Soil microbiological research with thrust to soil health

SOILS 691
DOCTORAL SEMINAR-I
ON
RECENT TECHNIQUES IN SOIL MICROBIOLOGICAL
RESEARCH WITH THRUST TO SOIL HEALTH
Presented by
Mr. Bagmare Rakesh Ramesh
Reg. No: 2019A/26P
VASANTRAO NAIK MARATHWADA KRISHI VIDYAPEETH, PARBHANI
Seminar In Charge
Dr. P.H. VAIDYA
Professor
Dept. of Soil Science and Agril. Chemistry
Research Guide
Dr. SYED ISMAIL
Associate Dean & Principal and Head,
Dept. of Soil Science and Agril. Chemistry
Submitted to,
DEPARTMENT OF SOIL SCIENCE AND AGRIL. CHEMISTRY
COLLEGE OF AGRICULTURE
VASANTRAO NAIK MARATHWADA KRISHI VIDYAPEETH
PARBHANI 431 402 [M.S.]
2021

CONTENT
❖ INTRODUCTION, PHILOSOPHER LOOK AT SOIL, HISTORICAL
OVERVIEW OF SOIL MICROBIOLOGY
❖ Soil health
❖ SOIL BIODIVERSITY
❖ WORKERS IN SOIL BIODIVERSITY
❖ CONCEPT OF BIOFERTILIZERS, PLANT GROWTH PROMOTING
RHIZOBACTERIA AND ITS MECHANISMS
❖ ISOLATION METHODS FOR PURE CULTURE
❖ MICROBIAL ISOLATES AND ITS ROLE IN NUTRIENT MOBILIZATION
❖ MICROBIAL BIOFORTIFICATION
❖ DROUGHT TOLERANCE MECHANISMS BY RHIZOBACTERIA
❖ CROP RESIDUES DECOMPOSITION BY MICROORGANISMS

❑ WHY WE STUDY SOIL MICROBIOLOGY ???
❑ SOIL MICROBIOLOGY is branch of soil science dealing with study of
soil microorganisms and their activities in the soil, their functions,
and how they affect soil properties
❑ A knowledge of soil microbiology is essential to understanding agricultural and
environmental science. Without soil microorganisms, life as we know it couldn’t exist
on earth.
❑ For instance, without soil microorganisms, we’d be overwhelmed by undecomposed
organic matter. Virtually everything we do is influenced by soil microorganisms and
their activity in soil.
INTRODUCTION
(Source: Book of Soil Microbiology: An explolatory Approach by
Mark Coyne)
Compiled and Edited by
Rakesh Bagmare (Ph.D) VNMKV
Parbhani

A PHILOSOPHICAL LOOK AT SOIL
Take a handful of soil
What is it? What does it represent to us?
Soil ??
Dirt ??
Minerals ??
On first site, it seems rather ordinary, But takes a closer look. What’s
in soil? There’s some air, some water, some minerals, clay and organic
matter, which is generally terms for compounds that contain carbon.
INTRODUCTION
(Source: Book of Soil Microbiology: An explolatory Approach by Mark Coyne)

HOW DO YOU TELL IF SOMETHING IS ALIVE
Living things move.
▪ Soil also move in wind and water, and part of organic matter in soil is mobile
Living things specially mammals like us, get hot because they generate heat
▪ Soils also generate heat because they respire, which means they convert energy that
is available in chemicals bonds or light into energy that can be used for work and
that work produces heat.
Living things breath
▪ So does the soil. If We measure soil, We can observe soil absorbing oxygen and
releasing carbon dioxide and other gases.
INTRODUCTION
Rakesh Bagmare (Ph.D) VNMKV Parbhani

Living things change with time
So soil does. Change with time what we call development or evolution, is one factor in
soil formation. In addition , parts of living organic matter in soil are growing,
reproducing and dying all the time. Soil microorganisms are dynamic and different
microbial groups dominate the soil at different times and season
Living things eat
So Soil also does. Proteins, nucleic acid, simple and complex carbohydrate are its food.
Place anyone of this compound in soil and they will dissppear which we do everyday.
INTRODUCTION

For the microbiologist, soil is a living organisms- a mixture of living cells in an
organic or mineral matrix. Not every cell is identical, or carries out of the same
function, or is active at the same time. Soil is immensely complex and
intersecting organisms.
Our task is to dissect this complex into its constituents parts,
Its functions and different activities…..
INTRODUCTION
Parbhani

Unlike soil science whose origin can be traced back to Roman & Aryan times, soil microbiology is
emerged as a distinct branch of soil science during first half of the 19th century. Some of the
notable contributions made by several scientists in field of soil microbiology are highlighted
HISTORICAL OVERVIEW OF SOIL
MICROBIOLOGY
Soil microbiology is emerged as a distinct branch of soil science during first half of the
19th century. Some of the notable contributions made by several scientists in field of soil
microbiology are highlighted …..
❑ A. V. Leeuwenhock (1673) discovered and described microorganisms through his own made
first simple microscope with magnification of 200 to 300 times. He for the first time made the
authentic drawings of microorganisms (protozoa, bacteria, fungi).
❑ J. B. Boussingault (1838) showed that leguminous plants can fix atmospheric nitrogen
and increase nitrogen content in the soil.
❑ J. Von Liebig (1856) showed that nitrates were formed in soil due to addition of nitrogenous
fertilizers.
( Source: https://agriinfo.in/history-of-soil-microbiology-1600-1920-140/)

