mean high tide level.

1 -33 C.
The Aquatic Environment

c Tropical and subtropical regions tidal mar sh is the mar.sh grass, but ver34
such as Florida, the Gulf Coast, little of it is use& flreman as grass.
and Central America, are _frequented (11ab1, 1)
"'by mangrove swamps. This unique _
type of growth is able to establish The nutritionaLanalysis of several.
itself in shallow water and move out marsh grasses as compared to airy land"
into progressively' deeper areas hay is shown in Table 2.
' (Figure 6). The strong deeply
embedded roots enable the-mangrove
to resist considerable wave action
at times, and the tangle of roots
TABLE 1,, General Order's of Magnitude &Gross Primary Productivity in Terms
quickly accumulates a deep layer of of ,Dry Weight of Organic Matter Pitad Annually
organic sediment...Mangroves gms/M /year
in the south may-be considered to Ec9system (grarns/awkre meters/year) lbs/acre/year
be roughly the equivalen,t of the Land desekts, deep sceane v Tens Hundreds

Spartina marsh grassin the north Grass4ds, forests, cutrophic Hundreds Ttiousends
lakes, ordthary agriculture
as a land builder. Whenfully Estuaries, deltas, coral reefs, Thousands Tenthousands
developed, a mangrove swamp is an intensive agriculture (sugar
impenetrable thicket of roots over cane% rice)

the tidal flat affording shelter to an

assortment of semi-aquatic organisms
such as various molluscs arid
crustaceans,' and providing access
from the nearby land to predaceous
bir'dsa.reptiles, add mammals. ,
Mprigrov.es are not restricted to #
estuaries, buernay develop out into . TABLE 2. Analyses of Some Tidal Marsh Grasses
shallow oceanic lagoons, orupstream
into relatively fresh waters.-
T/A li'ercentage Composition
Dry Wt; Protein r Eat . Fiber Water Ash Nfree Extract
WACO. Tit=taruS,4
Divichln spicata (Ore stand, dry)
:resit 2.8 ' 5.3 1.7. 32.4 8.1 6.7 4S.5
Short Spartina alternillora and Sabccenia curopaea In standing water)
IN0001 6.6
1.2 7.7 2.5 31.1 - 12.0 37.7
Is= Spartina alterndlora (tall, pure stand in standing water)
7.6 ' 2.0 . 29.0 8.3 15.5 37.3
an 1.40.4111 Sparrinavalftns sport stand, dry) .:. .. .
3.2 , 6,0 ,.. 2.1 30.0 8.1 9.0 44.5
Sparrina a/remit/4ra and Sp.iffifla patens (mixed stand, wet)
3.4 - 6.6 1.9.4 .29.5 8.1 10.4 42.8
Spariniakernillura (short, wet)
Z2 13.0 . 2.4 30.4 81 13.3 36.3
warauravasor Comparable Analyses for Hal 0
El 0
1st rut 6.7 4.2 44.9
Figure 6 Diagrammatic transect of a mangrove swamp 2,,,roil
° 36.2
10.4 5.9 30.5
showing transition from marine to terrestrial a
habitat. ° , AnalrieeperfOrMed by Roland W.. Gilbert, Deptrtment
b. of Agricultural Chemistry, U. R. I.
4O .


A Measuring the productivity of grasslands Ow

not_easy,tbecause.today,grass is seldpm
used directly 4isisuch by man. Itis thus .
- usually 'expiessed-as production of meat,
milk, or bythe case of salt marshes, 'the
total crop'of animals that obtain food per :. .
unit of area. '4he.prfirnary producer in a:.
. .
The Aquatic Environment


B The actual utjlizaiion'of marsh grass is

accomplighed primarily by its decom- 11;"1
position-and ingestion by micro organ&ns. .
(Figure 7) A small quantity of seeds and
solids is consumed directly by hirds. VrTZtle



.eli4106A ek

Figure 8 Diagram of the life cycle- .

of white shrimp (after Anderson.and
.Lunz .1965).

3. An effort to make an indirect

, estimate,ofprogiuctivity in a ithOde
Figure 7 The nutritive composition of ^ Island marsh was made on a single
successive stages of decomposition of - dad by recording the numbers
Spartina marsh grass, showing increase
in protein and decrease pt carbohydrate 1= and kinds oebirdirthat fed on a
with increasing age and decreasing size, relatively -small area (Figure
of detritus particles. Betwepn 700 and 1000 Wild oirdsof
12 species, ranging from 100 least
1 The quantity of micro invertebrates sandpipers to uncountable numbers
which thrive on this wealth of decaying of seagulls were counted. One food
marsh has not been 'estimated, nor has requirement estimate for three-
the actual production'of small indigenous pound poultry in the confined inactivity
fishes and invertebrates such as the of a poultry yard is approximately one ft
top:minn.ows (Fundulus), or the mud ounce per pound of bird per day.
-snaili (Nesse), and others.
2 Many forms of oceanic life migrate
into the estuaries, especially the '
marsh areas, for important portiohs ; Ps.

of their life histories as is mentioned a

elsewhere (Figure 8)." It has been 0 .

estimated that in exgess of. 60% of the

marine commercia1:4nd sport fisherIe4e
are estuarine or marsh. dependent in
some way.
Greater Yellow legs (left) -
and black duck .
Great blue heron
Figure 9 SomeCommon Marsh Birds

0, V. 1-35-
',- -40;:
The Aquatic Environment

One-hundred black bellied plovers and geographic distribution, etc.

at approximately ten ounces each Included would be the familiar cattails,
would weigh on the order of sixty spike rushes, cotton grasses, sedges,
pounds, At4the same rate of food trefoils, alders, and many, many'.
consumption, this would indicate Others.
nearly fotfr pbunds of food required
for this species Lone. The much C Types of inland wetlands.
greater activity of the wild birds
would obviously greatly increase their 1 As noted above (Cf: Figure 1)
food requirements, as would their tidal marshes often merge into
relatiSely smaller size. freshwater marshes and bayous.
- Deltaic tidal swamps and marshes
-Considering the range of foods con- are often saline in the seaward
sumed, the sizes of the birds, and the portion, and fresh in the landward
fact that at certain seasons, thousands . . areas.
of migrating ducks and others pause
to feed here, the enormous productivity, 2 River bottom wetlands differ from
of such a rbarsh can be better under- -those formed from lakes, since wide
stood. flood plains subject to periodic
inundation are the final stages of
- the erosion of river valleys, whereas
lakes in general tend to be eliminated
V INLAND BOAS AND MARSHES byhe geologic proceSses of natural
eutrophication often involving
A Much of what has been said of tidal Sphagnum and peat formation.
marshes also applies to inland wetlands. Riverbottom marshes in the southern
As was mentioned earlier, not all inland United States, with favorable climates,
swamps are salt-free, any more than all have luxurient growths such as the
marshes affected by tidal rythms are canebrake of the lower Mississippi,
saline. ° or a characteristic timber growth
- .. such as cypress.
13 The specificity of specialtz'ed floras to
particula types'Of wetlands is perhaps 3 Aithoug\bird lift is the
more spectacular in freshwater wetlands conspicuoUs animal elener in the
than in the marine, where Juncus, fauna (Cf:- izigure 9), many mammals,
Spartina, and Mangroves tend to dominate. such as muskrats, beavers, otters,
t 1 Sphagnum, or peat mass,--is'
probably one of the most widespe d o.
Figure 12)
.2 ,,
and others areal marsh-oriented.

and abundant wetland plants on ea

Deevey Q.958) quotes an estimate tai
there is pt:obably upwards of 223
billions (dry weight) of tons of peat
in ti( world today, derived during
recent geologic time, from Sphagnum
bogs. Particularly in the northern
regions, eat moss t nds to overgrow
to ponds An shallow de fissions, eventually
forming the vast turd lains-and-
moores of the nort
2 Long lists of other bog and marsh plants
Might be cited; each with its own
special requirements, topographical,


The AcLuatiC Environment

VI POLLUTION 2 Marsh grasRes.may alsO be eliminated

- by smothering as, for example, by
A No "Single statement can summarize the deposition of dredge spoils, or the
of pollution on Marshlands'as spill or discharge of sewage sludge.
distinct from effects ,noted elsewhere on
other habitats. 3 Considerable marsh.area has been 4
eliminated by industrial construction
B .Reduction of Primary Productivity activity, such as wharf and dock con-
struction, oil well construction and
The primary producers in most wetlands operation, and the discharge of toxic
are the grasses and peat mosses. brines and other chemicals.
Production may be reduced or eliminated
by: C Consumer production (animal life) has
been dr a st Joan:), 're'auised by th'..dr. liberated'
1' Changes iri the- water level brought distribution of-pesticides. .lesome cases,
about by flooding or drainage. this has been aimed at nearby agricultural,. ,
lands for economic crop pest control, in
a Marshland areas are s5metimes other cases the marshes have been sprayed
diked and flooded t9 produce fre or dusted directly to control noxious
water ponds. This may bq for' insects.
4 aesthetic reasons, to ,suppress the
ingrowth of noxious marsh inhabitating The results have been universally
sects -such as mosquitoes or biting disadtrous for the marshes, and the
midges, to constructan industrial bene \ to the human community often
waste }folding pond; a thermal or a questio able.
sewagg stabilization pond, a .`
"convenient" result of highway .2 Pesticides designed to kill nuisance'
'causeway construction, or other insects, are atso toxic to other
reason., The result is the elim- arthropods sb that in addition to the'
ination of an area of marsh. A a target species, such forage staples as
sitallcompensating border of the various scuds (amphipods), fiddler
marsh may or may nor develop. crabs, and other macroinvertebrate
have either been drastically reduced
b r-High i'dal marshes were often or entirely eliminated in many places
ditche and drained in former days For example, one familiar with fiddle
to stab ze the sod for salt hay or crabs can traverse miles of marsh
"thatch' barvesting which was highly margins, still riddled with their burrows,)
sought a sr in colonial days. This without seeing a single live crab.
,inevitably changed the character
of the marsh, but it remained as a, DDT and related compouna have been'
essentially marshland. COnversion "eaten up the food chain" (biological
to outright agricultural land has Magnification effect) until fidh eating
been less widespread because of the and other predatory birds such as herons
necessity of diking to exclude the and egrets (Figure 9), have been virtually
periodic floods or tidal incursions,.. eliminated from vast areas, and the /
and carefully timed drainage to -accumulation of DDT in .nian himself
eliminate excess precipitation. is only too well known.
Mechanical tidal marshes
has_not been econo 'cal in this
country, althoUgh the succesb of
the Dutch andOthers in this regard
is well known.

The Aquatic Environmen

- .
D Most serious of marsh enemies is E Swimming birds uch as duckd, loons,
man himself. In his quest for "lebensraum" cormorants, pelicans, and many othervs
near the water, he has all but killed the are severely jeopardizedby floating
water he strives fo approach.. Thus up to - pollutants such as oil..
twenty percent of the marshestuarine
area in various pails of the country has
already beentutterly destroyed by cut and
. fill real estate development's (Figures,
`10, 11).

Yt t
4 rigure 10. Diagrammatic representation of cut-and-Sill for
real estatexclevelopment mlw = m ean 'low. water

. .
Figure 11. Tracings of portion of map of a southern
1, : .. city. showing, extent of cut-and-fill real
j.,, -f estate development. t .
) . 0 z
The Aquatic Environment


VII SUMMARY 5 Morgan, .J. P. Ephemeral Estuaries of

the Deltaic Environnient in :'Estuaries;
A "Wetlands comprise the marshes, swamps, pp. 115-120. Publ. No. 83, Am.
bogs, and tundra areas of the world. Assoc. =Adv. Sol, Washington, DC. 1967.
They are, essential to the well-being of
our surface waters and ground waters. 6 Odum, E.P. and Dela Crug, A.A.
They are essential to aquatic life of Particulate Organic Detritus in'a
all types living in the open waters. They Georgia Salt Marsh - Estuarine
are essential as habitat
for all forms of, Ecosystem. 'in: Estuaries, pp. 383-
wildlife. 388, Publ. No. 83, Am. Assoc. Adv.
Sci. Washington, DC. 1957.
B The tidal marsh is the area of emergent 0
= vegetation bordea.igthe,..ocean er ash 7 Redfield, A. C. The .Ontogeny of a Salt.
, estuary. Marsh Estuary. in: Estuaries, pp.
108-114. Pulil. No. 83, Arn, Assoc.
C Marshes are highly productive areas, Adv. Sci. Washington, QC. 1967.
essential to the maintenance of a well
rounded community of aquatic life. 8 Stuckey, 0. H. Measuring the Productivity
of Salt Marshes: Maritimes (Grad
D Wetlands may be destroyed by: School of Ocean. , U. R. I. ) Vol. 14(1):
9-11, February 1970.
1 Degradation of the life forms of
Aft, which it is composed in the name of Williams, -11.B. _Compartmental
nuisance control. Analysis of Production and Decay
of Juncus reomerianus. Prog.
2 Physical destruction by cut-and-fill Report, Radiobiol. Lab., Beaufort, NC,
to create more land area. Fiscal Year 1968, USDI, BCF, pp, 10-
12. ,

0 This outline was prepared by H. W. Jackson,

former Chief Biologist, National Training.
REFERENCES Center, and revised by R. M._ Sinclair, Aquatic
Anderson, VP. W. The Shrimp anti the Biologist, National Training Center, MOTD,I
Shrimp Fishery of the Southern " OWPO, EPA, Cincinnati, OH 45268.
United States. TJSDI, FWS, BCF.
Fiihery Leaflet 5,89. 1966. °
2 ,,DeeVey,- E. S. ,. Jr. ,BogS. Sci.' Am. Vol.
199(4):115-122. October 1958. Descriptors: Aquatic Environment, Biological
Estuarine Environment, Lento Enrironment,
3 "Emery, K. 0. and Stevenson. Estuaries Lotic Environment, Currents, Marshes,
and Lagoons. Part II, Biological Limnology, Magnification, Water Properties
0 Aspects by J.W. Hedgepeth, pp: -693-
"M. in:. Treatise on Marine Ecology
and Pleoecology. Geol. Soc. Am.
Mem. 6 ?. Washington, DC.' 1957.

4 Hesse, R.;; W. C. Allee, and K. P.

Schmidt. Ecological Animal
6,4 GeOgraphyw John Wiley; & Sons. 1937.

( 1-39

I A COMMUNITY is an ashmblige of 1 Collectors strain, filter, or otherwise

populations of plants, animals, bacteria, collect fine particulate organic matter
and fungi that live in an environmental and from the passing current.
interact with one another, forming together
a destinctive living system with its own 2 Shredders feed on leaves, detritust
composition, structure, environmental and coarse particulate organic matter.
relations, development, and function.
, 3 Grazers feed on attached growths.
II An ECOSYSTEM is a community and its
environment treated together as a functional 4 Predators feed on other organisms.'
system of complementary relationships,
and transfer and circulation of energy and IV Taxonomic Groupings
matter. (A delightful litte essajr.on the
odyssey of atoms X and Y through an A TAXOCENES, a specific group of organisms.
.ecosystem is in Leopold's, A Sand County Ex. midges. For obvious reasons most
Almanac. ) systematists (..xonomists) can specialize-
in only one group of organisms. This fact
III TROPHIC levels are a convenient means is difficult for the non-biologist to grasp:
of classify ing organisms according to
nutrition, or food and feeding. (See B Size, which is often dictated by the inves-
-Figure 1. )' tigator's `sampling equipment and specific
A PRODUCER, the photosynthetic plant or
first organism on the food chain sequence. V Arbitrary due to organism habitat prefer-
Fossil fuels were produced photosynthe- ences, available 'sampling devices,
personal preference of the investigator,
and mesh sizes of nets and sieves.
B Herbivore or primary CONSUMER, the
first animal which feeds on plant food.. A PLANKTON, organisms suspended in a
body of water and at the mercy of currents.
C First carnivore or secondary CONSUMER, This group has been subject to numerous
an animal feeding on a plant-eating animal. divisional schemes. Plants are PHYTO-
D. Second carnivore or tertiary CONSUMER Those retained by nets are obviously, MET
feeding on the preceding. PLANKTON. Those passing thru even the-
finest meshed nets are NANNOPLANKTON:
E Tertiary carnivore.
B PERIPHYTON, the community of micro-
F Quaternary carnivore. organisms which grow on submerged
objects (substrates). -Literal meaning
DECOMPOSERS OR REDUCERS, bacteria "to grow around plants", however
which break down the above. organisms. standard glass microslides are sub-
Often called the middlemen or stokers of mersed in the aquatic habitat to
the furnace of photosynthesis., standardize results.
H Saprovores-ox! DETRITIVORES which feed C BENTHat. is often used to mean
on bacteria and/or fungi. MACROINVERTEBRATES; although there
....-- are tenthic organisms in other Plant,
I Macroinvertebrates have been subdiVided ariiinal, and protist grolips. Benthic
into trophic levels according to feeding refers strictly to the bottom substrates of
habits ,(See Figure 1 from Cummints); lakes, - streams, and other water bodies.
42 2-1
BI. ECO, 25a. 3.80
Classification of Communities, Ecosystems and Trophic Levels

0.5' METERS)

,. 2 (1-2 METERS)

Az4!re P/R°<I


401 00.
W 4 (10 METERS)




4. P/R < 1



- 12 METERS)

Classification of Communities, Ecosystems andTrophic Levels


retained on a No. 30 mesh screen (approx- TIES. Like the preceding community,
imately 0.5 mm) and thus visible to the; theses are manipulated by man. Placing
naked eye. artificial or natural substrates in a body
of water will cause these communities
E MACROPHYTES,. the larger aquatic plants to appear thereon.
which are divided into emersed, floating,
and submersed communities. Usually K We will again emphasize ARBITRARY,
vascular-plants but may inclUde the larger because organisms confound our neat
algae and "primitive" plants. These have little schemes to classify them. Many
posed tremendous economic problems in move from one community to another
the large man-made lakes, especially for variousreasims. However, all
in tropical areas. these basic scheme do have intrinsic
value, provided they are used with
F NEKTON, in freshwater, essentially fish, - reason:
salamanders, and the larger crustacea.
In contrast to PLANKTON, 4\ese organisms
are not at the mercy of the-current. REFERENCES

G NEUSTON, or PLEUSTON, -are inhabitants 1 Cummins, Kenneth W. The Ecology

ofthe surfaCe film (meniscus organisms), - of Running Waters, Theory and
either supported by It hanging from, or " Practice in Proc. Sandusky River
breaking through. it. Other organisms Symposium. International Joint
are trapped by this neat little barrier of Commission. 1975.
nature. The micro members of this are
easily sampled by placing a clean cover. 2 Leopold, Aldo. A Sand County Almanac.
slip on top of the surface film then either Oxford University Press. 1966.
leaving it a specified time or examining:
it immediately,under_the microscope. ' 3 Peters, Robert Henry. The Unpredictable
Problems of Tropho-dynamics.
H DRIFT, macroinvertebrates which drift Env. Biol. Fish 2:97-101. 1977.
with the streams current either periodically
(diel or 24 hour), behaviorally, catastro-
phically or incidentally.
This outline was prepared by R. M. Sinclair,
BIOLOCIAL FLOCS, are suspended Aquatic Biologist, National Training and
'microorganisms that are formed by , . Operational Technology Center, MOTD,
various means. In wastewater treatment OWPO, USEPA, Cincinnati, Ohio 45268.
plants they are encouraged in concrete aer
aeration basins 'using diffused air or Descriptors: Biological Communities
oxygen (the,heart of the, activated sludge . ,

,2 -3


I INTRODUCTION br outside of a lake.

A Most Interference _Organisms are a That which comes in from

Small. outside (allochthonous) is
predominately inert solids
B Small Organisms generally have (ttripton):
Short Life Histories.
b That of internal origin (auto-
.0 POpulations of Organisms with chthonous) tends totbe bio-
Short Life Histories may Fluctuate 1ogieal in nature.
Rapidly in Response to Key Environ- 4'
, mental Changes. B Heat and Temperature Phenomena
are Important in Aquatic Ecology.
D Small Organisms are Relatively
at the Mercy of the Elements 1
. The total 4uantityof heat avail-
able to a body of water per year
E The Following Discussion will can be calculated and is known
Analyze the Nature of These'Ele- as the heat budget.
. ments with Reference to the Res-
. ponse of Important Organisms; 2. Heat is derived directly from in-
solation; alsO by transfer from
air, internal friction, and other
Density Phenomena
A Light is a Fundamlental SoUrce.of
Energy for Life and Heat," '1 Density And viscosity affect the
floatation and locomotion of
1 Insolation-is affected by geo- microorganisms.
graphical location and mete-
orological factors. a Pure fresh water achieves
its maximum density. at 4 °C
2 Light penetration in water is and its maximum viscosity
affected by'angle of .incidence at 0 °C.
(geographicalY, turbidity, and
color. 'he. propOrtion of light b The "rate of change of density
reflected depends on the angle ) increases with the temperature.
of incidence, the temperature,
color, and other qualities of 2 Density stratification affect
the water. In general, as the aquatic life and water uses.
depth increases arithmetically,
the light tends to decrease geo-: a In summer, a mass of warm
metrically, Blues greens, and surface water, the epilimnion,
yellows tend to penetrate most is usually present and separated
deeply while ultraviolet, vio- from a cool deeper mass, the
lets, and orange-reds are most hypolimnion, bYa relatively
quickly absorbed. On the order thin layer known as the
of 90% of the total illumination thermocline.
which penetrates the surface '41

film es absorbed in the first b Ice cover and annualspring

10 meters of even the clearest and fall overturns are due to
witdr. successive seasonal changes
in the relative densities of
3 TuXidity may originate within the epilimnion and the hypo-

45.. 3-1
Limno loley and EcologyAokPlankton

limnion, profoundly influ- D Shore development, depth, inflow -

enced by prevailing meteoro7 outflow pattern, and topographic
logical conditions. features affect the behavior of the water.
c The sudden exchange of E Water movements that may affect organ-
water masses having differ- isms include such phenomena as waves,
ent chemical characteris- currents, tides, seicheS, floods, and
tics may have catastrophic others.
effects on bertain biota, may
cause others to bloom.
d Silt laden waters may seek 1 Waves or rhythmic movement
certain levels, depending
on their own specific gravity The best luiolAn are traveling
in relation to existing layers waves. These are effective
already present. only against objects.near
the surface.. They have little
Saline waters will also 4. effect on the movement of
stratify according to the large masses of water.
relatiye densities of the ts
Various layers. b Standing waves or seiches
ocdur, in all lakes but are
3 Theoriseosity of water is greater seldom-large enough to be
at lower temperatures. observed. An "internal seich"
is an oscillation in a density
a This is important not Only mass within a lake with no
in situations involving the surface manifestation may
control of flowing water as in cause considerable water
a.sand filter, but also since moVement.
overcoming resistance to
. flow generates eat, it is
significant in the heating
- of water by internal friction
from waive and current ac-
tion and many delay the
establishment of anchor
ice under critical conditions.

b It is easier for plankton

to remain suspended in cold
viscous (and also dense)
water than in less Viscous
warm water. This is re-
flected in differences in the
appearance of winter vs
summer forms of life (also
arctic vs tropical).


Limnology and Ecology of Plankton

2 Currents There are typically two tidal

cycles per lunar day (approx-
a Currents are arhythmic imately 25 hours), but there is
water movements which have continuous gradation from this
had major study only in ocean- to only one cycle per (lunar) day
ography. They primarily in some places,
are concerned with the trans-
location of water masses. Estuarine plankton populations
They may be generated inter- are extremely influenced by local
nally by virtue of density . tidal patterns.
changes, or externally by
wing or runoff.. d Flood waters ranmfrom torren-
. tial velocities which tear away
Turbulence phenomena or and transport vast masses of
eddy currents are largely re- substrate to quiet backwaters
sponsible for lateral mixing which may inundate normally dry
in a current. These are of and areas for extended periods
far more importance in the o time. In the former case, ..._

economy of a body of water, pl ktonic life is flushed away

than mere laminar flow. co pletely; in the latter, a local
pla on bloom may develop which
4c Tides, or'rathetidal may of immediate significance,
currents, are reversible or which may serve as an inoculum
(or oscillatory) on a relative- for receding waters.
ly long and iiredictable period.
They are closely allied to F Surface Tension and the Surface Film
seiches. For all practical
purposes, they are restricted 1 The surface film is the habitat of
to oceanic (especially-coastal) the "neuston", a group of special "
waters. -significance.
If there is no freshwater 2* Surface tension lowered by surfactants"
inflow involved, tidal currents may eliminate the neuston.' This can
are basically "in and out;" be, a significant biological observation.
if a significant amount of
freshwater is added to the III DISSOLVED SUBSTANCES
system at a constant rate, the
outflowing current will in general A Carbon dioxide is re eased by plants and
exceed the inflow by the amount animals in respir on but taken in by
of freshwater inpvt. plants in photosynthesis.
Oxygen is the biblogical complement of
carbon dioxide, 'a.nd'necessary for all
animal life.
C ,Nitrogen and phosphorus are fundamental
nutrients for plant life.
Occurin great dilution, Concentrated
by plants.

47' 3-3
Limnology and Ecology of PlanktOn 1111.01

The distribution of nitrogen V BIOTIC COMMUNITIES (OR ECOSYSTEMS)
compounds is generally corr ,
ed with the oxygen curve, esp A A biotic community will be defined.here
cially in oceans. j as an assemblage of organisms living in
a given ecological niche (as defined
D Iron, manganese, sulphur, and silicon below). Producer (plant - like); consumer
are other minerals important to aquatic (animal-like) and reducer (bacteria and
life which exhibit biological stratification. fungi) organisms are usually .included.
. A source of energy (nutrient, food) must
E Many other minerals are present but their also be present. The essential concept
biological distribution in waters is less in that each so-called 'community is a
well known? fluorine, .tin, and vanadium relatively independent entity. ArCtually
shave recently been added to the "essential" this position is only tenable at any given
list, and more may well follow. ,
instant, as individuals are constantly
shifting from one community to another in
F Dissolved organic matter is present in response to stages in their life cycles,
even the purest of lakes. physical conditions, etc. The only one
tosbe considered in detail here is the
A Isttritional Classification of Organisms B Plankton are the macroscopic and
microscopic animals, *ants, bacteria,
1 Holophyt ic or independent or- etc. floating free in the open water.
ganisms, like green plants, pro- Many clog filters,. cause tastes, odors,
duce their own basic food elements and other troubles in water supplies.
from the physical environment. ,
1 Those that pass !through a plankton
2 Holozoic or dependent organisms, net (No. 25 silk bolting cloth or
like animals, ingest and digest equivalent) or sand filter are often
solid food particles of organic known as nannopankton (they
usually greatly exceed the "net"
plankton in actual quantity).
3. Saprophytic or carrion eating
organisms, like many fungi and 2 Those less than four microns in
bacteria, digest and assimilate length are sometimes called
the dead bodies of other organ-- ultraplankton.
isms or their products.
There are many ways in which
B The Prey-Predator Relationship is plankton may be classified: taxo-
Simply one Organism ating Another. nomic,- ecological, industrial:
Toxic and Horm c Relationships 4 TheMitconcentration of plankton varies
markedly in space and time.
1 Some organisms such as certain
blue green algae apd some ar- Depth, light,. currents, and
mored flagellages produce s 4 water quality profoundly affect
stances poisonous to others. plankton distribution.
2 Antibiotic action in nature is The relative abundance of
' not well understood but has been plankton in the; various sea-
shown,to play a very influential sons is generally:
role in the economy of nature. 0
1 sprjng, 2 fall, 3 summer,
4 winter
. 1.

