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One of the most important systems for the identification of microorganisms is

observed for growth in artificial food substances prepared in the laboratory . The
food material in which microorganisms grow is the culture medium and the growth
of microorganisms is the culture .
For bacteria to grow properly in artificial culture medium must meet a number of
conditions including: temperature , humidity and pressure of oxygen adequate and
proper degree of acidity or alkalinity. A culture medium must contain nutrients and
growth factors required and must be free from any organism pollutant.

General conditions for the cultivation of microorganisms

The proper development of microorganisms in a culture medium is affected by a
number of factors of great importance and, in some cases, are entirely devoid
medium itself.
Suitable nutrient availability

1- A suitable culture medium for microbiological research must contain at least

carbon, nitrogen, sulfur, phosphorus and inorganic salts. In many cases certain
vitamins and other growth inducing substance they will be required.
Certain bacteria have specific nutritional needs so many media added substances
such as serum, blood, ascites, etc. Likewise certain carbohydrates and minerals
such as calcium, magnesium, manganese, sodium or potassium and growth
promoting substances, generally of vitamin nature may be necessary. Very often
certain dyes, as well as certain metabolic indicators or by their ability to exercise
selective inhibitors of certain microorganisms activities are added to the culture

2- suitable medium consistency From a liquid medium can change its consistency
by adding products such as albumin, gelatin or agar, which would obtain media
semisolid or solid state.

3- presence (or absence) of oxygen and other gases lot of bacteria can grow in an
atmosphere with normal oxygen tension . Some may get oxygen directly from
various substrates. But strict anaerobes only properly developed in an atmosphere
without oxygen environment .

4- adequate moisture A minimum humidity level , both in the environment and in

the atmosphere, conditions is essential for proper development of vegetative
microbial cells in cultures . We must provide for the maintenance of these minimum
conditions stoves culture at 35-37 ° C providing an adequate supply of water to
maintain the necessary moisture for crop growth and thus prevent the medium
from drying
Ambient light

5- Most microorganisms grow much better in the dark than in the presence of
sunlight . There are obvious exceptions as in the case of photosynthetic

6- pH The hydrogen ion concentration is very important for the growth of

microorganisms . Most of them develop better in media at neutral pH , although
there are requiring more or less acidic media. One should not forget that the
presence of acids or bases in quantities not prevent bacterial growth can however
inhibit or even alter their normal metabolic processes.

7- The mesophilic microorganisms grow optimally at temperatures between 15

and 43 ° C. Others as psychrophilic grow at 0 and temófilos at 80 or even at higher
temperatures ( hipertemófilos ) .

8- Sterility of media All culture media must be perfectly sterile to prevent the
emergence of life forms that may alter , mask or even prevent normal or specimens
in the media inoculated microbial growth.
semisolid media They are prepared from liquid media , adding a solidifying agent
thereto in a minor proportion to prepare solid media . One of its uses is
investigating the mobility of bacteria.


1) Common means: Those who possess the minimum components for the growth
of bacteria that do not need special requirements can be produced
2) Media enrichment: Those who, in addition to the normal nutrients, incorporate a
number of essential factors for the growth of fastidious microorganisms. This
enrichment is made by addition of blood or other biological products (blood, serum,
milk, egg, bile, etc.) that provide such factors

3) Selective media: media are used to promote the growth of certain bacteria in
polymicrobial population. The foundation of these means is nutritionally facilitate
the growth of a specific microbial population. An example of selective medium is
selenite, which is used to promote the growth of salmonellae

4) Media inhibitors: When substances added to a selective medium tot prevent a

microbial population is called inhibitor.

5) Differential Media: Used to highlight biochemical characteristics that help

differentiate 11 genera or species. Adding a fermentable sugar or a metabolizable
substrate used for this purpose. MacConkey medium is a differential medium
because it allows to distinguish the germs that ferment lactose of those who do not

6) Means of identification: They are designed to test a specific quality that can help
us to recognize the identity of a microorganism. These means must possess the
necessary elements to ensure the growth of microorganisms, the specific substrate
that will be metabolised and the indicator showing us the result.

