AGM 301 - After Mid Semester Materials

AGM 301 Agricultural Microbiology (1+1)
Lecture 10. Sulphur cycle - sulphur oxidizers; microbial transformation of K, Zn and Si I.
Sulphur cycle
Sulphur like N, is an essential element for all living systems because of it’s inert nature, is not
utilized by plants. Most agricultural soils contain S in the range 20–2000 μg S/ g. It occurs both in
organic (S containing amino acids, vitamins) as well as inorganic forms (Sulphur, sulphates etc.,) .
Of the total Sulphur present in soil, only 10-15% is in the inorganic form (sulphate) and about 75-90 % is
in organic form. Soil microbial biomass S represents 2–3% of the total organic soil S. To be used first S
has to be oxidized or reduced in soil.
S cycle
Cycling of sulphur is similar to that of nitrogen. Transformation / cycling of sulphur between

organic and elemental states and between oxidized and reduced states is brought about by various
microorganisms, specially bacteria. The conversion of organically bound sulphur to the inorganic state by
microorganisms is termed as mineralization of sulphur. The sulphur / sulphate, thus released are either
absorbed by the plants or escapes to the atmosphere in the form of oxides. When plant and animal
proteins are degraded, the sulphur is released from the amino acids and accumulates in the soil, which is
then oxidized to sulphates in the presence of oxygen and under anaerobic condition (water logged
soils) organic sulphur is decomposed to produce hydrogen sulphide (H 2S). H2S can also accumulate
during the reduction of sulphates under anaerobic conditions, which can be further oxidized to sulphates
under aerobic conditions
Four distinct transformations are recognized during the cycling of S
1. Decomposition/Mineralization of larger organic S compounds to smaller units and their

conversion into inorganic compounds
2. Microbial associated S - immobilization
3. Oxidation of inorganic ions and compounds such as sulphides, thiosulphates, Sulphurr
4. Reduction of Sulphates and other sulphides
AGM 301 Agricultural Microbiology (1+1) 1

S transformation in soil
1. Mineralization
The conversion of organically bound sulphur to the inorganic state by microorganisms is
termed as mineralization of sulphur. The breakdown / decomposition of large organic sulphur
compounds leads to production of smaller units and which will be converted into inorganic mineral
compounds (sulphates) by the microorganisms. The rate of sulphur mineralization is about 1.0 to 10.0
percent / year. When Plant and animal proteins are degraded and the S is released from the amino
acids and accumulates in soil. Conversion of organic bound S into inorganic state, mediated through
microorganisms. The released S in either form Sulphur / Sulphate absorbed by plants or escaped into
atmosphere in the form of oxides. Much of the soil organic sulfur is present as sulfate esters, aryl,
alkyl and choline sulfatase enzymes play a key role in sulfur mineralization.
2. Immobilization
S released by mineralization in inorganic form such as Sulphur/Sulphate will be assimilated by
microbial cells, there by converted into organic from, called Immobilization
3. Oxidation of inorganic ions and compounds (S oxidation)

The conversion of inorganic sulfur compounds of lower oxidation state to a higher state, called S
oxidation. (Oxidation is the loss of electrons during a reaction). Conversion of elemental sulphur
and inorganic sulphur compounds such as H2S, sulphite and thiosulphate to sulphate (SO4) occurs
during oxidation.
HS-→ S0 → S2O32- → S4O62- → SO42-.

Oxidation process
Sulphite Sulphate
3 SO 4
Thiosulphate tetrathionate
S + SO 3 S 2O 3 S 4O 6
S + SO 3
SO 3 SO 4
S SO
S oxidizers are categorized into three groups
1. Photoautotrophs
2. Chemolithotrophs
3. Heterotrophs
1. Photoautotrophs
This group included the species of green and purple sulfur bacteria. Based on the colour of the
carotenoid pigment they are named as purple and green S bacteria. They are reported to oxidize
sulphur in aquatic environment. Theses organism uses H2S as electron donor during photosynthesis
in contrast to plants which uses H2O as electron donor and produces Oxygen. They are commonly
found in mud and stagnant waters containing H 2S and exposed to light. They are also found under
extreme conditions of high salinity or temperature and occur in S hot springs and saline lakes as a
colored layer under salt deposits.
Purple S bacteria:
Belong to Chromatacea. Generally found in illuminated anoxic zones of lakes and other aquatic
habitats where hydrogen sulfide accumulates. Also found in "sulfur springs" - where geochemically
or biologically produced hydrogen sulfide can trigger the formation of blooms of purple sulfur
bacteria. Anoxic conditions are required for photosynthesis. They can use S, H 2S, Thisulphides as
electron donors.
 Chromatium, Rhodopseudomonas, Rhodomicrobium
Green sulfur bacteria:
Belong to cholorobiacea. They use sulphide ions, hydrogen or ferrous iron as an electron donor.
Elemental sulphur deposited outside the cell may be further oxidized.
 Chlorobium, Chloropseudomonas, Chlorobacterium, Ancalochloris, Chlorobaculum

2. Chemolithotrophs
They are the members of Thiobacilli.
 Thiobacillus, Thiomicrospira. Beggiatoa, Thiothrix and Thioloca. T. ferrooxidans and T. thiooxidans

are important organisms involved in oxidation of elemental
sulphur to sulphates. Thiobacillus denitrificans can carry out the S
oxidation with NO3 reduction
3. Heterotrophs - including a wide range of bacteria and fungi
 Arthrobacter, Bacillus, Micrococcus, Mycobacterium, and Pseudomonas; some actinomycetes;

and fungi such as Alternaria, Fusarium, and Trichoderma ,Penicillium
 Hydrogen sulfide oxidation has been observed in many unicellular and filamentous cyanobacteria
under anaerobic condition
 The first two groups occur generally in extreme environments such as hot sulfur springs, the last
groups are largely responsible for oxidizing S0 in aerobic soils
Oxidation
 Occurs both in aerobic and anaerobic condition - Bacteria

 Nonfilamentous forms- Thiobacillus
 Filamentous forms – Beggiatoa, Thiothrix and Thioloca
 Fungi and actinomycets - Aspergillus, Penecillium and Microsporium
Importance of S oxidizing bacterium (or) Application of S oxidation
 Produces Sulphuric acid, lower down the soil pH – Hence used in controlling plant disease
 Apple and Potato scab –Streptomyces scabis, Sweet potato rot – S. ipomea are controlled by
addition of Thiobacillus. S+ Thiobacillus application is used for the control
 Use for the remediation of alkali soil
 Increases the solubilization of other nutrients (P,K,Ca,Mn,Al and Mg )
 Preparation of biosuper - is developed in Australia
Addition of Rock phosphate + T. thiooxidans and S enhances the solubilization of P and hence P
availability is increased
 Lipman’s process of Compost preparation is developed in US.
Here the Soil is mixed with manure + elemental S + rock phosphate to get compost.
 In both the cases, the thiobacillus oxidizes S and produce Sulphuric acid, which inturn
solubilizes P present in rock phosphate

4. Sulphate reduction : (Gain of electron is called reduction)
 The conversion of inorganic sulfur compounds of higher oxidation state to a lower state, called
Sulphate. Sulphates and other sulphides are converted to elemental S or H 2S. This reduction
process occurs by two ways
a. Assimilatory sulphate reduction

Sulphate in the soil is assimilated by plants and microorganisms and incorporated into cellular
proteins. This is known as "assimilatory sulphate reduction". (Immobilization)
b. Dissimilatory sulphate reduction
Sulphate can be reduced to hydrogen sulphide (H 2S) by sulphate reducing bacteria which
diminishes the availability of sulphur for plant nutrition. This is “dissimilatory sulphate
reduction” or Respiratory Sulphate reduction. This H 2S produced will be further oxidized to
elemental S by Chlorobium and Chromatium
SO4 reduction is not at all desirable from soil fertility and agricultural productivity view
point. All sulphate-reducing bacteria excrete an enzyme called “desulfurases” or "bisulphate
Reductase".
II. Transformation of Potassium in soil

Potassium plays an important role in the growth and development of plants. It activates enzymes,
maintains cell turgor, enhances photosynthesis, reduces respiration, helps in transport of sugars and
starches, and is essential for protein synthesis. In addition to plant metabolism, potassium improves crop
quality because it helps in grain filling and kernel weight, strengthens straw, increases disease resistance
and helps the plant better to withstand stress.
Potassium in soils
• Soils usually contain Potassium in larger amounts than any other nutrients. Total K content in soil is
3,000- 100,000 kg/ha in top layer of the soil, 98 % bound in the mineral form and 2 % is in soil
solution. Generally, potassium in soils exist in four forms (often called fractions), their
availability to crop roots is considerably different. The order of availability is as follows:
1. Soil solution K (K+ ion)
2. Exchangeable K (easily exchangeable and slowly exchangeable K with other cations )-

readily available to plants
3. Non exchangeable K (reserves of K)

4. K minerals
Fractions/Forms of K in soil
• Exchangeable K - The K adsorbed on soil clay complex and replaceable with neutral salts in
relatively short time, available to plant roots.
• Non exchangeable or Fixed K - Added K which is firmly bound by the soil and not immediately
replaceable with neutral salts
• Mineral K - Muscovite, biotite and K feldspar. The capacity of soils to release lattice K by
weathering depends on the content of K minerals and soil texture.
K cycle in soil
 The potassium (K) cycle is almost entirely inorganic. The major role of K in living organisms
is the osmotic control. K is taken up, retained and excreted in ionic form (K +). The amount of
K in soil solution is relatively small but is in near equilibrium with the much larger amount of
exchangeable K from which it is replenished. Soils also contain K in more slowly exchangeable
forms (fixed K) which act as sources for crops. K present in clay minerals becomes available as
these minerals weather. The immediate source of K for plants is the small amount which is in the
soil solution. The concentration of potassium in the soil solution is of the range from 1 to 2
%. When K in soil solution and exchangeable form are removed by plants uptake, the
equilibrium is disturbed, K in the non-exchangeable and soil mineral fraction will be drawn upon.
K RELEASE BACTERIA IN TRANSFORMATION

K release organisms- Role in transformation process
• Microorganisms like Bacillus extroquens and Clostridium pasteurianum, Bacillus

mucilogenoses, Frateuria aurentia were found to dissolve potassium, silicates and aluminium
from insoluble minerals.
• Aspergillus niger, Pencillium frequentans and Cladosporium, were found to grow on muscovite,
biotite and mica in vitro and also releases K.
Mechanism of K release
• Potassium was solubilized by production of inorganic and organic acids (citric, oxalic, alpha
keto glutaric acids) production of mucilaginous capsules containing of Exopolysaccharides (by
Bacillus, Clostridium and Thiobacillus)
• These organic acids produced, enhance the dissolution of potassium compounds by supplying
protons and by complexing Ca2+ ions
• Microbially produced organic ligands include metabolic by products, extracellular enzymes,

chelates and both simple and complex organic acids enhance the dissolution.
III. Transformation of Silica in soil
Occurs in various forms as soluble silica (monosilicic acid & poly silicic acid), crystalline and
amorphous silica. Silica is generally found in soils as Silicic acid or its ionic form at concentrations
ranging from 0.1 to 0.6 mM. About 30 - 35 number of silicate minerals are found in soils (Mostly of K
containing minerals- biotite and muscovite).