Limnology and EColbgy of Plankton


5 Marine plankton include many .deeper, more turbid, and usually

_larger animal forms than are warmer water, sand, mud, silt,
found in freshwaters. or clay,bottorn -materials which
shift with increase in flow.
C Th benthic community is generally
cgixsidered to be the macroscopic life 4 In old age, streams have appi-oa-
living in or on the bottom. ched base level. During flood
es, stage they scour their bed and de-
D The peon community might be posit materials on the flood plain
defined as the microscopic benthos, which may be very broad an/ flat.
except alai they are by no Means confined During normal flow the etteneris.._
to the bottom. Any surface, floating, or I.
'refilled and many shifting bars are
not, is usually covered by 'film of living developed.
organisms. ,There is frequent exchange
betWeen the periphytdn and plankton (Under the influence of man this
communities. patternmaygle broken up, or tem-
porarily interrupted. Thus as essen-
E The nekton is the comMunityof larger, tially "youthful" stream might take
free-swimming animals (fishes, shrimps, On some of the characteristics of a
etc. ), and so is dependent on the other .'.'mature" stream following soil -
communities for basic plant foods. erosion, organic enrichment, and
increased surface runoff. Correction
F "Neuston or Pleuston of these conditions might likewise be
followed by at least a partial rever-
This community inhabits the air/water sion to the "original" condition.)
interface, and may be suspended above
or below it or break it. 'Naturally this C Lakes have a developmental history
interface is a very critical One, it being which somewhat parallels that of streams.
micro molecular and allowing interchange
between atmospheric contaminants and 1 The method of formation greatly
the water medium./ Rich in bacteria, influences the character and sub-
metals, protozoi, pesticides etc. sequent history of lakes.;

TIIE EVOLUTION OF WATERS _- 2 Maturing or natural eutrophication

VI of lakes
A, The history of a body of water determines e,
its present condition. Naturalswaters haye a If not already present, shoal
evolvedin the course of geologic time areas are developed through
to what we know today.. ro.?iop. of the shore by wave
. , -action and undertow.
B In the. course of their evolution, streams b Currents produce bars across'
in general pass through four general bays and thus cut off irregulars
stages of development which may be called:
birth, ypvth, Maturity, and old age. areas.
. ,
Estallishment of birth. In an ° c Silt brought in by tributtry
.extant' stream, .this might be streams settles eut in the quiet
a "dry run's or headwater lake water.
streambed, before it had eroded
down to the level of ground Water. d Rooted aquatics grow on shoals
and bars, and in doing so cut
Youthfultreams; when the off bays and contribute to the
stream bed is eroded below the filling of thelake.
1 grouhd water level, spring water . ,
e Dissolved carbonates and other
esters and the stream becomes materials 'are precipitated in the
,,.permanent. deeper portions of the lake in
Mature streams; have wide part through the action of plants.
valleys, a developed flood plain,

Limno logy and of


f When filling is well' ad.anced 1 .z Youthful streams, especially on

. sphagnum mats extend Nt- rock or sand substrates are low
ward from the shore. Ttibse- in essential nutrients. Tempera-
mats are followed by sedges tures in mountainous regions are
and grasses chfinalky usually low, and due to the steep
convert thelak Into a gradient; time for growth is short.
marsh. Although ample light is available,
growth of true plankton is thus
3 Extinction of lakes. After lakesA' greatly limited.
reach maturity their progress
toward -hp is accelerated. 2 As the stream flows toward a
They become extinct through: more 4'mature" condition nutrients
14 .4.4,1%tend tp accumulate, and gradient
'a The downcuitink Of the out diminishes 91 so time of flow ,
let. increases, temperature tends to
increase, and plankton flourish.
b° Filling with detritus eroded, '
from the shores or brought _Should a heavy load of inert silt
in by tributary streams. develop on the other hand, the
Q turbidity would reduce the light
c Fillingby the acca tion of ' penetration and, consequently the
.s.the remains orvegetable- general plankton production would
geowinen the
'e itself. 's
As the stream approaches base
t(Oftentiro or 'three pro- ." level (old age)' and the time avail-
cesses may act concurrently) able for plankton growth increases, .
o . L. the balance between turbidity,
When mth hastens tie abode : .," nutrient levels, and temperature
process, leis Often called and other_seasonal conditions,
"cultural eutrophication. " detdmines-the overall peoduc-
CI tivit37.- '`
Q. C.' F-actors Affecting the Productivity
A Thebiological resultant of all physical of ,Lakes i
sand ctlemical factors is the quantity of .

life that may actually be present. The '''10" The size, shapeand depth
ability to produce this "biomass" is ,of the lake basin: Shallow water
often referred to as the "produCtivity" is more productive than deeper
1' of a body of water. This-is neither good water .since more light will reach
nor bad per se. Aater of low producti- stile bottom to stimulate rooted
vity is a "poor" water. biologically, and plant growth. As a corollary,
also a relatively "pure" or "clean" water; lakeslvith more shoreline, having
hence desirable as a water supply. A more shallow water, are in general
productive water on the other nand may more productive. Broad shallow
be a 'nuisance to man or highly desirable. Vekes and reservoirs have the
Some of the factors which influence the sreatesttproduction potential (and
productivity of-waters are as ..follows: hence should_ be- avoided for water
B Factors affecting stream productivity.
-A*To be productive of plankton, a, stream 2 Hard waters are generally more
must provide adequate nutrients, light, prodhdtive than soft waters as
a suitable temperature, and time for there are more plant nutrient
growth to take place. mtperaleavailable. This is often


Geographic Location


Human Geological Latitude

Influence Pormatign Longitude

Sewage Competition She of Basin Climate
Agriculture of Substrate


Primary Drainage Dept"... Area Bottbm Precipitation Insolation

Nutritiv - Area Conforination
I I a II h 04W.

Nature of Inflow of Trans- --31....Light edt Penetration 4,2 Penetra Develop o Seasonal Cycle
Bottom A llochthon oils pa. r en cy Penetration and Stratification-01.- and Littoral Circulit. Stagnation
Deposits Materials ptilizatiop Region Growing Season:.

Trophic Nature of a Lake


'Limnology and Ecology of Plankton

greatly influenced by the 3 Reservoir discharges also pro-

character' of the soil and rocks formdly affect the DO, temperature,
iiithe watershed, and the quality and turbidity lr the stream below 1
and quantity of ground water a dam. Too much°fluctuation in
entering thd lake. lir general, flow may permit sections of the
pliTanges of 6.8 to 8.2 appear stream to dry periodically.
to be most productive.
3 Turbidity'reduces produCtivity RESERVOIRS
as light penetration is reduced.
A The productivity. of lakes and impound-
4 The presence or absence of ments is such a conspicuous feature ,
thermal stratification with its that it is often used as a means of
semi-;annual turnovers affect classification.
Productivity by distributing
nutrients throughout the water 1 Oligotroxhic lakes are the
mass. a
geologically younger, less produc- °

tive lakes, which are deep, have

5 Climate, temperature, pre - clear water, and usually support 6
valance of ice and snow., are Salmonoid fishes.
also important.,
2 Mesotroac lakes are generally
Di Factors AffeCting the Productivity of intermediate between oligotrophic
Reservoirs andeutrophic lakes. They are
moderately productive, yet
1 The productivity of reservoirs pleasant.to be around.
is governed by much the same
principle's as that of lakes, with 3 Eutrophic lakes gre more mature;
the difference that the water more turbid, and richer. They
level is much more under the are -usually shallower. They are
control of man. Fluctuations richer in dissolved solids; N; p,4;
r in water levelican be used to ap d Ca are abundant. Plankton is-
deliberately increase of decrease abundant and there is often a-
productivity. This can be dem- * ricli bottom'fauna. Nuisance
onstrated by a comparison of conditions often appear.
the TVA reservoirs which practice
a summer drawdown with some of 4 Dystrophic lakes - bog lake?:
those in the west where a winter .11
low in pH, wate yellow to brown,
drawdown is the rule. I dissolved solids; N, P, and Ca
scanty but humic materials abun,
2 The level at whiir water is re- . dant; ,bottom fauna and plankton
moved from the /reservoir is also poor, and fish species are limited.
important( The upper epilimnion dr.
may have a high plankton turbi- B Reservoirs may be classified as storage,
dity while lower down the plankton or run of the rider.
count may be less, but a taste
and odor causer {such as M_all_o- 1 Storage reservoirs have a large
I be present. There 4 volume in relation to .their inflow.
maybetwo thermoclines, with - 4'Run of thel river reservoirs have
a mass of muddy water flr3wing 2
between-a clear upper epilimnion a large flow throUgh in relation,
and a clear hypolimnion. Other to their storage value. .
combinations ad infinitum may A .,,


0 N.11A

Irmamman........ Limnology and Ecology of Plankton


'According to location, lakes and to maintain minimum limiting nu-

reservoirs may be classified as polar, tritional factors.
temperate, or tropical. Differences
inclimatic and geographic conditions 2 Shading out the energy ofinsola-
result in aiffvences in their biology. tion by roofing or inert turbidity;
r suppresses photosynthesis.
ENVIRONMENTAL FACTORS 3 Introduction of substances toxic
to some fundamental patt of the
A Liebig's Law of the Minimum states food chain (such as copper sul-
that productivity is limited by the phate) tends to temporarily inhibit
nutrient present iri the least amoung productivity..
at any given time relative to the
.assimilative capacity of-the organism. X .SUIVIMARY

B ShelfOrd's Law of T oler.ation: A A body of water such as a lake rep-

resents an Intricately balanced system
Minimum Unlit --Wange of Optimum' Maximum limit of in a state of dyhainic equilibrium. `
toleration Modification imposed at one point in
of toleration of factor
Absent Greatest abundance -t-ia- the system automatically results in
Abundance ' compensatory adjustments at associated
B The more thorough our knowledge of
C The artificial introductio, of nutrients the entire systeM, the better we can
(sewage pollution or Pert zer) thus judge where to imp`ofe control measures
tendsto eliminate existing iting to abhieve a desired result.
minimums for some specie and create
intolerable maximums for o her species. REFERENCES .

1 Known limiting min umsimay 1 Chamberlin, Thomas C., and SalisburgiO4.

S. sometimes be delib rately Rollin P., Geology Vol. 1, "Geolo
maintained. gical Processes:and Their Results",
pp irzlix, 1.nd 1-654, Henry Holt and_
2 As the total available en gy Company, , 904.
supply is increased, ductivity
; tends to increase. meat. ..r.aperties of
Ordinary Water - SUbstario .
3 As productivity, increases,, the Reinhold Publ. Corp., 17ew York.
whole character o; the water pp: 1-673. 1940.
may be changed from a Meagerly
. productive clear water lake 13 Hutchesoni-George E. A Treatise on
(oligotrophic).to a highly pro- LirnnOlogy. John Wiley Company.
ductive and Usually turbid lake
.11 4 Wuttner, Franz. Fundamentals of
. 4 Eutrophecation leads to tesatment Limnology. University of Tprontq
troutRes. "Press. pp. 1-242. 1953.

D . Control of eutrophication may be accom- 5 Tarzwell, Clarence M. ExPerimezital

.plisbed by various means Evidence on the Value of Trout 1937
Stmettm Improvement in Michigan.
- Watershed management, ade- . American Fisheries Society Trans,
quate preparation of .reservoir. 661177-187. 1936.
sites, and Pollution control tend.'-
. .1 .:317..
Limnology and EcOlogy of Plankton

,.6 US DHEW. PHS. Algae and Metropolitan

Wastes, transactions of a seminar
held April 2I-29, 1960 at the
Robert A,;' Taft Sanitary Engineer-
ing Center, Cincinnati, Ohio.
No. SEC TR IN61-3.
7 Ward and Whipple. Freshwater Biology
(Introduction). JAn Wiley
Company. 1918.,

8 Whittaker, 'R. Ir Communities and

Ecosystems. Macmillan
NeW York. 162 pp.' 1970.
9 Zhadin, V. I. and perd, Sr. Fauna
and Flora of the lakes and
- Reservoirs of the USSR., Avail-
able from the Office of Technical
Ser4ices, U. S. Dept. Commerce,
Washington, DC.
'41 o

10 Josephs, lifelvin and Sanders, Howard J.

Chemistry and the Environment
ACS Publications, WashAgton, DC.
11 Sournia, A. (editor), Phytoplankton
Manual. UNESCO. 1978. .

This outlike"was prepared by IL W. Jackson,
formai Chief Biologist,- National Training
Center, and z;evisect by 'R. M. Sinclair,
Aquatic .Biologist,' NationarTrairiing and
Operational Technology Center, ()WYO.' '
USEPA, Cincinnati, Ohio 45268.

Descriptors: Plankton, Ecology,. Limnology



I CLASSIFICATION E In this presentation, a minimum of clas-

sification and taxonomy is used, but it
A The planktonic cornmanity is composed of should be, realized that each group is ,
organisms that are relatively independent typified by adaptations of structure on
of the bottom to complete their life history. physiology that are related to the plank-
They inhabit the open water of lakes tonic mode of existence. These adapta-
(pelagic zone). Some Species have inactive tions are reflected in the classification.
or resting stages that lie ()Ole bottom
and carry the species through periods of
stress; e. g. , winter, A few burrow iri II FRESHWATER ZOOPLANKTON
the mud and enter the pelagic zone at night,
but most live in the open water all thb A The, freshwater zooplankton is dominated
time that the species is present in an active by representatives of three groups of
form. animals, two of them crustaceans:
Copepoda, Cladocera, Rotifera. All have
B Compared to the bottom fauna and flora, feeding mechanisins that permit a high
the plankton, consists of relatively few degree of selectivity of food, and two can
kinds of organisms that are consistently produce resting eggs that can withstand
and abundantly present. Two major 'cat- severe environmental conditions. In
egories are often called phytoplankton general the food of usual zooplankton pop-
(plants) and zooplanktonAanimals), but \ ulations ranges from bacteria and, small
this is based on an outmoded classification algae to small animals.
of living things. The modern tendency is
to identify groupings according to their B The C.opepoda reproduce by a normal
function in the ecosystem: Primary pro-. biparental process, and the females lay
ducers (photosynthetic organisms), consum fertilized eggs in groups which are carried
(zooplankton), and decomposers (hetero- around in sacs until they hatch. The
trophiC.bacteria and fungi). immature animals go through an elaborate
C, The primary difference ken is nutritional; development with many stages. The later
phytoplankton use inorganic nutrient 'stages have mouthparts that permit them
elements and solar radiation. Zooplankton to collect particles. In many cases, these
feed on particles, much of which can be are in the form of combs which remove
phytoplankton cells, but'can be bacteria or smallparticlps by a sort of filtration
particles of dead organisms (detritus) process. In others, they are modified to
originating in the plankton, the shore form grasping organs by which small
region; or the land surrounding the lake. animals or large algae are captured
'D The swimming powers of .planktonic C The Cladocera (represented by Daphnia)
organisms is so limited that their hort- reproduce much of the time by partheno-
zontal distributioniis determined mostly iesis, so that only females are present.
by movements of water. Some of the Eggs are held by the mother in a brood
viek animals are able to swim fast enough that
they can migrate vertically tens of meters chamber until the young are developed far
each day, but they are capable of little enough prrfepd for themselves. The newborn
horizontal navigation. At most some animals look like and do
species of crustaceans show a general not go through an elaborate series of
avoidance of%the shore areas during,calm ,developmental stages in the water as do
weather when the water is movinernore the copepods, oaphni.a, has
slowly than the animals can swim.47By filtering structures on some
some of its legs ,
definition, animals that are able to control that act as filters.
their horizontal location are nekton, not


BI..Aci.29. 6. 46 ,56
Biology of Zooplankton Communities

D Under some environmental conditions-the' 2 Inorganic materials

development of eggs is affected and males
are produced. Fertilized eggs are produced Freshwater lakes vary in the content
that can resist freezing and drying, and of dissolved solids according to the
these carry the population thrbugh geological situation. Tile total salinity
unsatisfactory conditions. and proportion of different dissolved
materials in water cartIffect the pop-
E The Rotifera are small animals.with'a ulation. Some species are limited to
ciliated area on the head which creates soft water, others to saline iaterl, as
currents used both for locomotion, and for the brine shrimp.. The maxim.= pop-
bringing food 'particles to the mouth. They ulation size developed may be related
too reproduce by parthenogenesis during to salinity, but thisris probably'an
much of the year, but production of males .indirect effect working through `the
results in fertilized, resistant resting eggs. abundance of nutrients and production
Most rotifers lay eggs one at a time and of food.
carry them until they hatch.
3 Food supply
III ZOOPLANKTON POPULATION DYNAMICS, Very strong corrylations have been
found between reproduction and food
A In general, zooplankton populations are at supply as measured by abundance of
a minimum in the cold seasons, although phytoplankton. The rate of food supply
some species flourish in cold water. Species can affect almost.,all aspects of pop -
with similar food requirements seern'to ulation .biology including rate of indi-
reproduce at different times of.the year or vidual growth-time of maturity, rate
are segregated in-di#e-nent layers of lakes. , of reproduction and length of life.
B There is no single, simple measurement 4 Apparently in freshwater, dissolved
of activity for the zooplankton as a whole organic materials are of little nutri-
that can be used as an.index of production tional significance, although some
as can the uptake of radioactive carbon for species can be kept -if the concentration.
the phytoplankton. However, it is possible of dissolved material is high enough.
to find the rate of reproduction of species Some species require definite vitamins
'that carry their eggs. The basis of the in the food.
method is that the number of-eggs in a
sample taken a #a given time represents 5 Effect of predation on populations
the number 9f animals that will be added
to the population during an interval that The kind, quantity and relative pro-
is equal to the length of time it takes the portions of species strongly affected
eggs to develop. Thus the potential growth' by grazing by vertebrate and inverte-
rate of.the populations can be determined. brate predators. The death rate of
The-actual growth rate, determined by Daphnia is correlated with the abun-
successive samplings and counting, is less dance of a predator. Planktivorous
than the potential, and the difference is a fish (alewives) selectively feed on
measure of the death rate.. larger species, so a lake with alewives
is dominated by the smaller species of
C Such measurements of birth and death rates crustaceads and large ones are scarce
permits a more penetrating analysis to be or absent.
made of the causes of population change
than if data were available for population 6 Other aspects of zooplankton
size alone.
Many species migrate vertically con-
D Following is an indication of the major siderable distances each day. Typically,
environmental factors in the control of migrating species spend the daylight
zooplankton. hours deep in the lake and rite 'toward
t-o the surface in late afternoon and early
1 Temperature has an obvious effect in 'evening.
its general control of rates. ,In addition,
the production and hatching of resting Some species go through a seasonal
eggs may be affected. change of form (cyclomorphosis) which
is not fully understood. It may have an
effect 4n reducing predation.

Biolo&Srof Zooplankton Communikies


1 Baker, A. L. An Inexpensive Micro- 7 Lund, J. W. G. 1965. The Ecology of

sampler. Limnol. and Oceanogr. the Freshwater Plankton. Biological
15(5) :. .158-160. 1970. Reviews, 40:231-293.

2 Brooks, J. L. and DOdson, S. I. 8 UNESCO. Zoolplankton Sampling.

Predation, ,Body, Size, and Corn- UNESCO Monogr. Oceanogr. Methodol.
pqSition of Plankton. Science 150: 2. 17.4 pp. 1968. (UNESCO. -Place
28 -35. 1965. de Fortenoy, 75, Paris 7e France).
3 Dodson, Stanley I, Complementary
Feeding Niches Sustained by Size.-
Selective Pl-edation, Limnology
and Ocenography 15(1): 131-137.
4 Hutchinson, G. E. 1967. A Treatise
on Limnology. Vol. II. IitrodUction
to Lake Biology and the Limnoplankton.
xi + 1115. John Wiley & Sons, Inc.
New York.
5. Jossi, Jack W. Annotated Bibliography
of Zooplankton Sampling peyices.
USFWS. Spec, Sci. : Rep. -Fisheries.
609. 90 pp. 1970.
6 Likens, Gene E. and Gilbert, John J. 'This outline was prepared by W. T. Edmondson,
Notes on Quantitativ,e Sampling of Professor of Zoology, University of
Natural Populations of Planktonic Washington, Seattle, Washington.
Rotifers. Limnol. and Oceanogr.
15(5): 816-820. .
Descriptor: Zooplankton

53 4-3

Biology of Zoopl nkton Communities 1111


Keratella hiemali

Kellicottia longispi

Polyarthra vulgaris

Daphnia longispina
Ceriodaphnia quadrangula

Bosmina coregoni

a a a a a a a

J P M M -J J A S 0 N D

Each panel shows the abundan4 of a species of animal.- Each

mark on the vertical axis represents 10 individuals/liter.
Nadwerck, A. 1963.11 Die Beziehungen zwischen Zooplankton and
Phytoplankton im See Erken. Symbolae Botanicae Upsaltensis, 1,7:1-163.


.. .

7 .
* .

. 7 .
Young per :Total young
0.20, 9
Brood 223,
. Temperature o
., more than 10
0 132

. .
, Temperature
co less thanI00
Number of young producedin each brood by Daphnia living in
ae four different concentrations of food organisms, rewewed
: .
daily. The total numbetaproduced during the life of a '
mother is shown by the numbers at the right. The Daphnia
at the two lowest concentrations produced their first batch
. of eggs on the same day as the others, but the eggs degen-
. .
erated, and the first viable eggs were released two days
later. Richman, S. 1958. The transformation of energy by
'0 Daphnia pulex. Ecol. Monogr. 28: 273-291.
0 200 400 600

Abundance of food organisms ugm /1, dry weight

4Mean rate.of'laying eggs by the planktonic

rotifer Keratella cochlearis in natural
populations ap a function of abundance of
food'organisms and temperature. W. T.
Edmonton. 1965. ReprOducEive rate of
planktonic rotifers as related to food
and temperature in nature.
Ecol. Mmogr.

Biology of Zooplankton Commuriities







fi `Stentor Epistyks

Biology of Zooplankton Communities


Polygarthra Brachionus

. .


Cladocera Copepoda

Nauplius larva of copepod
I 44.



Ins'ecta Chaoborus


. . .
..,. . .
.- a

*I.*, .":1
Biology of Zooplankton Communities


1,1 0

380p. 864.

2 3°
spinet (fin nttached).. sim le (gill attached)

Glochidia '(Unionidae) Fish Parasites-



Veliger Larva6 (Corbiculidae) Free LiArAng Planktonic

a . ..(4-5)

Pedivpliger' attaches byssus lines)



4 ?*i^



I OPTICS Strictly, speaking, of course, all reflected

light, even diffuse, obey/4 Snell's Law.
An understanding of elementary optics is Diffuse reflected light is made up 8l many
essential to theproper use of the microscope. specularly .reflected rays, each from a
The microscopist will find th4t unusual pro- a tiny element of surface, and appears
blems in illumination and photomicrography diffuse when the reflecting elements are
can be handled much more effectively bnce 'very numerous and very small. The terms
the,underlying ideas in physical opti7 are diffuse and .specular, referring to reflection,
understood. describe not so much a difference in the
nature of the reflection but rather a differ-
A Reflection ence in the type of surface. A polished sur-
face gives specular reflection; a rough
A food place to begin is-with reflection at surface gives diffuse reflection.
a 'surface or interface. Specular (on
regular) reflection results when d beam It is also important to note and remember
of light leaves a surface at-the same angle that specularly reflected light tends to'be
at which itreached it. This type of .strongly polarized in the plane of the reflect-
reflection occurs with highly, polished ing surface. This is due to the fact that
smooth surfaces. It Is stated more pre- those rays whose vibration directions lie
cisely as Snell's Law, is e. ,'the angle of closest to the plane of the reflection surface
incidence, i, ,is equal to the angle .of are most strongly reflected. This effect is
reflection, r (Figure1). Diffuse (or strongest when the angle of incidence is
-scattered) reflection results when abeam such that the tangent of the angle is equal
of light strikes a rough or irregular sur- to the refractive index of the reflecting sur-
face and different portions of the incident face. This particular angle of incidence is
light are reflected from the surface at called the Brewster angle.
different 'angles. The light reflected from
a piece of white paper or a ground glass is B Image Formation on Reflection.
an'example of diffuse reflection. ,

Considering reflection by mirrors, we find

(Figure 2) that a plane mirror forms a
virtual image behind the mirror, revered
right to left but of the same size as the
object: The word virtual means that the
,image appears to be in a Oren plane but
that a ground glass screen or a photographic
film placed in that plane would stiOw no
image.' The converse of a virtuariniage is
a real image. '

Spherical 'mirrors are either convex or con-

cave with the surface of the mirror repre-
senting a portion of the surface of a sphere.
The center of Curvature is the...center of the
Sphere, part orwhose surface forms The
. Figure 1 mirror: The Tocus,lies halfway bb'tween the
SPECULAR REFLECTION SINTELLIS ' -center of curvature 1144:14he milk9F surface.
,., - .7,
,BI. MIC. 18..2. 79
Opttr..5 and the Microscope


3 A ray of light which passes through the

focus is reflected parallel to the axis
of the mirror. .
- The image from an object can tie located
using the familiar lens formula:
it I 1
°bloat Virtual
Imago where p = distanCe from the object to - .
the mirror
q = distance from the image to
t the mirror
*Figure 2 f = focal length

Construction of an image by a concave No spheyical surface can be paeftct in its

mirror follows from. the ,two premises image-forming ability. The most serious
given below (Figure 3): of the imperfections,' spherical aberration,
occurs in spherical mirrors of large
aperture (Figure 4). The rays of light
,making up an image point from the outer
zone of a sphericd.1 mirror do not pass
through the same point as the more central
rays. This type of aberration is reduced by
blocking the outer zone rays from the image
area or by-using aspheric'surfaces.



c_ ( Figure -3

1 A ray of light parallel to the axis of

the mirror must pass through thd'
foCus after: reflection.
Figure 4
2 A ray of light which passes through the SPHERICAL ABERRATION BY

center of. curvature mast return along

the sarrie pat11.-
11. N
A corollary of the firstpremiseis:

`' .3

Optics and the Micrpscope

D Refiaction Of Light into focus and the new micrometer reading

is taken. Finally, the microscope is re-
Turning now.to lenses rather than mirrors focused until the surface of. the liquid appears
we find that the inost`ininortant t liaicte- in sharp focus. The micrometer reading.,
istic is refraction. Reratilni releys to is taken again and, with this information, .
the change of dirlction and/or velocity Of the 'refractive index may be calculated from
light as it passed from one 111..diutil to the simplified equation:
another. The ratio of the veloc it) in air
, (or more corrcu.tly in a vacuum) to, the actual depth
active index
velocity ip-th«.-; 111 rdit1111 i, called the apparent depth C

Lte frac ti've index. Some.. typical values of

refractive index measured with mono-
, chromatic' light (sodium I) line) are listed 'Table IREFRACTIVE INDICES OE COMMON

.Refraction causes an object inimer,s(el in

a medium of higher refractive index than V a't uu m 1.0,000000 Crown glass. 1.48 to 1.61
air to appear Olosert.to the surface thah it Air 1.00.029 i 8 Rock salt)oirs:.1.5443
actually is (Figure 5). This effect CO2, L0004498 Diamond 2.417
Wate 'I. 3330 Lead sulfide 1E3.912

When the situation' is reversed, add a ray

of light from a medium of high refractive
index p'a4es through the interfaCe of a
medium of lower index, the ray is refracted
until a critical angle.is reached beyond which
Air all pf the ,light is reflected frOm the interface
(Figure 6). This critical angle, C, has the
folldwing relationship to the refracti've irdices
Apparent Medhim of the two Media:
Actual-. depth
depth , 'TOO
sin C= ni
, where\n2 <n1.
Object-.0. 4

re When the second medium is air, the formula


Figuse 5
. 'six; C =
11 4 s. F 1.

be used to dete'rimine th,e,gpfractive index

or a.liquid:with the nlivipscope. A flat
'. ;,t . vial`with a. scHtcl).eiti Otte bottom cipside)
cr ,... is Placed,ern the' stage of the mic"rbscope.e.
The' microscope iS fo'cubed on ale stratch
and the fine adjustment micrometer reading
. 1. .. is noted. A small amount of the unknotArn.
liquid is added:
. ,
the scratch is again brought .
. ,

-... . a, A I

67 t .3
t ,

Optics and the Microscope


E Dispersion F Lenses
Dispersiodis another important property .There are two cPasses of lenses, con-
of transparent materials. This is the verg ing and diverging, called also convex
variation of refraetive index with color, and concave., respectively. The focal
(or wavelength)of light. When white'light point of a converging lens is defined as
passes` thro6gh a glass prism, the light the pqint at which a bundle of light rays
rays are-reffraCted'hy different amounts ...parallel to the axis of the lens appearA to
and separated into the colors of the .-,- 'converge aftter,passing through the lens.
spectrum.' This spreading of light into The focal lerigth of the lens is the distance
itg component, colors is due to disperses from the lens to the focal point (Figure 7):,
which,. in turn' is due to the fact that the
.refractive index of transparent substances,
. liquids and solids, is lower for long wave-
lengths than for short wavelength.
BeCause of dispersion, determination of
the refractive index of a substance re-
quires designation of the particula, ye-
length'used. Light from a sodium lamp
has a strong, closely spaced doublet with
an average wavelength of 5893A, called
the D line, which is commonly used as a
reference wavelength. Table 2 illustrates
the change, of refractive index with wave -'
length for a few common substrCes.
f. '
Figure 7

Fefractive index
F line D line C line.
blue (yellow) '(red)
4861A 5893A 6563A
G Image FoAnation by Refraction
Carbondisulfide 1.6521 1,6276 1.6182 Image formation by lenses (Figure 8)
Crown glass 1.5240 '1%5172 -.1.5145 .follows rules analogous to those already
given above for mirrors:
Flint glass 1.:6391 1.6210 \..1.t221/
Water 1. 3372 1.'3330 1. 3312 1 Light traveling parallel to the axis of
the lens Will he refracted so as to pass
through the foeus of the lens.
mr - .