MICROBIOLOGY
❑ S. N. Winogradsky (1890) discovered the autotrophic mode of life among bacteria and
established the microbiological transformation of nitrogen and sulphur. Isolated for the first
time nitrifying bacteria and demonstrated role of these bacteria in nitrification, further he
demonstrated that free-living Clostridium pasteuriamum could fix atmospheric nitrogen (1893).
Therefore, he is considered as "Father of soil microbiology".
❑ M. W. Beijerinck (1888) isolated root nodule bacteria in pure culture from nodules in
legumes and named them as Bacillus radicola Considered as ”Father of Microbial ecology".
He was the first Director of the Delft School of microbiology (Netherland).

MICROBIOLOGY
❑ Beijerinck and Winogradsky (1890) developed the enrichment culture technique for
isolation of soil organisms, proved independently that transformation of nitrogen in nature is
largely due to the activities of various groups of soil microorganisms (1891). Therefore, they are
considered as "Pioneer’s in soil bacteriology”.
❑ S. N. Winogadsky (1891) demonstrated the role of bacteria in nitrification and further in fill
1983 demonstrated that free living Clostridium pasteurianum could fix atmospheric nitrogen.

emerged as a distinct branch of soil science during first half of the 19th centSome of the
SOIL HEALTH
❑ Soil health is a state of a soil meeting its range of ecosystem functions as
appropriate to its environment.
❑ Health of soil arises from favorable interactions of all soil components
(living and non-living) that belong together, as in microbiota, plants and
animals.
❑ Soil health depends on soil biodiversity (with a robust soil biota), and it can
be improved via soil management, especially by care to keep protective
living covers on the soil and by natural (carbon-containing) soil
amendments.
❑ Inorganic fertilizers do not necessarily damage soil health if
1) used at appropriate and not excessive rates and
2) if they bring about a general improvement of overall plant growth which contributes
more carbon-containing residues to the soil.

SOIL HEALTH
The term soil health is used to describe the state of a soil in…….
Sustaining plant and animal productivity (agronomic focus)
Enhancing biodiversity (Soil biodiversity) (ecological focus)
Enhancing water and air quality (environmental/climate
focus)
Supporting human health and habitation
Sequestering Carbon
(Source: https://en.wikipedia.org/wiki/Soil_health)
Parbhani

SOIL HEALTH
The term soil health is used to describe the state of a soil in…….
Sustaining plant and animal productivity (agronomic focus)
Enhancing biodiversity (Soil biodiversity) (ecological focus)
Enhancing water and air quality (environmental/climate
focus)
Supporting human health and habitation
Sequestering Carbon

SOIL BIODIVERSITY
❑ The great strength of soil comes from the life that exists within soil – soil
biodiversity – ranging from genes and species to communities.
❑ A single teaspoon of garden soil may contain thousands of species, millions
of individuals and a hundread meters of fungal networks.
❑ Scientists estimate that at least about one-quarter of species on planet Earth
live in soils.
(Source: Soil Biodiversity broucher-European Commission 2010)
Compiled and Edited
by

SOIL BIODIVERSITY
❑ Diverse ecosystem performs a variety of functions. It processes waste organic matter
to sustain life above the ground, from plants to animals to humans
❑ It regulates the carbon flux and the water cycle; it keeps pests at bay and
decontaminates polluted land; and
❑ It provides raw materials for new pharmaceuticals to tackle infectious diseases.
❑ This is the factory of life. Its workers are micro-organisms, small and large
invertebrates, small mammals, even plant roots – their workplace is the dark or dim
layers of topsoil beneath grasslands, fields, forests and green spaces in towns
Compiled and
Edited by
Rakesh
Bagmare
(Ph.D) VNMKV
Parbhani

SOIL BIODIVERSITY
This diverse ecosystem performs a variety of functions. It processes waste organic
matter to sustain life above the ground, from plants to animals to humans
It regulates the carbon flux and the water cycle; it keeps pests at bay and
decontaminates polluted land; and
it provides raw materials for new pharmaceuticals to tackle infectious diseases.
This is the factory of life. Its workers are micro-organisms, small and large
invertebrates, small mammals, even plant roots – their workplace is the dark or dim
layers of topsoil beneath grasslands, fields, forests and green spaces in towns.
Rakesh Bagmare (Ph.D)
VNMKV Parbhani

Soil organisms can be classified into three main groups which describe the principal
function they perform in the soil:
(Source: Soil Biodiversity broucher-European Commission 2010)
Chemical engineers are like workers on
the factory floor, the smallest organisms in
soil. They break down organic matter, like
leaf and plant litter, and transform residues
into nutrients, such as nitrogen,
phosphorus and sulphur.
Biological regulators can be likened to factory supervisors.
They regulate the abundance and activity of chemical
engineers, mainly through feeding, which enhances or
reduces the productivity of the system.
Ecosystem engineers are the factory’s
architects and builders. They design the
structure of soil that enables the other groups
to thrive, building passages, tunnels and pore
networks and transporting particles around the
bustling subterranean community.