Silicon uptake in plants  Plants can absorb Si in the form of orthosilicic acid along with
water
 As water is lost through transpiration it polymerizes into silica gel through non enzymatic
reaction
 Plants deprived of Si are often weaker structurally and more prone to abnormalities of growth,
development and reproduction and it is the only nutrient which is not detrimental when collected
in excess
Occurrence of silica in soils

 The plants can take up silica in the form of soluble silica as ortho silicic acids from the
soil
 Si is found in plants at concentrations ranging from 0.1 to 10% (103– 105 mg/ kg ; dry
weight basis), an amount equivalent to, or even exceeding, several macronutrients
 Si accumulation has been found to a greater extent, but not exclusively, in
monocotyledonous plants but also in other plant families
 Poaceae, Equisetaceae and Cyperaceae show high Si accumulation (4 % Si), the
Cucurbitales, Urticales and Commelinaceae show intermediate Si accumulation (2– 4 %
Si), while most other species demonstrate little accumulation
 The species which are the richest in Si are monocots.
 Dicots are generally poorer, but there are exceptions.
 “Accumulators” have a Si concentration over 1% and a [Si]/[Ca] ratio >1
 “Excluders” have a Si concentration below 0.5% and a [Si]/[Ca] ratio <0.5
 Plants that do not meet these criteria are called “intermediates.”
Silicon cycle in soils
 In soils, silicate minerals undergo chemical and physical weathering, resulting in the release of Si
in solution
 which is either combined with other elements to form clay minerals or released towards the
streams and the oceans or absorbed by the vegetation
 During litter decomposition, plant Si is re-injected into the soil and can constitute an important
pool of Si in soil
 Crops can take up Si at a much faster rate , 300 kg ha −1 year−1 by sugar cane and 500 kg ha −1
year−1 by rice

 Non-reincorporation of the straw to the field may lead to a depletion of plant-available Si in soils
with a decline of cereal yields
Silicon cycle
Mechanism of silica solubilization
 Several bacteria can solubilize the silica from minerals as well as from plant residues
 Production of organic acids, exopolysaccharides, H2S production and CO2 production enable
the bio dissolution of silicon
 Organic acids play major role in weathering of Alumino silicate minerals
 These bacteria can colonize mineral surfaces, organic acids can supply H + ions and promote
hydrolysis on one hand and make complexes with cations on other hand
 Citric, oxalic, keto, hydroxyl and carboxylic acids can from complexes with cations and promote
their removal
 Biodissolution of Si can also releases K from minerals
 Paenibacillus muciloginosus, Achromobacter xylosoxidans, Bacillus altitutinis, Pseudomonas

gessardii have been identified for silicate solubilization
Benefits of SSB
 Increase available silica content in soil as well as in plants
 Offers resistance to plants against pests to diseases
 Facilitated the rapid breakdown of siliceous residues and release silica
 Silica application through silicate minerals and siliceous rice residues increase the grain
yield of rice

 Dissolution of K containing silicate minerals helps in K nutrition of the crop
AGM 301 Agricultural Microbiology (1+1)

10
IV. Zn in soils
 The native zinc in soils occurs in different chemical pools
 Availability of soil Zn to plants is governed by dynamic equilibrium among the different forms
(which differ in solubility and thus availability to plants) rather than the total content of Zn in
soils.
 These forms include soluble Zn present in soil solution (water soluble), adsorbed on exchange
sites (exchangeable), associated with organic matter, co-precipitated as secondary minerals or
associated with sesquioxides (14 to 38 per cent of total Zn) and clay minerals (24 to 63 per cent)
and as structural part of primary minerals
 The water soluble zinc applied as zinc sulphate gets transformed into different forms like Zn
(OH); zinc carbonate in calcium rich alkali soils ; zinc phosphate in near neutral to alkali soils of
high phosphorus application; and zinc sulphide under reduced conditions.
Zn solubilizing organisms
 Zn solubilizing microorganism are capable of solubilizing insoluble zinc compounds through the
production of gluconic acid.
 Inoculation of zinc solubilizing bacteria either alone in soils inherently rich in native zinc or
along with cheaper insoluble zinc compounds, like ZnO or ZnCO 3, will be useful for enhancing
Zn nutrition of crops
 Inoculation of this organism in zinc deficient soils will help for the alleviation of zinc deficiency
 Bacillus cereus, Bacillus sphaericus, Bacillus subtilis, Gluconacetobacter diazotrophicus,

Pseudomonas sp.
Importance of Zn solubilizing microorganism
 Alter the Zn availability in soil upon application
 Thiobacillus thiooxidans, helps in leaching of Zn from the ore
 Bacillus subtilis helps in adsorption and accumulation of Zn in soil
 Organic acid production by the Zn solubilizing microorganism helps in increasing Zn

concentration in the rhizosphere of crop plants
 Solubilizes the insoluble compounds like, ZnO, Zn CO3, ZnS and Spharelite ore

Lecture 11. Role of microbes in reclamation of problem soils. Microbes in solid waste management
Problem soil
The ‘problem soil’ means the soil that has agricultural problems due to the soil’s unsuitable
physical and chemical properties, or less suitable for cultivation, resulting in that crops are not
able to grow and produce yields as normal.
Types of problem soils

1. Physical problem soils
2. Chemical Problem soils
3. Biological Problem soils
4. Nutritional problem soils as a result of above constraints
Soils with Physical problems

• Slow permeable soils
• Soil surface crusting
• Sub soil hard pan
• Shallow soils
• Highly permeable soils
• Heavy clay soils
• Fluffy paddy soils
Chemical Problem soils
• Salt - affected soils
• Alkali / Sodic soils
• Saline-alkali/ sodic soils
• Acid soils
• Acid Sulphate soils
• Calcareous soil
Biological problems in soils
• Soil organic carbon (SOC) and microbial population
• Earthworms
• Soil Respiration
• Soil Enzymes
Role of microbes in reclamation of problem soils
Reclamation methods
• The application of gypsum (Calcium sulphate) is effective for the improvement of alkaline
or sodic soil. It reacts with the exchangeable sodium present in the soil and converts it into
Sodium sulphate. Sodium sulphate is leached out from the soil to reduce the soil pH.
• Saline soil can be improved by ensuring effective drainage system.
• Scrapping off surface salts from highly saline patches is beneficial for the soil.
• Use of acidifying fertilisers, e.g., superphosphate and Ammonium sulphate is also beneficial.
• Green manuring with dhaincha, sunhemp, mung bean, or addition of organic matter
reduces the salinity of the soil.
• Use of Plant Growth Promoting Rhizobacteria (PGPR) enhances the plant growth by
improving plant growth, nutrient uptake and stress mitigation.
Plant growth in problem soil can be improved by Plant Growth Promoting Microorganism (PGPR) by
following mechanisms
• PGPR elicited induced systemic tolerance (IST) through phytohormone secretion in salt
affected soil for root stimulation so as to confer plant defence against phytopathogens.
• The plant inoculated with PGPR that express ACC deaminase activity facilitate extensive root
growth in seedlings due to reduction in ethylene and thus, imparts resistance to crop plant
against various stresses including salinity.
• Certain microbes modify physical barriers around the roots through secreting exopolysacchrides
(EPS) under salinity stressed conditions. EPS may bind to soil particles to form micro- or
macroaggregates and thus, facilitate plant roots and fungal hyphal network to fit in the soil
pores.
• Microbes alter some of the biochemical pathways and biochemical activities like nitrogen cycle
under stressed conditions of salt and alkaline pH.
Waste:
• Wastes are unwanted or unusable materials. Waste is any substance which is discarded after
primary use, or it is worthless, defective and of no use.
Solid waste
• Generally, solid waste can broadly be categorized into biodegradable and non-biodegradable.
Biodegradable solid waste

• The biodegradables (biowastes) are those solid wastes generated, which could be decomposed by
microorganisms and does not constitute major sources of pollution for a long period of time.
• They include paper products and wastes of plant origin, wastes of animal origin (faecal matter,
carcass, droppings, and poultry waste products).
• These groups of solid waste even though they are easily degraded by microorganism in minimal
time, give off offensive odour and constitute nuisance to the aesthetic environment more than the
non-biodegradable solid wastes.
• They can also constitute a good habitat for the thriving of pathogenic microorganisms which
could easily pollute fresh food product and sources of fresh water.
Non-biodegradable solid wastes

• Non-biodegradable solid wastes are not degradable by microorganisms.
• This implies that other means of treatment such as incineration, landfill, and recycling are
employed as ways of disposing them.
• Examples of this group of solid wastes are solid wastes of metallurgical and smelting industries
(abandoned vehicles, motor cycles, vehicle part and scrap metals, iron, zinc, aluminium sheets
and other metals, machine parts); solids wastes of construction industries (sand, gravel,
bitumen wastes, concrete and waste building materials); solid waste of plastic industries (plastic
buckets, cable insulators, tyres, chairs, tables, cellophane bags, plastic bottles, cutleries, sachet
water containments, etc.) and glass products.
3 R’s of Solid Waste Management
If you have heard of something called the “waste hierarchy” you may be wondering what that
means. It is the order of priority of actions to be taken to reduce the amount of waste generated,
and to improve overall waste management processes and programs.
The waste hierarchy consists of 3 R’s as follows:
• Reduce
• Reuse
• Recycle
Called the “three R’s” of waste management, this waste hierarchy is the guidance suggested for
creating a sustainable life.
The First ‘R’ – Reduce

The concept of reducing what is produced and what is consumed is essential to the waste
hierarchy. The logic behind it is simple to understand – if there is less waste, then there is less to
recycle or reuse. The process of reducing begins with an examination of what you are using, and
what it is used for.
Here are some of things you can do to reduce the waste:
1. Print on both sides of the paper to reduce paper wastage.
2. Use electronic mail to reach out to people instead of sending paper mail.
3. Remove your name from the mailing lists that you no longer want to receive.
4. Use cloth napkins instead of paper napkins.
5. Avoid using disposable plates, spoons, glass, cups and napkins. They add to the problem and
result in large amount of waste.
6. Avoid buying items that are over-packaged with foil, paper, and plastic. This excess packaging
goes to waste.
7. Buy durable goods that have long warranty. They generally run longer and save landfill space.

The Second ‘R’ – Reuse
Learning to reuse items, or re-purpose them for a use different then what they are intended for is
essential in waste hierarchy.
You may either reuse those items for your own use or donate so that others can use them. You
can reuse below items like:
1. Old jars and pots: Old jars and pots can be used to store items in kitchen. They can also be used
to store loose items together such as computer wires.
2. Tyres: Old tyres can either be sent to recycling station or can be used to make tyre-swing.
3. Used wood: Used wood can be used as firewood or can be used woodcrafts.
4. Newspaper: Old newspapers can be used to pack items when you’re planning to move to
another home or store old items.
5. Envelopes: Old and waste envelopes can be used by children to make short notes.
6. Waste paper: Waste paper can be used to make notes and sketches and can be send to recycling
center when you don’t need them anymore.
The Third ‘R’ – Recycling

The last stage of the waste hierarchy is to recycle. To recycle something means that it will be
transformed again into a raw material that can be shaped into a new item. Recycling of waste can
be done by composting and other methods.
Methods of solid waste Management

• Composting
• Windrow composting
• Vermicomposting
• Incineration
• Landfill
• Recycling
Solid waste management: Biodegradable waste management

• Management of solid waste reduces or eliminates adverse impacts on the environment and human
health and supports economic development and improved quality of life.
• Composting is the most frequently used biological solid waste treatment method which is the
controlled aerobic decomposition of organic waste materials by the action of small invertebrates
and microorganisms.
• Composting is a technique in which organic waste materials (food, plants, paper) are decomposed
and then recycled as compost for use in agriculture and landscaping applications.
• The most common composting techniques include Coimbatore method, Bangalore method,
Indore method, windrow method of composting and vermicomposting.
Composting
Composting is the natural process of 'rotting' or decomposition of organic matter by

microorganisms under controlled conditions. Raw organic materials such as crop residues, animal wastes,
food garbage, some municipal wastes and suitable industrial wastes, enhance their suitability for
application to the soil as a fertilizing resource, after having undergone composting.
Advantages of Composting
• Volume reduction of waste.

• Composting temperature kill pathogen, weed seeds and seeds.
• Reduces the risk of pollution  Pathogen reduction
• Additional revenue.
• Suppress plant diseases and pests.
• Reduce or eliminate the need for chemical fertilizers.
• Cost-effectively remediate soils contaminated by hazardous waste.
Compost enrichment
Compost is poor in P content (0.4-0.8 percent). Addition of P makes the compost more
balanced, and supplies nutrient to micro-organisms for their multiplication and faster
decomposition. The addition of P also reduces N losses.
Methods of composting
In Coimbatore method, composting is done in pits of different sizes depending on the waste
material available. A layer of waste materials is first laid in the pit. It is moistened with a suspension
of 5-10 kg cow dung in 2.5 to 5.0 lit of water and 0.5 to 1.0 kg fine bone meal sprinkled over it
uniformly. Similar layers are laid one over the other till the material rises 0.75 m above the ground
level. It is finally plastered with wet mud and left undisturbed for 8 to 10 weeks. Plaster is then
removed, material moistened with water, given a turning and made into a rectangular heap under a shade.
It is left undisturbed till its use.
In the Bangalore method of composting, dry waste material of 25 cm thick is spread in a pit
and a thick suspension of cow dung in water is sprinkled over for moistening. A thin layer of dry waste is
laid over the moistened layer. The pit is filled alternately with dry layers of material and cow dung
suspension till it rises 0.5 m above ground level. It is left exposed without covering for 15 days. It is
given a turning, plastered with wet mud and left undisturbed for about 5 months or till required.
In Coimbatore method, there is anaerobic decomposition to start with, following by aerobic