The dispersion Qt a material can be defined t Light traveling through the geometrical
quanittativel§ as: center of the lens will be unrefracted.
n (yellow) - 1
v dispersion z-b1
n (blue)-- n (red)
he,pos ition of the, image c e determined'
. by remembering that a light r y passing
n (593mii) - 1' through itcus,',F, will be parallel to
n (486mv) - n(656mv) the axis of the lens.ori the opposite "sine' of
the lens and that a 'ray passing through the
where, n is the refractive index, of The geometrical. center of the lenswill be
material at the par+icular wavelength unrefracted. w
noted( in the parentheses.
Optics and.the Microscope


object points are riot located on the optical

axis of the lens and results in the formation-
of an indistinct image. The simplest
remedy for astigmatism is to place ?tie '
objey.t close to the dxis of the lens system.
I IntCrft.rence Phenomena
Interference and diffraction are two phe=-
nomena which are due *to thq wave character-
isties of light. The superposition of two .
light rays arriving simultaneously At a given
point will give rise td interference effects,
whereby the intensity at that point will vary
Figure 8 from. dark to bright depending on the, phase
different es between the two light rays..
The first requirement for interference is
The magnification,. M, of an image° of an that the light must come from a single
object,prothiced by a lens is given by the source. The light may be split into any
relationship: number of-paths but must originate from
the same poipt (or .coherent source). Two
image size image distance q light waves from a coherent source arriv-
.M = object digtance p
object size ing at .a.'point in phase agreement will
reinforce each other (Figure 9a). Two
where q = distance.; rom image to lens light waves,from a coherent source arriv-
and p =. distance from object to lens. ing at a point in opposite phase will cancel
each other (Figure 9b).
H Aberrations of Lendm
Lenses have aberrations of several types
which, unless corrected, cause loss of
detail in the image. Spherical aberration
appears in lenses with-spherical surfaces.
Reduction bf spheriCal aberration cane
accomplished by.diaphragming the outer
zones of the lens ,or by designing special
asplferical surfaces in the lens system.
Chromatic aberration is -a lifienomkwrion
caused by the vaiiation of refractiVe. index
with wavelength (dispersion). Thus' a lens
receiving white light from an object will
farm a violet image closer to the lens' and
a red one father away. AchroznA'CiC'
lenses arb,employed to minimize this . Figure 9a. Twolight rays, I and .2, of
effect...The lenses ar4"cornliiratizins of the same:frequency'but dif-
two or more lens elements made tip.of, ferent amplitudes, are in phaie .1

materials having different dispersive in the upper diagram. in the

titers: The use of monochromatic light- lower diagram, rays l 'and 2
.44.t interfere constructively to give
fs. obvious way of eliminating
chromatic aberration.. , , . a' single wave of the same fre-
quency aricP.with an, amplitude
Aistigmatism-is a third aberration of equal to the summation of the
spherical lens. syStems. It occurs' when two former waves.,

'Optics and the Microscope


Figure 9b. Rays 1 and 2 are no.v 180° Figur,e 19

-"out of phase and interfere
destructively. The resultant, INTERFERENCE IN A THIN FILM
in the bottom diagram, is of
the same frequency but is of A simple interferometer can be made,by
reduced amplitude (a is partially silvering a microscope slide and
negative and is subtracted cover slip. A preparation between the two
from b). partially silvered surfaces will show inter-
ference fringes, when viewed with mono-
chromatic light, either transmitted or by
The reflection of a monochromatic light vertical illuminator. The fringes will be
beam by a thin film results in two beanis, close together with a wedge-shaped prep-
one reflected from the top surface and one aration and will reflect-refractive index
from the bottom surface. The distance. differences due to Temperature variations,
traveled by the latter beam in excess of concentration differences, different solid
the first is twice the ,thickness of the film phases, etc. The method has been used to
. and equivalent air path is: measure quantitatively the concsrgration of
solute around a growing crystalui(Figure 11).
2 nt
where n is.the refractive index and
t is'the thickness-of the film.
50% Morrom/..
The second beam, however, upon reflection
at the bottom surface, undergoes a half
wavelength shift and now the total retaril- --Cover s-p -
tapon of the second beam with respect to Specimen
°the-first is given as:
retardation = 2 nt +4
0 b,

Where k is the wavelength of the light

e 4
Figure 11
When .retardation is_ exactly an *odd -Timber a Examination is by transmitted light.
of half wavelengths, destructive interfer- Light ray undergoes multiple .
ence takes place resulting in darkaess. ' reflections and produces dark and
When it is zero di an even number-of half light frifiges in the-field. A speci-
wavelengths,, constructive interference men introduces a phase 'shift and
results in brightness (Figure 10). Changes the fringe pattern.
b Illumination is froni the top. The
principle is the same but fringes
show greater contrast.
Optics and the Microscope

2 44 f X
Each dark band represents an equivalent d
air thickness of an odd number of half
wavelengths. Conversely, each bright where f is the focal length of thelens,
band is the result of an` even number' of
X the wavelength, and D the., diamete
half wavelengths. ' of the lens.
Witlyinterference illumination, the effect
of a transparent object of different re- It is seen that in order to maintain
fractive index than the medium in the small diffraction disc at a given wave-
microscope fittia is: '4 length, the diameter of the lens should
be as large as possible with respect to
a change of light intensity of the object the focal length. It should be noted,

if the background is uniformly illumi- also, that a shorter wavelength produces

nated (parallel cover slip), or a smaller disc.
If two pin points of light are to be distin-
2 a shift of the ipterference bands within guished in an image, their diffraction discs
the object if the background consists must not overlap more than one 'half their
of bands ( tilted cover slip). diameters. The ability to distingvish such
image points is called resolving power and
The relationship of refractive indices of is expressed, as one half of thepreceding
the surrounding medium and the object is expression:
as folloWs:
resolving power = 1.22 f
ex D
nm( 360t

where ns = refractive index of the II THE.°C.OMPOUND MICI:PSCOk'E

nITI = refractive index of the The compound m Ccoscope is an extension in
surrounding_medium 'principle ofhe imple magnifying glass;
= phase shift of thostwo hence it is e ntial-toutidefstand "fully the
beams, degrees properties of this simple lens system.
X = wavelength of the light
t = thickness of the specimen. A Image Formation by the Simple Magnifier
J Diffraction' The apparent size of an object is determined'
In geometrical optics; it is assumed that, by the angle that is, formed at the eye by the
light travels in straight lines. This is not extreme rays of the object. By bringing the
always true. We note that a beam.passing 4 object closer to the eye, that angle (called
through a slit toward a screen creates a the visual angle) is increased. This_also
bright band wider than the slit with alter- increases the apparent size. HoWever a
nate bright and dark bands appearing on limit of accommodation of the eye is reached,
either side of the central bright Band, at which distance the eye can no longer focus.
decreasing in intensity as a function of :a .
This limiting distance is about 10-inches or 25
the distance from the center. Diffraction centimeters. It is at this distance that the
describes this phenomenon and, as one of magnification of an object observed, by the
its-practical'consequences, limits the unaided eye is said:to be unity. Theeye can,
lens in its ability to reproduce an image. of course, be focused at shorter distances but
For example, the image of a pin point of not usually in a relaxed condition,
light produced by a-lens is not a pin point
but ,is revealed to be a somewhat larger A positive, or converging, lens can be'used
patch of light surrounded by dark and to'permit placing an object closer than 10
bright rings. The diameter, d, of this inches to the eye (Figure 12). By this means
diffraction disc ( to the first dark ring) the visual angle of the object is increased
is given as: (as is its apparent size) while the image of
5 -7°

Optics and the Microscope


1. 111060

r Eyepiece
4 IMO.. ObjeCI

Ilast414, EV,
4 t %

Figure 12
the object appears to be 10 inches from
the eye, where it is best accommodated. 4
ob,.... 111

B Magnification by a Single Lens System

The magnification, M, of a simple magni- Object t

fyinass is given Py:
25 ,
M= + 1

v 'tug Image
where f = focal length of the lens in
centimeters. Figure 13

Theoretically the magnification can be IMAGE FORMA ION IN

increage-d-with-shorterlocal-lengtirienses7 COMPOUND MIC OSCOPE
However such lenses require placing the
eye very close to the lens surface and lengths to give different agnifications
have much 'mage distortion and other (Table 3). The magn cation is calculated
optical aber ations. The practical limit from the focal 1 by dividing the latter
for a simple magnifying glass is about into the tube length, usually 160 mm.
The numerical aperture (N. A.) is a measuz4
In order to go to magnifications higher of the ability of an objective to resolve detail''
than 20X, the compound microscope is This is more fully discussed in the next
required. Two lens systems are used section. The working distance is in the free
to form an enlarged image of an object space between the objective and the cover
..,.(Figure 13). This is accamplished in slip and varies slightly for objectives of the
two steps, the first by a lehs called the same focal length depending upgn the degree
objective and the second by a lens known of correction and the manufacturer.
as the eyepiece (or ocplar).
There are three basic classifitations of
C The Objective objectives: achromats, fluorites and
apochromats, listed in the order of their
The objective is the lens or lens system) complexity. The achromats are good fOr
closestito the object. Its function' is -to. routine work' while the fluorites and apo-
reproduce an enlarged image of tht object chromats offer additional optical corrections
in the body tube of the Microscope. to compensate for spherical, chromatic and'
.Objectives are available in various focal other aberrations.

5 -8
Optics and the Microscope,


Nominal ' Working Depth Diam. of, Resolving Maximum Eyepiece
focal length Nominal N. A. distance focus field power, white useful for niax,
magnif. mm mm. light, p. magnif., useful magnif.
mm ii

56 2. 5X 0.08 .40 50 8.5' 4..4 80X 30X

32 4 1 4... 'O. 10 ,25* 16 5 3.9 el )X 20X
16 \ 10 0.25 7 8- 2. 1.4 250X 25X
8 20 0.50 1.3 2 1 0.7 500X 25X

\ 43 .
45, /
0.2 ,


Another tem of objectives employs at one time (there are some nosepieces
refytti surfaces in the shape of concave that accept five and even six objectives).
and vexx mirrors. Re Reflection optics, In this system, the objectives are
. ..1.--eSuse they have no refracting elements, usually nccncenterable and the stage is
do not suffer from chromatic aberrations centerable. Several manufacturers pro-
as ordinary refraction objectives do. Based vide centerable objective mounts so.that
. entirely on reflection, reflecting objectives each objective on the nosepiece need be
are extremely useful in the infrared and centered only'once to the fixed rotating
'ultraviolet regions of the spectrums They stage. The insides of objectives are
also have a much longer working distance better protected from dust by the rotating
than the refracting objectives. nosepiece. This,yas well as the incon-
venience of the so-called "quick-change"
The body tube of the microscope supports objective holder, makes it worthwhile
the objective at the bogom (over the object) to have one's microscope fitted with
and the eyepiece at the top. The tube rotating nosepiece.
length i's maintained at 160 mm except for
Leitz instruments, which have a 170-mm .D The Ocular
tube length.
The eyepiece, or ocular, is necessary in
The objective support may be of two kinds, the second step of the magnification process.
an objective clutch changer or a rotating The eyepiece functions as a simple magni-
nosepiece: fier viewing the image formed by the
1 The objective clutch changer ("quick-
change" holder) permits the mbunting There are three classes of eyepieces in
of only one objective at a time on the common use: huyghenian,,compensating
microscope. It has a tentering arrange- and flat-field. The huyghenian (or huyghens)
nient, so that each objective need be eyepiece is designed4to, be used with
centered only once with respect to the achromats while the compensating type is
stage rotation. _.The changing of objec- used with fluorite and apochromatic
tives with this system is somewhat objectives. Flat-field eyepieces, as the
awkward compared with the rotating name implies, are employed in photo-
nosepiece., -micrography or projection and can be used
with most objectives, It is best to follow
2' The revolving nosepiece allows mounting the recoramendations ofthe manufacturer
three or four objectiV,es on the microscope as to the proper combination of objective
and eyepiece. - .


Optics and the Microscope

The usual magnification available in eyepiece gives insufficient magnification

oculars run from abouttBiC up to 25 or for the eye to see detail actually resolved
30X. The 6X is generally too lo;,v to be of by the objective. ,
any real value while the ?5 and 30X oculars
have sligh y poorer imagery than medium F Focusing the Microscope
powers an have avery low eyepoint. The
most useful eyepieces lie in the 10 to 20X The coarse adjustment is*used to roughly
magnification range. positron the body tube (in some newer
microscopes, the stage) to bring the image
E Magnification of the Microscope into focus. The fine adjustment is used
after the coarse adjustment to bring the
The total magnification of the objective- image into perfect focus and to maintain
eyepiece combination is simply the product the focus as the slide is moved across the
of the two individual magnifications. A stage. Most microscope objectives are
convenient working rule to assist in the parfocal so that once they are focused any
proper choice of eyepiecesl states that the other objective can be swung into position
maximum useful magnification (MUM) for without the necessity of refocusing except,
the microscope is 1,000 times the numeri- with the fine adjustment.
cal apei-ture (N. A.) of the objective.
The MUM is related to resolving power The student of the microscope should first
in that magnification in excess of MUM learn to focus in the following fashion, to
gives little or no additional resolving prevent damage to a specimen ox objective:
power and results in what is termed empty
magnification. Table 4 shows the results 1 Raise the body tqbe and place the speci-
of such combinations and a comparison men on the stage.
with the 1000X N.A. rule. The under:.
lined figure Shows the magnification near- 2 Never focus the body tube down,(or the
estto the MUM and the eyepiece required stage up) while observing the field
with each objective to achieve the MUM. through. the eyepiece.
From this table it is al_parent that only
higher power eyepieces can give full use 3 Lower the body tube (or raise the stage)
of the resolving power of the objectives. with the coarse adjustment while care-
It is obvious that a 10X, or even a 15X, * fully observing the space between the


Focal Magni- Eyepiece MU Ma
length fication 5X 10X 15X 28X 25X (1000 NA)
56mm 3X 1.5X 30X 46X .-60X 75X 80X
32 5 25X 50X 75X 100X 125X 100X .
16 10 50X 100X 150X -200/ 250X 250X
8 20 . 200X . 300X 400RI .500X 500X
4 40 . 200X 400X OOX 800X 1000X 660X
1.8 90 . 450X 900X 135')X , 1800X 22504 1250X

'aMUM= maximum useful magnificati9p

Optics and the Microscope

objective, and slide and permitting the while in research microscopes, the
two to come close together wiothout poWrizer can be rotated. - Modern instru-
touching. A,
Inints have larizing filters (such as
Pblaroid) eplacing the older calcite
4 Looking through the microscope and prisms. Polarizing fgters are preferred
turning the fine adjustment in such a .,N beause they:
way as to move the objective away from
the specimen, bring the image into I are low-cost;
sharp focus.
2 require no maintenance;
The fine adjustment is VR ua11y calibrated
in one- or two-micron steps to indicate, 3 permit use of the full condenser
th vertical movement of the body tube. aperture.
ThN feature is useful in making depth
measurements but should not be relied An analyzer, of the same construction as
upon for accuracy. the polarizer, is fitted in the body tube of
the microscope on a slider So that it may
G The Substage Condenser - be easily removed from the optical path.
It is oriented with its plane of vibration
The substage holds the condenser and perpendicular to the corresponding direction
polarizer. It can usually be focused in a of the polarizer.
vertical direction so that the condenser can
be brought intethe.correct position with J The Bertrand Lens
respect to the specimen for proper
'illumination. In some models, the conden- The Bertrand lens is usually found only on
ser is centerable so that it ay be set the polarizing microscope although some
exactly in the axis of rotati of the stage, manufacturers are beginning to include it
otherwise. it will have been centered at on phase microscopes. It is located in the-
the factory and should be permanent. body tube above the analyzer on a slider
(or pivot) to permit quick removal from
H The Microscope Stage the optical path. The Bertrand lens is used
to observe the back fooal plane of the objective.
The stage of the microscope supports the It is convenient for checking cgickly the type
specimen between the condenser and and quality of illumination, for observing
objective, and may offer a mechanical stage interference figures of crystals, for adjust-
as an attachment to provide a means of ing t phase annuli in phase microscopy
moving the slide methodically during obser- and fo adjusting the annular and central
vation. The_polarizipg mica °scope is stops in aspersion staining.
fitted with a circular rotating stage to
which a mechanical stage may be added. K The Compensator Slot
The rotating stage, which is used fowbject
orientation to observe opticakeffects, will The compensator slot receives compensators
have centering screws if the objectives are (quarter:wave, first-order red and quartz-
not centerable, or vice versa. It is un- wedge) for observation of the optical prop-
desirable to have both objectives and stage erties Of crystalline materials. It is usually
centerable as this does not provide a fixed placed at the lower end of the body tube just
reference axis. above the objective mount, and is oriented
45° from the vibration directions of the
i The Polarizing Elements polarizer and analyzer.
A polarizer is fitted to,tlie`condenser of all L The Stereoscopic Microscope
polarizing microscopes. In routine instru-
ments, the polarizer is fixed with its The stereoscopic microscope, also called
vibration direction oriented north-south the binocular, wide-field, dissecting or
(east-vr-st for most European instruments)
Optics and the Microscope

Greenough binocular microscope, is in good illumination system for the microscope
reality a combination of two separate . is to have uniform intensity of illumination,
compound microscopes. The two micro- over the entire field of view with independent .
scopes,_ usually mounted in one body, havi control of intensity and of the angular aperture
their optical axes inclined from the vertical of the illuminating cone.
by about 7° and from each other by twice'
this angle. When an object is placed onthe A Basic Types of. Illumination
stage of a stereoscopic microscope, the
optical systems view it from slightly There are three types 'of ilfuminatiop
different angles, presenting a stereoscopic (Table 5) used generally:
pair of images to the eyes, which fuse the
two into a single three-dimensional image.,, 1 Critical. This is7used when high levels
of illumination intensity are necessary
,The objectives are supplied in pairs, either for"oil immersion, darkfield, fluores-
*.,T4 as separate units to be mounted on the cence, low birefringence or photo-
microscope or, as in,the"new instrtments,, micrographic studies. Since the lamp
built into a rotating drum. Bausch and filament is imadd in the plane of the'
Lomb was the first manufacturer to have a specimen, a ribbon filament or arc
zooni Iens system which gives a continuous lamp is required. The lamp must be
Change in magnification over the full range. focusable and have an iris diaphragm;
Objectives for the stereomicroscope run the position of the filament must also
from about 0.4X to 12X, well below the be adjustable in all directions.
magnification range of objectives available
for single-objective microscopes. 2 KOhler. Also useful for intense illumi-
nation, Kohler.illumination may be
The eyepieces supplied with stereoscopic obtained with any lamp not fitted with'a
microscopes run from 10 to 25Xand have ground glass.' The illuminator must
fAt.wider fields than their counterparts in the however, be focusable, it.must have an
single-objective microscopes. adjustable field diaphragm (iris) and the
lamp filament position must b\adjust-
Because of mechanical limitations, the able in all directions.
stereomicroscope is, limited to about 200X
magnification and usually does not permit 3 "Poor man's". So-called because a low-
more than about 120X. It is most useful priced illuminator may be used, this
at rela,tively low powers in observing method gives illumination of high quality
shape and surface texture, relegating the although of lower intensity becauseqf the
study of greater detail to the monocular presence of a ground glass in the system.
microscope. The .stereomicroscope is No adjustments are necessary on the
also helpful in manipulating small samples, illuminator or lamp filament although
separating ingredients of mixtures, pre- an adjustable diaphragm on the illuminator
paring specimens for detailed study at is helpful.
higher magnifications and performing
various mechanical operations under micro- All three types of illumination require that
scopical observation, e. g. micromanipulation. the microscope substage condenser focus
the image of the illuminator aperture in the
plane of the specimen. In each case, then,
HI ILLUMINATION AND RESOLVING POWER the lamp iris acts as a field diaphragm and
should be closed to just illuminate the field
Good resolving power and optimum specimen of view. The difference in these three
contrast are prerequisites for .good microscopy. types o'f illumination lie the adjustment
Assuming the availability of suitable optics of the lamp condensing le . With poor
(ocular, Objectives and substage condensei) man's illumination there is'no lamp conden-
it is still of paramount importance to use ser,.hence no adjustment. The lamp should
proper illumination. The requirement for a be placed close to the microscope so that

Optics and the Microscope°



Critical Kohler Poor man's

Lamp filamnent ribbon filament any type any type °
.Lamp condensing leris required t. required none
Lamp( iris ° required required useful.
Ground `glass at lamp none none . present
Image of light source in object plane. ° at substage none
Image of field iris nedr object in object neap object
plane plane cplane
IMage.of substage iris back focal Plane back focal' plane back focal plane
of objective of objective of objective

the entire field of view is always the substage condenser iris (also coincident
illuminated. If the surface structure of the with the anterior Meal plahe of the substage.
ground glass becomes apparent in he field condenser). The functions of the lamp
of view the substage condenser very condenser iris and the substage condenser
slightly defocused: iris in controlling, respectively, the area .
of the illuminated field of view and the
Critical Illumination angular apertureof the illuminating cone
are precisely alike for all three types of
Withcritical illumination the lamp conden-; illumination. a
ser is focused to give parhllel rays; focur
ing the lamp filament on a far wathis. Critical illumination is seldom used because
sufficient. Aimed, then,. at the substage it requires a special-lamp filament and be-
Mirror, the substage condenser will focus cause, wtren used, it,phows no advantage
the lamp filament in the.object plane. 'The over well-adjusted-Kohler illumination.
substage condenser iris will now be found 8 II
imaged in the back focal plane of the ob- Kohler Illumination
jective; it serves as a control over con-
v'ergence.of, the illumination. Althdligh the To arrange the microscope and illuminato.°
substage4ris also affects the light intensity for Kohler illumination it is well to proceed
over the field of view it should most decid- through the following steps:
edly not be used for this purpose. The
intensity of illumination may be varied by a ,Removelffediffusers and filters
the use of neutral density filters and, unless from the lamp:
color photomicrography is anticipated, by
the use of variable voltage on the lamp b Turn the lamp on and aim at-a con-
filament. venient wall'or vertical screen about
'19' inches away. Open the lamp
Kohler illumination (Figure 14) differs diaphragiri.
from critical ilhitmination in the use of the
lamp condenser. \With'critical illumination c By moving the lamp condenser, focus
the lamp condenser focuse§ the lamp a sharp image,of the filament. It
filament at infinity; with Kohler illumination should be of such a size as to fill,
the lamp filament-ts focused in the plane of not necessarily evenly, the microscope

,Optics and the Microscope

"Critical Poor man's



Focal plane

Focal plane .1)


O .
/NJ 'I
Substage' ,

Lainp iris

Light source,


Optic's and the Microscope

. .
substage ,condenser opening. If it as the center of movement (again
does note -move the lamp away ("von endeavoring to keep the lamp dia-
the wall to enlarge the filament image; phragm in the cienivred position), -c
refocus. If you have mastered this step, you
4 have Zieromplishd the must difficult
d Turn the lamp zuurain it attlie micro- portion. (}letter tilierosopd-larits
scope mirror-so as,to maintain th.: have adjustments to move the bulb,
same, 18 inches (oradjuided lamp 'indepindently of the lamp housing to
distance).` simplify this step,.)
. -
e Place a specimen oil' the iirospe. Put the specinulri in place, replace
`stage and focus sharp1.wilh.0 I ti -mm the eyepiece and the desired olSjec-
(lox) objective. Open fully the live and refocus.
aperture diaphragm in the substage
condenser. Irthe light is too bright, k Open orlose. tIm.rfield diaphragm
temporarily place a neutral density until it jtist disappears fi:om.the field.
filter or a diffuser in the lamp.
Observe -the back aperture of the
f Close the lamp diiphragm, or field. objective, preferably with the Bertrand
diaphragm, to about a 1-cm operringi lbns or the auxiliary telescope,, and
Rack the Microscope substage con- close the apeiture diaphragm on the
denser up and down to focus the substage condenser until it is about
field diaphragm sharply in the same four-fifths the diameter of the back
plane as the.speeimen.. aperture-. This fs the best position
for the aperture diaphragm, a posi-
I g. Adjust the mirror to center the field tion which minimizes glare and maxi-,
diaphragm in The field Of view. mizes:the resolving power. It is
instruttiVe to°.vary the aperturedia-
solro h Remove the 16-misr,,objectiveand phragm'and observe the image criti--
replace with.a4-mm objective, Move cally during the manipulation:
the specimen sb that a clear area is
under observation, -Place the m If the illumination is too great, : '
Bertrand lens in the optical path, or insert an appropriate neutradennity
remove the eyepiece and insert an filter between the illuminator and .
auxiliary telescope (sold with phase the condenser. Do not use the con.
contrast accessories) in itefilace, -denser aperture d.iaphragm or .the
orkremove the eyepiece and observe, lamp field diaphrAgm to controLthe
theback aperture of the objective intensity of illumination.
au ;
directly. Remove any ground glass
diffusers from the limp. Now Poor Man's Illumination
observe the lamp filament through
the microscope. . Both critical ana 'Kohler illumination re-
quireexpens illuminatorg' with adjust-
.i If the filament does not appear to be able focus, lamp 'iris and adjustable lamp
centered,'-swing the lamp housing in © mounts. Poor man's illumination requires
a horizontil,arc whoire center is at- a cheap illuminator although an expensive
Effie field-diaphragm. The putpose illuminator may be used if its expensive
is.to maintain the field diaphragm on features are negated by inserting a ground
"the lamp in_jti3.centered position, If glash diffuser or by rising arosted bulb.
a -.vertical movement of the filament Admittedly an iris diaphragm on the lamp
is required, loblieri the bulb base and would be a help though it is not necessary.
slide it up or down. If the base is'. ;, -
fixed,7-2tilt-theTlamP housing a The illuminator must have a kOsted
'vertical arc with`thefieI8diaphragm bulb , or a ground glass diffuser.

7' 5-15 ta

Orlties.and the Miernope .

should be possible to.direet it in li ItcsoWing Power
the general direction of the substage
mirror, very close thereto or in The resolving pow r of the microscope is
place thereof. its ability to distinguish ,separate details
of closoly tpaced microscopic structures..
b Focus on ahy preparation after
tilting the mirror to illumin9te the
The theoretical limit of resolving two'
dim:rely points, a. Ilistance X apart,',is:
X1. 22 k
c RemOve the top lens of the condenser 2 N. A. ;,
and, by racking the condenser pp*,
maze often, down, bring into focus wIn.re = waveleniftlmtf light, used to
(in the same plane as the specimen) illuminate °the specimen
a finger, iclneil or other object placed N, A. = nktnericalaperture of the
in the same general region as the objective ?

groundtlass diffuser on the lamp.

The glass surface itself can ern be Substituting a wavelength of 4,500
focused in the plane of the specimr.n. Angstroms an a numerical aperture of
0 1. 3, about est that can be done with
. visible light, we find that two points about
d Ideally the grotknd glass surface will ,
just fill `the field of view when centered' 2, 000A (or 0.2 fnicron) apart can be=seen
by the substage mirror; adjustment as two separate points. Further increase
may be made by moving the lamp in resolving power can be achieved far the
closer to or farther from the micro- light microscope by using light or shorter
scope (the position 'Might be marked wavelength. Ultraviolet light near 2; 000,
for each objective used) or by tutting Anqtroms lowers the- limit to about 0.1
paper diaphragms of fixed aperture micron, the lower limit for the light
(one for each objective used). In this microscope.
instance a lamp iris would be usejul.
The.numerical aperture of an objective is et

e .Low 9i condenser just sufficiently usually engraved on the objective and is

to defocus the ground glass surface related to the angular aperture, AA
and render the field of illumhtation (Figure 15), by the formula:
even. AA
N.A. r n sin 2
f,' Observe the back aperture of the
objective and open the substage con- where n = the lowest index in the spacp
denser iris about 75-percent of the between the object and the
way... The final adjustment of the objective.
substage iris is made 'while observing
the preparation; the iris should be
open as far as possible, still giving
good contrast.
g The intensity of illumination should
-be adjusted °WI, with neutral density
filters or by changing the lamp voltage.
Proper illumination is one of the most im-
portant operations in microscopy. It is
easy to.41:dge microscopist's ability by 4
a glance at his field of view and the objec-
tive back lens. Figure 15


s 2"
Optide and the Microscope
e '

1 Maximum useful magnification.

A helpful rule of thumb is that the use-
ful magdificatio'n will not exceed 1,000
times the numerical aperture of the
objective (see Tables 3 and 4). Although
somewhat higher magnification may be
used in specific cases, no additional 10111 1111114,-

detail will be resolved.

It is curious, considering,the figures a b

. in the table, that most, if ilot all, manu- Figure 16

facturers of microscopes furnish a 10X
eyepiece as the highest power. 4-10X ABBE THEORY OF RESOLUTION
eyepiece is useftiltlit anyone interested e
in critical work should use a 1-L2SX eye-
piece; the 5-10X eyepieces are best for b The conde.nser shotild be well-
scanning purposes. .
ra) corrected and have a numerical
aperture as high as the objective to
2 Abbe's theory of resolulion be used.
One of the ,mostcdgent theories,bf (c An apochromatic oil-immersion
resolution is due to Ernst Abbe, who objective should be used with a come-
suggested that mtroscopic objects act` pensating eyepiece of at least 15X
like diffraction gratings (figure 16) and magnification. The immersion oil
that the angle of diffraction, therefore, should have an index close to 1.515
increases' withthe fineness of the detail. and have proper dispersion for the
He proposed,tiltit a given microscope objective being used.
objective Would resolve a particular
detail if at least two or three' transmitted (cf--"Immersion oil should be placed
rays (one direct and two diffracted rays) between the' condenser and slide and
"°.entered the objective; In Figure )6 the between cover slip and objective._
detail shown would be resolved in A and The preparation itself should be
C but not in B. This theory, which can surrounded by a liquid having a
be,borne ougy simple experiment, is refractive index of 1.515 or more.
'useful in shoWing how to improve resolu-
tion. Since shoker wavelengths will e The illumination should be reasonably
give a smaller diffraction angle, there monochromatic and as short in wave-
is more chance of resolving fine detail length as possible. An interference
with short wavelengths.. Also, since filter transmittinga wavelength of
only two.of the transmitted rays are about 480-500 millimicrons is a
needed -Oblique lightmand a high N.A. suitable answer to this problem.
condenser will aid in resolving fine detail. Ideally, of course, ultraviolet light
4 0 3 should be used to decrease the wave-
o 3 Improv,ing resolving power length still further.
. .
The folToeing list summarize the The practical effect of many of these
practical approaches to higher resolu- factors is critically discussed by
tion with the light microscope: Loveland(2) in a paper on the optics of
a The specimen should be illuminated
by either critical or Kohler
- illumination.


Optics and the Microscope

IV PHOTOMICROGRAPHY curvature of field and depth of field. The

photographic plate, however, lies in one
A Introduction plane; hence the greatest care must be
used to focus sharply on the subject plane
-Photo (gra h , as distinct from micro- of interest and to select optics to give
photograp y," is tie art of ta iiii-TAC--tures---------minimtim_a.mounts of field curvature and
thrtugh microscope. A microphoto- chromatic aberrations.
graph is a small photograph; a photomicro-
graph is a photograph of a small object. With black and white film, color filters
Photomicrography is a valuable tool-ill may be used to enhance the contrast of
recording the results of microscopical \some portions of the specimen -while mini-
stud . It 6nables the microscopist to: mizing chrdmatic aberrations of the lenses.
In color work, however, filters cannot
1 describe a 'microscopic field objectively usually be used for this purpose and better
)bithout resorting to written descriptions, optics may be,required.
2 record a particular field for future Photomicrographic cameras which fit
reference, directly onto the microscope are available .
in 35-min or up to 3-1/4 X 4-1/4 inch sizes.
3 make particle sl,ze counts and counting Others are made which accommodate larger
analyses easily and without tying p a film sizes and which have their own support
microscope, independent of microscope: The former,
however, are preferred for ease of handling
4 enhance or exaggerate the visual micro- and lower cost. The latter system is-pre-
scopic field to bring out or emphasize ferred for greater flexibility and versatility
certain details not readily apparent and lack of vibration. The Polaroid camera
visually, . has-many applications in Microscopy and
can be ilised on the microscope directly but,
5 record images in ultraviolet and infra- becauSe of its weight, only when the micro-
red microscopy, which are otherwise scope his a vertically moving stage for
invisible to the unaidecl-eye. focusing rader than a focusing body tube.
There are two general approaches to photo- B Determination of COrrect Exposure
micrography; one requires only a plate or
film holder supported above the eyepiece ;Correct exposure .gitterntinatisin can be
of the microscope with a light -tight be,illows; accomplished by tfial and error, by. relating
the other utilizes any ordinary camera with new conditions to previously used successful
its own lens system, supported with a light- ,conditions and by photometry.
tight adaor above the eyepiece. It is
best, in the latter case, to use a reflex With the trial and error method a series of
'camera so that the image can be carefully trial exposures is made, noting the type of
foCused on the ground glass. photomi- subject, illumination, filters, objective,
crography of this type can be regarded eyepiece, magnification, 'film and. shutter
simply as replacing the eye with the camera /speed. The best 'exposure is Selected. The
lens system. The camera should be focused .fillowing parameters can be changed'and
at infinity, just as the eye is for visual The exposure time-adjusted accordingly:
observation, and it shoal be positioned
close to and over the eyepiece. 1 Magnification. Exposure time.vvies
as the square of the magnification.
. the requirements for photomicrography;
hOivever, are more rigorous than those Example: Good exposure was obtained
for visual work. The eye can norinally .; With a 1/19-second exposure
compensate for varying light intensities; .
and a Inagnification of 100)1.
If the'magnification is now:

5.48 'OP

4 '
i Optics and the Microscope '

° o ...
1 2004, the' correct exposure, Kodachrome II-Type A
__ ..._ I__ is calculated as follows.,, Professional is 40.
. -. 4
ifew exposure time= 'old exposure time new exposure time = old exposuie time
new ma gnification0 (20012 -
A. S. A. of old film 11100(400/40).

old magnification ' 100' A. S. A. of new film _

° .`
4/10 or say,' 1/2''sRcond. 10/100 or 1/10 second.