Bacteria live in the water-filled pores in soil.
They reproduce rapidly and can double their population in minutes. But they
can also enter a dormant stage and come back to life after a period of years. .
FUNCTIONS
1. Decomposition of cellulose and other
carbohydrates
2. Ammonification (proteins ammonia),
3. Nitrification (ammonia-nitrites-nitrates),
4. Denitrification (release of free elemental
nitrogen),
5. Biological fixation of atmospheric
nitrogen (symbiotic and non-symbiotic)
6. Oxidation and reduction of sulphur &
iron compounds.
(Source: Soil Biodiversity broucher
European Commission 2010)

HOSTORICAL OVERVIEW OF SOIL MICROBIOLOGY
Fungi comprise an electic group that vary from single celled yeasts to complex structures
visible to the naked eye. They inhabit the spaces around soil particles, plant roots. Some species
recycle dead or decaying organic matter, while others break up sugars, starches and cellulose in
wood.
FUNCTIONS
1. Fungi plays significant role in soils and plant
nutrition.
2. They plays important role in the degradation/
decomposition of cellulose, hemi cellulose, starch,
pectin, lignin in the organic matter added to the soil.
3. Lignin which is resistant to decomposition by
bacteria is mainly decomposed by fungi.
4. They also serve as food for bacteria.
5. They also plays important role in soil aggregation
and in the formation of humus

The supervisors
Protists are the smallest in this
group. They live in the water
layer around soil particles and
control bacteria populations
through feeding. They can propel
themselves by means of tiny oar-
or flipper-like body parts and can
be transported by wind as well as
water while in a dormant phase.
Nematodes are tiny worm-like creatures,
most of them measuring up to 1mm long.
They are highly adaptable and found in
all types of soil, including extreme
environments in Antarctica and the deep
ocean. Their prey varies from algae,
bacteria and fungi to plant roots, other
nematodes and protists.

PROGRAMME OF RESEARCH WORK
Microarthropods are small invertebrates
(animals without a backbone) which mainly
inhabit topsoil, feeding on decaying
vegetation, bacteria and fungi, as well as
their fellow ‘supervisors’.
They vary in size from microscopic
(certain mites) to several millimetres in
length (springtails, for example, which hop
around using their tail as a spring
(Source: Soil Biodiversity broucher European
Commission 2010)

The Architecture
Earthworms, termites, ants, isopods (such as
woodlice) and plant roots all fall into this group, as
do millipedes, centipedes, beetles, caterpillars and
scorpions.
Part-time soil residents such as voles, snakes,
lizards, mice and rabbits are also ecosystem
engineers, moving the soil as they dig for food and
shelter.

Practices that Improve the Soil Health
Source: Technical note-2019 Scottish government
Compiled and
Edited by
Rakesh
Bagmare (Ph.D)
VNMKV
Parbhani

Source:
Concept of Biofertilizer
Biofertilizers ( Microbial inoculants) can generally be defined as preparation containing live or
latent cells of efficient strains of nitrogen fixing, phosphate solublizing or cellulolytic
microorganism used for application to seeds, soils or composting areas with the objective of increasing
the number of such microorganisms and accelerate those microbial process which augment the
availability of nutrients that can be easily assimilated by plants
(Source: Kumar et al., 2017, Role of Biofertilizers in Agriculture, Popular Kheti, 5(4): 63-66)
Compiled and
Edited by
Rakesh Bagmare
(Ph.D) VNMKV
Parbhani

(Source: Trishna Mahanty et al., 2016 Environmental Science and Pollution Research · November)
Plant growth-enhancing
mechanisms of plant growth promoting rhizobacteria (PGPR)

(Source: Barman et al.,(2017), International Journal of Current Microbiology and Applied science ,6(11);1177-1186)
Biofertilizer
0
2000
4000
6000
8000
10000
12000
14000
16000
2008-09 2009-10 2010-11 2011-12 2012-13 2013-14 2014-15
1249.87
1861.33
2924
8743.69
5897.91
6218.607
14847.397
Biofertilizer production of Maharashtra (tones)
Biofertilizer
production in tones
Compiled and
Edited by
Rakesh Bagmare
(Ph.D) VNMKV
Parbhani

(Source:
Isolation Methods for Pure Culture

(Source: Microbiology Module Note Europe 2019)
Pure Culture Techniques
Culture : Act of cultivating microorganisms or the
microorganisms that are cultivated
Mixed Culture : more than one microorganism
Pure Culture : containing a single species of
organism
A Pure Culture is usually derived from a mixed culture (one containing many
species) by transferring a small sample into new, sterile growth medium in
such a manner as to disperse the individual cells across the medium surface or
by thinning the sample many times before inoculating the new medium.