7) Means of multiplication: Used to get a lot of cells from an already isolated

microorganism. Seemplean in obtaining vaccines, research and industry. The most
appropriate means for multiplying usually liquid.

8) Media storage: Used to keep a strain for various reasons keep us interested.
Mainly they used as quality control tests and reagents used in elLaboratorio
Microbiology. In the laboratory strains can be preserved in three ways:
a) making periodic passes from plate to plate,

b ) by lyophilization of a bacterial suspension

c ) freezing the strains in sterile 0.1% skim milk .

According to the substances that become part in its composition, the culture media
can be classified into :

1) Complex Media : They were the first used , and the employees are prepared
from animal tissue , and rarely vegetables.

2) synthetic Media : Those containing in its composition only known chemicals and
dissolved in distilled water in certain proportions , resulting in an average of well
defined composition.

3) semisynthetic Media : The large number of growth factors required for certain
germs makes manufacture of a synthetic medium for these organisms is
impossible or too expensive . In this case the growth factors in the form of a
complex organic extract (yeast extract , tissue extract , etc. ) is provided.


a) NATURAL : are prepared from natural substances of animal or vegetable origin

such as tissue extracts or infusions and whose chemical composition is not exactly
known .

b ) SYNTHETIC : they are the means which contain a defined chemical

composition and quantitatively . They are used to obtain reproducible results.

c ) semisynthetic are synthetic that are added growth factors 13 in a form of a

complex organic extract , for example yeast extract .
nutrient agar :
The nutrient agar culture medium is usually used as routine for all types of
bacteria. It is very useful because it remains solid even at high relative
temperatures. In addition, bacterial growth in agar makes this surface, so better
distinguish small colonies.
Blood agar: It is a rich medium is also used for research of various types of
hemolysis (α, ß or gamma). It is used for growth of streptococci. For the
preparation of blood agar can be used nutrient agar enriched with sodium chloride
or a preparation enriched with other substances

Agar C.L.E.D .: The average C.L.E.D. (Cystine Lactose Electrolyte Deficient) is

recommended for the enumeration and presumptive identification of
microorganisms urinary tract. Its low electrolyte content prevents invasion of crops
by Proteus.La presence of lactose in composition gives it the differential means,
but the interpretation is different from the previous medium by the addition of
another indicator: bromothymol blue. Lactose positive colonies appear yellow and
negative lactose will do so with a greenish, white or blue.

Agar Hektoen: It is a differential and less selective than the SS agar medium. It is
used to facilitate isolation of enterobacteria. The presence of three sugars (lactose,
sucrose and salicin) can extend the differential power of this medium. This medium
is capable of detecting germs SH2 trainers,
C.P.S. ID3. : Chromogenic medium developed by Biomerieux, which allows the
identification of E. coli, P. E. faecalis mirabilisy with simple visualization of the color
change of the colony on the medium (burgundy red, brown blue A).

Mueller-Hinton agar: Mueller Hinton Agar is a medium used for antimicrobial

susceptibility testing of aerobic microorganisms by the Kirby-Bauer method. This
medium is also known as M-H Agar. Bauer, Kirby, Sherris and recommended the
Tuck Hinton Agar Mueler to perform susceptibility testing to antibiotics, using a
single disk impregnated with a high concentration of an antimicrobial.

Agar Trypticase Soy: It is a medium used for the growth of fastidious germs, such
as Brucella, Neisseria or Streptococcus. It is a medium highly enriched, but not
differential. Thioglycollate broth with resazurin: It is a recommended for medium
sterility controls. It allows the cultivation of aerobic, microaerophilic anaerobiosy. It
has a low redox potential that increasing, is manifested by a medium pink.