1 should-hem- .hOWever, that the 4 Other parameters may be varied but the
above calculation carl:be made only when prediction of exposure time cannot be
there has been no change IQ the illumi- made readily. Experience and photo-
nation system including the condenser electric devices at'e the best guides to
orAthe objective. Only changes in magni- 'the proper exposur'e. ' .
fication due to changing eyepieces or ;!'
bellows extension distance
can hand- a Photoelectric devices are excellent for
led in the abovg manner. determining correct,exposure. Since
brdinary photographic.exposure meters
*4.- 'Numerical aperture:- Exposure time, \ 'Vre not sensitive enough for photomi-
varies inversely as the square ofm:the-% crography, more sensitive instruments,
smallest for king _hume B id ai!:"ap having a galvanometer'or
electronic .
of the condenser and objective. amplifyined'ircuit, are required. Some
.., photosensitive cells are inserted in the .
Example; Good exposure wad obtained body tithe in place of the eyepiece for
at 1/10 'geeorld wit)fi tide, light intensity roadings. This has the
lt° objective, N.A. 0.25, '°4t-0 advantage of delecting the light level at a
o full aperture., With a 204,r point of high intensity but &les not take 4
t objective, N,A. 0.25, at iqo account the eyepiece, the distance to
. . full aperture and the same ; thalm or the filar speed.
final tnagnificatioV what is 4
-the correct expcialire time? The cell May be placed just above the eye-
'Co P iece so that it. 5 egisters 'the total amount
new exposure time = old e. pure time alight leav,i0Vthe eyepliCk. 'Again, .the
;old N.A. 1 ..°1/19
effec,ts of film stgaetoatiinte44eid .
new N. A.' O. 0
11'40;or, distake are no "'r!..., The-04n
s ip.11dial,vbet.Fk with the total _light - , ,
/ , I -.1:=,%

measurihk-photomefer'is ip e difficulty' of ,.
, .1450 second.
taking into account the arkaof field covered.: -!`,,' .

It it -Oeen that moreeliglit reaches the - ake, for example, a.bright fieldin Mich , 1 ,,,t .°
photographiF filttkitytth highat humeri- P ly few crystalsappear,t perhaps 1 per-
CP cal apertures at tlic pagnemagnifiCation. derhf of the light entering the field, of view is
.1 scatter0,03/ the crystals and the photomete..y : ,. '"
3 Film. Etpbsure time varies inversely. shows closAto a ma9imum reading!' Now - .

.1. with the Aifierican Standard's Association asstime that evetrythihg r_e_2mains_constant- ,
speed index of the film. t-the hunib-Ei. of crYitals and, conse-

Exainple : A'gdeid.picture was obtained

. .quently-,,the amount of light.-scattered:'
The'pt6torpe'ter rdadini couId easily drop I
with Edstman'Tri-XfilrrYat by 501:lerdent., yet ttiii° proper,,xposure is
second. What is the' unchanged.. The.,:;ituation for
Correct ext)surefor photoTrdb.rograp y wi *th crb s,ed polarp since,

, Eastman Kodachrome .. . ' the photometer reading depends. oh thei
Type A. .The A. S.A. speed . intensity
-of lildrninationvOn the bire-
for Tri-X it 400 and for fringence andstlicckness of the,,crystals and
. . ;". ./ ,.
t t 44,
8 °
Optics and the Microscope e

on the number and size of the crystals in same film and projection
field or, alternatively, on the area of distance) and that the new
the field covered by birefringent crystals. meter reading is-16; therefore:
One of the best solutions to this problem
is to measure the photometer reading with expVsure time = k/meter
no preparation on the stage: A first-order reading = 8/16 = 1/2 second.
red compensator or,xqirartz wedge is in-
serted when crosoRfOlars are being used
illuminate -the entire field.- .V MICROMETRY

An alternative to place the cell on the A Particle Size Determination

ground,glass ere the film will be
located. ever, although all variables Linear distances and-areas can be
except m speed are now taken into measured with the microscope. This
account, measurements in the image plane permits determination of particle size 6

have the disadvantage of requiring a more and qUantitative analysis of physical

sensiOve electronic photoelectric apparatus. mixtures. The usual unit of length for ,
microscopical measurements is to micron
No matter what- m ?thod is used for placing' (1 X 10-3mm or about 4 X 10-5incffl.
the photocell, the exposure time can ,,W$ Measuring partici% in electron microscopy
determined by the general formula: requires an even smaller unit the milli-
micron (1 X 10-3 micron or 10 Angstrom
exposure time meter reading units). Table 6 shows 'the approximate
average size of a few common airborne
The constant k will'depend on the physical materials:'
arranerdnt and film used.; To determine.'
k for any rticular system, fillst set up
the microsccr totake a picture.- Record Table 6.. APPROXIMATE PARTICLE SIZE OF
the meter' reading anci take a series of N
S trial exposures. Pick out the best exposure
. and calcylate k.- Then the k which was Ragweed pollen
determaned holds as Iongas no change is 25 micr ons
made lit* right pathpeyond the photocell,- Fc.:,g droplets 20 microns
e. g: chAiging to a faster film or changing PnW Pr- plant flyash
'thF668atiOridratance.,. Thus the, objective, 5 rriic rons
(after prIcipitators)
'condenser position or illuminator may be
changed without affectinili if the cell is Tobacco smoke 0.2 micron
;,used as deicribed above. (200.millimicrons)
With one particular arrange- Foundry fumes .. ' 0.1 - 1 micron--
Example: (IUO =-1b00 rfaiiiinicrons)
ment of photocell and
found to
o be 40. A.serfes of photographs
arelakensat 1/2, 1/5, 1/10,
.IL2,5 and 1/N secon *. The. The practical lower Unlit of acaurate
ph9toinicrograpil taken at 1/5 particle size measurement withthe 4ght,
second is judged to be. the best; microscope is about 0.5,micron: The
',hence k is calculAea as follows: easurement of a particle,smaller than
this with the light microscope leads to
W4nieter.reading' X exposure errors which, under the best cirertm-
time = 40X 1/5,..= 8. stances, increase to about + 100,
,(usually +). 7
4t Assume,,novithat. a new picture .
.is to be takenat.sinot)ier Qne of the.princijIal-uses res.oltring
magnification (hut witty-the power is in the precise measurement of
r -* t
.;,t -4
Optics and 'the Microscope

*fr partide size.. There are, 'however, a in an'appropriate direction until the
variety otapproximategand useful proce- second trailing edge just touches the
dures -as well. cross-hair line. A second reading is
taken- and the difference in the two`
1 Methocid of particle size measurement readings is the distah-Ce mov- r the
size of the particle. This method is.
a Knowing he Magnification of the especially useful when the particle
microscope (product of the magni.- is larger than the field, or when the
fication of objective and ey'epicce), optics give a distorted image near the
the siie -of.particles-can be esti- - edge of the field.
mated. For example, with a.10X
. eyepieceind a 16-mm (or 10X10X)'
objective', the, total magnification
d The above method can be extended to
projection or photography. The image
is 100X. A particle that appears to .of the particles can be projected on a
be 10-mm at 10 inches from the eye screen with a suitable light source or
has an actual.size of ,l0 mm divided they may be photographed. The final
by 100 or 0.10 mm or 100 microns. magnification, M, on the projection
This is in no sense an accurate surface (or film plane) is given approxi-
method, but it does permit quick mately' by
estimation of partible size; the error
in this estimation is usually 10-25 M = D X O. M. X E. /25
where 0. M. = objective magnific4on
b' An9ther approxitrote method i's also E. M. =eyepiece magnifica ion
based on the use of known data'. If. D = projection distanC0
we know approximately the diameter from the' eyepiece'in
of the microscopelield, we can Centimeters.
.6- estimate the percentage of'the
diameter occupied by the object to The image detail can theft be measured,
be measured and calctlate from - ' in centimeters and the actual size com;)
these figures the approximate size puted by dividing by M. This methods
Aof the object. The size of the micro- is usually accurate to within 2-5 percent
scope field depends on both the objec- depending on the size range of the detail
tive and the ocular although the latter measured.
is. a minor influence. The size of
the field should be determined with e The stated magnifications and/or focal
a millimeter scale for each objective lengths of the microscopeoiltics_a.re
.lend ocular. If this is done, eiti- nominal and bit from objective
tiori .0.fizes-by-comparigiiii with- to objective or eyepiece to eyepiece.
entire field diameter carbe quite To obtain accurate measurements, .a
( accurate (5-10%). stage micrometer is used to calibrate
each Combination of eyepiece ant
c The movement of a graduated mechan- objective. T1* stage micrometer is
ical stage can also be used for rough a glass microscope slide that.has,
Measurement of diameters of largt accurately engraved in the center, a
larticles. Stages are usually grad scale., usually 2 millimeter slong,
aid (with vernier) tO'read to 0.1 divided into goo parts, each part repre-
mittimeter, or 100 microns. In senting 0.01 millimeter. Thus 'when .
..,practb-b,' the leading 'edge of the this scale is observed, projected.or
particlelis brought to one of the lines ,..,photbgraPhed, the exact image inagni-.
of the cross hair in.thp eyepiece and fication can be:determined. For
a.reading iSe,t4100 bf the AtAkeposition. 'example, if 5 spaces of theg,stage micro-
Then the particle if mimed across the meter measure 6 millimeters when '
fieldby moving the mOf tartirCailttage
' 'projected, the'actual magnification is
\\'' `


O .


4 '4 .4
tics and the Microsc

6 The calibration consists, then, of

120 times.
5 (0. 01) calculating. the number of microns per
This magnification figure can eyepiece Scale division. To make the
used to improve the accuracy of comparison as accurate as possible': a
method 4 above. large part of each scale must be used
(see Figure 17). Let's assume that
The simplest px.iXedure and the most with the-low power 16-mm objective.
accurate is based. on the use of a 6 large divisions of the stage scale
_ micrometer eyepiece.: Since the (s.m.d.)-are equal to 38'divisions of
eyepiece, magnifies a real image the eyepiece scale. This means that
from the objective, it is possible 38 eyepiece micrometer divisions (e.m..
to place a transparent scale in the are equivalent to 600 microns. Hence:
same plane as the image from the
objective and thus have a scale 1 e. m. d. = 600/38
superimposed over the image. This = 15. 8g.
is done by first piecing an eyepiece
micrometer scale disc in the eyepiece.
The eyepiece micrometer has an
arbitrary scale and mast be cali-
brated with each objective used. The
simplest way to do this.is to place . a
the stake micrometer -on the stage
and note a convenient whole number Step litcrentieler Scale
of eyepiec4 micrometer divisions., 14./4,14.A.16L4.1.1 .1.4.1411,
The value inmicrons for each eye-
piece micrometer division is then 0
easily computed.. When the stage Eyepiece blieremeise lids .
micrometer is removed and replaced
by the specimen; the ,superimposedi
eyepiece scale can be used for accu-
, rate measurement of any feature in
the spec:igen by direct observation,
photogrFWor projection. 4
Figure 17
2 Calibration of eyepiece micrometry .
Each znicgometer 'stage scale has SCALE WITH EYEPIECE MICROMETER SCALE
divisions 100g (0. 1mm) apa t; qne t
or two of these are gusuall subdivided 'Thus tvhen that micrometer eyepiece
into '10g (0. 01-mm) div ions. These is ased.with that 1.6 -nim objective each
form the standard against which the division of the eyepiece scale is eqUivalent
arbitiary divisions in the micrometer to 15.8µ, and it can be used to 'make.an
eyepiece are to be calibrated. Each accurate measurement of any object on
objective must be "calibrated-separately the microscope stage. A particle, for
by noting the correspondence between example, 'observed with the 16-mm objec-
the stage scale and the eyepiece scale. tive and measuring 8.5 divisions do the
Starting with the lowest power objective eyepiece scale is 8.5 (15.8) or 135g in
focus -on the stage/scale, arrange/ the diameter,
two scales parallel- and in goodldQus.
It should be pcishible to determine the Each objective on your microscope angst
number of eyepiece Azisions exactly be calibrated in tis manner. 0.
,equal to some whole umber of
divisibns of the stage scale, a distance A- convenient way to record_the. necessary:
readily expressed in microns. data and tct calinlate g/emd is by. means
of a table.


Optics ancrtlie Microscope

Table The more pafticles counted, the more

accurate will be the average particle
Objective nb. emd- no. emd size. Platelike and needlelike particles
1 emd should have a correctioir factor applied
32-mm 18 = 44 1800 = 44 40.9F.,
, to account for the third dimension since
all such particles ale restricted in their
16-mm 6 = 38 600 = 38 15.. 8Fi orientation on the rrTicroscope slide.
4-mm 1 = 30
100 30 3.33µ Whet; particle site is report11, the
'general shape of the particles as well as
the metboditied-to-delefrifine- the
"diameter" should be noted.
3 Determination of particle size Particle size distribution is determined
.distribution routinely by moving a preparation of
particles past an eyepiece micrometer
The measurement of particle size can scale in such a way that their Martin's
vary in complexity pending on parti- tdiameter-.can be tallied. All particles
cle shape'. Ttfe ze of a sphere ma e whose centers fall within two fixed
denoted by divisions on the scale are tallied. Move-
cube may be'expressed by engt Ment of the preparation is usually
4n edge or diagonal. Be d these two accomplished by means of a mechanical
configurations, the p- cle "size" must stage but may be carried out by rotation ,
include information' out the shape of of an off-center rotatingstage. A sample
the particle in quest on, and the tab
tabulation rn rTea .obnloe s8. sTohe ear -
expression'ofthis s ape .takes a more piece
complicated form. t .at least-six, but not more than tweive,,
size classes are required and sufficierit
Martin'S diameter is e simplest means pArticies are counted to give a smooth
of measuring and expr ssing the dia- curve. The actual number tallied 200
meters of irregular pa icles and is 000) depends on particle shape
sufficlVtly accurate wb n averaged for re laOty and the' range of sizes. The
a largeltAnber of partic es. In this ze tallied fqr each Particle is that 0

methodi the horizontal east-west number of eyepiece micrometer divisions

dimension of each icle which _divides most closely approximating Martin's
the projected,area into halves-is taken as diameter for that par rticle.
Martin's diameter (Figure 18):
4 4 C alculation of.size averages

The size dat'a maybe treated in a variety

1-74 I 1----4 1 1 of ways; one simple, straightforward
i 1
I treatment .is shown in Table 9. For a
I i
more complete discussion of the treat-
ment of particle size data see Chamot
and Masdn;s, bybook of Chemical
/Vlicroscopy(aage 26. re
Figure 18 The averages with resperugiiier,
9 7:11-, surface, d3; and weight or volume,
. MARTIN'S DIAMETER , d4, are calculated as follows for the
data in Table 9.

5 -23 ,

87 ,-

Optics and the Microscope

&so class
. (sods) Numb** of particles ° Total
4 1 r1=4.1 r-r44 rt.4.4 1$

! -
rt44 214..1
11-14 r-144
r144 ri-44

rt-1-4 ft4.1#1'S-1.1 11,1

rt-i4 1-14.4 1"-11-4.1 110
re-44 rt-44 2444 ri--1.4 r-24.4
r-1-44 2-14.1 rt-4-4

4 11-44
-2-1-4.1 11-541 1S7
r 1-4.1 1-1-14 rt-4-1 1-t-4-1
r1-4.3 rt-44 lit -4.l
ri-44 .7-1

rt44 rt-s4 rt.44


1-144 rt-4.1 45 S
7 t71-1..1 I-I-14 14-3-1 -1

*find eiralide microrneteL dizialeas

di = En'd/En = 1758/470 ,

= 3. 74 emd X 2. 82* =7. 10. 5p.

.713 = End3/ End2 = 37440 /7662

= 4. 89 emd X 2. 82 = 13. 81.1.

d4 = End4/End8 = 1991941 37440 .

= 5: 32 emd X 2. 82 = 15. OR
*2:82 micronSper emd for the. cumulative;wei4ht or volume
(determined by calibration of the curve, are plotted against d. Finally,
.eyepiece - objective combingtion the.specific surface, Sm, .in square
used for the determination). meters per gram, m, maykbe calculated
if thedensity, D, is known; the surface
Cumulative percents by number, average ;53, Is used.
surface and weight (or volume) may be -
plotted' frOm the data in Table 9. The If D 1, Sm = 6 /d3D = 6/13. 8(1. 1)
calculated percentages, -e.g. = 0, 395m2ig.

5 83
Optics and the Microscope

/ '
(Aver. diam. n nd nd2 nd2 nd4
in emd)
1 16 16 ^ 16 16 16
98 196, 392 784 . 1588
3 -.1.10 330. 990 2970 . 8910
4 '107 428 1712 6848 27392
5 355 1775 8875 "'"44375
,1' 6 45 270
-..4 "\
1620, , 58320
7. 2f 147 1029 '''7203. 50421
8 2 16 128 1024 8192

-470 1758 7662 37440. 199194,

B Counting Analysis 4 .e..g.seounting the percent of raxon fibers
, ip a sample of "silk".
\of .
Mixtures of patteulates can often be
quantitatively aniprzed by counting the Example 1:"A dot-count of,a mixture ,of
total number of particulates from each fiberglass and nylcin shows:
component in a' representative sample. '
The calculations are,' however, comPli.- nylon 262
cated by three factors: average particle fiberglass . 168
,size, particle shape and the density
of the componints: If all of tht compon- Therefore:
ents were equivalent in particle size, % nylon = 262/(262 + 168) X
shape and density then the weight.per- = 60.9i by number.
centage would'be identical tothe number_
perceritige. llsally, however, it is -However, although both fibers are smooth
necessary to determine correction factors cylinders, they do have different densities
to account for the differences. and usually differegt diqieters. To
correct for diameter one must measure. -
When pxoperly applied, this method cal the average diameter of each type of fiber
be-accurate to within + 1 pirced and, and calctilatethe volume of a unit length
in special cases, even better. It is often
applied to e analysis of fibefmixtures aver. diam. volume of
and is then ually called a dpt-count 4 1-p, slice, p.3
because the y of fibers is kept aathe nylon 185' 268
preparation is oved past a point or dot
in the epiece. fiberglass 13.2 117

A variety of methods n be used to "le percent by volume is, then:

simplify recognition of the different " 2620( 26,8
Components.-----Theseinclude chemical % nylon X 100
(262 X 268)+(168 X 1 .17)
staineor dyes and enhancementOrOptical- ---
differences such as refracWe indices, = 7; 1%-by-volume. _ _
dispersion or color. Often, however, one
.relled.cc.the differences, in morphology, , Still we must take into account the density of
each in order to calculate the weight percent.


Optics and the Microscope

If the densities are 4.6 for nylon and 2.2

efor glass then the percent by weight is:
262 X 268 X.]. 6.
% nylon :-.,(462 N 268 X L 4)+0 68>c,417 X 100
, = 72% by weight.

ElamP11%.2: A count of quartz and


gypsum. 467

To calculate the percent by weight we must

take into account the average particle size,
the shape and the-density'of
The average particle size with respect to
weight, d4,. must be measured for each
and the shape factor must be determined.
Since gypsum is more platelike than quartz
each particle of gypsuni is thinner. The
shape factor can be approximated or can be
roughly calculated by measuring the actual,
thickness of a number of particles. We
might find, for exaMple, that gypsum parti-
cles average 80% of the volttme of the aver-
- age-quartz particle; this is our shape factor.
The final equation for the weight percent is:
283 X itc14/6 X Dq
% quartz = 238 X it-a4/6 X f)q.+ 467 X it cit/6 X 0. 80 XIDg X ipo

where Dq and Dg are the densities of quartz

and gypsum respelcitrely;--0 : 80 is the shape,.
-.factor and d4 and d4, are the average parti-
cle sizes with respect to weight for quaytz
and gypsum respectively..

ACMIOWLEZIGNIEN. T: This ogtlih'e was 2 Loveland. R. P.. J. Roy. Micros.'OSc.
preparicl.by the U.S. Public Health Service. 79, 59. (1980).
Departmenfjctf-- Health. Education and Welfare,'
for use in its Training Program. 3 Chamot. Emile.Monnin and Mason.
Clyde Walter, Handbook of Chemical .
Microscopy, Vol. 1, third ed.
John Wiley and Sons. New York (1959).
lEhmn. C. W. Crystal Grovrth from
Solution. Discussions of the Faraday DESCRIPTORS: Microscope and Optical
eociety No.5. 132. Glittery and Jackson. propertieP
- g


. 5-26.


I INTRODUCTION membrane may be thought of as the

outermost layer f protoplasm.
What are cells? Cells may. be defined as the
basic structural units of life. The dell has b In plant cells the most-conspicuous-
many different parts which'carry on the protoplasmic structures are, the .
various functions of cell-life.- These are "chloroplasts", which contain-
called organelles ("little organs")..' . highly organized menibeane systems
bearing the photosynthetic pigments
A The branch of biology which deals With the (chlorophylls, carotenoids, and
form and structure of plants and animals xanthophylls) and enzymes.
is called "MorPho loci." The study of the .1

arrangement of their several parts is c The. "nucleus" is a spherical body

called "anatomy", and the study ,pf cells whicli regulates cell function by
,is called "cytology','.. controlling enzyme synthesis.

B "There is no "typical" cell, for cells differ d "Granules". structures of small

from ,each'other in detail, and these size and. be "living" or
differences are in part respdhAble for the non-living' material.
variety of life that exists on the earth.
. . e, "Flagella are whip-like structures
'found in both plant and animal cells.
II FUND'AMENTAI;S OF CELL STRUCTURE Thy flagella are used for locomotion,
or to circulate the surrounding
A tiow do, we recognize a structure as a cell? medium.
We must look for certain characteristi-ea.--
and/or -structures which have been found f "Cilia" resemble short flagella, found
to occur in cells. The cell is composed almost exclusively-on animal cells.
of a variety of substanc.es and structures, In the lower animals, tilts are used
some of which result from cellular for,Aocomotion and food gathering.
activities. These include both.living and -
non-living materialS. , g The "pseudopod", or false foot, is
an extension of the protoplasm of
Non-living components incltide: certain protozoa, in' which the -
colloidal state of the protoplasm
a. A "cell Wall" composed of cellulose alternates from a "soh' to a "jel"
may be found as the outermost condition from titne-to time to
covering a many plant cells. facilitate cell movement.

b t "Vacuoles" are chambers in .the . h "Ribosomesuare,protoplaarnic bodies

protoplatm which contain.fluidS of which
are the site of protein ;
different 'densities (i. e., *different synthesis. They are too small
from the surrounding protoplasm). (150 A in diameter)ito be seen with
7 a light microscope.' .

2 The "living" partii of the cell areicsilled

"protoplasm.", The following structures i "Mitochondria" are small. mem-
are, included: . branoUs structures containing .

enzymes that oxidize food to producel

a. A thin "cell Membrane" is located ienergy' transfer cdmppunds, (A TP ) yG
just inside the ref wall. This

BI. CEL. 6. 76 6-1

. -
Structure and Function of Cells

B how basic strutture is expressed in some 3 Thp green algae as a group include a.
major types of organisms. ' great variety of .Structural types, .
ranging from singleq-celled non-motile
We call better visualize the variety Of cell forms to large motile colonies. Some,
4 strUCture'by considering several spedifip types are large enough to resemble
cells-. higher aquatic plants.
o I ... a
1. li,4Cieria have fevrorganelle's;_ and are a. The' chloroplasts.aremoditied irkbos
so minute that under the light - A variety of: shapes and ire loomed
, MicroscOPe only general morpholOgical in different positions, Examples
types (ire., the three basic shapes; of dh3.oropla.st shape and position.aret
rods, spheres-, and spirals) can be
recognized: The following structures
have been defined: : 1) Parietal - located c5xrthe
r- a thick protective
a periphery of the cell;' usually
a The. "capsul e'',*s pup- shaped and may extend
cove0ng,of the cell ekterior., con- 1
completely around the inner
sisting of pArsacci-bifi, cie or surface of the plasnia membrane.
2) Discoid - also located oq the
b The dell watt and plasma membrane" periphery of the cell; but are.
- are present.. plate-shaped; usually many per
c Although no well.defined nucleus is . sokt,
visible in bacterial cells, the ; .3) Axial - lying in the central axis-
electron microscope has revealed of the cell; may be ribbon-like
areas pfdeoxyribone nucleic acid pr star-shaped.
(DNA).con8entration.' This sub- . .*-
stance is piese,nt within the nucleus of 4) Radial - have-os or processes
of higher cellk, and is'the genetic that extend' outwa-i'.rpd from the
or hereditary material: ;
scenter of the cell (radiate),
d Some types -Of pa-cttriaTkynt.iin er" reaching the plasma melnlYrane..
special type of chlorophyll 1 5) Reticulate - a mesh -like network _
(bacterioc rophyll ) and carry on that extends throughout Volume
itotosyn esis, ofthe cell.
2 ue-green algae are similar to the b Located in the chloroplasts may be
bactbria. in structure,. but contain the dense, proteinaceous, starch.
pho ospithgtic pigment chlorophyll) a. forming bodies call "pyrenoids".
a t ke the bacteria, these forms also 4 The flagellated algae .pbasess dne-to-
tlackan organized nucleus (the , eight flagella per cell., The chloro-
nuclear region is not bounded by a plants may contain brown and/or red
merhbrane). 7 pigments in addition to chlorophyll.
41. 4,1

+he Chlorophyll,b,ea.ring membranes a ReserVe food may be stored as

aide not localized in distinct bodies
. .
starch (Chlamydomonas)paramylon
0 IChloroplasts), but are diaperded- (EUglena), Or as oil. 4
throughout the. cell.
. .
.... ' 5 The protozoa are single-celled
Gas -filled structures called arlimals whidh exhibit a variety of
"pse'Udovacuoles" are found iii some cell structure.
type g.of blue-greens:,
/ '
.92 /.
Structure and Function of Cells

.:a higher concentration of a substance
. 4

a The
-, amoebae move.by means of
, pseudopodia, as described
- previously.
"(Such as, phosphate) inside the cell than
is found outside. Algae are able to
r- synthesis 'organic matter from inorganic
b The flagellated prbtozoa raw.materials (carbon dioxide and ,
(Mastigophora) possess one or more water), with the aid of energy derived
from light; whereW3 animal cells must
obtain their organic matterready- .
c The ciliates are the Most highly made" by consuming other organisms, ".
-friodifiedprotozdans. The cilia may organic debris,. or dissolved organics.
,be more or less evenly distributed . _

,bver:the entire surface. of, the cell,

or mapbeelocalized. IV SUMMARY
The cell is made up of many highly special-,
III FUNCTIONS OF CELLS izedgilbstructures. The types of sub-
structures present, and then. appearance
What are the functions of Cells and their (shape, color, etc, ) are very imnortant, in
structural components? Cellular function, understandinethe role of the organisth in
is called "life", and life is difficult to define. the aquatic comMunity4 and in classification.
Ltfe..is charadterized1:89moccsse ommonly %.-
referred to as rejarbduction, growth, photo- .
syntflegis, etc. ° \REFERENCES - °
'A" Microorganisms living in surface waters -1. Bold, H. C.. Cytology of algae. In: G.M:
are subjected to corratant fluctuations in 44 Smith, (ed. ), Manual of Phyvlogy.
. r .
the physical and chemical characteristic
of tthe
h e e n v i r ondAt, and mus t constantly
. .
Rona ld Press.
1951. 1

Im O d ifr t h e i r activities. " 2 B. otn. e) Geoff f ry H ed° : , Cytology aid

. 'Cll
, .
Physiology. .3rd ed.,' AAcademic
The. cell requtres'oe. source of chemical, 4
. o;
. .

energy to, carry On life proceies and .. .. °'

successfullZoompete with other-, 3 :lara_chet, Jean. The Living Cell:
° oFgarnsms. Plant cells may, obtain r
' Scientific American. 205(3): n61. .,..
1" this energy from light, ,which is,
absorbed by chlorOphyll and converted .4 Corliss, John O. Ciliated Protozoa. ,
into ATP or food reserves, 'dr from ,- ' ... Pegamon. 1961. -
the pxidation of food s tiffs. Animal
cells obtain energy on y from the i'rittch; Fre- The: structure and .
oxidation oflood.,- ° treproduction.of the algae. Cambridge
0 0e
F.' , ,. ,
Unik( Press. , 1965, . ,P
ells. must obtain *ravio5ixaci eriars, from
the *envirourpent In:ordeF to grovrand Frobisher, .M. I'luilamentals of
carryout otheilifeyfilnctiOris: Inorgaiitc, itliepdbiology .7th edition. W.B.
and'orgailic..inatelials may, be to:keit-up . Saunders Philaalphia; ;1962.0
by passive diffusion' or by . ...
44, .
.* ;-:.
transport". Irl,theaFter process, '7 Round, F'. E. The biology of-the algae.

energy Is used to,laulc1 up and maintain. St. Martin's .Prese. 'New York. 1999*

a ; `.14'4
.4 4 ; ' Ili

.. 4.. re
,, o o . :
1 ,. ca., '-; ... Tlis°outline,originally.prepared by ttilhaer .
° s
,,;:. , c
9' E. Bencier;Biologist, /orfnierlyth 9 . 4.1

.0 . Training ACtivities; FWPCA, SEC. and

revised by Cornelius I. Weber, March 19_700.