fermentation. It is the reverse in Bangalore method. The Bangalore compost is not so thoroughly
decomposed as the Indore compost or even as much as the Coimbatore compost, but it is bulkiest.
In the Indore method of composting, organic wastes are spread in the cattle shed to serve as
bedding. Urine soaked material along with dung is removed every day and formed into a layer of about
15 cm thick at suitable sites. Urine soaked earth, scraped from cattle sheds is mixed with water and
sprinkled over the layer of wastes twice or thrice a day. Layering process continued for about a fortnight.
A thin layer of well decomposed compost is sprinkled over top and the heap given a turning and
reformed. Old compost acts as inoculum for decomposing the material. The heap is left undisturbed for
about a month. The compost is ready for application in another month.
In windrow method large volume of compost is generated. Organic wastes are formed into rows
of long piles called windrows and aerated by turning piles periodically. Height of pile is 4-8 feet and
width is 14-16 feet.
Crop Residue (Agricultural Waste) Composting
Crop residues are the non-economic plant parts that are left in the field after harvest. The harvest
refuses include straws, stubble, stover and haulms of different crops. Crop remains are also from
thrashing sheds or that are discarded during crop processing. This includes process wastes like groundnut
shell, oil cakes, rice husks and cobs of maize, sorghum and cumbu. The greatest potential as a biomass
resource appears to be from the field residues of sorghum, maize, soybean, cotton, sugarcane etc.
Steps involved in compost making
1. Waste collection
Crop residues accumulated in different locations are to be brought to compost yard. The compost
yard is located in anyone corner of the farm with accessibility via good road. Water resource should also
be available in sufficient quantity. The crop residues that are brought to compost yard should be heaped in
one corner for further processing.
2. Shredding of waste materials
Particle size is one of the factors that influence the composting. It is advisable to shred all the
crop residues that are used for composting. Shredding the waste manually is labour intensive. Shredder
machine can be employed to shred all the crop residue biomass.
Particle size of 2 to 2.5 cm is recommended for quick composting.
3. Mixing of green waste and brown waste
Carbon and nitrogen ratio decides the initiation of composting process. If C:N ratio is wide
(100:1) composting will not take place. Narrow C:N ratio of 30:1 is ideal for composting. To get a narrow
C:N ratio, carbon and nitrogen rich material should be mixed together. Green coloured waste materials
like glyricidia leaves, parthenium, freshly harvested weeds; sesbania leaves are rich in nitrogen, whereas
brown coloured waste material like straw, coir dust, dried leaves and dried grasses are rich in carbon. In
any composting process these carbon and nitrogen rich material is to be mixed together to make the
composting quicker rather than putting green waste alone or brown color waste alone for composting.
Animal dung is also a good source of nitrogen. While making heap formation, alternative layers of carbon

rich material, animal dung and nitrogen rich material are to be heaped to get a quicker result in
composting.
4. Compost heap formation
Minimum 4 feet height should be maintained for composting. The composting area should be
elevated one and have sufficient shade. While heap formation, all the crop residues should be mixed
together to form a heterogeneous material rather than a single homogenous material. Alternate layers of
carbon and nitrogen rich material with intermittent layers of animal dung are essential. After heap
formation the material should be thoroughly moistened.
5. Aerating the compost material

Sufficient quantity of oxygen should be available inside the compost heap. For this external air
should be freely get in and comes out of the material. Normally to allow the fresh air to get inside, the
compost heap should be turned upside down, once in fifteen days. In this process top layer comes to
bottom and bottom layer goes to top. This process also activates the microbial process and compost
process is hastened. In some cases air ventilating pipe maybe inserted vertically and horizontally, to allow
the air to pass through. The wood chip that is available as waste in wood processing industry may also be
used as bulking agent in the composting process. This bulking agent gives more air space to the compost
material.
6. Moisture maintenance
Throughout the composting period 60% moisture should be maintained. On any situation,
compost material should not be allowed to dry. If the material becomes dry, all the microorganisms
present in the crop residues will die and the compost process gets affected.
7. Maturity
Volume reduction, black colour, earthy odor, reduction in particle size are all the physical
factors to be observed for compost maturity. After satisfying with the compost maturity index, the
compost heap can be disturbed and spread on the floor for curing.
8. Compost application
Organic manures are highly regarded as good source of material to maintain soil health and
increasing soil organic carbon content. Organic manures cannot be equated with inorganic fertilizers. But
organic manures deliver all the nutrients to the soil but with little quantity. For one hectare of land 5
tons of enriched biocompost is recommended. It can be used as basal application in the field before
taking up planting work.
Microorganism and temperature changes in composting
In the process of composting, microorganisms break down organic matter and produce carbon dioxide,
water, heat, and humus, the relatively stable organic end product. Under optimal conditions, composting
proceeds through three phases: 1) the mesophilic, or moderate-temperature phase, which lasts for a

couple of days, 2) the thermophilic, or high-temperature phase, which can last from a few days to several
months, and finally, 3) a several-month cooling and maturation phase.
• Different communities of microorganisms predominate during the various composting phases.

Initial decomposition is carried out by mesophilic microorganisms, which rapidly break down
the soluble, readily degradable compounds. The heat they produce causes the compost
temperature to rapidly rise.
• As the temperature rises above about 40°C, the mesophilic microorganisms become less
competitive and are replaced by others that are thermophilic, or heat-loving. At temperatures of
55°C and above, many microorganisms that are human or plant pathogens are destroyed. Because
temperatures over about 65°C kill many forms of microbes and limit the rate of decomposition,
compost managers use aeration and mixing to keep the temperature below this point.
• During the thermophilic phase, high temperatures accelerate the breakdown of proteins, fats,
and complex carboydrates like cellulose and hemicellulose, the major structural molecules in
plants. As the supply of these high-energy compounds becomes exhausted, the compost
temperature gradually decreases and mesophilic microorganisms once again take over for the
final phase of "curing" or maturation of the remaining organic matter
Vermicomposting
• Vermicomposting is the process of turning organic debris into worm castings. The worm
castings are very important to the fertility of the soil. The castings contain high amounts of
nitrogen, potassium, phosphorus, calcium, and magnesium. Vermicompost is nothing but the
excreta of earthworms, which is rich in humus and nutrients.
• The African earthworm (Eudrillus engenial), Red worms (Eisenia foetida) and composting
worm (Peronyx excavatus) are promising worms used for vermicompost production.
Management of oil spill (Degradation of oils/hydrocarbon)
• Crude oil (or petroleum): a liquid mixture of a variety of hydrocarbon compounds derived from
ancient algal and plant remains and found in reservoirs under the Earth’s surface. Nitrogen and
sulfur containing molecules (“resins”) are common constituents of some crude oils.
• Leaks and accidental spills occur regularly during the exploration, production, refining, transport,
and storage of petroleum and petroleum products.
• The process of bioremediation, defined as the use of microorganisms to detoxify or remove
pollutants owing to their diverse metabolic capabilities is an evolving method for the removal and
degradation of many environmental pollutants including the products of petroleum industry.
• There are at least 22 genera of bacteria that can metabolize petroleum hydrocarbons which
include- Pseudomonas, Aeromonas, Bacillus, Flavobacterium, Corynebacterium, Micrococcus
etc.
• Based on crude oil degradation capacity Pseudomonas aeruginosa is the most active
hydrocarbon utilizer in crude oil.
• Prof. Chakrabarty (Indian American microbiologist) genetically engineered a new species of
Pseudomonas bacteria ("the oil-eating bacteria") in 1971 while working for the Research &
Development Center at General Electric Company in Schenectady, New York.
• At the time, four known species of oil-metabolizing bacteria were known to exist, but when
introduced into an oil spill, they competed with each other, limiting the amount of crude oil that
they degraded.
• The genes necessary to degrade oil were carried on plasmids, which could be transferred among
species. By irradiating the transformed organism with UV light after plasmid transfer, Prof.
Chakrabarty discovered a method for genetic cross-linking that fixed all four plasmid genes in
place and produced a new, stable, bacterial species (now called Pseudomonas putida) capable of
consuming oil one or two orders of magnitude faster than the previous four strains of oil-eating
microbes. The new microbe, which Chakrabarty called "multiplasmid hydrocarbon-degrading
Pseudomonas," could digest about two-thirds of the hydrocarbons that would be found in a
typical oil spill.
• The bacteria drew international attention when he applied for a patent—the first U.S. patent for
a genetically modified organism. (U.S. utility patents had been granted to living organisms
before, including two pure bacterial cultures, patented by Louis Pasteur.

Principle of aerobic degradation of hydrocarbons by microorganisms
The initial intracellular attack of organic pollutants is an oxidative process and the activation as
well as incorporation of oxygen is the enzymatic key reaction catalyzed by oxygenases and
peroxidases.
Lecture 12. Biodegradation of agricultural residues and chemicals- processes involved in

remediation
Biodegradation of agricultural residue can be done by the process of composting. Agricultural

residues are materials left in an agricultural field or orchard after the crop has been harvested. These
residues include stalks and stubble (stems), leaves, and seed pods.
Composting
Detail information about composting is given in previous chapter (Lecture 11)
Xenobiotic
A xenobiotic is a synthetic chemical not produced by organisms in nature. Xenobiotics include
pesticides, polychlorinated biphenyls (PCBs), munitions, dyes, and chlorinated solvents, among many
other chemicals.

Bioremediation:
Process of detoxification of toxic chemicals / contaminants in the soil and other environment by using
microorganisms.
Phytoremediation:
Phytoremediation is a bioremediation process that uses various types of plants to remove, transfer,
stabilize, and/or destroy contaminants in the soil. There are several different types of phytoremediation
mechanisms.
1. Phyto-stabilization - plant immobilize contaminants
2. Phyto-accumulation - contaminats are stored in the plants aerial shoot
3. Phyto-volatilization - plants take up water containing organic contaminants and release the
contaminants into the air through their leaves.
Agricultural chemical residue biodegradation (Bioremediation)

Over 1000 pesticides have been marketed worldwide for pest control purposes. Pesticides include
herbicides, insecticides, and fungicides. Pesticides display a wide variety of chemistries, and include
chlorinated, aromatic, and nitrogen- and phosphorus containing compounds.
Some of these substances can be used as carbon and energy sources by microorganisms, whereas others
are utilized only poorly or not at all. Highly chlorinated compounds are typically the pesticides most
resistant to microbial attack. However, related compounds may differ remarkably in their degradability.
For example, chlorinated compounds such as DDT persist relatively unaltered for years in soils, whereas
chlorinated compounds such as 2,4-D are significantly degraded in just a few weeks.
Dechlorination
Many xenobiotics are chlorinated compounds and their degradation proceeds through dechlorination.
For example, the bacterium Burkholderia dechlorinates the pesticide 2,4,5-T aerobically, releasing
chloride ion (Cl-) in the process; this reaction is catalyzed by oxygenase enzymes. Following
dechlorination, a dioxygenase enzyme breaks the aromatic ring to yield.
Biodegradation of the herbicide 2,4,5-T. Pathway of aerobic 2,4,5-T biodegradation; note the importance
of dioxygenase enzymes in the degradation process.

Example of Biodegradation of Pesticides
Dichloro Diphenyl Tricholroethene (DDT):
• Probably the best known example of a chlorinated pesticide.
• This compound has been used in vast quantities as insecticide to control numerous insect pests.
• However, its use has been banned or restricted in many countries because of its deleterious effects.
• DDT present in the soil can be degraded in two years, while others have found that the process can take
from fifteen to twenty years or more.
• The degradation products of DDT are mainly the dechlorination products DDE and DDD.
• The pathway can be DDT DDE DDD, or from
DDT to DDD directly.
• Anaerobically, DDE is readily dechlorinated to DDNU and then to further products.
• Among microorganisms, bacteria comprise the major group involved in DDT degradation, especially
soil habitants belonging to genera Bacillus, Pseudomonas, Arthrobacter and Micrococcus.
Biodegradation of carbenazim Pseudomonas sp.
Microbial enzymes in the bioremediation of pesticides

Enzyme Organism Pesticide
Pseudomonas sp. LBr
Oxidoreductases (Gox) Glyphosate
Agrobacterium strain T10
Mycobacterium sp.
Monooxygenases Endosulphan and Endosulphato
Arthrobacter sp.
Pseudomonas putida Herbicides Trifluralin 2,4,5-T
Dioxygenases (TOD)
Burkholderia
Agrobacterium radiobacter
Phosphotriesterases
Pseudomonas diminuta Insecticides phosphotriester
OPH/OpdA
Flavobacterium sp.
Haloalkane Dehalogenases Sphingobium sp.
Hexachlorocyclohexane (β and δ isomers)
LinB Shingomonas sp.

CHEMICAL REACTION LEADING TO BIODEGRADATION

Lecture 13. Importance of soil and plant microbiomes – rhizosphere, spermosphere, phyllosphere,
epiphytic and endophytes. Plant growth promoting microbes-types and mechanism of action.
Rhizosphere
The term Rhizosphere was introduced by the German scientist Hiltner in 1904. It is region of the soil
which is subjected to the influence of plant roots. It is the unique environment under the influence of
plant roots and is characterized by greater microbiological activity than the soil away from plant roots.
Rhizosphere support the growth of microorganisms that root-derived compounds as sources of C, N,
Energy.
The term rhizsophere is often divided into three general areas
1. Endorhizosphere : inner rhizosphere (very root surface- cell layers of the root (cortex ) potentially
colonizable by microorganisms) - The microbial numbers are larger in this zone zone, where the bio
chemical interactions between microorganism and roots are most pronounced.
2. Ectorhizosphere : outer rhizosphere (immediate adjacent soil), the area surrounding the root
3. Rhizoplane: surface of plant root with strongly adhering soil particles; provides microenvironment-
soilplant interface – for microbial activity
In rhizosphere and rhizosplane, higher organism contributes excretory products and sloughed off tissue,
and certain benefits are derived from the microscopic organisms.
Rhizosphere effect
It indicates the overall influence of plant roots on soil microorganisms. It can be put on a quantitative
basis by the use of R-S ratio. The rate of metabolic activity of the rhizosphere microorganisms are
different from those of non-rhizosphere soil.