Chapman agar or saline mannitol salt agar or acronym MSA (Mannitol salt agar)
Mannitol is a culture medium which is normally used in microbiology. It allows the
growth of a particular group of bacteria while inhibiting the growth of others. This
medium is important in the clinical laboratory because it is able to distinguish
pathogenic microorganism in a short time
Baird-Parker Agar: Allows the growth of coagulase-positive staphylococci, while
the difference from the rest.
Loeffler medium: It is a specific medium for the culture of Corynebacterium

Deoxycholate agar Citrate Lactose Sucrose (DCLS): Similar to the SS agar. It is a

differential medium and inhibitor at once. Kligler agar: The agar medium Kligler
(TSI, Triple Sugar Iron) is a differential medium that can help us distinguish
between Enterobacteriaceae species according to their ability (or lack thereof) for:
a. Metabolise lactose, sucrose or both b. Produce acid by fermentation c. Gas
produced during fermentation d. Generate hydrogen sulfide

Typical reactions of some enterobacteriaceae

a) Shigella dysenteriae produces bacillary dysentery ferments glucose (but not

lactose) , it does not produce gas or H2S Far Far inferior.- superior.- Red Yellow

b ) Salmonella typhimurium food poisoning produces ferments glucose (but not

lactose ) produces gas and H2S Far Far inferior.- superior.- Red Black and yellow,
with rising average

c ) Salmonella typhi causes typhoid fever glucose (but not lactose ferments ) , does
not produce gas, produce H2S Far Far inferior.- superior.- Red Black and Yellow

d ) Aerobacter aerogenes similar to but is not Klebsiella respiratory ferments

glucose and lactose , produces gas , and does not produce H2S superior.- Far End
Yellow Yellow inferior.- elevation means 28

e) Escherichia coli is the most common bacteria of the intestinal flora ferments
glucose and lactose , produces gas , and no H2S superior.- Far End Yellow Yellow
inferior.- elevation means

f) Citrobacter freundii nonpathogenic glucose and lactose ferments, producing gas

and H2S Far Far inferior.- superior.- Yellow Yellow and black middle elevation
g) Proteus vulgaris causes infections of the urinary tract, highly mobile glucose and
lactose ferments, producing gas and H2S black superior.- Far Far inferior.-
Amarillo Amarillo and medium elevation
Levine agar or EMB (eosin methylene blue): The agar eosin methylene blue is a
medium used for the isolation and differentiation of Gram-negative enteric bacilli.
The use of eosin and methylene blue allows differentiation of lactose fermenting
colonies nonfermenters. Sucrose is included in the medium to detect coliform
group members that ferment faster sucrose lactose. This medium is considered
superior to ENDO Agar, as it turns out to be a more sensible, stable and secure
environment and allows an earlier differentiation between colonies and non-
fermenting lactose fermenting
Procedure: 1. Collect samples and plant them as soon reach the laboratory,
sowing the plates streak . 2. Incubate plates 35- 37 ° C for 18 to 24 hours and
observe growth. Colonies of Salmonella and Sigel are translucent, amber or
colorless. Coliforms using lactose and / or sucrose produce colonies of blue to
black with dark centers and metallic sheen.
Agar Cetrimida . Agar King:
It inhibits the growth of bacteria due to its action as a quaternary ammonium
compound. Cetrimide Agar base promotes the production of pyocyanin and
fluorescein observed with ultraviolet light in cultures of Pseudomonas aeruginosa ,
which can be considered presumptive evidence for identification. You can then
perform the oxidase test . Sow the test sample on the surface of the culture
medium by cross groove.

Schaedler Agar : A blood medium and vitamin K very suitable for growing
Gardnerella Agar : Agar with human blood over a mixture of antibiotics that allow
observation of beta-hemolytic colonies Gardnerella vaginalis Campylosel
characteristics : It is used for isolation of intestinal Campylobacter growing at 42 °
C in microaerophilic .
Agar BCYE (Buffer Charcoal Yeast Extract ): Used for the isolation of Legionella
Lowenstein Jensen : This is a basis for the preparation of various media for the
isolation , cultivation and differentiation of mycobacteria.