- $
Descriptor: °Cytological Studies
'' .. 4.04

- ' '44 9




2 Green photOsynthetic bacteria
Micvochloris, Chlorobium, and
The. WINOGRAD,SKy COLENIN is an excellent Chlorochromatum; methane bacteria;
simple classroom experiment as wklfas a and SO4 reducers.
miniature_ecosystem which yields a variety
of-photosynthetic and other prbtista -especially 13 Photosynthetic pUrple sulfur bacterfa -
the bacterial forms important in photosynthesis° Thiopedia and Thiosarcina. II
research. These photosynthetic bacteria, as
pointed out by Dr. Hui r, are ubiquitous in 4 Filamentous sulfur bacteria,
wet soils and natural waters but ordinarily Beggiatoa.
escape notice.
5 Non-sulfur photosynthetic bacteria,
- ,
6 Blue-green algae, Schizothrix and
The materials needed are simple laboratory Oscillatoria, .
items. \
Diatoms, Nitzschia.
A The'ino6ulurit la back sludge)*may be
easily obtained From a lOcal'sewage 8 Coccoid green algae, Ankitrodesmus;
treatment plant or the bottom of a pond and flagellate greens, Chlamy'domonas;
oil lake. Because the USPHS document, , similar to a stabilization'pond flora,
contairiing Dr. Hutner's paper, is out of
print\it reproduced, ere. 9 Filamentous green algae, Ulothrix.
B Directqls for preparing the cofiren,and C Besides the phbtosynthetic bacteria and'
other'useful inforination are given in that, ather.piotista there will be a variety of °
paper. peotozoa found in the aobic levels
(app. 6 through,9). Many of,these
01/4.12r. Butte 1s*bittliography Should be protozoans are typical fauna of activated
sufficient f r these who wish" more sludge.
. information:
D The possibilities are endless for further
experimentation. These ecosystems ax's
also conyeniehf and inexpensiye seurces
for protozoa and,ottterltiodsta for class,
arid laboratory instruction.
A yactoi' such as the substrate used, the ° .

inoculum, the overlying supernatant water,

end labOrafory, condiUsi- as temperature , 4 o.,
light: all;ittfltred440 p&rticul Ir. MICRIDAQUARIA
4 ,
type of biota forfaing successive layers
or zones.. The.4.cdtsmpanying figure is A Fenchel describes a microlquariunc
thier efore 'generalized, and is not intended '"° (1.5 X 54u) which may be observed under
, to be absolute. ,
a .compounernicroscope. (Figure 2y-

B Some. representative forms are listed for.' Be, The development of cpmrnunities of.
general inforination, The numbers - 'orkanisMs is, quits? similar-to the
correspondtO those,on the figur. Winogradsky Column..,(Figure 3)
. - o"

Inorganic substrate on towe/ing.-f.,

tit -

a.9.72 0 1. , 7:1
, Bacteria -aildIProtozoa as Toxicological Indifators

C The basic media consists of one liter of 3 Burbanck, W. D. and Spoon, D. M.

natural water 10 g CaSO4, k g glucose, The Use of Sessile Ciliates Collected
1 g of peptone utoclaved and stored at in Plastic Petri Dishes for Rapid
So C; Assessment of Water Pollution.
Jour, of Protozoology. 14(4):739-744.
\ Befo*re use agar is boiled with the media. 1967.
White hot7 the media is introduced into
one end of the "micr6 aquarium" with a 4 Curds, C.R. and Cockburn, A.
_pip et . _After. tiv agar _congeals, natural Protozoa in Biological Sewage Treat-
water samples are added. During , Pnent Processes. I. A Survey of the
incubation and When not being observed, Protozoan Fauna of BritiSh Percolating
the-micro aquaria are kept_in a moist . Filtermand Activated Sludge Plants.
chamber.. Water Research 4:224-236. 1970.
E Although Fenchel used a 'seawater medium 5 Curds, C.R. and Cockburn, A.
and inoculum, freshwater sources would II. Protozoa as Indicators in the
be equally useful. Activated Sludge Process. Water
Research 4:237-249. 1970.
F In these microaquaria simple ecosystems
develop when they are kept in complete. 6 Curds, C. R. An Illustrated Key to the
darkness. Complex photosynthetic British Freshwater Ciliated Protozoa
communities develop when they are Commonly Found in Activated Sludge.
illuminated, A natural ecosystem is Water Poll. Research Laboratory.
figured by FenChel (Figure 4), ald a Stevenage, England. 90 mi..
related food web is shown in Figure-5 .
(both from Fenchel). 7 Fenchelk Tom. The Ecology of Marine-- 4 "I

. . Microbenthom' IV. Structure and

G Microaquaria
e Using PAitic Petri Dishe's ..1i4=1._..onof the nenthic EeosAtem,
. its, Chemical and Phisical Factors
Sessile ciliates have been successfully and the`Micrpfauna Communities with
collected, cultured, and used for bioassay \ Special Reference to the Ciliated
'using the' same petri dish (membrane filter ' Protozoa, 01:Melia 6:1-182. 1969.*
type, with tight fitting lids).
, t

,. ,, rings
Botanical Gardens and
H. IVIicro*Aluartia,usindisilltorfe.cement , Hol-izong in Ald3.1.Rese,.arcIA. In Challenge
0,,*hilh Allow;diffiision of 'gases throtth the
for Survival. Pierre )5anseteau*ed.
-' pi/icohe: cultures win $12.erfbyflemp.in
active ipdelinirgy. 4 .G.
% .. s.

Columbia University Press. -1970. ,
. .

%tti ' , g 0 .
9-. Hutner, S.11. ; Baker, H. ;,031nColi, D.. .
. 4 . . ., ... Nutrition and Metabolism ro ozoa.
I ,
Chapter in ictlogy *pp: 85-177.
. - Adited
i rprganton.35*ress.
r Hutner, S.H. i?rotoxoa as.Toxicokgical 197.2. ' 15'

(roofs: The Jour. of Frotozoology. *". 4?* "*" .-*At :

11(1):1:6.' l9B4. :' ',, ft', ; 10 Hu S. H. The Urban.tiot°431ical:Giffae*A°
4, , ers A cadeknis Wildlife,.B..vesekirct. ,barden
2 Spoor; D. M. and.lierpanc It, , W. Dr n. 19L2V7,-40:
wpw 1969. 4;
A New Method X9r Collecting Sessile '4
Ciliates in Plastic Petri Dishes with
Tight - Fitting bids. Jour, of . Thisilyntlinevran prepared by Ralph M. Sindlair,
Protozoology J4(10:735:730. 1967.' 4quati4 Biologist, National Training Center,
- ,Water Pi'ograms Operations,,.EPA, Cincinnati,
01# a45268.-

,'7 -2 .95

Bacteria and Protgzoa Toxicolngical Indicators .

du 1


o. 4
... .
' 0 a %.
. tl ' ',40 A

' ' ^7. %




vtt t
., 411.
c ,

SUPERNATE , 7 sis -
41' a° 4 1 ' .` ' s. °
green .' CPCCOID GREEN' akbdA,E '.41/4 )V ! ... /
6- : ,tt / "
'''' .'

6 . 14*. ..; ..i e' - , +. A


I -iipt1.01041%

.. 6
C %
' A.

. . '


, 74. . =P.
r m'clge.n

- 04

gyttja .


, 4


Bacteria and Protozoa aS Toxicological Indicators'

t f


4 A micro aquarium fitted with electrodes and the redox conditions through 17 days.

, ,

/T c I ,---, . , j:,,..4.-- .., .,,,, ,, `2

o, '... i?":-.-,:i,i
.ezt',_'r/-A-,-4,1- r-
-4; ),) / ". is-'----,1
--,to 221 c"' :'t t %
V C. r---1.,V\ / t1,,,,..,-,,r,, - "---
" -......., (<
-----,__. ,, .2, ,-- 4,j,
/.-.....,-::-. :.--/'4.'d ,

- >1.
\ -";


brawing made by tracing a micrograpirof the bacterial plate in a micro aquarium
(same experiment as shown pn Figure t.) Most conspicuous are the filaments of
Beggiatoa and thg ciliates byclidium citrpllus, Euplotes elegans and Holosticha
Bp: Below the Oscillatoria filament (lower left) a Plagiogogon loricatus is seen.
Eafteria (exctpt Beggiatoa) 'arse not shoWn,
, .

Bacteria and Protozoa as Tokicolosica.1 Indicators


300 it

The microflora and fauna in the surface of the Beggiatoa patches. (Oscillatoria,
Beggiatoa, Thiovolum, diatoms, euglenoids, nematode, Tracheloraphis sp.,
Frontonia marina, Diophrys scutum, Trochiloides recta).

4.1ACaff LA S. ;ONO' (9STom S.

v. F...tv-7nus
etczettu LA.& Kt.tAlt.


etaramAti2 V
F ht),0*.4A
Li. T
n ClEvA '
5AMtatayjiczpauyi isSrf PO, sPe.
=4=S Lif

The food relationships ofthe most common ciliates in "estuarine" sediMents and in sulphureta.



L. A. Chambers, *Chairman
Bacteria:- Protozoa as Toxicological Indicators in Purifying Water
S. H. Hutner,, 'Herman Bakers S. Aaronson and A. C. Zahalsky+

There is a cynical a ge that all travelers The food -chain pyramids of sewage plants
become entomologi ts. But now with DDT and polluted waters haverbeen adequately
and detergents, t 4Velars and stay-at-homes described (Hynes, 196e; Hawkes, 1960).
_ alike are beco 4 g toxicologists. We have A problem treated here is hoW to scale those
an immediate in Brest in pollution problems: microcosms to experimentally manipulable
Our laboratory receiiies, like the Eait Riyer microcosms yielding dependable predictions
and adjoining United Nations buildings, a for the behavior of sewage-plant microcosms.
generous sootfall from a nearby power plant.
Also, we have'seen a superb fishing ground,
Jamaica Bay in New York City, become a THE WINOGRADSKY COLUMN AS 'SOURCE
sewer. (Jainaica Bay is, however, being OF INOCULA FOR MINERALIgATIONS AND
restored to its pristine cleanlines§but not AS TOXICOLOGICAL INDICATOR SYSTEM
the U. N. 'read. )" We take our'theme
neverthel ss not from aesthetics but from Total toxicity °depends on intrinsic toxicity- k
stateme s by Berger (1961): {1) It is an persistence relationship,. Techniques for
expensive, timq- consuming project". .. t testing the degradability, of organic compounds-
predict with confidence -a'new waste's and so their persistenoe in soil arilwater--
probable iinpact on certain important' down- are' still haphazard. The enrichment culture
! strewn water uses." And (2) "The technique, in which one seeks microbes that
toxicological.phase,octie study is p1rhaps use the compound in.question as sole substrate- -
its Mod perplexing aspect' The specialized hence degrades it and even "mineralizes"
Services and cost necessary for determining it - -was developed by the Dutch school of
the effect of repeated upoSure -to low con- microbiologists. Enrichment cultures are/
centrations of the waste forlong periods of- used routinely by biochemits wishing to.
dine would inevitably plaCe this job out of work out the microbial catabolic metabolic
reach of most public agencies. Equally pathway for a compound of biochemical
discouraging, ,perhaps, is the probability interest. Since the compounds dealt with by
that the, toxicological study may take as long' biochemists are of biological origin, micro-
as two years." bial degradability 'can be assumed. Still,
finding a microbe to degrade a rare biochemical
As describtd here, advances in niicrobiology, is not always easy: Dubos, in a classical
offer Hopes, of lightening this burden. The
first question is: What kind of microcosm
hunt for a microbe able to live off the capsules
of pneumococci, found the bacterium only,
wilAserve'for toxicological surveys, especially after a long search which ended in a cranberry
foripredictinithe pOisOning of biological means bog. Stkh.difficulty in finding mierobes that
of waste disposal? The second is: Cap the degrade raz biochernrcale implies an even
protozoa of this microcosm predict toxicity greater difficulty in finding microbes that
to highei aninials?,, degrade many products of the synthetic
*Iiirect6r,' Allan Hancock Foundation and Head, Bio. Dept.-, Univ. South. Calif.
+Haskins Laboratories, 305 E. 43rd St., New York 17, K. y.,; a/id Seton Hall College
of 1Viedidine and Dentistry, Jersey City, N.J. Pharmacological work from Haskins
LabdrafoPies discussed here vitas assisted by grant R6-9103, Div. of General Medical.
Sciences, of the Nationtillnstitutes of Health. Paperresented by Hutner.

't 1,

Environmental Requirements of Fresh-Water Invertebrates

organic chemicals indu"Stry, since such As-pointed out by our colleague, Dr. L. ,

compounds may embody4oiochemically rare Provasoli41961), the heterotrophic, ,

or biochemically nonexistent linkages,. capacities of algae are very imperfectly
Intimations of the frnpTaante of inoculum known. This is underscored by recent
abound in the literature, e.g., Ross and studies of the green flagellate Chlamydomonas
Sheppard (19561 could not at first obtain . mundane. as a dominant in sewage lagoons in
phenol oxidizerslfrom ordinary inocula the Imperial Valley of California (Eppley and
(presumably soil and water); but manure MaciasR,' 1962); other than that it prefers
and a trickling filter from a diemicar_plant______ acetate among the few substrates tried; its
proved abundant sources of active-bacteria.- \ heterotrophic capacities are unknown.
Onelvonders how extensive a study underlies \ More unexpectedly, some- strains of the
photosynthetic bacterium Rhodopseudomona
the statement mloted by Actixander (1961)tilat
Z "soils treated witI 2,4, 5-T (trichlorophenoxy- palustris use benzoic acid arferobically as
acetic acid) still retain the pesticide Meng the reduclant in photosynthesce--(SchF,_Scher,
after all vestiges of toxicity due to equivalent and Hiitner, 1962) narrowing the gap between
quantities of 2, 4-D have disappeared.',' the photosynthetic pseudomonads and the
ubiquitous pseu,domonada-so often represented ,

What then is a reasonable inoculum for testing among bacteria attacking resistant substrates
a compound's susceptibility to microbial (e.g.', hydrocarbons) as well as highly
...attack? The size range is wide: from the _vulnerable subtra s. For the widely
traditional crumb or gram of soil or mud to studied, strongly h erotrophic photosynthetic
the scow-load ot activated sludge.contributed flagellates Euglena. gracilia'aind E. viridis,./
by New York City to inaugurate the Yonkers, common in sewage, no speceic enrichment
..)_ sewage-disppsal plant. We suggest that to procedure is known, .meaning that their
' strike a practical mean in getting a profile ecological niches are unknon but labotatoiy
of Soil, mud, or sludge to bye used as inoculum data provide hints. , ,i
the uses of the Winogradsky column should be
explored. Directions for Winogradsky column The increasing use of oxidation ponds would
and bacteriological enrichments have been in any case urge a greater use of Sealed-
detailed (Hutner, 1962) and so only an outline' down ecological systems in which development
is given hei.e. The column is prepared by of none of the photosynthesizers present in
putting a paste of shredded:paper, CaCO3, the original inoculum was suppressed;
and CaS04jarat ine bottom of a hydrometer jar, Conceivably, some of the rare microbes
Ming the with mud smelling H2S, coveving attacklikg rare substrates - -and such micrObes
with a..shallow layer of water, and illuminating are likely to represent a sourcei of degiaders
from the side With an'incandescent lamp. In of resistant non-biochemicals--are specialists
2 or 3 weeks sharp zones, appear: a green- in attacking products of photosynthetic
and-black anaerobic zone at,the bottom organisms.
(a mixture of green photosynth 'tics bacteria
along with SO4-reducers, meth roducers, Traditionally, the inoculum for a Winogradsky
and the like); over tha0. red zonesk '....sf column is a ma.rincorgbrackish mud (as
(predominantly photosinthetic bacteria); New Yorkers we Would be partial to mud from
above this garnet or magenta spots or zone flats of the Harlem River). Little is known
(pre dominantly non- sulfur ,photo synthetic about the effectiveness as inocula of freshwater
bacteria); above this a layer rich in blue- muds or water-logged soils, It would be
green algae (the transition to the aerobic valuable to know how complete a cohimn
zones); above this aerobic bacteria along with '',coUld develop from material from a trickling
green algae, diatoms, other algae, and an filter or an activated-sludge plant.' A practical
-assortment of protozoa. This makes an iattue is: Might the poisoning of 'a sewagea
excellent siTple classroom experiment to oxidation system be paralleled, by the poisoning
defhonstrate the different kinds of photo- of a Winogradsky column}-whgre'the poison
synthetic organisnis, especially the bacterial Was Mixed with the inoculum for the column?
forms that are important in photosynthesis Might tile, variously 5t111 photosyntlietic-
research and that ordinarily escape notice zones of the column and the.'aerobic population
yet are ubiquitous in wet soils and natural on top procride sensitive indicators for the
waters. .
0 Environmental Requirements of Fresh -Water Invertebrates

,performance of a sewage-oxidation system The.anticonvulsant primidone provides a

subjected to chern,ical wastes? clekr indication of how protozoa can be used
to pinpoint the location of a metabolic lesion.
If a,particular compoundmixed with inoculation Primidpne had been known to cause folic acid-.
mud Fr sludge suppressed development of the responsive-anemias. It is therefore ea+to
lull Winogradsky pattern, one might assume find that with joint use of a thymine-dependent .

that the compound at the test concentration Escherichia coli and the flagellate Crithidia
was poisonous and persisteirt. Biological Nv/ fasciculata reversal of growth inhibition by
,destruction of such poisons, if at all possible, folic acid and related compounds permitted
might demand a long hunt for Suitable micro- the charting of interferences with the inter-
organisms, tHexi buildup of the culture to a connected folic acid, biopterin, and'DNA
pActical scale. This might best be done with function (Baker et al., 1962), which amply
illuminateti shake or aerated' cultures, with accounted for the megaloblasic anemias.
the inoculg coming from a variety of environ- Lactiohacithis casei, a bacterium much used
ments.. Optimism that microbes can be found in chemotherapeutic research, was unaffected
capable Of breaking almost all the linkages of b§ the drug.
organic chemistry is fostered by the study of
antibiotics, which include a wealth of In another instance, Where the mode pf action
previously "unphysiological" linkages--azo of the drug in higher animals was Unknown,
compounds, oximes, N-oxides, aliphatic and growth inhibition property of the anticancer
. aromatic nitro and halogen compounds, and compound 1-aminocyclopentane-l-carboxylic
strang9 heterocyclic ring systems. Some acid was. reyersed for Ochromonas danica
natural heterocycles, e. g,, pulcherriminic by L-alanine and glycine, as wag-th-e'rnhibition
acid and 2-n-nony1-4-hydroxyquinoline, have property of 1-amino-3-methyl-cyclohexane-
a disquieting resemblancelto the potent 1-carboxylic acid by L-leucine (Aaronson
carcinogen 4-nitroquinoline N-oxide. and Bensky, 1962).

Growth irklvdbition of Euglena bythe, potent

PROTOZOA AS TOXICOLOGICAL TOOLS carcinogen 4-nitroquinoline N-oxide was
annulled by a combination of tryptophan,.
A difficult pfoblerri is one mentioned earlier: tyrosine, nicotinic acid, phenylalanine,
persistence joined with lr-grade toxicity to uracil (Zahalsky et al, ; .,1962) and, in more
higher animals. Recent developments in the recent experiments, the vitamin K relative
use of protozoa as pharmacological tools show phthiocol. These N-oxides are of interest
that protozoa can serve as sensitive detectors because of recent work indicating that perhaps
of metabolic lesions ("side actions " ?) of a the main way in which:the body converts such
wide assortment of "safe" drugs. The list compounds as the amino hydrocarbons to the
includes the "antich,olesterol" triparanol actual carcinogens m4y be by an initial
(MER/29). TriParanol toxicity manifested by oxylation of the nitrogen, e. g., work
. itself with severalprotozoa, including b Miller et al., (19610. Whether the
Ochromonas danica (Aaronson et al., 1962) pe oxides in photo - chemical smogs of the
and Tetral1mm (Holz et al., 1962). Los geles type action hydrocarbons to
. not acting simply as an anti- produ e carcinogen' N-oxides is entirely
cholesterol for its obvious toxicity to protozoa unkno n. Leighton ( 961) ,lists, an array of
was annulled by fatty,acids as well as' by peroxy reactions pro uced by sunlight in
sterols. The connection between the protozoan polluted ail..
results and the-ttside actions" of triparanol--
baldness, impotence, and cataracts- -are of Our aforementioned experience with primidone,
course Unclear, but protozoal toxicity might a ketonic, heterocycle, le us to test the
serve as an initial warning That it might not sedative thalidomide. It as toxic for
,be as harmless as assumed from short-term Ochromonas danica O. m. emensis and
experiments with higher animals. Tetrahymenapyriformis; this toxicity was

ry 101
. 7-8
Environmental Requirements of Fresh-Water Invertebrtes

annulled by nicotinicacid (or nico inam de) Since the embryos appear to lack the
or vitamin K (menadione) (Frank et al, 1963). detoxication mechanisms of the adult animal
We do not know whether a similar protection (Brodie, 1962), toxicity for protozoa (which _-
could have been conferred on human embryos presumably lack these detoxication
or polynedritis in the adult. mechanisms) should be compared with that
fOr the embryo, not the adult, as emphasized
Many widely used herbicides of the dinitrophenol by the thalidomide disaster.
type are powttrful poisons for higher animals.
We do not know how sensitive protdzoa would There 'are further limitations on the use of
be Rix- detecting their persigtence. However, microbes as detectors of toxicity. High-
since somewhat similar thyro-,active com- molecular toxins seem inert to microbes,
pounds can be Sensitively detected by their- and antihormOnes (with the exception of
exaggeration of the B12 requirement of anti-thyroid compounds) are generally inert.
Ochromonas malhamensis (Baker of al, 1961), 'The main usefulness of microbial indices bf
this flagellate might be a useful test object toxicity would-appear, then, to be for detecting
for dinitrophenols and congeners. low-molecular poisons acting on cellilar
targets rather than on cell` systOms and
The PararneCium (ai-id perhaps too the. , organs. These are precisely the poisons
Tetrahymena) test for polynuclear benzenoid like to'put out of: business a pollution-
carcinogens has remarkable sensitivity and contr 1 installation primarily dependent on
spedificite (Epstein and Btirroughs, 1962; Hull, microbial activity.
1962): This test depends on the carcinogen-
sensitized phot9dynamic destruction of
paramecia by ultraviolet light. This test is REFERENCES fr
approaching practicality for air, and there is
.noTeason to suppose it cannot be applied to Aaronson, S. and Bens*, B. 1962.
;benzene extracts of foodstuffs and water. _protozoological: studies of the cellular
'action of drugs. I. Effect of
1-aminocyclopentane-1- carboxylic acid
CONCLUSIONS and 1-4amino -3 -methylcyclo-hexanel-1
carboxylic acid on the phytoflagellate
We have suggested here new procedures for Ochromonas danica. Biochem.` Pharmacol.
examining the intrinsic toxicity-persistence 11:983-6.
relationship, using as test organisms the, 4.
protists represented conspicuously in a Aaronson; S., Bensky, B., Shifrine, M. and
Winogradsky column. The new field of Baker, -H. 1962. Effect of hypotholes-
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This is not the place for a detailed discussion.., AleXander, M. 1961. "Introduction to Soil
of the medical implications of phis area of Microbiologir", John Wiley & Sons, N. Y. .'
research, but it should be eplphasiied that (see p. 240).
chronic toxicities and carcinogenesis are'
related. Conversely, Umezawa (1961) has Bhker, H., Frank, O., Hutrier, S. H.,
remarked that most antitumor subStance's Aaronson, S., Ziffer, H. and Sobotka,. 1t.
have chronic toxicities and that elaborate 1962. Lesions in foljc acid ,metabolism
testing procedures for toxicity are required induced by sprimidone 'Ekperientia
tq, fix the daily,tolerable dose; appa'rently this 18:224-6.
problem is a central theme in medical as well
as pollution research. Inhibition of girth of Baker, H., Frank, O., Pasher, , Ziffer, H.,
anarray of protozoa is now in practiCal use Hutner, S.H. and Sobotka, H, 1961.
Usi a means of detecting anticancer substances Growth inhibition of microorganisms by
in antibiotic beers (Johnson et al, 1962). thyroid hormones. Proc. Soc. Exptl. .

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1 f .
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Enviromtental Requirements of Fresh-Water Invertebrates

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P. 109-37. -
Hynes, H. B. N. 1960. The Biology of 13olltited "Biological Problems in Water Pollution"
Waters. Liverpool Univ. Press. (Third Seminar 1962) U.S. Public Health
Service-Pub. No. 999-WP-25, Cincinnati,
OH . 1965, II


1-10 110 3

-\ I /INTRODUCTION Transverse separations within the sheath

indicate that a row of 'cells is included in
There are a number of types of filamentous one sheath. The sheath may be clearly
bacteria that occur in the aquatic environment. visible or so slight that only special-staining
They include the sheathed sulfur and iron will indicate that it is present.
bacteria such as B_ esgiatoa, Crenothrix and
Sphaerotilus, the actinomycetes-which are The organism grovis as a ii,hite slimy or.
unicellular microorganisms that form chains felted cover on the, surface of various objects
of cells with special branchings, and undergoing decompdsition or on the surface
Gallionellae a unicellular organisrli that of stagnant areas of a stream receiving
secretes a long twisted ribbon-like stalk. sewage. It has also been observed.on the
These filamentous forms have at times base of a trickling filter and in contact
created serious problems in rivers, aerators.
reservoirs, wells, and water distribution
systems. It is most commonly found in sulfur strings
or polluted waters where H2S is present.
3eggiatoa is distinguished by its ability to
II BEGGIATOA deposit sulfur within its cells; the sulfur
deposits appear as large retractile globules.
Beggiatoa is a sheathed bacterium that grows (Figure 2)
as a long filamentous form. The flexible
filaments may beias-large as 25 microns wide
and 100 microns long: (Figure 1)
Figure 2
Filaments-of Beggiatoa
contaihing granules of

When H2S is no longer present in the environ-

ment, the sulfur deposits disappear.
Dr. Pringsheim of Germany has recently
proved that the organisin can groiv as a true
Stitotroph obtaining all its energy from the
oxidation of H 2S and using.this energy to fix , _
CO,,' into alkmaterial. It can also use
Begglatoa albk
certain t Materials.if they are present
along with. he H2S.

151.1 X Faust and k\rolfe, and Scotten and Stokes
have grownpe organism in pure culture. in
up to 1.000pi, this country'... Beigiatoa exhibits a motility
that is quite different,from the typical
flagellated motility of most bacteria; the
filaments have a flexible gliding motion.

,=,13.A.: 8a. 5.80 8-1

Filamentous Bacteria

The only major nuisance effect of Beggiatoa

known has been overgrowth on.trickling filters
receiving waste waters rich in H2S. The Figure 4
normalinicroflora of the filter was suppressed
and the filter failed to give good treatment. Egg albumin isolation plate.
Redi Oval of the H 2S from the water by blbwing 'A' an actinomycete colony.
air through the water before it reached the and 'B'.a bacterial colony
filters caused the slow decline of the
Beggiatoa and a recovery of the normal ,
microflora. Beggiatoa usually indicates
polluted conditions with the presence of H2S
rather than being a direct nuisance.

Actinomycetes are unicellular microorganisms, `11

lsmicron in diameter, filamentous, non- 4viof
sheathed, branching monopodially, and Appearance:
reproduced by fission or by meanstof special
coniqia.... (Figure 3) du II and powdery /smooth. and MUcOld
These organisMs may be considered as a
group intermediate between the fungi and
the bacteria. They require organic matter
for growth but can use a wide variety of
substances and are widely distributed.
Actinomycetes have been implicated as the
cause of earthy-odors in some drinlqg
waters (Romano and Safferman, Sily and.
Roach) and in earthy smelling substance has
been isolated from several members of the
Figure 3 Filamente. of Actlnomycetes. group by Gerber and Lechevalier. Safferman
and Morris have reported on a method for the
- "Isolation and Enumeration of Actinomycetes
Related toyWateeSupplies. " But the actino-
Their filamentous habit and method of mycetes are primarily soilinici-oorganisms
sporulation is reminiscent of fungi. However, and often grow in field's or on the banks of a
-their size, chemical cbrigiosition,4and other. river or lake used for the water supply.
characteristics are more similar to,bacteria. Although residual. chlorination will kill the
(Figure 4) organisms in the treatment plant or distribqion


. Filamentous 'Bacteria

system, the odors often are present-before

the water enters the plant. Use of perman-
ganate oxidation and activated carbon filters
shave been most successful of the methods
tried to rJmove the odors from the water.
control procedures to prevent the odorous
ma rial from being washed into the water
sup151 rains or to prevent possible develop-
men of t e actinomycetes in water rich in
deca ing organic matter is still'needed.