R:S Ratio
It is defined as the ratio of microbial numbers per unit weight of rhizosphere (R), to the numbers in
a unit weight of the adjacent non rhizosphere soil (S). The rhizosphere effect is consistently greater for
bacteria than for other microbe.
Microflora of the root zone

• Bacteria are quite abundant and the greatest rhizosphere effect is observed with bacteria (R: S
values ranging from 10-20 or more) than with actinomycetes and fungi.
• Fungi and Actinomycetes are observed but not at higher frequency
• Protozoa are relatively prominent, particularly small flagellates and large ciliates.
• Gram-negative, rod shaped, non-sporulating bacteria which respond to root exudates are
predominant in the rhizosphere (Pseudomonas, Agrobacterium), which almost make upto larger
% of the rhizosphere
• While Gram-positive, rods, Cocci and aerobic spore forming (Bacillus, Clostridium) are
comparatively rare in the rhizosphere.
• No selective stimulation or inhibition of gram variable rods, cocci, non spore – formers etc.
• The most common genera of bacteria are: Pseudomonas, Arthrobacter, Agrobacterium,
Alcaligenes, Azotobacter, Mycobacterium, Flavobacter, Cellulomonas, Micrococcus and others
have been reported to be either abundant or sparse in the rhizosphere.
• Bacterial density is 109/g rhizosphere soil. It may cover 4-10% of the root area, occurring
profusely on the root hair region and rarely in the root tips.
• Fast growers and metabolic active population are dominating in the rhizohsphere. This suggest
that the rhizosphere flora has a greater ability to effect rapid biochemical changes than the
organisms of fallow land.
• Qualitative changes / selective influence of the root system is exhibited.
Ex: Preferential stimulation of G-ve, non spore forming short rods.
- Abundance of N2 fixing P solubilizing organism
- Preponderance of amino acid requiring (bacteria)
- Preferential stimulation of vitamin requiring (bacteria)
• In contrast to their effects on (bacteria), roots do not appreciably alter the total counts of fungi
and the influence is selective on fungal genera. The spectrum of fungal genera varies with plant
species and age and kind of the soil.
• Actinomycetes and algae are not significantly benefited by their proximity to roots, and the R:S
ratio may rarely exceed 2 or 3:1.

Root exudates
One of the most important factors responsible for rhizosphere effect is the great variety of organic
substances available at the root region by the way of exudates from roots, which directly or indirectly
influence the quality and quantity of the microorganism in the root region.
The substances exuded by plant roots contains amino acids, sugars, organic acids, Fatty acids
vitamin, nucleotides, flavonones, enzymes and many other unidentified substances. The
microorganisms uses the plant derived carbon compounds as well as the root derived rhizodeposits
include sloughed-off root cells or lysates as energy sources.
Influence of the rhizosphere microflora on plant growth
The rhizosphere community have either a favourable or a detrimental influence of plant development.
Because the microflora is so intimately related with the root system, partially covering its surface, any
beneficial or toxic substance produced can cause an immediate response.
• Vast microorganisms demands a variety of anions and cations for its own development and
immobilization may takes place
• Aerobic (bacteria) remove O2 from the environment and add CO 2 and either by lowering O 2 or
increasing CO2 tension, may reduce root elongation and development and reduce the water and
nutrient uptake.
• It may however favour plant development by producing growth stimulating substances,
contributes to the formation of a stable soil structure, releasing elements in organic forms
through the mineralization of organic complexes, and by entering into symbiotic root
associations.
• Play a major role in nutrient transformation by the way of involving in solubilization and
mineralization reactions
• Rhizosphere community is mainly of nonpathogenic microorganism but, the biological
interactions in rhizosphere may lead to harmful and beneficial effect
• Rhizosphere may be considerable as a microbiological buffer zone in which the microflora serves
to protect the plant from the attack of the pathogen. Antibiotic production by the root microflora
is often cited.

Ephiphytic organisms
Epiphytic bacteria are bacteria which live non-parasitically on the surface of a plant on various
organs such as the leaves, roots, flowers, buds, seeds and fruit. They generally don’t harm the plant,
some produce an auxin hormone which promotes plant growth and plays a role in the life cycle of the
bacteria. Different bacteria prefer different plants and different plant organs depending on the organ's
nutritional content, and depending on the bacteria's colonization system which is controlled by the host
plant. Bacteria which live on leaves are referred to as phyllobacteria, and bacteria which live on the root
system are referred as rhizobacteria. They adhere to the plant surface as cluster, individual bacterial cell
and as biofilm.
Phyllosphere and Phylloplane
The term phyllosphere was first introduced by the plant pathologist F.T. Last in 1955.The microbiologist
J. Ruinen drew the same analogy at nearly the same time (Ruinen 1956). Five years later (In1961),
Ruinen wrote: “The external surface of the leaf, as an environment for microorganisms, has been
termed as ‘phyllosphere’ by analogy with the rhizosphere of roots...and the term has been
accepted”
The phyllosphere is a term used in microbiology to refer to the total above-ground portions of plants
as habitat for microorganisms. The phyllosphere can be further subdivided into the caulosphere
(stems), phylloplane (leaves), anthosphere (flowers) and carposphere (fruits) .

Microorganisms in phyllosphere and phylloplane
• Filamentous fungi are considered transient inhabitants of leaf surfaces, being present
predominantly as spores, where as rapidly sporulating species and yeasts colonize this habitat
more actively.
• Bacteria are by far the most abundant inhabitants of the phyllosphere.
• Bacterial communities are generally dominated by Proteobacteria, α-Proteobacteria (e.g.,
Methylobacterium, Sphingomonas) are often most abundant, although γ-Proteobacteria (e.g.,
Pseudomonas) may sometimes reach high levels.
• Fungal communities typically are dominated by Ascomycota. Within Ascomycota, the genera
Aureobasidium, Cladosporium, and Taphrina are often among the most common.
Basidiomycetous yeasts belonging to the genera Cryptoccoccus and Sporobolomyces are
generally abundant
Pink pigmented Facultative Methylotrophs (PPFM)
• The phyllosphere is a well-known habitat of methanol-utilizing methylotrophs, and leaf surfaces
are colonized by a large population of these bacteria, which include the genera
Methylobacterium
(PPFM), Methylophilus, Methylibium and Hyphomicrobium. In general apart from Pink Pigmented
Methylotrophic bacteria (Methylobacterium), other genera such as Pseudomonas,
Pseudobacterium, Phytomonas, Erwinia, Sarcina and other unidentified ones have been
encountered on plant surface, especially on leaf surface.
Plant leaf colonization by Methylotrophs
Naturally occurring epiphytic methylobacteria as coevolved symbionts of the plants. Growing plant cells
produce waste products (methanol; leakage of amino acids etc.) (1) that are taken up and metabolized by
the bacteria (2) which may release ammonium-ions (3). The extracellular prokaryotic “waste managers”
produce and secrete cytokinins and auxin (4). These bacterial signals (5) may indicate to the plant that
epiphytes are present and active. The exogenously produced phytohormones stimulate the growth of the
plants (6).

Naturally occurring epiphytic methylobacteria as coevolved symbionts of the plants. Growing plant cells
produce waste products (methanol; leakage of amino acids etc.) which are taken by and metabolized by
the bacteria and release ammonium-ions. The bacteria can produce signals and secrete cytokinins and
auxin. These signals are received by the plant, there after the exogenously produced phytohormones
stimulate the growth of the plants Spermosphere.
Spermosphere
The spermosphere is the region of soil directly under the influence of seeds and the critical interface
between plants and microbes where beneficial and detrimental interactions occur. This is area of
increased microbial activity around seed (imbibing, germinating) in soil- 5 to 10mm zone. The
spermosphere has only a temporary existence during germination : before germination, there is no
interaction between the mature dry seed and the soil, while after the emergence of the radical, the soil
environment surrounding the seed is defined as the rhizosphere.
When seeds imbibe, a variety of compounds are passively exuded from them into the spermosphere. The
composition of these seed exudates varies with plant species and with other factors such as soil type, soil
pH, soil moisture, temperature, available nutrients in the soil and the presence of micro-organisms.
Exudates release from germinating seeds are usually ‘normal products’ of seed metabolism and they
generally consist of carbohydrates, amino acids, flavonoids, sterols and salts.
Volatile compounds such as aldehydes, alcohols, ethylene, CO 2 and volatile carboxylic acids are also
released from germinating seeds. Most of the carbohydrates exuded by seeds are simple sugars such as
sucrose, glucose, fructose and maltose, but the ratios of these sugars are strongly dependent on the
developmental stage. Seed exudates are crucial to initiate the establishment of rhizobacteria, which can
affect plant development and health
Endophytic bacteria
Endophytes are bacterial or fungal microorganisms that colonize healthy plant tissue intercellularly
and/or intracellularly without causing any apparent symptoms of disease. They are ubiquitous,
colonize in all plants, and have been isolated from almost all plants examined till date. Bacteria enter
plants through roots and spread to other parts of the plant.

Their association can be obligate or facultative and causes no harm to the host plants. They exhibit
complex interactions with their hosts which involves mutualism and antagonism. Plants strictly limit the
growth of endophytes, and these endophytes use many mechanisms to gradually adapt to their living
environments. In order to maintain stable symbiosis, endophytes produce several compounds that
promote growth of plants and help them adapt better to the environment. Endophytic bacteria have
more advantage over rhizospheric bacteria because they get the opportunity to stay in direct contact
with the plant tissues. Also, they offer more beneficial effects to the plants as compared to bacteria
residing outside plants.
Bacterial root colonization often starts with the recognition of specific compounds in the root exudates by
the bacteria. The most common mode of entry of endophytic bacteria into plant tissues is through
primary and lateral root cracks, and diverse tissue wounds occurring as a result of plant growth. As
soon as their cells are inside the plant, competent endophytes respond to plant cues to enable further
induction of cellular processes necessary for entering the endophytic life stage and spreading to other
(intercellular) tissues of the root cortex and beyond. Production of enzymes, such as cellulases
(endoglucanases and endopolygalacturonidases) seems to be obligatory in this process to degrade the cell
wall components. At this point, competent endophytes can quickly multiply inside the plant often
reaching high cell numbers. Ex. Bacillus, Gluconacetobacter, Pantoae etc.,

Plant growth promoting microbes - mechanism of action
Phytohormone Production
PGPRs microbial population mainly promote plant health, they stimulate growth promoting activity
directly by changing and producing the plant growth regulators concentrations like cytokinins, gibberellic
acid (GA) and Indole-3-acetic acid (IAA) etc. In that case, auxins are the main key hormones which
control plant health and growth.

Phosphate Solubilization
These bacteria secrete different types of organic acids (e.g., carboxylic acid) thus lowering the pH in the
rhizosphere and consequently release the bound forms of phosphate like Ca 3 (PO4)2 in the calcareous soils.
Utilization of these microorganisms as environment-friendly biofertilizer helps to reduce the use of
expensive phosphatic fertilizers.
Siderophore Production
PGPR are reported to secrete some extracellular metabolites called siderophores. For the first time
Kloepper (1980) reported the significance of siderophores produced by certain genera of PGPR in plant
growth promotion. Siderophores are commonly referred to as microbial Fe-chelating low molecular
weight compounds. The presence of siderophore-producing PGPR in rhizosphere increases the rate of
Fe3+ supply to plants and therefore enhance the plant growth and productivity of crop. Siderophore
producing PGPR helps in bio-control of root pathogen by creating competition for Fe 3+.
Nitrogen fixation
PGPR has the ability to fix the atmospheric nitrogen by using nitrogenase enzyme. Rhizobium,
Azospirillum, Azotobacter and Gluconacetobacter are involved in nitrogen fixation by establishing
symbiotic, associative symbiotic, free living and endophytic association with plants respectively.
Stress ethylene reduction

PGPR has the ability to produce ACC-deaminase (ACCD) is to reduce the ethylene production level in
the roots system of the growing plants. ACCD breakdown 1-aminocyclopropane-1-carboxylic acid (ACC)
in to ammonia and alpha ketobutryrate. 1-aminocyclopropane-1-carboxylic acid (ACC) is the precursor of
ethylene. Particularly Pink Pigmented Facultative Methylotrophs (PPFM) Methylobacterium involved in
the stress ethylene reduction in plants grown under salt, drought and heavy metal stress conditions.
PGPR as Biocontrol Agents
Besides siderophore production, the biocontrol abilities of PGPR strains essentially depend on aggressive
root colonization, induction of systemic resistance in the plant, and production of antifungal
antibiotics.
The production of one or more antibiotics is the mechanism most commonly associated with the ability of
plant growth-promoting bacteria to act as antagonistic agents against phytopathogens. Antibiotics, such as
polymyxin, circulin and colistin, produced by the majority of Bacillus ssp. are active against Gram-
positive and Gram-negative bacteria, as well as many pathogenic fungi.
Non-pathogenic rhizobacteria have been shown to suppress disease by inducing a resistance mechanism
in the Plant growth-promoting rhizobacteria plant called “Induced Systemic Resistance” (ISR).
Induced Systemic Resistance (ISR) is a resistance mechanism in plants that is activated by infection. Its
mode of action does not depend on direct killing or inhibition of the invading pathogen, but rather on
increasing physical or chemical barrier of the host plant. ISR instead relies on signal transduction
pathways activated by jasmonate and ethylene.