Pure cultures are important in microbiology for the following
reasons-
Why So Important…….
1. Once purified, the isolated species can be cultivated with the knowledge that
only the desired microorganism is being grown.
2. A pure culture can be correctly identified for accurate studying and testing,
and diagnosis in a clinical environment.
3. Testing/experimenting with a pure culture ensures that the same results can
be achieved regardless of how many time the test is repeated. Compiled and Edited
by
Rakesh Bagmare
(Ph.D) VNMKV
Parbhani

Common Isolation Techniques of Pure Culture
Streak plate method
Pour plate method
Spread plate method
Compiled and Edited
by
VNMKV Parbhani

1. Streak Plate Method
▪ Streaking is the process of spreading the microbial culture
with an inoculating needle on the surface of the media.
▪ Sterilize the inoculating needle by flame to make red hot
And allow it to cool for 30 seconds.
▪ The sample is streaked in such a way to provide series
of dilution.
▪ Purpose-thin out innoculum to get seperate colonies.
▪ Subculturing can bed one by streaking well isolated colonies from streak plate to new
plate Compiled and Edited by
VNMKV Parbhani

2. Pour Plate Method
❑ The bacterial culture and liquid agar medium are
Mixed together.
❑ After mixing the medium, the medium containing
the culture poured into sterilized petri dishes
( petriplates), allowed solidifying and then incubated.
❑ After incubation colonies appear on the surface.
VNMKV Parbhani

(Source:
Compiled and Edited
by
VNMKV Parbhani

e Techniques
(Source: Microbiology Module Note Europe)
Why to Preserve Bacteria????
❑ In nature there are only 1% bacteria which is pathogenic and harmful to
Animalia and Plantae group.
❑ 99% of bacterial populations are of economic importance for human beings
and plants.
❑ In soil for nutrient uptake in food industry, in sewage treatment, In medical
industry.
❑ So the preservation of bacteria is one of the most profitable practice
economically as well as environmentally.
VNMKV Parbhani

Why We Isolate the Anaerobic Bacteria
from Aerobic Soil Condition??????
e Techn
Parbhani

❑It is difficult to identify bacteria using phenotypic methods, However It can
be accurately identified using 16S rRNA gene sequence analysis.
❑Identification of common bacteria, identification of bacteria difficult to
distinguish, identification of rhizobacteria, identification of many bacteria existing
in the microenvironment
❑ Campared with traditional classification that is based on the morphology and
physiology of bacteria and the biochemical charecteristics on the basis of
phenotypic charecteristics.
❑ 16S rRNA sequence analysis has the advantages in the analysis and the
identification of evolution and affinity.
16S rRNA Sequence Analysis Technique in the
Taxonomic Identification of Bacteria
Parbhani

16S rRNA Sequence Analysis Technique in the
Taxonomic Identification of Bacteria
Source: Janda and Abbott (2007) Journal Of Clinical Microbiology,76, 2761–2764
VNMKV Parbhani

(Source : Book Agriculture Microbiology bu TNAU and Soil Microbiology by Singh , Purohit and Parihar,(Agribios India)
Microbial Isolates and its Role in
Nutrients Mobilization
Nitrogen
Phopherous
Potassium
Iron
Zinc
Sulphur
Nutrients mobilized by
microorganism
VNMKV Parbhani

(Source : Book Agriculture Microbiology by TNAU and Soil Microbiology by Singh , Purohit and Parihar,(Agribios India)
Biological Nitrogen Fixation
❑ Reduction of N 2 to NH 3 by living organisms is called
as a Biiological Nitrogen Fixation.
❑ That is done with the help of Nitrogenase enzyme
complex
❑ The Nitrogen fixing bacteria could be
❑ Free living- Azotobacter ( Aerobic ) and Rhodospirillum
( anaerobic )
❑ Symbiotic N 2 fixation: Rhizobium symbiotically
associated with legumes Compiled and Edited
by
VNMKV Parbhani

(Source : Soil Microbiology by Singh , Purohit and Parihar,(Agribios India)
Compiled and
Edited by
Rakesh
Bagmare
(Ph.D)
VNMKV
Parbhani

(Source : Soil Microbiology by Singh , Purohit and Parihar,(Agribios India)
Parbhani

Is More Number Of Root Nodules Will Be
Criteria For More Biological Nitrogen Fixation
?????
VNMKV Parbhani

Source: https://www.slideshare.net/JayvirSolanki1/role-of-microbes-in-nutrient-mobilization-transformation-
Nitrogen Mobilization/
Transformation
VNMKV Parbhani

Source: https://www.slideshare.net/JayvirSolanki1/role-of-microbes-in-nutrient-
mobilization-transformation-
Phosphorus Mobilization/
Transformation
Parbhani

(Source: https://en.wikipedia.org/wiki/phophorus transformation)
Phosphorus Mobilization/
Transformation
Some important microorganism for P Solubilization
➢ Bacillus sp.
➢ Beijernckia sp.
➢ Burkholderia sp.
➢ Enterobacter sp.
➢ Flavobacterium sp.
➢ Microbacterium sp.
➢ Pseudomons sp.
➢ Mesorhizobium cicero sp.

https://www.sciencedirect.com/science/article/pii/S0176161713003611
Potassium Mobilization/
Transformation
1. 98% of this K is bound within the Phyllosilicates structures .
2. The remaining 2% exists in soil solution or on exchange sites to become available
for plants
3. Hence, soil fertility is decreased due to low availability of this nutrient .
4. Many microorganism in the soil are able to solubilize unavailable forms of K-
bearing minerals, such as micas, feldspar, illite and orthoclasses by excreting
organic acids which either directly dissolves rock K or chelate silicon ions to bring
the K into solution .
Parbhani

https://www.sciencedirect.com/science/article/pii/S0176161713003611
Transformation
Some important microorganism for K Solubilization
1. Frateuria sp.
2. Acidothiobacillus ferrooxidans
3. Bacillus mucilaginosus
4. B. edaphicus
5. Burkholderia sp.
6. Pseudomonas sp.
7. Rhizobium sp.
Parbhani