Crenothrix polyspora
The filamentous iron bacteria of the
cells are very variable in
Sehaerotilue- Lept othrix group, Crenothrix,
and Gallionella have the ability, to either size ttom small cocci or
oxidize manganous or ferrous ions to innic polyspores ta cells 3x12/.4
or ferricsalts-or are able to
precipitates of these compounds within the
sheaths of the organisms. Extensive growths
or accumulations of the empty, metallic
encrusted sheaths devoid of cells, have
created much trouble in well's or water dis-
tribution systems. Pumps and back-surge
valves have been clogged with masses of
material, taste and odor problems have
occurred, and rust colored masses of
material have spoiled products in contact
with water.
Crenothrix polyspora has only been examined
under the microscope as we have never been
able to grow it in the laboratory. The orga-
nism is 'easily recognized by its special
morphology. Dr. Wolfe of-the University-of
Illinois has published photomicrographs of
the organism. (Figure 5)
Orgapisms of the S haerotilus-Leptothrix
group have been extensivel studied by many
investigators (Dondero . al., Dondero,
Stokes,- Weitz and 1,0. ey, Mulder and van
,Veen, and Amberg an Cormack.) Under
jdiffereht ,environmental conditions the mok-
.phological se- ance of the organism varies,
, -The usual farm f in polluted streams or
bulke vated sludge is Sphaerotilus natans; Figure 6
(Fly re 6) Sphaerotllus natans

3-8 x 1.2 1.8fp

. cells

..10G. 8-3
Filamentous Bacteria

This is a sheathed bacterium consisting of ,

long, unbranched filaments', .whereby individual

rod- shaped bacterial cells are enclosed in a
linear order within the sheath. The individual
,cells. are .31 microns long and 1.2-1.8
microns wide. Sphaerotilus grows in great
masses; at times in streams or rivers that
receive wastes from pulp mills,' sugar
refineries; distilleries, slaughterhouses,
or ilk prOoessing plants. In these conditions,
it ap ars as large masses or tufts attached
to rocks, twigs, or other projections and -the
masses may vary In color from light grey to
,reddish brOwn. In sortie rivers large tnasses
of , Sphaerotilus break loose and clog watet
intake pipes or foul fishing nets. When the Figure 7
cells 'die, taste and odor proble4 may also Galionella furrugfnea
occur in the wa ,

OA X PI -1.1p
Amberg, Cormack, and Rivers and McKean Cells
have reported on methods to try to limit 'the
development of Sphaerotilus in rivers by
intermittant discharge of wastes. Adequate
control will probably only be achieved once REFERE ES
.the wastes are treated before discharge to
such an extent that the growth of Sphaerotilus
is no longer favored in the river. Sphaerotilus Beggiatoi.
grows welt at cool temperatures-and slightly
low DO levels in streams receiving these r- Faust, L. and Wolfe; R. $. Enrichment
wastes and domestic sewage. Growth is slow .*; and Cultivation of Beggiatoa Alba.
where the only nitrogen present is inorganic Jour. Bact., 81:99-106. 1961.
nitrogen; peptones and proteins are utilized 2 Scotterrr-H. L. and Stokes, J. L.
Isdlation and. Propertifs of Beiatoa.
Gallionella is an iron bacterium which appears Arch Fur. Microbiol.
. .
as a kidney-shaped cell with a 'twisted ribbon-
like stalk emanating from the concavity of the 3 -Kowallitk, U. and Pringsheim, .B. G.
cell. Gallionella obtains its energrby
oxidizing ferrous ironlnerric iron and uses The Oxidation of Hydrogen Sulfide by
only CO2 and inorg)cnic salts to forth all- of a Beggiatoa: Amer. Jour. of Botany:
the. cell material; it is an antotroph. Large. 53:801-805. 1966, .

masses of Gallionella may clause problems

in wells or accumulate in low-flow how=
pressure water mains. Super chlorination
Actinomycetes and Earthy Odors

(up..to-p0, ppm of sodium hypochlorite for ,.4 Silvey, J.,K. G: et al. A ctinomycetes and
48 hours) followed by flushingill often Common Tastes and Odors. JAWWA,
rer hove tile masses of growth and'periodic 42:1018-1026. 1950.
treatmen will preirentthe nuislince,effeCts 5 Safferman, R. 5; and Morris, M. E%
of the extensive masses1/4& Gallionella.
(Figure 7) A Method for the Isolation and,
Enunieration of ActinomyCetes Related
4 to Water Supplies. Robert A. Taft
Sanitary Engineering Center Tech.
Report W-62-10. 1962.


Filamentous Bacteria


.6 Gerber, N. N. and Lebhevaliert, H.A. 15 Donlero, N. C. Sphaeretilust Its Natiire

_j Geosmin, an Earthy-Smelling Substance and Economic Signifirianee. Advances
Appl. Microbiol. 3:717-107. 1961.
Isolated from Actinomycetee. Appl.
Microbiol. 13:935-938. 1965. ,
16 Mulder', E. G. akid van Veen, W. L.,
Filamektous Iron Bacteria Investigations on the SphaerotilUs
' Leptothrix Group. Antonie van
7 Wolfe, R. S. Cultivation, Morphology, and Leewenhoek 29:121-153. 963.
Classification of the Iron Bacteria.:
50 :1241 -1249. 1958.
r .-
Amberg, H.R. and Cormac , J. F. _
kip). .

Factors Affecting Slime Growth in
.E1 Kucera, S. and Wolfe, R. S. A Selective the Lowers Columbia 'River and
Enrichment Method foi Gallionella Evaluation of Some Possible ContrOl.
ferruginea. Jour. Bacteriol. 74 :344- Mea'sures. Pulp and Paper Mag. of
349. 1957. Canada. 61.;T70:-T80. 1960.
9 Wolfe, R. S. Observations and Studies 18 Amberg, H.R. , Cormabk, J. F. and
of Crenothrix polyspora. JAWWA, Rivers, M. R. Slifne. Growth Control
52:915-918. 1960. by Intermittant Discharge of Spent
Sulfite Liquor.. Tappi. 45':770-779.
10 Wolfe, R. S., Microbibl. Concentration 1962.
of Iron and Manganese in Water with .

Low Concentrations of theiefrElements. 19 McKeown, 4. J. - The Sonttol of

JpWAr. 521335=1337. 1860. Sphaerotii.us natans. Watchr
and Wastes, 47:137 19-22 and
11 Stokes,c4.,L. Studies on the Filamentous 8:(4)30-33'. 1963.
Sheathed Iron Baetiriurn Sphaferotilus
natans. Jour; Bacteriol. 67:278-291. -20 CtiPtis, ,L J. C:, Sco,a3e Fun-,:us;
41954. : Nature.and Effects. Wat. Res.
.3:289.4311. 1969,
1Z, 'Waltz, S. and Lackey; J. B. Kerphologicfti
and Biochemical Siudies on the _ 21 Lechevalier, Hubert A., P:ctinomycefes
OrIganismSphaerotilus natans. arf. of Sewage 'treatment Plants. Envir.
Joy. Fla. Acad. -Sci. 24(4)13 Prot. Tech. Series USEPA,
1958. '600/r-75-031. 1975.
" o I °

13' Dondero, N.C., Philips, R.A. anr . This outline was prepared by R. F. 'Lewis
Henkelkian, H. Isolatidn and Bacteriologist, Advanc aste Treatment
Preservation of Cultures of Sohaerotill ResearchLaboratory, NER,C, 'USEPA,
Appl. 9:2191-227% 1961. Ciricinniliz-OhiO 45268.

14 Eikelboom D. H. , Identification-QS__ Descriptors: Aquatic Bacteria, Sphs.erotilusl

Filamentous Organisms in Bulking Actinomycetes, Nocardia
Activated Sludge.- Frog. Water
Te0.,h. 8(8):155-161.' . 4
Pergamon Press.



Descri tion A Distribution

Fungi a e heterotrophicachylorophyllous
plant-like organiSms which possess-true Fungi are ubiquitous in nature and mem-
nuclei with nuclear membranes andnu- bers of all classes may occur in large
cleoli. Dependent upon the species and numb aquatic habitats. Sparrow
in some instances the environmental (1968) has riefly reviewed the ecology
conditions, the body of the fungus, the of fungi in reshwaters with particular
thallus, varies from a microscopic emphasis on the zoosporic phycomycetes.
single cell` to an extensive plasmodium The occurrence a.p.d ecology of fungi in
j .
or mycelium. Numerous forms produce r 'marine and estuarine waters has been
. ,macroscopic fruiting bodies. examined recently by a number of in-
vestigators (Johnson and Sparrow, 1961;
B Life Cycle Johnson, 1968; Myers,1968; van Uden
and Fell, 1968).
Thelife cycles of fungi vary from simple
to complex and may include sexual and
asexual stages with varying spore types B -Relation to Pollution
as the reproductive units'.
Wm; Br'cllxe Cooke, in a series of in;
C ClairSifica.tion -
vesti ions (Cooke, 1965), has estab-
, lished that fungi other than phycomycetes
Traditiorially, true fungi are classified _occur in high numbers 'in sewage and
within the Division Etnnycotina of the pollutedAters. His reports on organic
Phylum Mycota of the plant kingdom. pollution of streams (eooke, 1961; 1967)
Some authorities consider the fungi an show that the variety of the Deuteromy-
essentially monophyletic group distinct cete flora is decreased at the .immediate (
from the classical plant and animal sites a pollution, but dramatically in-
kingdoms. creaseddownstream from these regions.

U AC' IVITY Yeasts,- in particular, have been found

innarge numbers in organically enriched
I gen eral, -fungi possess:btoad enzymatic waters (Cooke, et al., 1960;- Cooke and.
apacities; Various species are able- to .Matsuura, 1963; 'Cooke, 1965b; Ahearn,
a.ctiOely, degrade such compounds as. et al., 1968). > Centain yeasts are of
complex polysaccharides (e. g., cellulose, special interest. due-to their potential°
chitin, and glycogen), proteins (casein, use as "indicator" organisms and their
albumin, keratin), hydrocarbons (kerosene) ability to degrade or utilize proteins,
and pesticides. 'Most species, possess an various hydrocarbons,. straight and
oxidative or microaerophilic metahq4111, branch, chained alkyl-benzene sulfonates,
bit anaerobic catabolism is not uncommon. Tats, .metaphosPhates, and wood sugars.
A fespeCie.s.iihow anaerobic metabolism
and growth.

BI. FU. 6a. 5. 80 109 9-1*


C "Sewage Fungus" Community (Plate. I) 2 Le om. itus lacteus also ,produces

ex ensi, e slimes and fouling flocs ,
A few microorganisms have Tong been In es water's. This speciesiorrns
termed "sewage fungi. " The mbst thal ified by regular constrictions.
common microorganisms includsidin
this group are the iron bcterk a Morphology
Sphaerotilus natans and the phycomy-
cete- Leptomitus' lacteus. Cellulin plugs may be present *
\ near the constrictions and there
1 Sphaerotilus natans is nota fungus;' may be nurrterous granules in
rather it'is a sheath bacteritlyn of the cytoplasm. The basal cell
the order -chlamydobacterialts. of the thallus may possess
This polymorphic bactersium occurs rhizoids.
.clommonly in organically enriched'
streams where it may produce b Reproduction
extensive slimes.
The segments delimited by the..
a Morphology partial constrictions are con-
verted basipetally to sporangia.
Characteristically, S. natans The zoospo'res are diplanetic
forms chains of rod shaped (i.e., dimorphic) and each
o cells (1. - 2.04 x 2.5 - 174). possesses one whiplash and one
within 'a clear sheath or tri- tinsel flagelluth. No sexual
clome composed of a protein- stage has been demonstrated
poly saccharidae-lipid complex. for this species.
The rod cells are frequently
motile upon release from the . c Distribution
sheath; the flagella are lophO-
trichous. Occasionally two For further information on the
row.s-of cells may be present distribution and systematics
in a single sheath. Single tri- of L. lacteus see Sparrow (1960),
chomes may be several mm Yerkes I1h66) and Emerson and
in lerigth and bent at various Weston (1967). Both S. natans
angles. Empty sheaths, ap- and L. lacteus appear to thrive -
pearirig like thin cellophane in organically enriched cold
straws, may be present. waters (59-22°C) and both seem
- $ incapable of extensive growth at
13' Attached growths temperatures of about 30°C.

The trichomes are cemented d Gross morphology

at one end to solid substrata
such as stone or metal, and Their metabolism is oxidative
their cross attachment and and growth of both specie ay
bending gives a .superficial appear as reddish brown flocs
similarity to true fungal hYphae. or stringy.slimes of 30 cm or
The ability to attach firmly to more in length.
solid substrates gives S. natans
a selective advantage in the e NUtritive requirements
population of flowing streams. Sphaerotilus natans is able to
For more thorough reviews of utilize a wide variety of organic
S. natans see Prigisheim (1949) compounds, whereas L. lacteus
and Stokes (1954). does not assimilate simple

Fungi .



(Attached "filamentous" and slime growths)

14. 3
Sphaetotilus natans

Begeatoa alba BACTERIA


Fusarium aqueductum Leptomitus lacteus


9 OpercUlaria


Fusarium aquaeductuum
(Radlmacher and
Rabenhorst) Saccardo Figure
Microconidia (A) produced Leptomitus lacteus (R
from phialides as in Cephalo- Agardh
IPOriUM. remaining in slime
balls. Mac:soma& (B), with Cells of theqy hae show.
one to several cross walls, ing_constrictionVivith cellulm
produced from collared phial. plugs. In ,one/cell large zoo.
ides. Drawn from culture. spores hive been deliraid.
. Redrawn from Coker, 1

Figure .3 Figure 4).-

Geouichum candulum Zoophagus in:idiom
Link ex Persoon Sommerstorff
Mycelium with short cells Mycelium with hyphal pegs
and arthrospores. Young by (A) on which rotifers will
pha (A) ; and mature arthro- become impaled; gemmae (B)
spores (B). Drawn from cul produced as conidis on short
ture. hyphal branches; and rotifer
impaled on hyphal peg (C)
from which hyphae have
grown into the rotifer whose
thell will be discarded after
the contents are duisumed.
Drawn from culture.

Figure S
nichlya americana Humphrey
Oeogonium with three oo-
spores (A); young rmosier- r wrier/dub rolm M11111111111
121011M with delimited Zoo-
spores (R) and zeosporangia
(C) with released zoospores
that remain encysted in dus-
ters at the month of the dis-
charge tube. Drawn from ma-

FIGUR); 7 Ilapinsporideum ccWale. spots;

Figures 1 th;ough 5 from Cpoke; FigUres 6 and 7 from Galtsoff.


sugars and grows most lu,iniriantly:in IV CLASSIFICATION

the pres1,-nce of organio nitrogenous 4,.
wastes. In recent classification schemes, classes
of fungi are distinguished primarily onthe
3 Ecological roles basis of the morphology of the sexual and
zoosporic stages. In practical schematics, '
Although the "sewage fUgg?"_olt however, numerous fungi do not dernonstVe
occasion attain visdally notic.eable these sta s, -.Classification must therefore
concentrations,,the less obitious be based nlhe sum total of the morphological
populations. of cfeuteromycetes may arid/or physiological characteristics. The
be more important in the ecology of extensive review by Cooke (1963) on methods
the aquatic habitat. Investigations of of isolation and classification of fungi from
the past decade indicate that numerous sewage and polluted waters precludes the
fungi and of primary importance in the need herein of extensive keys and species
mineralization of organic wastes; the -illustrations. A brief synopsis key of the
overall significance and exact roles of fungi adapted in part from Alexopholous
fungi in this process are yet to be (1962) j.s presented on the following pages.
D Predacious Fungi

1 Zoophagus insidians
(Plate H, Figure 4) has been observed
. to impair functioning of laboratory
activated sludge units (see Cooke and
'Ludzack). ,

2 Arthrobotrys is usually found along

with Zoophagus in laboratory activated
sludge units. This fungus is pretiacioua This outline was prepared by Dr. Dontld G.'
, upon nematode's. Loops rather than Ahearn, Professor of Biology, Georgia State
"pegs" are used in snaring nematodes. College; 'Atlanta, Georgia 30303.

Des Cr iPtbr41, Aquatic Fungi

PLATE II (Figure 4) -1



Definite cell walls lacking. somatic phase a free li;)ing Plasmod ium : :$41=n

Sub-phylum Myxomycotina . (true slime cnolcfs)..Class Myxomycete'

Cell walls usuallyiwell defined, somatic phase not a free-living Plasvmodikm. a

(true fungi) Sub-phylum Eumycotina ..... . .2 t.

2 Hyphal filaments usually coenoctytic, rarely septate, sex sElls when pciesent farming
oospores or zygospores, aquatic species propagating asexually by zoospores, terrestrial
species by zoospores, sporangiospores conidia or conidia-like sporangia .VPhycomycetes".: . 3

The phycomycetes, are generally considered to include the most primitive of the true
fungi. As a whole, they encompass S wide diversity of forms with some ,showing relation-
ships to the flagellates, while others closely resemble colorless algae, and still others
are true molds. The Vegetative body (thallus) maybe non-specialized and entirely con-
verted into a reproductive. organ (holocarpic), or it may bear tapering rhizoids, or be
mycelial and v?ry extensive. The outstanding characteristics of the thallus is a tendency
to be nonseptate and in most groups, multinuNiate; cross walls are laid down in vigorously
growing material only to delimit the rAfeorductive organs. The spore unit of nonsexual re-
production is borne in a siporangium, and, in aquatic and semiaquatic orders, is provided
with a single postdrioror anterior flagellum or two laterally attached ones. Sexual activity
in the phycomycetes characteristically results in the formation of resting spores.
2.t1'1 Hyphal filaments when present sefitate. without zoospores. with or without sporangia.
usually with conida; sexual reproduction absent pr culminating in the formation of asci
or basiaia 8

3 (2) Flagellated cells characteristically produced 4

3' Flagellated cells lacking or rarely produced .7

4 (3) Motile cells uniflagellate 5

. 4'. Motile cells biflagellate 6

5 (4) Zoospores posteriorly uniflagellate, formed inside the sporangium class .Chytridiomycetes

The Chytridiomycetesoroduce asexual zoospores with a single posterior whiplash

flagellum. The thallus is highly variable, the Most primitive forms are unicellular and
holocarpic and in their early stages of development are plasmodial (lack cell walls). more
advanced forms develop rhizoids and with further evollikonary progress develop mycelium
The principle chemical 'component of the cell wall is chitin, but cellulose is also present.
Chytrids are typically an atic organisms but may be found in other habitats. Some species
are re chitinolytic and/or k ratinolytic. Chytrids may be isolated from nature by baiting (et. g.
hemp seeds or-pine pollen) ytrids occur both in marine and fresh water habitats and are
of some economic importance due to their parasitism of algae and animals. The genus
Dermocystidium may be provisionally grouped with the chytrids. Species Of this genus
cause serious epidemics of oysters and marine and freshzater fish,
5' Zoospores anteriorly uniflage4late, formed inside or outside the spbrangium. class
Hyphochyt r iciiomycetes

These fungi are. aquatic-(fresh water or marine) Chytfid-like force whbtZe motile cells
posses; a single anterior flagellum of the tinseThYpe (feather-like). They are parasitic on
algae and fungi or may be saprobic. Cell walls contain chitin with some species also demon-
strating cellulosecontent. Little information is available on the biology of this class and
at present it is limited to less than 20 species.
a ', '
. ,re
claws... ...Oomycetes
' 6 (4!) Flagella nearly equal, poe whiplash the other tinsel
. .
A number of representatives of the Oomycete; cAve been shown tp have cellulosic cell
* rt
walls. The mycelium is coenocytic, branched and well doieloped in most cases, The sexual
process results in the formation of a resting spot4 of the.00gamous type, i:e., a type of
fertilization.in which two heterogametangia cqme fri contact'and fuse their contents through...,
a po4s or tube. Tke-thalli in this class r e from unicellular to profusely branched
filamentous types. Most forms arc eucarpi zoospores arc produced throughput the class
except in the more highly advanced species Certain species are of economic ithportance due
e, 'ec: their destruction of food crops (potatoes and grapes) while others cause serious diseases of
a fish (e. g. Saprolegina parasitical. Members of the family Saprolegniaceae are the common



water molds and are among the most ubiquitous fungi in nature, The order Lagenidta es
includes only a few species which are parasitic on algae, small'inimal, and other aqua is
life. The somatic structures of this taxon are holocarpic and endobiotic. The sewage f ngi4
are classified in'the order Leptomitales. Fungi of this order are ch:aracte rued by the
formation of retractile constrictions. cellulin plugs ' occur throughout the thalli or, at 1 ast,
at, the bases o( hyphae or to cut off reproductive structures. Leptomitus lacteus may
produce rather extensive fouling floes or slimes in organically enriched waters.

Flagella,of unequal size, both whiplash ...... class....Pfasmodiophoromycetes

Members of this cla e obligate endoparasites of vascular plants, algae. and fungi
The thalls consists of a pla chum which develops within the host cells. Nuclear division
at some stages of the life cycle of a type found in no other fungi but known to occur in
protozoa. Zoosporangia which ari e directly from the plasmodium bear zoospores with two"
unequal anterior falgella. The cell waifs of the fungi apparently lack cellulose.
7 13') Mainly .sapro,bic. sex cell when present a zygospore.. ..... class . 7%vomycetes

This class has well developed mycelium with septa developed in portions of the
older hyphae, actively growing hyphae are normally non-septate. The asexual spore,. at E
non-motile sporangioipores (aplanospore,$). Such spores lack flagella and are usually
aerialy disseminated. Sexual reproduction is initiated by the fusion of two gametangia
with resultant formation of a thick-walled, resting spore, the zmospore. In the more
advanced species. the sporangia or the sporangiospores are conidia -like Many of the ,
Zygomycetes are of economic importance due to their ability to synthesize commercially
valuable organic acids and alcofiols. to transform steroids such as cortisone, and to
parasitize and destroy food crops. A few species are capable of causing disease in man
avid animals (zygomycosis)
Obligate commensals of arthropods, zygospores usually lacking chss ...Trichomycetes
The Trichomycetes are an ill-studied group of fudgi which appeao be obligate
commensals of arthropods. The trichomycete are associated with a wide variety of insecta.
2111iplopods, and crirstacea of terrestrial and aquitic (fresh and marine) habitats. None of
the members of this class have been cultured in vitro for continued periods of times with any
success. Asexual reproduction is by means of sporangiospores Zygospures have been
observed in species of several orders.
8.(2'1 Sexual spores borne in asci class ..fikscOmycetes

In the Ascomycetes the products'of meiosis; 'the ascospores, are borree in sac
.like structures termed asci. The ascus usually contains eight ascospores. but the number
produced may vary,with the species or strain. Most sp6cles produce extensive sepiate
mycelium. This large class is divided into two subclasses on the presence or absence
of an.ascocarp. The Hemiasconcetidae lack arLascocarp and do not produce ascogenous
hy'phae; this subclass includes the true yeasts. The Euascomycetidae usually are divided
into'three series (Plectomycetes, Pyrenomycetes, and Discomycetes) on the basis of
ascocarp structure.
8' Sexual spores borne on basidia .
The Basidiomycetes generally are considered the most highly evolved of the fungi.
Karyogamy and meiosis occur in the basidium which bears sexual exogenous spores,
basidiospores. The mushrooms. toadstools, rusts, and smuts are included in this class.
_Sexual stage.14cking . .Form clarri.(Fungilmperfecti).Deuteromycetes
The Deuteromycetes is a form class for those fungi (with orphological affinities
to the Ascomycetes or Basidiomycetes) which have,not demon trated a sexual stage.
The generally employed classification scheme for these fung is based on the morphology iY
'and color of the asexual reproductive etages. This scheme is briefly outlined be'low
Nehver concepts of the classification based on conidium development after the classical
atoirk_of S. J. Hughes (1953) may eventually replace the gross morphology system (see
Barron 1968).



1 . Reproduction by means of conidia, oidia, or by budding 2
1' No reproductive structures present Mycelia Sterilia
2 (1) Reproduction by means of conidia borne in pycnidia Sphae rolisidales
2' Conidia, when formed, not in cycnidia 3 ,
'3 (2') Conidia borne in acervuli t;',. ,. , . Melanconiales 1

3' Conidia borne otherwise, or rep'roduction by oidia or by budding, Moniliales


1 Reproduction mainly by unicellular budding, yekst-like; mycelia' phase, if present,

secondary, arthrospores occasionally produced, manifest nielanin pigmentation lacking 2
Manus mainly filamentous; dark melanin pigments sometimes produced 3

2 (1) Ballistospores produced .'Sporobolornycetceae

2' No ballistospores Cryptococeaceae
3 COnidiophore*, if present, not united into 8porodochia or synnernata 4
3' Sporodochia present Tuberculariaceae
3" Synnemata present Stilbellaceae

4 (3) Conidia and conidiophores or oidia hyaline or brightly colored Monies liacCae
4' Conidia and/or conitliophores, containing dark melanin pigment Dematiaceae

O 4


Fungi and the Sewage Fungus Community'


SELECTED REFERENCES , Stagnant. waters. I. Morphology and

I. Occurrence in Nature. Arher. J.
Ahearn, D. G., Roth,F. J. Jr . ; Meyers; S.P. Botany '54:702-719." 1967 t.
Ecology and Charact erization Of Yeasts
from Aquatic Regions of South Florida. Hughes, S. J. Conidiophores, Conidia and
Marine Biology. 1:291-308. 1968 Classification. Can. J. Bot. 3 r:577 -
659. 1953
Alexopoulos J. C.' Introductory Mycology.
2nd ed. John Wileyand Sons, New York, Johnson, T.W., Jr. Saprobic Marine Fungi.
613 pp. 1962 In Ainsworth, G. C. and
Sussman, A. The Fungi, III.
Harron, G. L. The Genera of Hyphomycetes _Academic Press, New York. 1968
frbm Soil. Williams and Wilkins Co., -
Baltimore. 364 pp. 1968 and Sparrow, F. K., Jr. Fungi
Cooke, W. B. Population Effects on the in ceans and Estuaries. Weinheim,
Fungus Population of a Stream.
Germany. 668 pp. 1961
Ecology 42:1-18. 1961 Meyers, .S.P. Observations on the Physio-7
'1 logicalltcology of Marine Fungi. Bull.
. A Laboratory Guide to Fungi in Misaki Mr. Biol. Inst. 12:207-.228.'1968
Polluted Waters, Sewage, and Sewage
Treatmept Systems. U. S. Dept: of .
E.G. Iron Bacteria. Biol. Revs.
Health, Education and Welfare, Cincinnati, Cambridge Phil: Soc. 24:200-245. 1949
132 pp. 1963 1
Sparrow, F. K., Jr. A quatic Phypomycetes.
Fungi in Sludge Digesters. 2nd'ed. lUniv. Mich. press, AnnArbor.
\ Purdue Univ. Proc. 20th Industrial 1187 pp: 1960.
Waste Conference, pp 6-17.. 1965a
. Ecology of Fr_Fshwatertungi:
4The Enumeration oreast pp. 41 -93k.Ainsworth G.C. and
Populations in a Sewage Treatment' }dart._ SusSman, A.S. The Fungi., III. Acad
Mycologia 57:6 6-103. 1965b Jress,'New York. 1968'
. Fung 1 Populations in. Relation
Stokes, J.L. Studies on thetFilamentous
to Pollution of he Bear River,:ldaho-Utah. Sheathed Iron BaCterium Sphaerotilus
Utah Acad. Pr c. 44(1):298-315. 1967, natans. J. Bacterio1.67:278-291. 1954
and Matsuura, George S. A Study van'Uden, N. a,rid Fell, J.W. Marine Yeasts.
of Yeast Populations in a Waste Stabilizatipn 167 -201. In Droop, M.R. and Wood,
Pond System. protoplasm 57:163-187.. E. J_. F. Advan-cealn-Microbiology of
the Sea, Academic Press, New York.
, Phaff, Miller, M.W.,
Shifrine, M., and Knapp, E. Yeasts Yerkes, D. Observations on an Occurrence
in Polluted ater and Sewage: of-Leptomitus lacteus in Wisconsin.
Mycologia 5:210 -230. 1960 Mycorogis. 58:976-978. 1966
EmersonTlialpTrandVe'ston,. W; H. 4.,Cooke-,---William B. and Ludzack, F. J.
:Aqualinderella fermentans Gen. et Sp. Predac s Behavior in
Nov., A PhyComyceie Adapted to c hated Sludge Systems. Jour. Water
Poll. Cont. Fed.s30(12):1490-1495. 1958.
. Fungi and the Sewage Fungus Corn pity


Curtis, E. J. C. Review Paper'Sewage

Fungus:_ Its Nature and Effects.
Wet. Res. B:289.-311 1969.
. Curtis, E. J. C. and, Curds, C. R.
Sewage Fungus in Rivers in the
United Kingdom: The Slime
Community and It&Consituent O
Organisms. Wafer Res,'
5:1147-1159. 1971.



I GENERAL CONSLDE4ATIONS considered as indigenous to natural

waters. Sulfur and iron bacteria are
A Microbial quality ?onstitutes only one more common in the bottom mud.
aspect of water sanitation; microchemicals
and radionuclides are attracting increasing C,Actinomycetes,' Bacillus sp. Aeroares
amount of attenticn sp., and nitrogen-fixation bacteria are
primarily soil dwellers and may be washed
bi Microbes considered:here include bacteria, into the 'water by runoffs.
protozoa, and microscopic tnetazoa; algae a
and fungi excluded.. E Nematodes are usually of aerobic sewage
treatment origin..
C Of the free-living fOrmi, some are
members of the flora and fauna, of surface D E. coli, streptococci, and Cl..perfr iugea
waters; others washed into the water from are True indicators of fecarpollyon..
air and soil; still others of wastewater
origin; nematodes most commonly from III PROTOZOA
sewage effluent.
A Classification
D Hard to separate "native" from "foreign"
free-litring 'microbes, due to close 1 Single- cell animals in the most
association of water with soil and other primitive phylum (Protozoa) in the
environments; generally speaking, bacteria animal kingdom.i
adapted to water are those that can grow
on very low concentrations of nutrient
and zoomicrobes adapted to water are
2 A separate kingdom, Protista, Aci in-
elide protozoa, algae, 'fungi, and
those that feed on algae, and nematodes, bacteria proposed in thp 2nd edition'
especially bacteria eaters, are uncommon ofWard-Whipple's Fresh-Water
2.3.191210,) 10)
water but in large numbers in sewage
3 Four subphyla or classes:

E More species and lower densities of MattigdPhora (flagellates)- Subclass

.mic`robes in clean water and fewer species Phytomastigina dealt with under
and higher densities in polluted water. algae; only subclass Zoomastig a
F Pollution-tolerance or nontolerance of included here; 4 orders:'
microbes closely related to the DO level, 1) Rhizdmastigina - with flagell m
required in respiration. or flagella and psetidopodia
G From pollution viewpoint, the following 2) Protomonadine. -. With 1 to 2
groups of microbes ate of importance: flagelmostly free-living man
Bacteria, Protozoa, Nemateda, and apamsttic
Rotifera; 3) 'Polymastigina - with 3.to 8
flagella;..mostly parasitic In
II BACTERIA , elementary tract 9f animals
and man
A No ideal,method for studying distribution
. _
and ecology of bacteria in freshwater,
4) Hyperinaktigina - 1 inh itants
, of-alimentary 'tract o sects.
B According to Collins,
Achrombacter, Alcaliguies Chromobac- .

Tterium, Fla..7/212usteriurn, and,Micrococcus

- '", are the most widely distributed and may be 9:

. W. BA. 45c. 5.80 10-1


Protozoa, Nematodes) anti Rotifers

b Ciliophora-or Infusoria (ciliates) present in many for feeding. .

no pigmented members; 2 classes;

1) Ciliate - cilia present during they_, 2 Ciliophora:
whole trophic life; containing
majority of the ciliates Most highly' developed protozoa; with
few exceptions, 'a macro and a micro-
2) Suctoria - cilia present while ._ nucleus; adoral zone of membranellae,
young and tentacles during p-ophic -F mouth, .and groove usually present in
swimmingand crawling forms, some
with conspicuous ciliation of a disc-like
Anterior region and little or no body
c Sarcodina (amoebae) - Pseudopodia cilia (stalked and shelled forMs);
(faise feet) for locomotion andfood- Suctoria nonmotile (attached) and with-
capturingi 2 subclasses: out cytostome cysts formed in most.