Systemic Acquired Resistance (SAR) a plant can develop defences against an invader such as a
pathogen or parasite if an infection takes place. SAR which is triggered by accumulation of
pathogenesis-related proteins or salicylic acid.
Lecture 14. Soil microorganisms and their interactions – positive and negative interactions. An
overview of industrially important microorganisms and products.
Microorganisms live in soil as complex populations. Nutrient limitations and environmental stresses
make many soil microorganisms to interact with one another to circumvent the difficulties. Through
these interactions, varieties of relationships occur between different microorganisms either between
themselves or with plants. The composition of the micro flora of any habitat is governed by biological
equilibrium created by the associations and interactions of all individuals found in the community.
Microbial interactions in soil are considered as one of the most important activities that occur in
the terrestrial ecosystem. They affect all the dynamic processes of plants and other living organisms that
live near from them either directly or indirectly.
There are two types of microbial interaction that occur in soil. The interactions that occur between
individuals within the same species are called intraspecific interaction, and those that occur between
organisms of different species either two microbial populations or microbial population and plants or
animals are called interspecific interactions. Each microorganism could perform more than one type of
interaction depending on the sounding environmental conditions, its partner in the interaction. Microbial
interactions are very essential for plant growth and health.
The interactions occurring in soil can also be classified according to the partners involved in association.
a. Plant microbe interaction
b. Plant - microbe – microbe interaction
c. Microbe – microbe interaction and

a) Plant microbe interaction
It mainly constitutes the association of microorganism with plants little in a positive way or in a negative
way. The positive approach is mainly the symbiotic relationships and the negative approach
constituents mainly pathogen plant interactions.
b) Plant microbe – microbe interaction
Also called tripartite symbiosis. Interaction between the macrobiont and a microbiont
Eg: Alnus – Frankia –Mycorrhiza
Casuarina – Frankia – Mycorrhiza
c) Microbe – microbe interaction
Different species or genera interact in a positive or negative way and exhibit various types of
intererationship.
Interrelationship between microorganisms: Beneficial and harmful relationship
Microbial ecosystem of soil is the sum of the biotic and the abiotic components of soil. Many of these
organisms depend upon one another for direct and indirect nutrients.
The microorganisms that inhabit the soil exhibited many different types of associations or interactions.
Some of the associations are indifferent or neutral, some are beneficial type of interactions and some are
detrimental or negative.
Beneficial interactions Harmful interactions
Neutralism Competition
Symbiosis / mutualism Amensalism
Protoco-operation Parasitism
Commenalism Predation

I. Beneficial / positive interactions
A. Neutralism
B. Symbiosis / mutualism
C. Protoco-operation
D. Communalism
A. Neutralism
It is a type of neutral association, in which two dissimilar organisms inhabiting the same
environment without impacting each other microorganisms as entirely independent. Each could
utilize different nutrients without producing metabolic end products that are inhibitory.
B. Symbiosis / Mutualism
It is an obligatory or highly specific interaction between two populations in which both of them benefit
from each other. It usually requires close physical connection in which both partners can act, as if they
are one, when in separate their metabolic activities are different.
Based on the partner selection it is classified as obligate symbiosis and facultative symbiosis. Based on
the purpose of interaction also it is classified. Based on the purpose it is classified as service-resource
type, resource-resource type and service to service type
1. Obligate symbiosis: It occurs when both microorganism live together in close proximity, and both
species cannot survive without its mutualistic partner.
Symbiotic association is evident in soil among several groups of organisms algae and fungi in lichens,
bacteria residing within protozoan cells, bacteria and roots in the Rhizobium legume symbiosis, fungi
and roots in mycorrhizae.

a. Lichens
In lichens, the algae and fungi are in intimate physical and physiological relationship. The alga
benefits from the protection afforded by the fungal hyphae that envelop and protect it from environmental
stresses. While the fungi gains benefit by making use of the CO 2 fixed by its photosynthetic partner as
well as the oxygen. Where ever the BGA are the participants, the fungi also benefits from the fixed N 2.
b. Mycorrhizae
It is a mutualistic association among mycorrhizal fungi and plant roots, in which plants provide
fungus with carbohydrates and offer it protection. In turn the fungus increases the surface area of plant
roots for absorbing water, nitrogenous compounds, phosphorus, and other inorganic nutrients (e.g.,
phosphate) from the surrounding soil and delivers them to the plant which improves plant growth and
health. c. Symbiotic N2 Fixation
The nitrogen-fixing bacteria provide the plants with nitrogenous compounds, while in return the plants
provide the nitrogen-fixing bacteria with carbohydrates. This mutualistic association improves plant
growth and health, and it has different types which include Rhizobium spp. with root nodules of legume
plants and Frankia which is an actinomycete (nodule-forming filamentous bacteria) with the roots of
Alnus and Casuarina trees which are “nonlegumes”
2. Facultative mutualism : It occurs when one of the two partners can survive without its mutualistic
partner by itself in some conditions. Membership in this association is not usually specific and one
organism can be replaced by the other. It is also termed synergism. It is a loose association
C. Proto co-operation
Synergism (protocooperation) is a relationship that occurs between two or more populations at which
both or all of them benefit. In this relationship microbial populations perform a function which may not
be performed individually or produce a new product that neither each population can produce alone.
This relationship is different from mutualism because as it is not an obligatory interaction, none of the
species depend on the relationship for existence, as each member can live and produce its own food
individually.
Types of Protocooperation
There are different types of protocooperation relationship that could be found in the terrestrial ecosystem
which is considered very useful in agriculture:
• Nutritional protocooperation: It is the most popular relationship between terrestrial populations

at which the populations exchange nutrients between each other. Such a cooperation is also called
syntrophism protocooperation.
Nutritional proto co-operation has been demonstrated in cultures. Eg: In a medium deficient in
nicotinic acid and biotin, neither Proteus vulgaris nor Bacillus polymyxa will multiply as the former
bacterium requires nicotinic acid and the latter needs biotin. In mixed culture, in the same medium
however both grown since the partner bacterium synthesizes the missing vitamins.

Protocooperation also occurs between higher plants growing in the soil and bacteria or fungi living in the
rhizosphere. Neither each of them is dependent on this association, since bacteria and fungi get benefits
from the exudates of plant roots and interact with each other to form the essential nutrients
necessary for plant’s growth such as decomposed organic materials, production of phytohormones,
minerals, water, vitamins, and amino acids which in return improve soil fertility as well as the plant
health and growth
• Metabolism of toxic end products: In this type of association one organism establishes its
association by eliminating toxic substances from the habitation versus obtaining carbon products made by
the other associate partner.
• Production of derivative enzymes: Arthrobacter and Streptomyces (soil flora) produce enzymes
which collectively degrade diazinon which is an organophosphate pesticide (useful in the degradation of
xenobiotics or recalcitrant compounds).
D. Commensalisms
It is a relationship at which one population benefits, while the other population is unaffected
(neither harmed nor benefited). It is a very common relationship between different microbial populations.
It is the type of beneficial association, in which only one population benefits, while the other population
is unaffected (neither harmed nor benefited). This is a very common relationship between different
populations.
This occurs commonly in soil with respect to degradation of complex molecules like cellulose and lignin.
One patter can attack a substrate not available to the second organism, but the decomposition results in
the formation of products utilized by the second. The one which offer benefit is called commensal. eg:
Many fungi able to degrade cellulose and yield glucose and organic acids. This can serve as a
nutrient source for many bacteria and fungi, which are noncellulolytic
III. Negative / harmful / deleterious interactions
Detrimental effects of one species on its neighbours are quite common in soil, and they are demonstrated
by the decreases in abundance or metabolic activities of the susceptible organisms. It consists of different
relationships between different populations either two or more, at which one population at least is harmed
while the other is either harmed, benefited, or not affected.
This include
a) Competition
b) amensalism
c) parasitism and predation
a. Competition
It is a relation that occurs between different populations in the soil which use the same
limiting resources that are insufficient to support all the individuals. The rivalry for limiting nutrients
or other common needs. These resources include raw materials important for life such as water, light,
nutrients, oxygen, and space for occupying or any other resources, which is essential for survival and
reproduction. In this relation, the best adapted microbial species will predominate or infact, eliminate

other species which are dependent upon the same limited nutrient substances. Also, organisms which
have the capability to grow faster are considered good competitors.
Resource Competition It occurs when the growth rates of both populations are limited by the same resource
and one population has the ability to diminish the availability of that resource for the other populations.
Interference competition It occurs between two populations in which one of them damages the other
population’s habitat either physically or chemically and excludes it from the habitation. This relationship
is also called direct competition or active competition.
Few Examples The chlamydospores of Fusarium, oospores of Aphanomyces, as well as conidia of

Verticillium dahliae need exogenous nutrients to germinate in soil, while some other fungi bacteria that
inhabit the soil have the ability to deplete these important nutrients that are required for spore germination
and thereby delay its germination which results in decrease in the population of these plant pathogens in
the soil
b. Amensalism (Antagonism)
It is the most common negative relationship in nature at which one microbial population suppresses or
adversely influences the growth or the activities of the other population in the same environment by
producing inhibitory substances either directly or indirectly. The release of products by one species is
toxic to its neighbours. The population that produces the inhibitors is not affected by them and therefore
gains the antagonistic edge.
These inhibitors may be antibiotics, toxins, organic acids, alcohols, or other allelochemicals, lytic
enzymes, as well as harmful gases like methane, ethylene, HCN, nitrite, or sulfides or other volatile
sulfur compounds. The population that adversely affects the other is called antagonistic species, and it
constantly has great practical importance. Antagonism is a type of ammensalism. Types of Antagonism
There are diverse types of antagonism according to the nature of substances that is used in the
antagonism.
• Antagonism by Antibiosis: This process is called antibiosis in which the antibiotics or other
allelochemical metabolites are produced by one organism to inhibit another organism. Antibiotics is
common among Streptomyces isolates, but numerous strains of Micromonespora and Nocardia are also
active. Bacillus and Pseudomonas species are producing antibiotic substances against plant pathogeneic
fungi. Other than antibiotic some inhibitory compounds are secreted by several soil microorganisms
• Antimicrobial compounds against fungi are present in the soil, which inhibit the germination of
fungal spores. This phenomenon is termed as fungistasis.
• Cyanide is produced by certain fungi in concentrations toxic to other microorganisms
• Algae elaborate fatty acids which exhibit and marked antibacterial activity
• Bacillus thuringiensis toxin to lepidopteran insect pests
C. Parasitism
It is a relationship between two dissimilar organisms that is called host-parasite relationship in which
one of them (parasite) lives in or on the other organism (host). The parasite lives in close contact with
the host and forms metabolic association with the host and feeds on their cells, tissues, or fluids in which
the parasite is profited, while the host is adversely affected. Sometimes the relation between the host and
parasite could be diverged from parasitic relationship to a pathogenic relationship.
This relationship is widely spread in soil communities and characterized by its long period of contact and
the specialization between parasite and host. Also, parasite is usually smaller than the host (in most
cases). This relationship has two sides, one is useful while the other harmful. If the parasitism is
accomplished on bacteria that are considered pathogenic to plants, it is considered as a useful relationship
for plant growth and health. While if the parasitism is accomplished on bacteria that are considered
profitable to plants, it is considered as a harmful relationship for plant growth and health Types of
Parasitism
Parasitism could be classified according to its nature of parasitism and its infection type.
Types of Parasitism according to Parasitism Patterns
• Obligate parasitism: Occurs when the parasite cannot live without its host.
 Facultative parasitism: Occurs when the parasite can survive by itself without its host cells in some
conditions.
Types of Parasitism According to Infection Form •
Ectoparasitism: The parasite remains outside the host cells.
• Endoparasitism: The parasite penetrates the host cells.
Examples of Parasitism
(a) Viruses which attack bacteria (bacteriophages), fungi, algae, or plants are strict endoparasites
(intracellular parasites) as they obligate parasite and cannot be cultivated on the media as free-living
forms.
(b) Chytrid fungi parasitize on algae as well as other fungi by penetration into the host.
(c) Daptobacter spp. penetrates and degrades the cytoplasm of several genera of Chromatiaceae. It grows
and propagates in the cytoplasm of the host
d. Predation
Predation is one of the most dramatic interrelationships among the microorganic in nature, at which
predator organism directly attacks a prey organism and feeds on it. This relationship has short
duration, at which predators may or may not kill their prey prior to feeding on them, but the normal result
is generally absorption of the prey’s tissue through ingestion and subsequently the death of prey. Prey
may be larger or smaller than predator.
The most numerous predators of bacteria are protozoans, which by feeding on the billions of
bacteria, undisputedly affect their populations. Protozoans are a key factor in limiting the size of bacterial
populations.
Bdellovibrio bacteriovorus is a predatory bacterium, which penetrates the cell wall and
multiplies between the wall and the plasma membrane, which causes lysis of the prey and releases its
progeny. It attacks and consumes different bacterial strains, including Escherichia coli and Aquaspirillum
serpens, Salmonella typhimurium, and Helicobacter pylori.
Trichoderma spp. could be considered as the most essential biological control agent in that soil because
it utilizes different mechanisms to fight various disease-causing agents including parasitism, which
directly attack the pathogens especially fungi; competition, as it has the ability to colonize the soil or to
compete for nutrients which is causing relegation of pathogen from plant rhizosphere; antagonism
(antibiosis), as it is able to produce secondary metabolites, which have a lethal or depressant effect on
the plant pathogen as well as it has ability to make a repressive environment by diverse relations in the
soil community to create unfavorable ecological conditions that limit the development or
multiplication of pathogenic populations; and the secretion of numerous compounds that induce the
mechanisms of plant resistance to combat pathogens attack.
An overview of industrially important microorganisms and products