Source: https://www.slideshare.net/JayvirSolanki1/role-of-microbes-in-nutrient-mobilization-transformation-
Transformation
VNMKV Parbhani

Source: https://en.wikipedia.org/wiki/Siderophore
Siderophore Production by Microbial
Isolates
❑ Siderophores are compounds from ancient Greek words, sidero ‘iron’ and
phore ‘carriers’ meaning ‘iron carriers’. These are low-molecular-weight
iron-chelating compounds, produced by ‘rhizospheric bacteria’ under iron-
limited conditions.
❑ They are small, high affinity iron chelating compounds secreted by
microorganisms such as bacteria, fungi etc. Siderophore usually form a
stable hexahendate, octahedral complex with Fe3+
Compiled and Edited
by
VNMKV Parbhani

Siderophore
Siderophore receptor site on cell
Outside the Cell
Inside the Cell
Fe
3+
HOW SIDEROPHORES WILL FORM ??
Parbhani

Outside the Cell
Fe
3+
Fe
3+
Inside the Cell
HOW SIDEROPHORES WILL FORM ?
VNMKV Parbhani

Fe
3+
Fe
2+
Siderophore
Outside the Cell
Fe
3+
Inside the Cell
HOW SIDEROPHORES WILL FORM ?
Parbhani

Source: Soil Microbiology by Singh , Purohit and Parihar,(Agribios India)
Zinc Solubilization by Microbial
Isolates
❑ Zinc is required in relatively small concentration although prevalence of Zn
deficiency in crop is due to low solubility of Zn rather than low Zn
availability.
❑ 50% of Indian soils are Zinc deficient.
❑ Solubility of Zn decrease with.
Increase in pH
High organic matter
Bicarbonate content
High magnesium to calcium ratio
High availability of P and Fe
VNMKV Parbhani

Zinc Solubilization by Microbial
Isolates
▪ The Zinc applied to agriculture fields as Zinc sulphate (Soluble) get converted to
different insoluble forms like -
❑ Zinc hydroxide [Zn(OH)2] at high soil pH,
❑ Zinc Carbonate [ZnCo3 ] in calcium rich alkali soils,
❑ Zinc phosphate [Zn3(PO4)2] in near neutral to alkali soil with large application of P
fertilizers and
❑ Zinc sulfide [ZnS] under reducing conditions particularly during flooding .
▪ The soluble form of Zn fertilizers applied to the fields become readily insoluble
forms that cannot be assimilated by plants which leads to the Zn deficiency in crops.
▪ The microbes solubilize the Zn by lowering the pH by gluconic acid and indole
acetic acid production.
VNMKV Parbhani

Microbial Biofortification
➢In developing countries, up to ~50% of pregnant women and ~40% of preschool children
suffer from anaemia.
➢Microbial biofortification is the process of adding essential micronutrients and other
health-promoting compounds to crops or foods to improve their nutritional value with the
help of plant growth promoting rhizobacteria.
.

Microbial Biofortification
➢ In developing countries, up to ~50% of pregnant women and ~40% of preschool children suffer
from anaemia.
➢ Microbial biofortification is the process of adding essential micronutrients and other
health-promoting compounds to crops or foods to improve their nutritional value with the help
of plant growth promoting rhizobacteria.
➢ Iron (Fe) is undisputably one of the most important micronutrients in the human diet, the basis
for the correct function of haemoglobin, and deficiency in iron can lead to anaemia.
➢ Zinc (Zn) plays a critical role in strengthening of the immune system, male fertility and
human health.
Source: Yee-Shan Ku et al., 2019 Agronomy, 9, 764;
VNMKV Parbhani

Mechanism of Microbial
Biofortification
The Zinc applied to agriculture fields as Zinc sulphate (Soluble) get converted to
different insoluble forms like:
✓ Zinc hydroxide [Zn(OH)2] at high soil pH,
✓ Zinc Carbonate [ZnCo3 ] in calcium rich alkali soils,
✓ Zinc phosphate [Zn3(PO4)2] in near neutral to alkali soil with large
application of P fertilizers and
✓ Zinc sulfide [ZnS] under reducing conditions particularly during
flooding
The soluble form of Zn fertilizers applied to the fields become readily insoluble forms
that cannot be assimilated by plants which leads to the Zn deficiency in crops.
The microbes solubilize the Zn by lowering the pH by gluconic acid and indole acetic
acid production .
Source: Yee-Shan Ku et al., 2019 Agronomy, 9, 764;
Parbhani

❑ Several mechanisms have been proposed for PGPR mediated drought stress
tolerance in plants.
❑ It includes phyto hormonal activity, volatile compounds, alteration in root
morphology, ACC deaminase activity, accumulation of osmolytes, EPS
production, antioxidant defense and co-inoculations.
❑ The term Induced Systemic Tolerance (IST) has been coined to accommodate the
microbial induced physical and chemical changes in plants, which result in enhanced
tolerance to abiotic stresses
Drought Tolerance Mechanism By
Microbial Isolates
Source: Sai Shiva Krishna Prasad Vurukonda et al., (2016) Microbiological Research 184 13–24
VNMKV Parbhani