1) Rhizopoda - Pseudopodia without

axial filaments; 5 orders: Sarcodine:

a) Proteornyxa - with radiating Cytoplasmic membrane but no cell wall;

pseudopodia; without test or endoplasm and ectoplasm distinct or i-,
shell distinct; nucleus with small or farge
nucleolus; some with test or shell;
b) Mycetozo f ming plasmodium; eying by protruding pseudopodia; few
resembilin in sporangium apabIe of flagella transformation; fresh-
formatIon water actinopods usually sperical with
many radiating axopodia; some Testacea
c) Amoebina - true amoeba containing symbiotic algae and mistaken.
forming lobopodis2 N. for pigmented amoebae; cysts with single
\or double wall and 1 or 2 nuclei.
d) Testacea amoeba with single
test or shell of chitinaus
material 4 Sporozoa: to be mentioned later'.

e) Foraminifera amoeba with 1 ...

or more shells of calcareous C General Physiology

nature; practically all marine 1 Zoomastigina:
Free-living forms normally holozic;
.food supply 'mostly-bacteria in growth 's

d Sporozoa - no organ of locomotion; film on surfaces'or clumps relatively

amoeboid in asexual-ohase; all aerobic, th -refore the first protozoa to
parasitic disappear -in naerobic conditions and
re-appearing at recovery; reproduction
. by sim-Aelission or occasionally by
B General' Morphology budding. ',/,

1 Zoomastigina:'
2 Ciliophora:
.Relatively small size (5 to 40 p);*with
- the exceptionOUthizomastigina, the Holozoic; trt# ciliate's concentrating
body has a definite shape (oval, leaf- food particles by ciliary movement.
like, pear-like, .etc.); common members around the mouth part; suctoria sucking
with 1 or 2 flagella and some ,with 3, 4, through teycles; bacteria and small -
or more; few formpit colonies; ..;:torne

120 1
Protozoa, Nematodes, and Rotifers

algae and protozoecon titute main 3 Tylenchida - Stylet in-mouth; mostly

food under natural condi 'ons; so plant parasites; some feed on
shown in laboratory to th ixe otyflead nematodes, such as Aphelenchoides.
organic matter and serum protein; not as
aerobic as flagellates -.some surviving 4 Rhabditida - No stylet in Mouth or caudal
under highly, anaerobic conditions, such glands in tail; mostly bacteria-feeders;
as Metopus; reproduction by simple common genera: Diplogaster,
fission, conjugation or encystment. MT.hoides
DiplospAte...:2ALest MRhabiditist Peisferat
1.fa.12..a.......ellus, and Turbatrix.
( 3 Sarcodina: 5' Dorylaimida - 'Relatively large nematodes;
stylet in mouth; feeding on other nematodes,
Holozoic; feeding through engulfing by algae and probably zoomicrobes; Dorylaimus
pseudopodia; food essentially same as common genus.
for ciliates; DO requirement somewhat
similar to Ciliates - the small amoebae 6 Chrom dorida - Many marine forms;
and Testacea frequently present in large some f, e'shwater dwellers feeding on
numbers in sewage effluent and polluted algae; ch acterized b strong orna-
water; reproduction by simple fission tio of knobs, bristles or
and encystation. !punc1 atio s in cuticle.
M nhysteridi. - Freshwater dwellers;
IV NEMATODES e ophago-intestinal valve spherical to
elongated; ovaries, single or paired,
A Classification usually straight; common genus in
water - Monhystera.
1 All in'the phylumffiemata (non'egment-
ed round worms); subdivided by some 8 Enoplida - Head usually with a number
authors into two classes: of setae; Cobb reported one genus, .
Mononchulus, in sand filters in
Secernentea - 3 orders: Washington, D. C..
Tylenchida, Rhabditida, Strongylida, B General Morphology
a Teratocephalida; with papillae on
male tail, caiudal glands absent. Round, slender, n nsegmented (transverse
markings. in.cuticle of some) worms;
some small (about a mm long, as Tri-
o cephalobus), ma 1 to 2 mm long
Adenophora - 6 orders: (Rhabditis, Dinlogaster, and tiplogasteriodee
for instance), and some large (2 to 7. nnn,
( aphasmids) such as Dorylaimus)rsex separated but few
Araeolaimida, Dorylaimida, parthenogeristic complete alimentary; canal; -
Chromdorida, Monhysterida, Enoplida, with elaborate mouth parts with or without__
and Trichosyringida no papillae on stylet; complete. reproductive system in
male caudal glands absent. each sex; no circulatory or restiratory
system; complex nervous system with
conspicuous nerve ring across oesophagus.
2 Orders encountered in water and sewage
treatment - Free-likring fot6s inhabitat- AK,

ing. sewage treatment plants are usually C General Physiology

bacteria-feeders and those,feeding on
other nematodes; those inhabitating clean 1 Feeding - Most sewage treatment plant
'waters feeding on plant matters; they dwellers feeding bn bacteria; others
fall into the following orders: preying on protozoa, nematodes, .rotifers,

Protozoa, Nematodes, and Rotifers

. etc., clean-water species apgarently Keratella, Monostyla, richocerca,

vegetarians; those with stylet in mouth Asplanchna, Polyarthrta, Synchaeta,
use the latter to pierce the body of/anithal- Midrocodon; common genera under the
or plant and suck contents; metabolic order Ffosculariaceae: Floscularia,
wadte mostly liquid containing ammoniiun and Atrochus. Common genera under
c rbonate or bicarbonate; enteric order Melicertida: Limnias and
pa ogens swallowed randomly with Conochilus.
suspending fluid, hence remote possix 5 Unfortunately orders and families( of
taffy of sewage effluent-borne nematodes A

being pathogen-carriers. rotifers partly based on character of -

corona and trophi (chewing organ),
which are difficult to study, -esp. the
2 ,OxYge n requirement - DO, amiarently latter; the foot and cuticle much ea.4ier
,to study.
diffused through cuticle into body; DO
requirement somewhat simgar to General Morphology and Physiology
protozoa; Rhabditis tolerating reduced
DO better than other Rhabditida members;, 1 Body weakly differentiated into head,.
all disappear under sepsis in liquid; some neck, trunk,- and foot, separated by
thrive in drying sludge. ,
folds; in some, these regions are
3 Reproduction Normal life cycle requires merely gradual changes in diameter
mating, egg with embryo formation, of body and without a separate neck;
- hatching of eggs inside or outside femals, Segmentation external only.
4-larval Stages, and adult; few repro-
duce in the Operice of males. , 2 Head withccirona, dosal antenna, and
ventral mouth; mastax, a chewing organ,
located in head and neck, connected to /
V ROTIFERS - mouttLanteriorly by a ciliated/gullet an
posteriorly to a large stoma.01 occupy g
A Classificatiolii ,mucli of the trunk..
- 1 Classified either as a crass of the 'phylum 3 Common rotifers reproducing partheno-
Aschelminthes (various forms of worms) genetically by diploid eggs; eggs laid in
or as a separatel phylum (fiotifera); cbm-, water, cemented to plants, or carried
monly called wfirelanimalcules, on on female until hatching. b
account df apparent circular movement of
cilia arbund head (corona); corona con- 4 Foot, a prolongation of body, usually
tracted when 244a4ing or swimming and 'with 2 toes; some with one toe; some °
expan. dedwithenched to catch food. with one toe and an extra toe-like
_structure (dorsal spur).
2 Of the 3 classes,-.2 (Selsooidea and
Bdelloidea) rollped by some authors ome, like Phlilodina, concentrating
under Digon8nta (i °Varies) and the bacteria ariciother microbes and minute
other beibOlonogimont (1 ovary); particulate organic matter by ciliary
eisonideacontaining rn stly marine movement on corona larger-microbes
forms. /- chewed by mastax; some such ors
Monostyla feeding on clumped matter,
3 Class Digononta containing 1 order such as Cacterial growth, fungal massed,
(Bdelloida) with 4. families, 'Philodinedae- -- etc. at bottom; virus generally not
being the most important. ingested - apparently undetected by
4 °Class Monogononta comprising 3 ordetst-:'
Notommatida (mouth not near cente -Of 6 DO requirement somewhat simil ar to
corona) with 14 families, Floscularida protozoailome disappe ring under
Melicertida (corona with two wreaths of reduced DO, others, lik Philodina,
cilia and furrow between them) with 3 .
surviving at as little as ppm DO,
families; most import .genera included
in the order Notommatida: Brachionus,
eia0111=0. ProtozolLIImatodes.1_and Rotifers

VI SANITARY SIGNIFICANCE B Protozoa and rotifers 4 should be included

in examination for planktonic microbes.
A Pollution tolerant and pollution non-
toledigat species - hard to differentiate - C Nematodes
re/411Eing specialist training in protozoa,
- nematodes, and rotifers. D Laboratory Apparatus(3)
B Significant quantitative difference in clean 1 Sample Battles - One-galron glass or
and polluted waters - clean waters con- plastic bottles with metal or plastic
taining large variety of genera and species screw caps, thoroughly washed and
but quite low in densities. rinsed three times with distilled water.
C Aerobic sewage treatment processes 2 Capiplor Pipettes and Rubber Bulbs -
(trickling filters and activated sludge Long (9 in.) Paaeur capillary pipettes
'processes, even primary settling) ideal and rubber bulbs of 2 ml capacity.
breeding grounds for those that feed on
bacteria, fungi, and mintztelLotozoa and 3 Filtration Unit - Any filter holder
present in very large numbers; effluents assembly usetd bacteriological
from-sticifiCocesses carrying large num- examination. 'The funnel should be
bers of the-fie zoomicrobes; natural waters at least 650 ml and the filter flask at
receiving such effluents show.ing significant least 2 liter capacity.
increase in all 3 categories.
j 4. Filter Membranes - Millepore SS (SS
D Possible Pathogen and Pathogeri Carriers iiirgiM) type membranes or equivalent.
1 Naeleria causing swimming associated 5 MicrOsc..._me - Binocular microscope,
meningoencephalitis and Acanthameoba with 10X eyepiece, 4X, 10X, and 43X
causing,nonswimming associated cases. objectives, and mechanical stage.
2 Amoebae. and nematodes grown E Collection of Water Samples
pathogenic enteric bacteria in lab;.tione
alive in amoebic cysts; very few alive Samples are collected in the same mannero
in nematodes after 2 days after ingestion; as those for bacteriological examination,
,virus demonstrated in nematodes only except that a dechlorinating agent is not:
when very high virus concentrations... 41* needed. One-half to one gallon samples are
present; some"TreeliNiing amoebae collected from raw water and one-gallon
palasitizing humans. samples from lap water. Refrigeration is
not essential and samples may be transported
5\ Swimming ciliates and some rotifers without it unless examination is to be delayed
(Concentrating food by corona) ingesting for more than'five days.
large numbers of pathogenic enertic
bacteria, but digestioh rapid; no . F Concentration of Sainples
evidence of concentrating virus; crawling
ciliates and flagellateei feeding on- clumped 1 One gallon of tap water can usually be
organisms,' , filtered through a single 8-u membrane
within 15 minutes unless the water has
,4 Nematodes concentrated from sewage high turbidity: -"At least one gallon of
effluent in Cincinnati area showing sample should be used in 'a single examina-
live E. coli and streptococci, but not tion. Immediately after the last of the
human enertic pathogens. water is disappearing from the membrane,
the suction line is disconnected and the
VII EXAMINATION OF WATER FOR MICROBES membrane placed on the wall of a clean
50 to 100 ml beaker and flushed repeatedly
A Bacteria - not dealt here. with about 2-5 ml of sterile distilled water

Pr'otozoa, Nemato4s and Rotifers

with the aid of acapillary pipette and a sluggishly motile nematodes may be
rubber bulb. The concentrate is then confused with root fibers, plant fila-
pipetted into a clean Sedgewick-Rafter ments of various types,' elongated
Counting Cell and is ready for examina- ciliates such as Homalozooji vernii-
tion. culare, or segments of appendages_of
small crustacea. To facilitate a,
2 1p concentration of raw water samples geAral identification of nematodes, the
having visible turbidity, tem to :our gross morphology of three of the free-
87rriicron membrapes. may be required living nematodes that are frequently
per sample, with filtration through each found in water supplies is sh9wn in the
mellibrane being limited to not more attached drawing. The drawing provides
than 30 minutes. Samples ranging from not only the gene ral anatomy for recogni-
500 ml to 21iters may be filtered With tion of nematodes but also most of the
one membrane, depending on degree of essential structures for guidance to those
turbidity. After filtration the membranes who want to use the "Key to Genera" in
are placed M the walls of separated chapter No. 15 on Nemata by B. G.
beakers and washed as above. To Chitwood and M. W. Allen in the book,
prevent the particulates from obscuring Fresh Water Biorogy. (10)
the nematodes, the washing from each
filter is examinedsin a separate counting 2 Under normal conditions, practically
chamber. all nematodes seen in samples of
finished water are in various larval
G 'Direct Microscopic Examination, stages and will range from 100 to 500
L, microneiri length and 10 to 40 microns
Each counting chamber containing the in width. Except in the fourth (last)
filter concentrate is first examined under stage, the larvae have no sexual organs
a 4X objective. Unless the concentrate but show other structural characteristics.
contains more than 100 wormso they whole
Fell area is surveyed for nematodes, with 3 If identification of genera -i's desired,
respect to number, develbpmental stage, the filter washings are 'centrifuged at
and motility. When an object having an 500 rpm for a few minutes. The
outline. resembling that of a nematode is supernate id-discarded, excapt a few
observe it is re; examined under a ipx drops, and the sediment is resuspended
objective for anatomical structures, ukless in the remaining water. A drop of the
the object exhibitg typical nematode move- final suspension is examined under both
ment, which is sufficient' for identifying the 10X and 43X objectives for anatomical
object as a nematode. When the concentrate characteristics, without staining, and for
contains more than 100 worms, the worm supplementary study of structures the
density can.be estimated by counting the rest is fixed in 5% formarin or other
number ofworms in representative Micro- fixation fluid and stained according to
scopic fields and multiplying the average instructions given in Chitwood and
number of worms per field by the number ,Chapter 'on Nemata, (7):
of fields in the cell'trea. The nematode Goodey's Soil and Freshwater Nema-
density may be expressed as number of todes(11) or other bboks on nematology.
worms, per gallon with or without differenti-
ation as to adult or larval stages or as to

H' General Identificatiotrof Nernt.todes A Idea not new, protozoa suggested long ago;
many considered impractical because of
1 While actively motile nematodes can be the need of identifying pollution - intolerant
readily recognized by any person who and poIlution-tolerant specie's.- proto-
has some general concept of micro- zoologist required. Method'also time
scopic animals, the nonmotile or consuming. . .


Protozoa; Nematodes, and Rotifers

B Can use them on a quantitative basis FLAGELLATA

nematodes, and nonpigmented Bodo caudatus
protozoa present in small numbers in
clean water. Numbers greatly increased Pleuromonas jaculans
when polluted with effluent from aerobic Oikomonas termo
treatment -plant or recovering from sewage
pollution; no significanrerror introduced Cercomonas longicauda
"when clean-water members included in the Peranema trichophorum
enumeration if a suitable method of com-
puting the pollution index developed.
Swimming type
C Most practical method involves the Ciliophora:
equatibn; A + B + 1000 C = Z. P. I. ,
A Colpidium colpoda
where Colpoda cuculus
A = number of pigmented protozoa,
B = non pigmented protozoa, and Glaucoma pyriformis ,r0
C = nematodes in a unit volume of sample, Paramecium candatum; P bursaria
azd Z, P. I. = zoological pollution index.
For relatively clean water, the value of Stalked type
Z.P.I. close to 1; the larger the value
above 1, the greater the pollution by aerobic a2rcularia sp. (short stalk dichotomous)
'effluent (see attached report on zoomicrobial
indicator of water pollution). Vorticella sp. (stalk single and contractile)
CONTROL aistylis plicatilis (like opercularia, more
colonial, stalk not contractile)
A _-Chlorination of effluent
Carchesium sp. (like vorticella bat colonial,
B Prolongation of detention time of of individual zooids contractile)

C Elimination of slow sand filters in foothamnium sp., I entire colony contracts)

nematode control.
Crawling type
PROCESS Euplotes patella
Stylonychia mylitus
PROTOZOA UrOstyla sp.
Sarcodina - Amoebae Oxytricha sp.
Amoeba proteus; A radiosa
Hartmanne lla
Amelia Vulgaris Dipluaster sp. Doryhrr...itis sp.

Noegleria gruberi Manochoides sp. Chlindrocorpus sp.

A ctinophrys
2klorosteroides sp. agtloblis sp.
Rhabditis sp. Rhabditolaimus sp.

Pelodera,sp. NIonhystera.sp.
AphelelichoideS sp. r-..24lolaus sp.
125 imp*,
Protozoa,. Nematodes, °s.nd Rotifers

ROTATORIA 3 Chang, S. L. Interactions between Animal

Viruses and Higher Forins of Microbes.
21..glena Proc. Am. Soc. Civ. Eng. J1. San. Eng.
96:151. 1970
, 4 Chang, S. L. Zoomicrobial Indicators
° of Water Pollution presented at the ---
Polarthra Annual Meeting of Am. Soc. Microbial,
Philadelphia, April 23 -28, 1972.
5 Chang, S. L. Pathogenic Free-Living
Ke rate lla Amoebae and Recreational Waters.
Presented at 6th International Confer-
Brachionus , en%e of Water Pollution Research
OLIGOCHAETA (bristle worms) Association, Jerusalem, Israel,.
June 19-24, 1972.
Aelosomh hempachi..
> 6 Chang, S. L. Proposed Method for
,Aulophonts limosa Examination of Water for Free-Living
Nematodes. 5.A.W.W.A. 52:695-698.
Tubifex tubifex
Lumbricillus lineatus 7 Chang, S. L., et al. Survival and Protection
17=11D 111 all.a Against Chlorination of Human Enteric
Pathogens in Free-LivingNematodes
Isolated from Water Supplies. Am. Jour.
Chironomus e Trop. Med. and Hyg. 9:136-142. 1960.
8° Chang, S. L. Growth of Small Free-Liiiing
Psychoda sp. (trickling filter fly) Amoebae in Bacterial and Bacteria-Free
ARTHRoPODA Cultiires 4. Can. J. Microbial.
6:397-40 s'1960. '
, 'f-
Lessertia sp 9 ChaneSs. L. and fabler, /1. W. tree- Living
Porrhomma sp. Nematodes in Aerobic Aeatment Plant
Effluents. J.W.P.O.F. 34:1256-2161. 4
Achoratua subuiata s ('collembola).
10 Chitwood, B. G. and Chitwood, M. B. An
Folsomia sp. (collerhhola) Introduction to'Nernatolozz. Section I
Anatomy: 1st e71. Monumental Printing
Tomocerus sp. (collembola) Co. Baltimore. 1950. pp 8-9.
REFERENCES 11 Cobb, N.A. Contributions to the Science of
1 American Public Health Association, ' Nematsology VIt. "Williams and Wilkins Co.
4merican Water Work's Association and Baltimore. 1918.
Water Pollution Control Federation. 12 Collins, V. G. The Distribution and Ecology
Standard Methods for the Examination of Bacteria in Freshwater, Pts. I &
of Water a Warftewater, 13th ed. Proc: Soc. for Water Treatment and Exam.
New York. 1971. 12:40-73. 1963: (England)
2 Chang, S. L., et al. SurveY of Free-, 13 Edmondson, W. T., et al.. Ward-Whipple's
Living Nematodes and Amoeba in Fresh Water Biolm. 2nd ed. John
Muicipal Supplies. J. A. W. W. A.. 52: Wiley & Sons, New York. 1959. pp 368-401.
613 -618. .
Protozoa, Nematodes, and Rotifers

14 Goodey, T. Soil \and Freshwater This outline wa prepared by S. L. Chang,

Nematodes.' (A Monograph) 1st ed. Chief, Etiology, Criteria Development
Methuen and Co. Ltd.' London. 1951. Branch, Water Supply Research Laboratory,
NERC, USEPA, Cincinnati, Ohio 45268.
Descriptors: Protozoa, Ne atodes, Rotifers




Piotozoa, Nematodes, and tifers j

Effluent 1-7-4-
Raw Sewage
;1 TR RR
OligochaeteS-& Suspe)ed o ank matter
insect larvae-
(by h crolysis)

& rotifers
DisSolved organic matter
AERATION Nonpimented
TANKS protozoa
I It A (respiration,
de amination,
Heterotrophic decarboxylation, etc.)

Inorganic C, P, N,
Fungi S comp.

Autott'ophic bacteria (Nitrification, sulfur

& iron bacteria)
Pathogenic organiSms

Food Chain in Aerobic.Sewage Treatnient Processes

The same sequences and processes occur

in polluted streams and streami'receiving
treated wastes.

1 28


Larggr animals (worms, snails, fly larvae, 1

etc. ) dominate trickling filters. Why are

these always absent from the activated
sludge process?
Why are there numerous micro- species "2
common to both trickling filters and
activated sludge?
What Organisms besides protozoans and 3
animals are present in activated sludge?
What is (he advantage(s) of microscopic 4
examination of activated sludge?
One sampling site would be the one of choice 5 .
in sampling Ni activated sludge plant for
microscopic analysis. Why? Where?
What organisms predominate in activated 6
sludge? a 4.

Why are photosynthetic green plants (in 7

contrast to animals) basically absent from
the activated sludge process in general and
mixed liquor specifically?
hy are the same identical species of protozoa 8
f d in activated sludge plants all over the
What is, the significance of a microscopic 9
examination of mixed..)liquor?

Define and characterize: 10

Activated Sludge
.Mixed Liquor
What is the relation between bacterial) . 11
protozoan populations in activated sludge
and the proces,s itself?
At what total magnificatiori wereyou able to 12
believe the smallest cells observed.w.ere in
fact bacteria?
Activated sludge is a dynamic (although 13
man-manipulated) ecosystem. How-does,
it differ from a natural ecosystem?

Activated-Sludge PiOtozoa -AO

What is the greatest problem(s) with a wet 14

mount slide preparation? . I

How do you overcome these disadvantages? 15

How do yoU slow down fast moving protozoans 16

on a wet mount?
Why are quantitative counts of protozoa (like 17
number/ ml) generally meaningless?
What' is the significance of proportional 18
Scanning a slide (in making a count) should 19
generally be done at X. (Total
magnification) .
The iris diaphrUm on thenlicroscope is 20
used to adjust light intensity (true-false).
Why sample the surface film of the 21-
Why is the thinnest film most ideal for a 22
wet mount? 63

What did you learn from the microscopic 23
examination' of the activated sludge?
What is the physical nature of the flocs 24

What filamentous organisms were 25°

Why are "rare" species of no practical 26
significance in microscop analyses of
activated sludge? o

Why are there no protozoan indicator species 27

of process efficiency in activatedillludee?
.0 4
Activated sludge-biological communities 28
are temporal in contrast to biological
.communities in trickling filters which
are spatial-(TRUE/FALSE).
"Seeding" a newly started activated sludge 29
plant with cultures or material from other O

plants is only a. wasted effort (TRUE/FALSE).

Justify,y.our answer. .

co . .
4- .1 . ti

Activated Sludge Protozoa

if a wet mount slide` of mixed liquor is 30

prepared and placed in 'a petri dish with a
wet blotter underneath and allowed to sit
for sgveralhours, what will be the distri-
bution of the protozoa under the cover slip?
In a mixed liquor sample nearly all of the 31
stalke,d ciliates have""broken off. the stalks
and are frerswimmineas utelotrcichs."
What does'this indicate? tto

What are Monags? And are they good, tad, 32

4or indifferent in activated sludge?
What are hypotrichs or crawling ciliates, 33
and are they good, bad or indifferent ii
activated sludge? A %. Q

What are swimming ciliates, and are they 34 _

goodbad or indifferent in activated sludge?
What are flagellAtes, and are they good, bad '35 .
or indifferent in activated sludge?o
What are amoebae, and are they good, bad, 36
A or indifferent in activated sludge?
What are the ideal characteristics of a wet 't 37
mount slide preparation?
Why does total community givea- betfbr 38
indication of process efficiency in activated ..

In observing and identifying protozoa one looks 39

for what, characteristics of an individual
What is the role of bacteAa in activated a ;10

Vihatsis the role or,protozoa in activated 41;

Microscopic analysis ofAmbeed liquor 42
ot, 4&)

sample can be very quick, simple, and - .

meaningful (TRUE /FALSE). .


Activated Sludge Protozoa f

Protozoan communities present in activated 43

'sludge reveal:
a. Plant efficiency
b. Settlfabil4
c. ROD removal
d. Solids removal
ei.; Plant ling o
(Circle applicable .descriptions)
Pr.otozob.n-communities in activated sludge 44
c. °reveal complete-and instantaneous to,nditions;
average of physical and_chemical-conditions;
-egtremes of chemical and physical conditions.
(Draw a line through phrases true.)
Rank in increasing plant efficiency the 45
°following ptotoioan group which would pre-,
Stalked ciliates ,
Swimming ciliatt s
9 Crawling ciliates
(For exa ple, use number 1-6. One would be
startup co ditiops or least efficient, and six
would be t e.most efficient.) ..
Identification is usually done at X and 46
sometimes requires X.

Immersion oil should be used sparingly at 47

what two points on a slide?
'''''''4:9 Which comes first in microscopic examina- 48
lion; s anning at low power to pick out. 1<
unkno ns or higher power to identify ?,

In making proportional count, which total. 49

number to count would be better; total of ten
_organisms or a total of 100 organisms? Why?
Why not kill the organisms so you can 50
identify and count them'on the slide?
,What simple chemical solutions are useful 51
to immobiliie protozoa if methyl cellulose
or polyvinyl alcohol is hot-available?'

Activated Sludge Protozoa

Initially the° wet mount slide should be 52

racked up close to the low power objective:
by your eye on the eyepiece through the
scope; or by glancing at the actual distance .

with the naked eye While you rotate the

coarse adjustment knob. (Underline which)
What are par -focal objectives on the 53
Why should water on the microscope and all 54 lj

its parts be carefUlly avoided?

If activated sludge is a mah manipulaiied 55
system, 6re there comparable,na.tuPal,
ecosystems? Exainple?
What is the " community '; concept in exami- 56
nation of aptivated sludge?
What a're the appliCations of direct micro- 57
scopist examination of activated sludge?
What are rotifers, and are they good, bad - 58
* or indifferent in activated sludge? . o*.

List the five kingdomS of organismsnd give 59

a specific example for each.

What techniques are most useful in 60

identifying an unknown Organism, and why
is -correct identification Important?
Scanning and counting is done at * X 61
magnification. Identification of most
PROTOZOA usually requires . X
-magnification and occassionally X
- magnification. -5' ,


43.5 X
10 X I.

40 X ".

boo° x

(1,0 X eyepiece's)

Activated Sladge Protozoa

hand is constantly operating the 63

hand issconstantly, operating the

The microscope is manually operated and

requires skill and understanding on the part
of the operator. A microscope, no matter
how costly is only as good as the micro-
scopist operating it.
List the basic skills required in utilizing the 64 .7
optimum capabNty of your microscope.
A h,

This outlinerv/as prepared Isr,R. M. Sinclair,
.fiational Training tenter, MOTD, OWPO,
USEPA, Cincinnati, Ohio 45268.

Descriptors: Zlicroorganisms, Protozoa

;.... .
Rotifers Activated Sludge. Riota
.? 1,


". 3



I FREE - LIVING AMOEBAE C Morphological Characteristics of

Small, Free-Living Amoebae
Importance of Recognizing Small,
Ftee- Living Amoebae in Water .1 Morphology of Trophozoites'-
Supplies Ectoplasm and endoplasm usually
,dIst ot; mar:dens-with large nucleolus.
1 Commonly found in soil, aerobic is
sewage effluent and natural, fresh 2 Morphology of cysts - Single or
waters - hence,- frequently en- double wall with or without pores
countered in examination of raw
watek. ' D Cultural Characte ristics of Small;
Free - Living- Amoebae
stenot infrequently found in
wits]. al supplies - not pathogen 1 How to cultivate these amoebae -.
arrie- s. plates with bacteria; cell cultures,

\ :-.
axenre culture.
lage 41,e-a,moebae Naeolerla
involved-140 some cases of 2 Growth characteristics on plate,
meningoencephalitis, about half . cell, and axenic culture
in the U:S.; associated with
swimming in small warm lakes. ,3 Complex growth requirements
Acanthamoeba rhisodes parasitizing for most of these amoebae
hyman throats and causing (3 cases)
nonswimming-associated meningo- E Resistance of Amoebic Cysts to
encephalitis. . Physical andChemical Agents
4 Cysts not to be confused with those
of Endamoeba histoljtica in water -

borne. epidemics. II FREE- LIVING NEMATODES

B Classification of Small, Free-Living A Classification of Those Commonly

Amoebae Found in Water Supplies

1 Recognized classification based 1 Phasmidia (Secerneutes):

on characteristics in mitosis. Genera Rhabditis, piploga.ster,
Dipl.oacqeroidesi Cheilolms
2 k Common species fall into the Panalal
following families and genera:
Aphasmidia (Adenophoro): Genera
Family Schizopyrenidae: ,Genera moL2-il st1., Aehelenchusr. Turbatibc..
Naegleriaj Didascalusi and (vinegar eel), l2c2D22.imusj and
Schizopyrenu.s.- first-two being Rhabdolaimus ,
flagellate amoebae.
B "MorphOlogical F4tures
Family Hartmannellidae: Genera
Hartmanella (Acanthamoeba) 1 Phasnaids: papilla on tail 'at males,
mouth 'adapted to feed on bacteria,
3 H9w to prepare material's for few exceptions.
studying mitosis Feulgen stain
2 Aphasmids: no papilla on male ts.- -

glandular cells in male.