Enzyme producing microorganisms Organic acid producing microorganisms
Microorganisms in Alcohol industry Microorganisms in Antibiotic production
Yeast - Saccharomyces cerevisiae Fungi - Penicillium chrysogenum Bacteria

Bacteria - Zymomonas mobilis - Bacillus sp.
Actinobacteira - Streptomyces griseus
Microorganisms in Food industry

Lecture 15. Silage production. Bioinoculants – types; biofertilizers - bacterial, fungal (AMF) and
algal biofertilisers. Biopesticides – types and mechanism of action Silage production
• Silage is the green material produced by controlled fermentation of the green fodder crop
retaining the high moisture content, contain 20-40% DM.
• Silage involves natural fermentation, which produces lactic and other acids, which ‘pickle’ or
preserve the forage.
Crop suitable for silage making
1. The fodder crops such as Maize, Sorghum, Oats, Pearl Millet and hybrid napier grass rich in
carbohydrates are most suitable for silage making.
2. Procedure of silage making
3. Construct a surface/ trench silo (silage storage structure). One cubic meter space / silo can store
500- 600 kg of green fodder.
4. Harvest the crop at 30-35% dry matter stage.
5. Wilt the harvested fodder to bring down dry matter to 30-35%, if required 6. Chop the fodder into
small pieces of 2-3 cm
7. Fill the chopped fodder into the silo.
8. Press the chopped fodder in the silo layer by layer of 30-45 cm.
9. Filling and pressing should be completed as fast as possible.
10. Use additives (Sprinkle Molasses solution over it at 2-3% of the material weight) during
filling of fodder in silo, if required.
11. After filling and pressing, seal the silo with thick polyethylene sheet.
12. Put weight through mud layer/sand bags/tyres on the sheet to prevent air flow beneath the sheet.
13. Open the silo for feeding, minimum after 45 days, as per the need.
Feeding of Silage
• Silage can be taken out as per the requirement. Initially silage can be fed 5 kg/animal

Characteristics of good quality silage
• Bright, light green yellow or green brown in colour
• Lactic acid odour with no butyric acid and ammonia odour.
• Firm texture with softer material
• Moisture should be in range of 65-70%
• Lactic acid 3-14 % • Butyric acid less than 0.2% • pH - 4.0 to 4.2.
Fermentation process of silage

The first stage (Respiratory stage)
• The packed raw materials are still respiring immediately after chopped and consumes oxygen.
• The temperature will rise to about 32ºC around 4 days after packing.
The second stage (Early fermentation)
• Production of acetic acid, formic acid and other organic acids as a result of the growth of
facultative aerobic bacteria such as Enterobacteria.
• The silage pH slowly changes from about 6.0 to 4.0
The third stage (Lactic acid fermentation)
• Lactic acid fermentation begins by lactic acid bacteria witch are strictly anaerobic about 3 days
after packing chopped materials and acetic acid production declines.
The fourth stage (Cont.… of Lactic acid fermentation) •
Lactic acid production continues for about 2 weeks.
• The temperature goes down slowly to about the normal atmospheric temperature and pH is
maintained at 4.0
The fifth stage (Stabilization phase)
• Due to the presence of lactic acid, further degradation is inhibited, as bacterial and fungal growths
are checked.
• The lactic acid fermentation completes in about 20 days, and the silage product is finished.
Figure. Temperature, pH changes and bacterial changes during fermentation process of silage
Biofertilizer

Biofertilizers are defined as preparations containing living cells or latent cells of efficient strains of
microorganisms that help crop plants’ uptake of nutrients by their interactions in the rhizosphere when
applied through seed or soil.
Classification of biofertilizer
4. Endophytic Glucanoacetobacter
2. PPFM Methylobacterum
Biopesticides
Biopesticides are living organisms which can intervene the life cycle of insect pests in such a way that
the crop damage is minimized. The agents employed as biopesticides, include parasites, predetors and
disease causing fungi, bacteria and viruses, which are the natural enemies of pests.
Biopesticides type
• Microbial pesticides which consist of bacteria, entomopathogenic fungi or viruses (and
sometimes includes the metabolites that bacteria or fungi produce). Entomopathogenic nematodes
are also often classed as microbial pesticides, even though they are multi-cellular.
• Bio-derived chemicals. Four groups are in commercial use: pyrethrum, rotenone, neem oil, and
various essential oils are naturally occurring substances that control (or monitor in the case of
pheromones) pests and microbial diseases.
• Plant-incorporated protectants (PIPs) have genetic material from other species incorporated
into their genetic material (i.e. GM crops). Their use is controversial, especially in many
European countries.

Major advantages of bio pesticides
Bio-pesticides are preferred over chemical pesticides for the following reasons:
• no harmful residues;
• target specific and safe to beneficial organisms like pollinators, predetors, parasites etc.;
• growth of natural enemies of pests is not affected, thus reducing the pesticide application;
• environmental friendly;
• cost effective;
Control Mode of Action Examples Control Agent
Agent
Bacteria Produce toxins that are detrimental to certain insect Bacillus thuringiensis Lepidopterans
pests when ingested Bacillus popilliae Japanese beetle
Viruses Kills insects when ingested. Insect's feeding Baculoviruese: Nuclear Lepidopteran
behavior is disrupted thus it starves and dies. polyhedrosis virus (NPV) and
Hymenopteran
Baculoviruses: Granulosis Lepidopteran
virus (GV)
Fungi Controls insects by growing on them secreting Metarhizium anisopliae Lepidopteran
enzymes that weaken the insect's outer coat, and insects
then getting inside the insect and continuing to Entomophaga praxibulli Grasshoppers
grow, eventually killing the infected pest Zoophthora radicans Aphids
Neozygites floridana Cassava green
mite
Protozoa Kills insects when ingested. Insect's feeding Nosema Grasshoppers

behavior is disrupted thus it starves and dies. Vairimorpha Lepidoptera
Malamoeba Locusts
Major types of bio-agents available for commercial production

There are different types of bio-agents which can be commercially mass produced for large scale distribution among
the farmers for control of insect pests. They are:
Parasitoids Predators Insect Pathogens

• Trichogramma chilonis, • Cryptolaemus montrouzieri • Virus: Nuclear Polyhedrosis
T.brasiliensis and T.pretiosum (Austrtralian ladybird beetle) for Virus (NPV) - for major
(egg parasites) - for tomato control of several species of polyphagous pest like
fruit borer mealy bugs and soft scales Helicoverpa armigera (gram pod
• Chrysopa spp. (green lacewing borer) and Spodoptera litura
• Trichogramma chilonis - for bug) - for the control of aphids, (Tobacco caterpillar)
brinjal shoot and fruit borer, white flies etc. • Bacteria: Bacillus thuringiences
shoot borers of cotton, (B.t) - for control of
lepidopterous pests
sugarcane, rice etc.
• Fungi: Trichoderma viride and
Trichoderma harziarum against
soil borne fungal diseases

Lecture 16. Mass production and quality control of bacterial and fungal bioinoculants. BIS
standards– methods of application of bioinoculants.
Biofertilizer/Bioinoculants:
Biofertilizers are defined as preparations containing living cells or latent cells of efficient strains of
microorganisms that help crop plants’ uptake of nutrients by their interactions in the rhizosphere when
applied through seed or soil.
Mass production and quality control of bacterial and fungal bioinoculants
The mass production of carrier based bacterial biofertilizers involves three stages.
1. Mass culturing of microorganisms in fermentor

2. Processing of carrier material
3. Mixing of broth culture with the carrier and packing
1. Mass culturing of microorganisms in fermentor

The media used for mass culturing are as follows:
Rhizobium : Yeast extract mannitol broth.
Azospirillum : Dobereiner's malic acid broth
Azotobacter : Waksmann No.77 broth
Phosphobacteria : Nutrient broth
Gluconacetobacter : LGI broth
• Prepare appropriate broth in 50 ml flasks and inoculate the mother culture in to the flasks.
• Grow the culture under shaking conditions at 30±2 OC until maximum cell population of 1010 to
1011 cfu/ml is reached
• Under optimum conditions this population level could be attained within 4 to 5 days for
Rhizobium; 5 to 7 days for Azospirillum; 2 to 3 days for Phosphobacteria and 6-7 days for
Azotobacter & Gluconacetobacter. The culture obtained in the flask is called starter culture.
• Use the starter to inoculate the broth in large size flasks of 250 ml, 500 ml, 3 liters and 5 liters
and grow until required level of cell count is reached.
• For large scale production of inoculant, inoculum from starter culture is transferred to large
flasks/seed tank fermentor.
2. Processing of carrier material

The use of ideal carrier material is necessary in the production of good quality biofertilizer. Peat
soil, lignite, vermiculite, charcoal, press mud, farmyard manure and soil mixture can be used as
carrier materials. The neutralized peat soil/lignite are found to be better carrier materials for
biofertilizer production.
• Powder the carrier material (peat or lignite) to a fine powder so as to pass through 212  IS sieve.
• Neutralize the pH of the carrier material with the help of calcium carbonate (1:10 ratio), since the
peat soil / lignite are acidic in nature (pH of 4 - 5)
• Sterilize the neutralized carrier material in an autoclave to eliminate the contaminants.
3. Mixing of broth culture with the carrier and packing

• Add the bacterial culture drawn from the fermentor to the neutralized and sterilized carrier
material to the moisture content of 35 to 45% on wet basis. The carrier and broth can be mixed
either manually (by wearing sterile gloves) or mechanically.
• After mixing, pack the inoculants in 1kg quantities in polythene bags, seal with electric sealer and
allow for curing for 2 -3 days at room temperature (curing can be done by spreading the inoculant
on a clean floor/polythene sheet/ by keeping in open shallow tubs/ trays with polythene covering
for 2 -3 days at room temperature before packaging). Curing improves the cell count to 10 9 to 1010
cells /g. After curing it is then packed in low density polythene bags. The inoculants may be
allowed for curing even after packing for 3- 4days at room temperature.