Role of bacterial ACC deaminase in stress agriculture
➢ The overproduction of ethylene in response to abiotic and biotic stresses leads to
inhibition of root growth and consequently growth of the plant as a whole.
➢ Ethylene synthesis is stimulated by a variety of environmental factors which hamper
plant growth.
➢ ACC deaminase PGPR boost plant growth particularly under stressed conditions by
the regulation of accelerated ethylene production in response to a multitude of
abiotic and biotic stresses like salinity, drought, water logging, temperature,
pathogenicity and contaminants.
➢ Applications of PGPR containing ACC deaminase in relation to the nature of stress
are salinity stress, drought, Temperature stress etc
Microbial Isolates
Source: Sai Shiva Krishna Prasad Vurukonda et al., (2016) Microbiological Research 184 13–24
VNMKV Parbhani

Microbial Isolates
Source: Sai Shiva Krishna Prasad Vurukonda et al., (2016) Microbiological Research 184, 13–24
VNMKV Parbhani

Crop Residues Decomposition By
Microorganisms
▪ On an Average 500 Mt of crop residues is
generated in India.
▪ While a majority of its used as a fodder, raw
material and energy production etc.
▪ Still there is a huge surplus of 140 Mt out of
which 92 Mt is Burnt each year, Mainly states
such as Punjab, Haryana and Uttar Pradesh
Source: wikipedia.org/stubble burning
Harmful Effects -
▪ Loss of Nutrients,
▪ Gasses release from it are very
harmful,
▪ Some causes the holes in ozone layer
▪ Pollution from smoke,
▪ Risk of fire Compiled and Edited by
Parbhani

(Source: Dotaniya, M. L., & Meena, V. D. (2015) Proceedings of the National Academy of Sciences, India
Section B: Biological Sciences, 85(1), 1-12.)
Table no. 1 Microorganism population in the Rhizosphere of wheat
(Triticum aestivum L.) and bulk soil.
Microorganisms Rhizosphere soil Bulk soil
(CFU g-1 soil)
Bacteria 1.2 ×109 5.3 × 107
Actinomycetes 4.6 × 107 7.0 × 106
Fungi 1.2 × 106 1.0 × 105
Protozoa 2.4 × 103 1.0 × 103
Algae 5.0 × 103 2.7 × 104
Ammonifiers 5.0 × 108 4.0 × 106
Denitrifiers 1.26 × 108 1.0 × 105

(Cheke, A. S., Patil, V. D., and Srivastava, A. K.(2018) Studies on rhizosphere hybridization and nutrient
dynamics in sweet orange seedling from pot culture experiment. JPP , P-ISSN: 2349-8234 SP1: 3077-3082)
Table no. 2 Effect of various rhizosphere soil on number of germinated
seeds (out of 10) and percent germination. ( sweet orange seed )
Symbol Soil Type
Number of
Germinated seed
N=10
Seed Germination
(%)
T1 Control (Non rhizosphere) 3 30 (%)
T2
Rhizosphere soil of Ficus
religiosa L. (Pipal tree)
4 40 (%)
T3
benghalensis L. (Banyan tree)
7 70 (%)
T4
racemosa L. (Umber tree)
10 100 (%)

Table no. 3 Effect of zinc mobilizing cultures on DTPA micronutrient in
soil after harvest of soybean
.
Treatments
DTPA Zn
(mg kg-1)
DTPA Fe
(mg kg-1)
DTPA Cu
(mg kg-1)
DTPA Mn
(mg kg-1)
Zinc solubilizers (S)
S0: Control
0.55 3.24 3.41 7.32
S1: Pseudomona striata
0.66 3.61 3.82 7.80
S2: Bacillus megaterium
0.61 3.56 3.66 7.79
S3: Trichoderma viride
0.62 3.55 3.66 7.76
S.Em.±
0.003 0.04 0.049 0.045
C.D. at 5%
0.008 0.13 0.142 0.13
Bhushan Pagar, Syed Ismail and Prashant H Rathod (2019). Effect of zinc mobilizing cultures and zinc
levels on chemical properties and nutrient availability in soybean grown Vertisols. Journal of
Pharmacognosy and Phytochemistry, 8(5): 1290-1295.

Table no. 4 Potassium solubilization activity of various microorganisms
under insoluble potassium source in solid media (Aleksandrov media-
Plate assay )
.
Nihala Jabin, P.P. and Syed Ismail,( 2017). Solubilization of Insoluble Potassium by Different Microbial
Isolates in vitro Condition. International Journal of Current Microbiology and Applied Sciences. 6(10):
3600-3607
Treatment
Colony diameter
(cm)
Halozone diameter (cm)
Microbial inoculants
T1: Bacillus sp (KSB-W1) 0.47 0.87
T2: Bacillus sp (KSB-PD-3-A) 0.83 1.90
T3: Bacillus sp (KSB-NP-3) 0.63 1.20
T4: Pseudomonas sp (KSB-PD-1-A) 1.17 3.00
T5: Pseudomonas sp (KSB-M-1) 1.07 2.70
T7: Sinorhizobium metallidans (KSB-PD) 0.73 1.53
T8: Sinorhizobium metallidans (KSB-1-B) 0.83 1.83
T9: Sinorhizobium metallidans (KSB-M-3) 0.67 1.33
SE+ 0.03 0.07
CD at 1% 0.13 0.23