BI. AQ."14b. 6. 76 I 2-1

'!$ (2.
Amoebae aid IsTematodes in Water Ues we, 4.
C Life Cycle 2 Chang, S. L., et al. IsSurvey of Free -
Living Nematddes and Amoebas' in
1 MetIods Of mating ' Municipal Supplies". J. A. W . W. A.
52.613-618, 1960. ,
2 Stages of developthent
3 Chang, S. L., "Growth of Small Free-
3 Parthenogenesis Living Amoebae in Various Bacterial
and in Bacteria-Free Cultures".., Can,
b Cultivation Jour. -.Microbiol:. 6 :397 -405, 1960.
1 Bacteria -fed, cultures Nematodes
2 Axenic' cultures 1 Goodey, T., "Freshwater Nematodes",
,,>; 1st. Edition, Methuen & Co., LondyA,
E Occurren-ie in Water Supplies 1951.

1. Relationship between their_.--- 2 Edmondson, W. T. ,. Ed., Ward & Whipple' s

appearance in finished,waer '''Fresh-Water Biology" 10th Edition,
and that in raw water. S'
page 397, 1955.
2 r Frequency of occurrence in f 3 Chang, S. L., et al. , "Occurrence of a
diffetent types of'raw water,,, 'Nematode Worm in a City-Water Supply ".
and sources. J.A.-W.W;A., 51:671:676, 1959.
3 Survival of human enteric path- 4 Chang, S. L., et al., "Suzvival, 3-11A
ogenic bac't'eria and viruses in Protection Against Chlorination, of ;
'- nematodes. HumanEnteric Pathogens in Free- '
Living Nematodes Isolated From. Water
4 Protection of human enteric Supplies". Am. Jour. Trop. Medicine
pathogenic bacteria and viruses & Hygiene, --9:136-142, 1960.
in lematode-carriers.
, Chang, S. L., et al., "Survey of Free-
F Control. Living Nematodes and Amoebas in
Municipal Supplies". J. A. W. W. A. ,
1 Chlorination of sitwa.ge--eltu-ent
52:613-618, 1980.
Floqdulation and sedimentation 6 Chang, Si L. , "Proposed Method for
of A4ater Exa-nination'of Water for Free-Living
Nematodes". J. A W. W. A. , 52:695-638,
3 Chlorination of water -1950->7 -4*

4 Other methods of destruction Ch, g, S. L.; "Viruses, Amoebas,

a d Nematodes and Public Water
S pplies". J.A9rW.W.A., 53 :288 -296, ,
8 Chang, \S. L. and Kabler, P. W., "Free4
Amoebae Living Nematddes 4n Sewage Effluent
f tom erobic Treatment Plants": To
1_ Sinkh, B. N. , "Nuclear Division in Nine ub shed. -
Species of Small, Free-Living Amoe- This outline as prepared by ShT,ii L7Charig,
bae and. its Bearing on the Classifica- M. D: , Chief, iology, Criteria Lievelopment.
tion of the -Order Amoebida", Branch, Water Supply Research Laboratory,
Trans; RcrialSoc. London; Series- B, _NERC, EPA, C cinnati. OH 45268.
236:405-461;, 1952. ,
Descriptons; Amebae, Nematodes


Subphylum: Sarcodina Hertwig and Lesser

ss: ,Rhizopoda von Siebold
Subclass: Amoebaea Butschli
'Order: Amoebida Calkins and Ehrenberg to
Superfamiky: Amoebaceae - free-living
(Endarooebaceae parasitic in animals)
Family: Schizopyrenidae - active limax form common; transcient
° flagellates present or absent; nucleonus-origin of
polar masses; polar caps and interzonal bodip-present
Genus: Schizopyrenus - no transcient flagellates; single-walled
cysts; no polar caps or interzonal bodies in mitosis
Species: S. erzthaenu,3a - reddish orange pigment formed In agar
cultures with gram-negative bacillary bacteria
S. russelli - no pigment produced in agarcultures
Genus: Didascalus - morphology and cytology similar toSchizopresms
buFsmall 'numbers' of transcient flagellates fdrmed at times
Species: D. thorntoni - only species described by Singh (1952)
Genus: %aerie. Alexeieff - double-walled cysts; transcient
fla,gellatcs formed readily; polar caps and interzonal
bodies present in mitosis
Species: N. gr.utieri (Schardin-ger) - only species established;
Singh (1952) disclaimed the N. soli he described in 1,951
Family: _Hartmiannellidae - no trans dent flagellate formed; motility
sluggish; no limax form; nucleolus disappearing, probably
Vh., forming spindle in mitosis; no polar caps or masses, aster,
and centrosome not known
Genus: Hartmannella - ectoplasm clear or less granular than
endoplasm; single-walled cysti3; single vacuole
Species: H. fasbae - clear ectoplasm,
agricola.- ectoplasm less granular than endoplasm
Genus: Acanthamoeba - filamentous processes from ecto- or
endoplasm; growing axenically in fluid bacteriological
a/Arne dta

137 12 -3'
Su p, estecassification of Small Amoebae ..


. , Species: A. rhLsodea
, .

Genus: Sinehella -,double-walled cysts; ecto- and endoplasm

indistinguishable; many vacuoles
I Species: Singilella latocherrius


* tr,
. ...

61> /
, z




. .. : .-'



, (Note Figure: Nonpigmehted, Non - Oxygen
Producing Protozoan Flagellates in the
A Planktonic animals or zooplankton are outline Oxygen Relationships.)
found in nearly every major group of
animals. 1 Commonly encountered genera
1 Truly pla:hktonic species (euplankton) Bodo
spend all or most of their active life
,Cycle- suspended in the ;water. Three .Peranema
groups are predominantly involved in
. fresh water; the protozoa, rotifers, 2 Frequently associated with eutrophic
"and microcru.stotea. conditions
2 Transient planktonic phases such as C Class Rhizopoda amoebOi.dprotozoans
floating eggs and cysts, and larval
stages occur in many Other groups.. 1 Forms commonly encountered as
B Many forms are strictly seasonal in
occurrence. Chaos fi4noeba)
C Certain rare forms occur in great numbers Arceila Centropyxis
at unpredictable intervals. 77
Difflusia Fieliozoa
D Techniques of collection, prese'rvation,
and identification strongly influence the Euglyphe.
, species reported.
2 Cysts of some types may be encountered
E In oceanographic work, the zoop ?Acton is in water plants bedistributiori systems;
considered to include many relath.vely large rarely in plahkton of open lakes or
animals such as siphonophores, ctenoplores,p reservoirs.
hepteropods, pteropods, arrowworms, and
etiphausid shrimp, D Class Ciliophora
F The plant-like or phytoplankton on the 1 Certain "attached" forms often round
other hand are essentially similar in all floating freely with plankton:
waters, and are the nutritional foundation
for the animal community.. Vorticella

2 Naked, unattached ciliates, Halteria
A The three typically free living classes, one of commonest in this grouirTa7Rous
Mastigophora, Rhizopoda, and Ciliophora, heavily ciliated forms (holotrichs) maY
all have planktonic representatives. As occur from time to time ailch as
a group however, the majority of the phylum Colpidium, Enchel, etc.
is benthic or bottom-loving:, Nearly any
ofothe,benthic forms may occasionally be 3 Ciliates protected by a shell or test
w.ashe'd up into the overlying waters and (testaceous) are most often recorded
thus be collected along with the euplankton. from preserved samples. Particularly
common inthe experience of the National
B Class mastigophora, the nonpigmented 'Water Quality Sampling Network are:
Codonella fluviatile a
These have frequently been confused with
the phytomastigina or plant-like flagellates. Codonella cratera
The distinction-is made here on the basis
of the presence or absence of chlorophyll Tintinnidium (usually with organic matter)
as puggested.by Earner 'and Ingram 1955.
- TintinnoRgis

13 -1
Animal PlanktA

III PHYLUM ROTIFERA 2) As unfavorable conditions develop,

,, males- appear, -and- thick walled
.i/N Some forms such as .Anuraea cochlearis sexual eggs are enclosed in egg
and Asplanchna pridor 'cases called ephippia which can
at all tames of the year. Others such as often endure freezing and drying.
lc,Totholca striate, N. Ion spina and Poly-
arthra platypTera are repor e to be essen- 3) Sexual reproduction may occur
tially winter torms.. at different seasons in different
B Species in approximate order of descending
frequency currently recorded by National 4) Individuals of a great range of *
Water. Quality Sampling Network are: sizes, and even ephippia, are
thus encountered in the plankton,
Keratella cochlearis but there is no "Larval" form.
Polyarthra vulgaris b Seasonal variation - Considerable
variation may occur between winter
Synchaeta pectinate and'summer forms of the same
species in some cases. Similar
Brachionus quadridentata variation also occurs between arctic
and tropical. situations.
Trichocerca longiseta
c Forms commonly encountered as
Rotariesp. open water plankton include:
Filinia longiseta. Bosmina longkostrisCand others
Kellicottia longispina Daphnia galeata ai.14.-erthers
Pompholp sp. a Other less common genera are:
C Benthic species almost without number may Diaphanosoma, Chydorus, Sida,
be collected witb4e plankton from time to 7croperus, Creriodaphnia, Byt 'ho-
time. trephes, analTeEataVorous
reYEZ'ara and Polyphemus.
IV PHYLUM ARTHROPOD.A d :Heavy blooms of Cladocerans may
build up in eutrophic waters.
A Claes Crustacea
3 The copepods (Order Copepoda) are the
1 The Class Crustacea includes the larger perennial microc'ustacea of open waters,
common freshwater euplankton. They both fresh and marine. They are the
are also the greatest planktonic consum- most ubiquitous of animal plankton.
ers of basic nutrients in the form of
phytoplankton, and are themselves the a Cyclomis the genus Most often
greatest planktonic contribution to the rdiirarby the National Water Quality
food of fishes. Most'of them are herb- Sampling Network activities. Eucr.
ivorous. Two. groups, the cladocera clops, Paracyclops, Diaptomue,
and the copepods are most'conspicuous. UrriffiocaraptirCiiiiiscTirra, an
LimnocIlanirrare other forms
2 Cladocera,(Subclass Branchiopoda, FFIWe7-1-5.-We planktonic.
Order Cladocerarar Water Fleas 4
b Copepods 'hatch into a minute char-
a Life History acteristic larvae called a nailius
which, differs considefably from the
1) During most of the year, 'eggs adults. After five r six moults, the
which will develop without fertil- copepodid stage is j'eached, and after.-
ization (parthenogenetic) are six more moults, the adult. These
deposited by the female in a dorsal larval stages are often encountered
brood chamber. Here they hatch arfd are difficult to identify.
ir}to minature adults which escape
and mini away. ,
Animal Plankton

Insecta yluirt-Nymathelminthes
1 Only a few kinds of insect that can be 1 N atodeslbr nemas) or roundworms
ranked as a truly planktonic. Mainly appr h the bacteria and the blue-green
. the phantom midge fly Chaborus: algae in, iquity. Thevare found in
the soil and in tke water, and in the air
2 The larva of this insect has hydrostatic as dust. 'In both marine and fresh waters
organs that enable it to remain suspendcd and from, the Arctic tb the tropics.
in the water, or make vertical asaens ii: 2 Although the majority are free living,
thewate column. some occur as parasites of plants,
animals, and Man, and some of these
3 It is usually benthic .during the daytime, . parasites are among our most serious.
but ascends to the surface at night.
3 With this dfstribution, it is obvious that
they. will occasionallz be encountered as
V. OCCASIONAL VLANKTERS 'plankton. A more complete discussion
of nematodes and their public health
A While the protozoa, rotifers, and micro- implications in water supplies will be
crustacea make up the bulk of the plankton, found elsewhere (Chang, S. L. ).
there are many other groups as mentioned
above that may also occur. Locally or E Additions.' 1 crustacean groups ,sporadically
periodically these may be of major impoit- met with in the plankton include the folloWing:
- ance. Examples are given below. 4.
1 Or er Anostraca or fairy shrimps
B Phylum Coelenterate , merly included with the two
f lowing orders in the Euphyllopoda)
T Polts of the genus Hydra may become primarily planktonic in nature.
detached and float aHourhadiging from
the surface film or floating detritus. a Extremely local and sporadic, but
when present, may be dominatihg.
I- . 2 The freshwater medusa Craspedacusta
occasionally appears in Tale% or reser- b Artemia, the brine shrimp,can
voirs in great numbers. tolerate very high salinitie.
C Phylum Platyhelminthes c Iiery widely distributed, poorly
1 Minute Turbellaria (relatives of the , understood. -
well known Planaria) are sometimes 2 Order Notostraca, the tadpole shrimps.
taken with tbe 'plankton in eutrophic-- Essentially southern and western in
conditions. They are often confused distribution.
with ciliate protozoa.
3 Order Conchostraca, the clam shrimps:
2 Cercaria larvae of Trematodes (flukes) Widely distributed, sporadic in occur-v
parasitic on certain wild animals, rence.' Many local species.
frequently appear in great numbers.
When trapped in the droplets of water 4 Subclass Ostracoda, the seed shrimps.
i on a swimmer's skin,' they attempt to Up to 3 in. in length. Essentially
bore in. Man not being their natural benthic but certain species of C ris,
host, they fail. The. resultant irritation and Notodromas may occur in consi -
is called "swimmer's itch". Some can erabTerniribiTs as plankton at certain
beridentified, but many unidentifiable times of the year.
species may be found.
5 Certain members of the large subclass
3 In many areas of the world, _cerparia Malacostraca are limnetic, and thus,
larvae of -human parasites such as the planktonic to some extent.
blood fliAce-Schistosorna japonicum may
Jive as plan orkr7-'1-fT-"IpEngfiirf. Fia-humah a The scuds, (odder Amphipoda) are
skin,directly on contact. essentially benthic but are sometimes
collected in plankton samples around

tr 13-3 ;0
'Animal Plankton

weed beds or near shore. Nekto- the uninitiated, they are sometimes
planktonic forms include Pontoporeia mistaken for fungi or other organisms
and some species of Gammarus. (and vice versa).
b The mysid, or opossum-shrimps are In flowing waters, normally, benthic
represented among the plankton by (bottom living) organisms are often found 41kP
Mysis-relicta, which occurs in the drifting freely in the stream. This
deeper waters,, large lakes, as far phenomenon may be constant or periodic.
north as the Arctic Ocean. ' When included in plankton collections,
_ .
theyamust be reported, but recognized
F The Class Archnoidea, Order Hydracarina for what they are.
(or A cari) the mites. Frequent in plankton
tows near shore although Unionicola crass-
ipes has been reported to-be yirtually films are especially rich in
planktonic. micro "biological garbage" and these
enrich the plankton.
G The phylum Mollusca is but scantily
represented in the freshwater plankton, REFERENCES
in contrast to the marine situation. Edmondson, W. F., ed. Ward and
Glochidia (ciliated) larvae are occasion- Whipples'aFreshwater Biology, 2nd
ally collected, and snails now and then . Edition, Wiley & Sons) Inc., New York.
glide out bn a.,quiel surface film and are ,
taken in a plankton net. An exotic 1959.-
bivalve Corbicula heave planktonic
veliger stage. 2 Hutchinson, G. Evelyn. A Treatise on
Limnology. Vol. 2. Introduction to
H Eggs and other reproductive structures . Lake Biology and the Limnopla.nkton.
of many forms including fish, insects, and Wiley. 1115 pp. 1967.
rotifers may tpe found in plankton samples.
Special-reproductive structures such as 3 Lackey, J. B. Quality and Quant y of
the statoblasts of bryozoa and sponges, . Plankton in the South End of ke
and the ephippia of cla.doCerans-may also Michigan in 1952. JAWW
be ineltided. 36:669-74. 1944.

I Adveptitious and Accidental Plankters 4 McGauhey, P. H. , Eich, H. F. , Jackson,

H. W., and Henderson, C. A Study
Many shallow ,water benthic organisms of the Stream Pollution Problem in the
may become accidentally and temporarily Roanoke, Virginia, Metropolitan ,
incorporated into the plankton. Many of District. Virginia Polytech.
those in the preceding section might be Engr. Expt. Sta.
listed here, in addition to such forms as
certain free living nematodes' small 5 Needham, J. G. and Lloyd, J, T. The
oligochaetes, and tardigrades,. Collembola Life of Inland Waters. Ithaca, New
and other surface film livers are also, York, Comstock Publishing Co., Inc.
taken at times but should not be ,Mistaken- j 1937.
for plankton. Fragments and molt. skins' , .

from a variety of arthrOpods are usually 6 Newell, G.E. and Newell, R. C.

observed. Marine Plankton. Ilutchinson duc.
Ltd. London. 221 pp. 1963.
Pollen from terrestrial or aquatic plants
is often unrecognized, or confused wit 7 Palmer, C. M. and Irlgram, W M.
one, of the above. Leaf hairs from Suggested Classification of Algae and
texrestrial plants are also confusing to Protozoa in Sanitary Science.
Sew; &''Ind. Wastes. 27:1183-88.
1955. ," ,

Animal Plankton

8 Pennak, R. W. Freshwater Invertebrates 10 Welch, P. S. Limnology, McGraw-Hill

of the United States. The Ronald Press, Book C'o., Inc., New York. 1935.
New York. 1953.

9 Sverdrup, IL W. , Johnson, M. W. , and

Fleming, R. H. The Oceans, Their
Physics, Chemistry and General This outline was prepared by-H. W. Jackson,
,Biology. Prentice-Hall, Inc., New York. former Chief Biologist, National Training
1 1942. Center, and revised by R. M. Sinclair,
Aquatic Biologist, National Training Center
MOTD. OWPO, ,USEPA,_ Cincinnati, Ohio

Descriptor: Zooplankton



Anim1 Plankton

3/4 - Free Living Representative.'

I. Flagellated Protozoa, -C sae Mistigophora

Pollution tollerant Pollution tollerant
6,u 19 A..
Colony of Poteriodeidron
Pollution tollerant, 35jz

II. Amsboid Protozoa; Class Saroodina


Dimastizamoeha tiuolearia,reported Diffluxia

Pollution tollerant to be intollerant of Pollution tollerant
1067,50)u pollution,-451u. 601563.14

_III. Ciyied-Protozoa, Class'Ciliophora

Coliod; Nolovhrva,reportod Aaitvlis, pollution

Pollution tollerant to'be intollerant of tollerant. Colonies often,
20-120)4 pollution, 35,E maprosoopio.

Ahimal Plankton


Peranema trichophorum

A r cella
vu garis A ctinosphad r him


J, A

Godonella _Tintnidium
Vorticeila cratera fluviatile

Animal Plankton


$ 'Various Forn of Keratella cechlearis

a 11.4'

Synchaeta PolyarthrA Brachionus
pectknata vulgaris
quadridentata I

Rotaria sp

Animal Plankton


_ A NatiPtiis larva of a Copepod

1-5 mm
, 14,
Copepod; Cyclops, Order Copepoda
2=.3 mm .

Water Flea; Order Cladocera
2-3 mm'


Left: Shell closed Right: Appendages extended

1-2 mm

- Aninial Plankton


A mysid shrimp - crustacean A water mite - arachnid


Chaoborus midge larva - Insect



/- Aspects in the life cycle of the human tapeworm,

Diphyllobothrium Tatum, Class Cestoda. A . adult as in human /
intestine; B. procercoid larva in copepod; C. plerocercoid
larva in flesh of pickerel (X-ray view).
. 9 H. W. Jackson





`I RECEPTION AND PREPARATION OF, The amount of sodium borate and

SAMPLES merthiolate may be vAried slightly to
adjust to different waiters, climates,
A Preliminary-sampling arid analysis is an . ana organic, contents.'
essential preliminarS, to the establish-
ment of a permanent or- semi-permanent' 3 "Prepae a saturated aqueous. Lugol''S
'program. solution- as follows:
B Concentration or sedimentation of pre- a Add 60 grams of potassium iodide
served samples 'may precede analysis. (IP) and 40 grams of iodine crystals
to 1 liter of distilled water.
1 Batch centrifuge
, 4 Prepare the preservative solution by1
2 Continuous centrifuge adding approximately 1. 0 ml of the
Lugol's solution to 1 liter of merthi-.
3 Sedimentation olate stock.
C Unpreserved (living) samples should be
analyzed at once or refrigerated fo III SAMPLE ANALYSIS
future analysis.
II PREPARATION OF MERTHIOLATE A Microsc'opic examination is most frequently
PRESERVATIVE employed in the laboratory to determine 7
what plankton organisms are present and -NI_
A The Vater Pollution Surveillance System bow many there .are: -
of the FWPCA haS developed a, modified
merthiolate preservative. (Williams, 1-Optical equipment need not be elaborate
1967) Sufficient stook to "make an approx- but should include.
imately 3. 5% solution in the bottle when
filled is placed in the sample bottle in a Compound microscope with the
the laboratory. `The bottle is then_ filled' following equipment: #
witli water in the \isteld and returned to
the laboratory for nalysis,. 1) Mechanical stage
2)- Ocular: 10X, with :Whipple type
B Preparation of Mertbiolare Preservative counting eyepiece or reticule
3) Objectives:
Merthiolate is availablelfrom many approx. 10X( 16mm)
chemical laborator supply houses; aaprox, 20X(8 mm).
one should specify he water 'soluble approx. 40X(4 mm) .
sodium salt. approx. 95X( 1. 8 mm)( optional)
2 Merthiolate stock: d, -solve approxi-- A 40X objective with a working
mately 1.5 dram of sodium borate distance of 12,8 mm and an erect .
(borax atd approximat ly 1 gram of image may be obtained as special
merthiolate in 1 liter o distilled water. eqUipment. A water, immersion
objective (in addition to oil) .might
be considered for use with water
mounts. k
Binocular eyepieces are 'optional.

MIC. enu. 15f. 6.76 14-4_


Preparation and Enumeration of Plankton -A

Stage micrometer (this may be 3 Direct 'counting of the unconcentrated'

borrowed, if necessary, as it is sample eliminates manipulation, saves
usually used only once, when the time, and reduces error. If frequency
-equipment .is calibrated) . of organis is low, more area may
. need to be xamined or concentration
Inverted microscopes offer certain , of the, samp may be in ortfe'r.
advantages but a r efoot widely
available. The same is true of 4 Conventional techniques emplpying
some of the newer optical systems, concentration of the sample provide more
such as phase contrast microscopy. organisms for observation, but because
These are often excellent but ex- they involve more m nipulations,
pensive for routine plant use. additional error and fake more
2 Precision made counting chambers 4--
are required for quantitative -work with
liquid mounts. C Several methods of counting plankton are
in 'general use.
a Sedgwick- Rafter cells' (hereafter
referred to as S-R 'cell) are used 1 The anumerical or clump count is
for routine counts of medium and regarded as tile simplest.
larger forms.
a Every organism observad must be'
b Extremely small forms or "nanno- enumerated. If it cannot be identi-
plankton" may be counted by use of fied, assign a symbol or number
the nannOplankton (or Palmer) cell, and make a sketch of it on the back
. a Fisher- Littman cell, a hemacytp- of the record sheet.
meter, the Lackey drop method, or
by use of an inverted microscope. b Filaments, colonies and other
associations' Of cells are counted
g Previous to starting serious analytical as units., equVr to single isolated
work, the microscope should be cali- cells. _Their identity as indicated
brated as described elsewhere. Di- on the record sheet is the key to
mensions of the S-R cell should also the significance' of such a count.
be checked, -e ecially th depth/
Individual cell 'count. In this method,
4 Autom is rticle cou ters may be every cell of every colony or clump .
useful for c9ccoid organisms, of organisms is- counted, as well as
each individual single- lled organism.
B Quantitative Plank-Ion Counts The_ areal standard unit method rs
L certain technical advantages, but als
1 All quantitative eounting techniques involves certaininherrent difficultie
involve the filling of a standard cell
of known dimension's with either a An areal standard unit is 400 square
straight -sample or a concentrate or microns. This is the area of one
dilution .theieof. of the smallest subdivided squares
in the center of the Shipple eyepiece
2 The organisms in a predetermined at a magnification of 100:X.
number. of microscope fields or other
known area are then observed, and by b In .operation, the nurnber f areal
means of a' suitable series of multi- r
units of each species is recorded on
plier factors, projected to a number or the record. sheet rather than'the
quantity per ml gallon, etc.' number of indiviiduals. Average
ar=eas of the common spe'cies are

14 -2
t Preparation And Epuinberation of Plankton

are sometimes printed on record after each group of fields are counted
sheets for a particular plant to and making up to 5 additional such
obviate the necessity of estimating counts. This may not be practical with
the area .of each cell- observed high counts.
7 The strip count. When a rectangular
c The advantage of they method lies in slide such as the S-R cell is used, a,.
the cognizance taken of the relative strip (or strips) the entire length (or
masses of the various species as- known portion Thereof) of the cell may
indicated by the area presented to be counted instead of separate isolated
the viewer. These arel.s., however, fields. Markinahe bottom of the cell.
are often very difficult to estimate. by evenly, spaced cross lines as ex-
.... plained elsewhere greatly facilitates
4 The ,cubia standard unit method is counting. .
logical extension of the areal met od,
but ha§ achievedness acceptance. a When the count obtained is multi-
plied by the ratio of the width of
5 Separate field "count the strip counted to the width of
the cell, the product is the esti-
a In counting separate fields, the mated number of organisms in the
-'question always .arisee as to how cell, or per ml.
to count Organisms touching or
crossed by the edge of the Whipple b Whenthe material in the cell is
field. Some 'workers *estimate the unconcentrated sample water, this
e proportion of, the organism lying count- represents the condition of
inside the, field as compared to -that the water being evaluated withobt
outside. °nit), those which are over further calculation,
half way inside are counted. 'Y./
8 Survey count. A survey count is an
b Another system is to select two examination of the entire area of a
adjacent sides of the square for volumetric cell .using a wide field low
reference,. such as the top and left power microscope. The objective is
boundaries. Organisms touching to locate and record the larger ,forms,
these lines in any degree, from especially zooPlankton such as copep
outsid e or inside, are then counted, or large rotifers whi2h may be present
while organisms touching the opposite in size. Special large capacity cells
sides fire ignored. It is important are often employed for this purpose.
to adopt some such system and For still larger marine forms; numerous
adhere to it consistently. special devices have been created. .

c It is suggested that if separate 9 Once a procedure for "concentration

101P 'microscopic fields are examined,, and/or counting is adopted by a plant
a standard number of ten be adopted. or other organization it should be
These should be evenly spaced in two used consistently from then on so
rows* about one-third of the distance . that results from year to year can be
down from the top and one-third of compared.
the distance lip frc he bottom of
the S-R cell.
rz Differential or qualitative "counts" are
6 Multiple area count., This is an ex- ' essentially lists of the kinds of organisms.
tension of the separate field count. found.
A Considerable increase in accuracy
has recently been shown to accrue by
entptying and refining the S-R sell

Preparation and. Enumeration of Plankton

E It9portional or relative counts of special e Difficulties include a predilec-

groups are often very us%itilth .For ex- tion of extremely fine membranes-
ample, diatoms'. is b always to clog rapidly with silt or .

count a standard numbers of cells. increase in plankton counts, and

the difficulty of making observations
F Plankton are sometimes measured by on individual cells when the
means other than microscopic counts. organisms are piled on top of each
other. It is sometimes necessary
1 Settled volume of killed plankton in an to dilute a sample to obtain suitable
Irnhoff-,cone may be 'observed after a distribution.
standard length of time. This will
evaluate primarily only the llsher forms. IV SUMMARY AND CONCLUSIONS
2 A gravimetric-method employs drying A The field' sampling program should be
at 60° C for 24 hours followed by ep carefully planned to evaluate all significant
ashing at 6000 C for 30 minutes. This location in the reservoir or stream,
is particularly useful for chemical giving e consideration to the capacity
and radiochemical analysis. of the la ratory.
3 Chemical and phySical evaluation of. B Adequate records and notes- should be
plankton pdpulations employing various made of field conditions and associated
instrumental techniques are coming - to lit h the laboratory analyses in a
be widely used. Both biomass and permanent file.
productivity rates can be measured.
Such determinations probably achieve C Optical equipment in the -laboratory should
their greatest utility when coordinated be calibrated.
with microscopic examination.
D Once a procedure for processing plankton
4 The membrane (molecular) filter has is adopted, it should be used exclusively
a great- potential, but a generally by all workers at the plant..
acceptable technique has yet to be
perfected. E Such a procedure should enable the water
plant operator to prevent plankton troubles
a Bacteriological techniques for or at least to anticipate them and have
coliform determination are corrective materials or equipment stockpiled.
widely accepted.
'b Nematodes and larger organisms REFERENCES
can readily be washed off of the
membrane after filtration. 1- Ely Lilly Company. Merthiolate as a
c 'els also being used to measure Preservative. Ely Lilly & Co.
ultraplankton that pass treatment Indianapolis 6, Indiana.
plant operations. -
2 Gardiner, A. C! Measurement of ,
Phytoplankton PopulationWY-the
d Membranes can -be cleared and .Pigment Extraction Method. Jour.
organisms depbsited thereoni'; Marine Biol. Assoc. 25(4):739-744. 1943.
observed directly, although 4.3
accessory staining is desirable. 3 Goldberg, E. D. , Baker, 1VI. , and Fox, D. L.
Microfiltration in Oceanographic
Research Sears. Foundation. 'Jour:
Mar. Res. 11:194-204. 1952.-

Preparation and Enumeration of Plankton

4' Ingram, W. M., and Palmer, C. M. '7 Weber, C. I. The Preservation of

Simplified Procedures for Collecting, Plankton Grab Samples.- Water
Examining, and Recording Plankton Pollution Surveillance Systems,
in Water. Jour. AWWA; 44(7): Applications and Develppment Report
617-6'24. 1952. No. 26, USDI, FWPCA, Cincinnati,
° Ohio. 1967,
5 Jacksti, H.W Biological Examination
(of plankton) Part III in Simplified 8 Williams, L. G. Plankton Population
Procedures for Water Examination. Dynamics. National Water Quality
,AWWA Manual M 12. Am. Water Network Supplement 2. U. S. Public
Works Assoc. N.Y. 1964. Health Sy.vice Publ. No. .663. 1962

6 Lund, J. W. G., and Talling, J. F. 9 Wohlschag, D.- D., and Hasler, As'. D.
Botanical Limnological Methods with Some Quantitative ASpects of Algal
Special References to the Algae Growth in Lake Mendota. Ecology.
Botanical Teview. 23(8&9):489-583; 32(4):581-593. 1951
October, 1957.

This outline was prepared by W. Jackson,