Mass Production of Fungal Bioinoculant
• One of the fungal bioinoculant used is VAM/AMF. The commercial utilization of mycorrhizal fungi
has become difficult because of the obligate symbiotic nature and difficulty in culturing on
laboratory media. Production of AM inoculum has evolved from the original use of infested field
soils to the current practice of using pot culture inoculum derived from the surface disinfected
spores of single AM fungus on a host plant grown in sterilized culture medium. Several
researches in different parts of the world resulted in different methods of production of AM
fungal inoculum as soil based culture as well as carrier based inoculum. Root organ culture is
being used for the production of soil less culture.
Mother Culture of AMF

• Mother culture can be produced from efficient spores using funnel techniques as well as
multiplication in small sized pots. 3-4 cycles will be made for developing the mother culture.
Mother culture should have 100 % root colonization and the minimum of 8-10 spores per gram
of the inoculum. Mother culture can also be maintained in maize roots continuously
• Form a trench (1m x 1m x 0.3m) is and line with black polythene sheet to use this as a plant
growth tub.
• Mix 50 kg of vermiculite and 5 kg of sterilized soil and pack in the trench up to a height of 20 cm
• Spread 1 kg of AM inoculum (mother culture) 2-5 cm below the surface of vermiculite
• Sow with surface sterilized Maize seeds (5% sodium hypochlorite for 2 minutes)
• Apply 2 g urea, 2 g super phosphate and 1 g muriate of potash for each trench at the time of
sowing seeds. Further apply 10 g of urea twice on 30 and 45 days after sowing for each trench
• Apply 1 g of micronutrient mixture when there will be a symptom for deficiency
• Test the quality of the inoculum by estimating the AM colonization in root samples on 30 th and
45th day
• Grow the stock plants for 60 days (8 weeks).
• Pull out the plants and cut the roots into small pieces
• Mix the roots thoroughly with vermiculite in the trench
• The inoculum produced consists of a mixture of vermiculite, spores, pieces of hyphae and
infected root pieces
Quality control
• 108 Viable bacterial Cells (CFU)/g of the Carrier within 15 days of manufacture and 10 7
CFU/g of Carrier within 15 days before the Expiry at Storage temperature 25-30°C.
• It should not have any contaminant at 10-5 dilution
• Liquid biofertilizer 1012 Viable bacterial Cells (CFU)/ml of the liquid biofertilizer
• Carried biofertilizer – 6 month storage ; liquid biofertilizer – 1 year storage

Example given above for the quality control of phosphate solubilizing biofertilizer.
BIS (Bureau of Indian standard) standards
(i) Viable Cell Count: 108 Viable bacterial Cells (CFU)/g of the Carrier within 15 days of
manufacture and 107 CFU/g of Carrier within 15 days before the Expiry at Storage temperature
2530°C. It should not have any contaminant at 10-5 dilution
Liquid biofertilizer 1012 Viable bacterial Cells (CFU)/ml of the liquid biofertlizer
(ii) pH: 6-7.5 (iii) Moisture Percentage: 35-40%

(iv) Shelf Life: 6 Months (Carrier based biofertilizer) 12 months (Liquid biofertilizer) (v)
Carrier:

Peat, Lignite, Peat Soil, Humus or Similar Material Neutralized with CaCO 3 and then sterilized.
Carrier should be in the form of powder capable of passing through 150 to 212 Micron (72-100
Mesh) IS Sieve
(vi) Marking on the Packet:
The Inoculatent Packet/bottle should have the following information:
(a) Name of the Product
(b) Name and Address of the Manufacturer
(c) Carrier Used
(d) Batch Number
(e) Date of Manufacture
(f) Date of Expiry
(g) Net Mass
(h) Crop for which intended
(i) Storage Instructions
Methods of Application of Bioinoculants
Bacterial biofertilizers are applied as carrier based inoculants. Peat or lignite is used as carrier material.
Carrier based bacterial inoculants are applied by the following methods.
1. Seed treatment or seed inoculation

2. Seedling root dip and
3. Main field application
Seed Treatment
Mix one kg of the inoculant with approximately one liter of rice gruel to make slurry. Treat the seeds
required for one hectare with the slurry so as to have a uniform coating of the inoculant over the seeds
and then shade dry for 30 minutes. The shade dried seeds should be sown within 24 hours. For small
seeds 600 g/ha can be used for seed treatment. For horticulture crops also it is recommended as
600 g/ha.
Seedling root tip
This method is used for transplanted crops. Mix one kg of the inoculant in 200 liters of water. Dip the
root portion of the seedlings required for one hectare in the mixture for 15- 20 minutes before
transplanting.
For horticulture crops, it is recommended as 800 g/ha seedlings
Main field application
Mix two kg of the inoculant with 25 kg of dried, powdered farm yard manure and then broadcast in one
hectare of the main field just before transplanting.
Recommendation of biofertilizers
Rhizobium:
It is recommended for all legumes as seed treatment.

Azospirillum / Azotobacter:
Recommended for all crops, other than legumes. For transplanted crops apply Azospirillum /
Azotobacter by seed treatment, seedling root dip and soil application methods. For direct sown crops
apply Azospirillum / Azotobacter through seed treatment and soil application methods.
Phosphobacteria:
Recommended for all crops. Apply phosphobacteria by seed treatment, seedling root dip and soil
application methods as that of Azospirillum/Azotobacter.
Gluconacetobacter diazotrophicus:
It is recommended for sugarcane. For sett treatment it can be used @ 2kg/ha setts and soil application @
2kg/ha at 30, 60 and 90 days of planting.
Azophos:
It is a combined formulation of Azospirillum and Phosphobacteria. Apply this formulation as
Azospirillum / Phophobacteria with the same dosage.
Points to remember
• Bacterial inoculants should not be mixed with insecticide, fungicide, herbicide and fertilizers.
• Seed treatment with bacterial inoculant is to be done at the last when seeds are treated with other
fungicides.
• Biofertilizer treated seeds should be sown immediately.
• Biofertilizer treated seeds should be dried under shade only.
Methods of application of liquid biofertilizers
Liquid biofertilizers can also be applied by the above three methods.

1. Seed treatment : 120 ml/ha seeds
2. Seedling root dip : 375 ml/ ha seedlings
3. Soil application : 500 ml/ ha
Biofertigation: All the bacterial inoculants are recommended through drip fertigation. It is recommended
at the rate of 500 ml/ ha.
Foliar application: PPFM alone is specifically recommended through this method @ 500 ml/ha twice in
vegetative and flowering stages.
Method of inoculation of AM fungal inoculant
Nursery application
100 g inoculum is sufficient for one meter square. The inoculants should be applied at 2-3 cm
below the soil at the time of sowing. The seeds/cutting should be sown / planted above the AM inoculum.
For polythene bag raised seedlings (Forest trees, Coffee & Tea)
About 10 g of inoculum is sufficient for each plant raised in poly bags. Mix 10 kg of inoculum
with 1000 kg of potting mixture and pack the potting mixture in polythene bags before sowing.

For out planting Twenty grams of AM inoculum is required per seedling. Apply inoculum at the
time of planting. For existing trees
Fifty to one hundred gram of AM inoculum is required for inoculating one tree. Apply inoculum
near the root surface at the time of fertilizer application.
Method of inoculation of BGA in rice field
• Powder the soil based algal flakes very well.

• Mix it with 10 kg soil or sand (10 kg powdered algal flakes with 10 kg soil / sand).
• BGA is to be inoculated on 7-10 days after rice transplanting.
• Water level at 3-4” is to be maintained at the time of BGA inoculation and then for a month so as
to have maximum BGA development
• A week after BGA inoculation, algal growth can be seen and algal mat will float on the water
after 2-3 weeks. The algal mat colour will be green or brown or yellowish green
Method of inoculation of Azolla to rice crop
The Azolla biofertilizer may be applied in two ways for the wetland paddy. In the first method, fresh
Azolla biomass is inoculated in the paddy field before transplanting and incorporated as green manure.
This method requires huge quantity of fresh Azolla. In the other method, Azolla may be inoculated after
transplanting rice and grown as dual culture with rice and incorporated subsequently
A. Azolla biomass incorporation as green manure for rice crop
• Collect the fresh Azolla biomass from the Azolla nursery plot.
• Prepare the wetland well and maintain water just enough for easy incorporation.
• Apply fresh Azolla biomass (15 t ha-1) to the main field and incorporate the Azolla by using
implements or tractor
B. Azolla inoculation as dual crop for rice

• Select a transplanted rice field.
• Collect fresh Azolla inoculum from Azolla nursery.
• Broadcast the fresh Azolla in the transplanted rice field on 7 th day after planting (500 kg / ha).
• Maintain water level at 5-7.5cm.
• Note the growth of Azolla mat four weeks after transplanting and incorporate the Azolla biomass
by using implements or tractor or during inter-cultivation practices.
• A second bloom of Azolla will develop 8 weeks after transplanting which may be incorporated
again.
• By the two incorporations, 20-25 tonnes of Azolla can be incorporated in one hectare rice field.

Lecture 17. Biofuel production – methane, hydrogen, alcohol and biodiesel production
Biofuels
Biofuel, any fuel that is derived from biomass—that is, plant or algae material or animal waste. Since
such feedstock material can be replenished readily, biofuel is considered to be a source of renewable
energy, unlike fossil fuels such as petroleum, coal, and natural gas.
First generation Biofuels: The first generation biofuels refer to the fuels that have been derived from
sources like starch, sugar, animal fats and vegetable oil. The oil is obtained using the conventional
techniques of production.
Second-generation biofuels: also known as advanced biofuels, are fuels that can be manufactured from
various types of biomass (cellulose). Biomass is a wide-ranging term meaning any source of organic
carbon that is renewed rapidly as part of the carbon cycle.
Third Generation biofuels: It is based on improvements in the production of biomass. It takes advantage
of specially engineered energy crops such as algae as its energy source. The algae are cultured to act as
a low-cost, high-energy and entirely renewable feedstock.
Bio-hydrogen
• Hydrogen is a noncarbonaceous fuel and energy carrier possessing higher net calorific value
compared to other fuels.
• It can be directly converted into energy in fuel cell or mixed with natural gas for use in internal
combustion and jet engines, as well as the gas power turbines.
• Combustion of hydrogen yields only water; thus it is considered as a clean energy source.
• The limitation in using hydrogen is its explosivity when mixed with oxygen, leading to difficulty in
its storage and distribution.
• Bio-hydrogen is a term used to call hydrogen produced via microbial fermentation. Dark- and
photofermentation are typically applied for bio-hydrogen production.
• Dark fermentation of organic carbon substrates is carried out by obligate or facultative anaerobic
bacteria yielding bio-hydrogen and other side products, such as volatile fatty acids (VFAs) and
alcohols.
• Photo-fermentation requires energy from light to aid the decomposition of organic substrates by
photosynthetic bacteria, mostly purple non-sulfur bacteria (PNSB).
• The dark fermentative bacteria are capable of utilizing various substrates with high rate of hydrogen
production.
• A drawback of dark fermentation is its low yield due to the large quantity of side products formed.
• The substrates for photo-fermentative bacteria are limited to simple sugars and organic acids, and the
hydrogen production rate by photo-fermentation is usually lower than dark fermentation.
Dark fermentation : C 6 H 12 O 6 → 2 CH 3 COOH + 2 CO 2 + 4 H 2

Photo - fermentation : 2 CH 3 COOH + 4 H 2 O + Light → 4 CO 2 + 8 H 2
• Dark fermentation - Enterobacter sp. and Clostridium sp.
• Photo – fermentation - Rhodobacter capsulatus, purple non-sulfur bacteria
Bio-methane
•Methane is a fuel gas mainly produced from anaerobic digestion process.
•Organic substrates are decomposed by diverse microbial communities through a series of metabolic
stages during anaerobic digestion, resulting in gaseous products called biogas and inorganic molecules
remaining in digestate.
•Biogas mainly comprises methane (50–75%), carbon dioxide (25–40%), nitrogen (<5%), hydrogen
(<1%), oxygen (<1%), and hydrogen sulfide (50–5000 ppm).
•Biogas is suitable for use in internal combustion engines and gas turbine generators.
•Methane has higher octane rating than gasoline, and its combustion produces less CO2 as compared
to fossil fuels.
•Methane production by anaerobic digestion process involved multiple steps performed by several groups
of microorganisms.
•Typically, anaerobic digestion is divided into four steps that are hydrolysis, acidogenesis, acetogenesis,
and methanogenesis.
•In hydrolysis step, complex organic matters (such as cellulose and protein) are converted into simpler
and soluble molecules (such as sugars and amino acids) by hydrolase enzymes excreted by facultative
and strictly anaerobic microorganisms called fermentative bacteria.
•The soluble molecules produced by the hydrolysis steps are then utilized by acidogenic bacteria to
produce short-chain organic acids (such as acetic, butyric, and propionic acids) along with hydrogen,
carbon dioxide, and alcohols in the acidogenesis step.
•These products are further consumed in the acetogenesis step to produce acetic acid by acetogenic
bacteria.