Table no. 5 Potassium solubilization activity of various microorganisms
under insoluble potassium source in solid media (Aleksandrov media-
Plate assay )
.
Nihala Jabin, P.P. and Syed Ismail,( 2017). Solubilization of Insoluble Potassium by Different Microbial
Isolates in vitro Condition. International Journal of Current Microbiology and Applied Sciences. 6(10):
3600-3607
Treatment Solubilization Efficiency (%) Solubilization Index
Microbial inoculants
T1: Bacillus sp (KSB-W1) 185 2.85
T2: Bacillus sp (KSB-PD-3-A) 228.24 3.28
T3: Bacillus sp (KSB-NP-3) 189.68 2.89
T4: Pseudomonas sp (KSB-PD-1-A) 257.32 3.57
T7: Sinorhizobium metallidans (KSB-PD) 208.93 3.08
T8: Sinorhizobium metallidans (KSB-1-B) 219.90 3.19
T9: Sinorhizobium metallidans (KSB-M-3) 200.79 3.00
SE+ 5.47 0.05
CD at 1% 17.01 0.16

Table no.6 Solubilization of insoluble zinc compounds (per cent) in solid
medium by different microbial inoculants
.
Pawar A , Ismail Syed, Mundhe Swati and Patil V.D. (2015) Solubilization of in soluble zinc compounds by
different microbial isolates in vitro condition International Journal of Tropical agriculture.33(2).
Zinc sources at 0.1%
Colony diameter(cm) Halo zone diameter (cm)
Microbial Inoculants ZnO ZnCO3 Zn3(PO4)2 ZnO ZnCO3 Zn3(PO4)2
T1 : Burkholderia cepacia 1.56 1.10 1.40 2.46 1.66 2.20
T2 : Burkholderia
cenocepacia
1.50 1.16 1.70 3.53 1.50 2.20
T3 : Pseudomonas
fluorescence
1.60 1.50 1.80 3.25 2.13 2.73
T4 : Pseudomonas striata 1.66 2.06 1.30 3.60 4.10 2.03
T5 : Trichoderma viridae 1.93 2.33 1.40 3.80 4.10 2.30
T6 : Trichoderma
harzianum
1.76 1.96 1.30 3.00 3.96 2.10
T7 : Bacillus megaterium 1.93 2.20 1.93 2.93 4.06 2.86
SE+ 0.01 0.01 0.028 0.02 0.03 0.05
CD at 5% 0.02 0.04 0.079 0.05 0.08 0.15

Table no.7 Effect of siderophore producing microorganisms along with
Rhizobium phaseoli on number of nodules, nodule fresh weight, nodule
dry weight per plant in green gram
.
Bagmare RR and Syed Ismail (2019) Siderophore production by microbial isolates and their effect on
nodulation attributes in green gram. International Journal of Chemical Studies; 7(5): 1287-1290
Sr. No. Treatments
No. of nodules
plant-1
Nodule
fresh wt
plant-1 (mg)
Nodule Dry
wt
plant-1 (mg)
T1 Absolute control 14.67 20.00 8.67
T2 Only RDF 21.67 35.00 13.00
T3 RDF+Rhizobium phaseoli 25.00 47.33 22.00
T4 T3 + Pseudomonas flurescens 39.00 74.67 39.33
T5 T3+ Pseudomonas striata 30.00 40.67 22.67
T6 T3+ Bacillus subtilis 25.33 40.33 19.33
T7 T3+Bacillus polymyxa 25.67 50.67 31.00
T8 T3+Bacillus megaterium 25.67 40.00 20.00
T9 T3+ Azotobacter chroococcum 30.00 46.67 31.67
T10 T3+ Azospirillum lipoferum 28.33 48.00 27.00
S.Em.± 1.62 2.64 1.75
C.D. at 5 % 4.81 7.84 5.19
C.V. % 10.58 10.31 12.90

Table no.8 Effect of zinc solubilizing microorganisms on enzyme
activities in soil after harvest of summer groundnut
.
Tr. no. Treatment Enzyme activities in soil after harvest of
groundnut(μg g-1)
Alkaline
Phosphatase
Acid
Phosphatase
Dehydrogenase
T1 RDF+Rhizobium 56.03 35.42 49.81
T3 T1+Burkholderia cepacia 65.33 38.85 53.03
T4 T1+Burkholderia cenocepacia 69.25 38.66 53.75
T5 T1+Pseudomonas fluorescens 72.76 39.67 57.43
T6 T1+Pseudomonas striata 75.58 42.87 57.75
T7 T1+Trichoderma viiide 75.18 39.50 53.38
T8 T1+Trichoderma harzianum 66.65 39.36 54.52
S.E.± 0.16 0.19 0.10
C.D. at 5 % 0.51 0.58 0.32
C.V. % 4.1 8.3 3.4
Sable Prasad*, Ismail Syed And Pawar Anuradha (2016) Effect Of Zinc Solubilizing Microorganisms In Enhancing Enzyme
Activity And Nutrient Availability In Groundnut Grown On Vertisol, International Journal of Agriculture Sciences. 49(8): 2099-
2102.