•In the last step, acetic acid, hydrogen with carbon dioxide, formic acids, and alcohols were utilized by
methanogenic bacteria to produce methane under obligate anaerobic condition.
•Methanogens are microorganisms that produce methane as a metabolic byproduct in hypoxic
conditions. • Methanogenic Bacteria are Methanosarcina, and Methanomicrobium
Bioethanol/ bioalcohol
Production of ethyl alcohol from sugary materials is one of the oldest known microbiological
process. Alcohol is an important solvent and raw material used in a variety of chemical industries.
Although today industrial alcohol is also produced synthetically from ethylene, production of alcohol by
fermentation of cheap sugary materials such as molasses by yeast is still an important industry.
Bioethanol is fuel alcohol manufactured from biomass
For ethyl alcohol production, selected strains of Saccharomyces cerevisae are employed since all the
strains are not equally efficient. The alcohol tolerance and sugar tolerance are important criteria used in
the selection of yeast strains. The yield of ethyl alcohol is about 50 per cent of the fermentable sugar
concentration. Further purification of ethyl alcohol is done by fractional distillation. In some distilleries,
the yeast is recovered and used as animal feed while in most, it is discarded into the effluents, a procedure
that is very undesirable.
In recent years because of the possibility of using ethyl alcohol as a fuel supplement and a
chemical feed stock, there is increased interest in increasing production but at a cheaper and economical
rate.

Benefits of bioethanol
• Less dependence on crude oil

• It is a renewable fuel
• Increase octane number (a standard measure of the performance of motor fuel)
• Reduces air pollution, cleaner environment due to cleaner combustion  lower net carbon
dioxide emissions
• Helping emerge a new market
• Expanded market opportunity in the agricultural field
Microorganisms for Bioethanol Production
Bacteria Zymomonas mobilis, Bacillus stearothermophilus, Escherichia coli
Klebsiella oxytoca
Yeast Saccharomyces cerevisiae, Pachysolen tannophilus, Candida shehatae
Pichia stipitis
Characteristics of Saccharomyces cerevisiae
• The most common and natural fermentative yeast for ethanol production
• Wild strains convert only glucose to ethanol
• Limitation of using lignocellulose
• Relative high ethanol yield
• Can be easily modified by metabolic engineering to ferment pentose Convertion involves the
enzymes Invertase and Zymase
Invertase - converts sucrose into glucose and fructose. Zymase - another enzyme present in yeast
converts glucose and fructose into ethanol and carbon dioxide.
The carbon dioxide formed is allowed to escape, but air is not allowed to enter. In presence of air ethanol
formed would be oxidized to acetic acid.
The fermentation will be completed in 3 days. The carbon dioxide obtained as byproduct is recovered and
can be sold
Combined hydrolysis and fermentation

Some species of bacteria capable of direct conversion of a cellulose substrate into ethanol.
Clostridium thermocellum, which uses a complex cellulosome (Cellulose degrading enzyme complex) to
break down cellulose and synthesize ethanol. However, it also produces other products during cellulose
metabolism, including acetate and lactate, in addition to ethanol, lowering the efficiency of the process.
Raw materials for ethanol production
Wheat Maize/Corn, Sugar Beet and Sugar Cane are widely used .

Production process
• Milling the cereal: meaning mechanical crushing of the cereal grain to release the starch
component
• Saccharification: after milling heating and addition of water and enzymes can be done for
conversion into fermentable sugar
• Fermentation: fermentation of the mash using yeast
• One glucose molecule is converted into two ethanol molecules and two carbondioxide molecules.
C6H12O6 → 2 C2H5OH + 2 CO2
• Distillation and Rectification: concentration and cleaning of the ethanol produced by the
distillation by removing by-products.
• Distillation: It is a thermal separation method that can be used to fractionate liquid mixtures. It
utilises the different volatility of the components of the mixture to be separated.
• Dehydration: drying (removing residual water) off the ethanol
• Possible secondary process: Animal Feed as a Co-Product
• Evaporation: The goal of evaporation is to vaporize most of the water from a solution containing
a desired product
• Mash coming out of Distillation goes into evaporators where most of the water is separated from
the solid parts in the mash, the stillage.
• If the stillage is dried and pelleted, the resulting storable animal feed is known as
DDGS(Distillers' Dried Grains with Solubles)

Overall Production Process
BIOETHANOL FROM LIGNOCELLULOSIC BIOMASS

Basically, the lignocellulosic biomass comprises of cellulose, hemicellulose and lignin Steps
of conversion of cellulose and hemicellulose to Bioethanol
1. Pretreatment 2. Hydrolysis 3. Fermentation 4. Distillation of the product mixture to separate
ethanol
E85 and Flexible Fuel Vehicles
• Ethanol can be blended with gasoline to create E85, a blend of 85% and 15% gasoline. Due to
the corrosive effects of E85, because of its high alcohol content, traditional vehicles cannot use it.
It needs flex fuel vehicles. Flex fuel vehicles (FFVs) have engines modified to accept higher
concentrations of ethanol. Such flexible-fuel engines are designed to run on any mixture of
gasoline or ethanol with up to 85% ethanol by volume.
Fuel properties of bioethanol
Bioethanol has much lower energy content than gasoline. About two-third of the energy content
of gasoline on a volume base
Octane number of ethanol is higher than petrol, since ethanol has higher antiknock characteristics.
It increases the fuel efficiency of the engines. Also ethanol has the oxygen content of 34.8 %,
which also leads to a higher efficiency, which results in a cleaner combustion process at relatively
low.
Temperatures
Reid vapour pressure (measure for the volatility of a fuel) is very low for ethanol, indicates a slow
evaporation. Hence the concentration of evaporative emissions in the air remains relatively low,
and reduces the risk of explosions.
Disadavantage : : low vapour pressure of ethanol ,gives starting problem during cold. Engines
using ethanol cannot be started at temp < 20ºC
Blending of ethanol:
Blening with a small proportion of a volatile fuel such as gasoline is more cost effective
Various mixture of bioethanol with gasoline or diesel fuels are used

– E5G to E26G (5-26% ethanol, 95-74% gasoline)
– E85G (85% ethanol, 15% gasoline)
– E15D (15% ethanol, 85% diesel)
– E95D (95% ethanol, 5% water, with ignition improver)
Advantages of using bioethanol
• Exhaust gases of ethanol are much cleaner – It burns more cleanly due to the more complete
combustion
• Reduction in emission of green house gases- ethanol-blended fuels such as E85 (85% ethanol
and 15% gasoline) reduce up to 37.1% of GHGs
• Decrease in ozone formation - The emissions produced by burning ethanol are less reactive with
sunlight than those produced by burning gasoline, which results in a lower potential for forming
ozone
BIODIESEL
Biodiesel is defined as non-petroleum-based diesel fuel consisting of alkyl esters (mainly methyl,
but also ethyl, and propyl) of long chain fatty acids. It could be produced from various animal and
plant sources by esterification of triglycerides with methanol.

Microbial lipids, namely single cell oils (SCOs), are produced by oleaginous microorganisms
including algae, fungi and yeast, and are considered to be promising alternatives to vegetable oils as
their oil properties are similar.
Oleaginous microorganism
Lipids (oils), proteins and carbohydrates are the three macromolecules occurring naturally in
microbial systems. Lipids are an important structural component of the microbial cell membrane. Certain
microorganisms (algae, yeast, fungi and bacteria) can accumulate more than 20% of lipids inside their
cells as triacylglycerol (TAG). Those microorganisms are called as oleaginous microorganisms. The oils
thus produced are popularly called SCOs (single cell oils).
Oleaginous microorganisms :
Bacteria : Rhodococcus opacus, Bacillus alcalophilus

Algae-Chlorella, Botryococcus
Yeasts : Rhosotorula glutinis, Cylindrotheca sp.
Biodiesel from microalgae
The production of biodiesel from microalgae has multiple advantages and has been termed the
third-generation biofuels. Unlike other oil crops, microalgae grow extremely rapidly and many are
exceedingly rich in oil. Microalgae commonly double their biomass within 24 h, and biomass doubling
times during exponential growth are commonly as short as 3.5 h. Oil content in microalgae can exceed
80% by weight of dry biomass, and oil levels of 20–50% are quite common
Biosynthesis of lipids in microalgae :
Free fatty acids are synthesized in chloroplast and triglycerides are accumulated in endoplasmic
reticulum
Advantages of producing biodiesel from algae
• The production of biodiesel from microalgae has multiple advantages and has been termed the
third-generation biofuels.
• Unlike other oil crops, microalgae grow extremely rapidly and many are exceedingly rich in oil.
• Microalgae commonly double their biomass within 24 h, and biomass doubling times during
exponential growth are commonly as short as 3.5 h.
• Oil content in microalgae can exceed 80% by weight of dry biomass, and oil levels of 20–
50% are quite common Production of biodiesel from algae
1. Culture of microalgae
The large scale production of microalgae is generally performed with solar energy
(photoautotrophic metabolism) in open ponds (raceways), closed systems (photobioreactors) or
fermentors.
2. Harvesting of microalgae
Microalgae biomass could be harvested by, centrifugation, flocculation, gravity sedimentation,
filtration methods
3. Extraction techniques
The lipids are extracted using chemical solvents, chloroform-methanol blends have been
extensively used with high extraction yields up to 83%. Some physiochemical as well as
biochemical techniques are used for extraction of lipids
4. Transesterification
It is the process of converting oils to biodiesel. In this process lipids can react with alcohol and
produce biodiesel
5. Purification of biodiesel and by-products
Microalgae biodiesel and by-products must be separated for increasing the biodiesel production.
The main separation processes used hot water (50˚C), organic solvents such as hexane and
waterorganic solvent for a liquid-liquid separation.

Production process of Oleaginous yeast and biodiesel

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AGM 301 Agricultural Microbiology (1+1)

Lecture 10. Sulphur cycle - sulphur oxidizers; microbial transformation of K, Zn and Si I.

Cycling of sulphur is similar to that of nitrogen. Transformation / cycling of sulphur between

Four distinct transformations are recognized during the cycling of S

1. Decomposition/Mineralization of larger organic S compounds to smaller units and their

AGM 301 Agricultural Microbiology (1+1) 1

3. Oxidation of inorganic ions and compounds (S oxidation)

AGM 301 Agricultural Microbiology (1+1) 2

S oxidizers are categorized into three groups

 Chromatium, Rhodopseudomonas, Rhodomicrobium

Green sulfur bacteria:

 Chlorobium, Chloropseudomonas, Chlorobacterium, Ancalochloris, Chlorobaculum

AGM 301 Agricultural Microbiology (1+1) 3

They are the members of Thiobacilli.

 Thiobacillus, Thiomicrospira. Beggiatoa, Thiothrix and Thioloca. T. ferrooxidans and T. thiooxidans

3. Heterotrophs - including a wide range of bacteria and fungi

 Arthrobacter, Bacillus, Micrococcus, Mycobacterium, and Pseudomonas; some actinomycetes;

 Occurs both in aerobic and anaerobic condition - Bacteria

Importance of S oxidizing bacterium (or) Application of S oxidation

AGM 301 Agricultural Microbiology (1+1) 4

a. Assimilatory sulphate reduction

b. Dissimilatory sulphate reduction

II. Transformation of Potassium in soil

1. Soil solution K (K+ ion)

2. Exchangeable K (easily exchangeable and slowly exchangeable K with other cations )-

3. Non exchangeable K (reserves of K)

AGM 301 Agricultural Microbiology (1+1) 5

K RELEASE BACTERIA IN TRANSFORMATION

AGM 301 Agricultural Microbiology (1+1) 6

• Microorganisms like Bacillus extroquens and Clostridium pasteurianum, Bacillus

• Microbially produced organic ligands include metabolic by products, extracellular enzymes,

III. Transformation of Silica in soil

AGM 301 Agricultural Microbiology (1+1) 7

Occurrence of silica in soils

Silicon cycle in soils

AGM 301 Agricultural Microbiology (1+1) 8

Mechanism of silica solubilization

 Organic acids play major role in weathering of Alumino silicate minerals

 Biodissolution of Si can also releases K from minerals

 Paenibacillus muciloginosus, Achromobacter xylosoxidans, Bacillus altitutinis, Pseudomonas

 Increase available silica content in soil as well as in plants

 Offers resistance to plants against pests to diseases

 Facilitated the rapid breakdown of siliceous residues and release silica

AGM 301 Agricultural Microbiology (1+1) 9

AGM 301 Agricultural Microbiology (1+1)

 The native zinc in soils occurs in different chemical pools

 Bacillus cereus, Bacillus sphaericus, Bacillus subtilis, Gluconacetobacter diazotrophicus,

Importance of Zn solubilizing microorganism

 Alter the Zn availability in soil upon application

 Thiobacillus thiooxidans, helps in leaching of Zn from the ore

 Bacillus subtilis helps in adsorption and accumulation of Zn in soil

 Organic acid production by the Zn solubilizing microorganism helps in increasing Zn

AGM 301 Agricultural Microbiology (1+1) 11

Types of problem soils

Soils with Physical problems

Role of microbes in reclamation of problem soils

Biodegradable solid waste

Non-biodegradable solid wastes

3 R’s of Solid Waste Management

The First ‘R’ – Reduce

1. Print on both sides of the paper to reduce paper wastage.

4. Use cloth napkins instead of paper napkins.

AGM 301 Agricultural Microbiology (1+1) 14