Table no.9 Effect of zinc solubilizing cultures and zinc levels on DTPA micronutrient in
soil after harvest of spinach.
.
YA Waghmare, AL Dhamak and Syed Ismail(2019) Impact of levels of zinc and various zinc solubilizing microorganisms on
changes in zinc and other soil nutrients in spinach, International Journal of Chemical Studies 8(1): 529-535
Treatments
DTPA Zn (mg kg-
1)
DTPA Fe (mg kg-
1)
DTPA Cu (mg
kg-1)
DTPA Mn (mg kg-
1)
Zinc solubilizers (S)
S0: Control 0.38 2.28 0.88 7.31
S1: Pseudomonas striata 0.64 3.05 0.96 7.74
S2: Trichoderma viride 0.51 2.53 0.92 7.58
S3: Bacillus megaterium 0.55 2.68 0.94 7.66
S.Em.± 0.01 0.03 0.007 0.06
C.D. at 5 % 0.03 0.10 0.02 0.19
Levels of ZnSO4 (Zn)
Zn0: ZnSO4 0 kg ha-1 0.38 2.43 0.85 7.14
Zn1: ZnSO4 10 kg ha-1 0.44 2.51 0.92 7.51
Zn2: ZnSO4 20 kg ha-1 0.57 2.67 0.94 7.73
Zn3: ZnSO4 30 kg ha-1 0.68 2.94 0.98 7.92
S.Em.± 0.01 0.03 0.007 0.06
C.D. at 5 % 0.03 0.10 0.02 0.19
S.Em.± 0.02 0.07 0.01 0.13
C.D. at 5 % 0.07 0.21 0.03 0.39

Table no.10 Effect of phosphorus and zinc solubilizing acidifiers on soil microbial
population after harvest of summer soybean
.
B Kranti Kumar and Syed Ismail (2017) Influence of different microbial inoculants on biological health of soil and economics
in Soybean crop grown on Vertisol, Journal of Pharmacognosy and Phytochemistry, 6(3): 198-201
Tr.
No.
Treatment Bacteria
(CFU X10-7)
Actinomycetes
(CFU X10-7)
Fungi
(CFU X 10-4)
T1 RDF+ Rhizobium 24.6 14.3 3.00
T6 T1+Trichoderma viride 28.0 24.0 6.67
S.E. ± 1.44 1.35 0.63
C.D. at 5 % 4.42 4.15 1.93
C.V. % 8.35 13.5 18.5

Table 11. Influence of bacterial isolates inoculation on sunflower germination and
growth in plate experiment
Raval, A. A., and Desai, P. B. (2015). Screening and Characterization of Several siderophore Producing
Bacteria as Plant Growth-Promoters and Biocontrolling agents. International Journal of Pharmacy & Life
Sciences, 6. 112-120 (Surat,Gujrat)
No Given Bacterial isolates Root length Shoot length Vigor index Germination%
C Control 5.6 3.1 609 70%
M1R2 Pseudomonas spp. 9.6 5.2 1480 100%
M7R1 Pseudomonas spp. 4.2 5.5 582 60%
M6S3 Enterobacter spp. 5.4 6.6 840 70%
M7S1
Stenotrophomonas
spp
9.0 5.2 1420 100%
M7ER1 Microbacterium spp. 9.9 5.3 1292 85%
M7ER2 Microbacterium spp. 6.0 3.1 728 80%
M12R2 Bacillus spp. 7.8 6.2 1120 80%
M12R1 Bacillus spp. 10.6 3.5 930.6 66%

Table 12: Siderophorogenesis by PGPR isolates.
Gupta A. and Murali G(2008),Indian journal of Agricultral Research (42) 153-156.
PGPR Isolates Isolate no. Incubation time(hr) % siderophore units
Bacillus coagulans BpN 48 -
Bacillus sp. BAC 32 -
Bacillus polymyxa BbP 48 -
Brevibacillus brevis BbC 52 21
Enterobacter sp. EB-RS-1 52 38
Pseudomonas sp. Z2 24 51
Pseudomonas fluorescens PF11 24 76
Pseudomonas striata PS1 32 -
Azospirillum brasilense SP7 72 24
Enterobacter sp. EG-ER-2 48 64

Fig: Effect of Siderophore on growth of Lens culinaris (Masoor Dal)
Parmar, H. Y., and Chakraborty, H. (2016). Effect of siderophore on plant growth promotion. International
Journal of Applied Pure Sciences, 2(3), 60-68.

Table no.13 Effect of inoculation with rhizobacteria containing 1-aminocyclopropane-1-
carboxylic acid-deaminase on the number of pods and the total grain yield of peas
exposed to drought stress at different stages of growth (average of six replicates)
.
Arshad M., Shaharoona B. and Mahmood T. (2008) Inoculation with Pseudomonas spp. Containing ACC-Deaminase Partially
Eliminates the Effects of Drought Stress on Growth, Yield, and Ripening of Pea (Pisum sativum L.). Pedosphere 18(5): 611–
620(Ravalpindi, Pakisthan)
Growth stages
exposed to
drought stress
Number of Pods/ plant Grain yield/ plant
Uninoculate
d
P. Putida P. fluorescens Uninoculate
d
P. Putida P. fluorescens
Control 3.5 4.7 5.3 11.0 17.6 17.4
Vegetative 2.8 3.3 4.8 7.9 8.4 10.2
Flowering 2.7 4.0
5.8 9.4 10.6
15.2
Pod Formation 2.1 3.2 3.3 7.2 8.7 10.1
Mean 2.8 3.8 4.8 8.9 11.3 13.2

Table no.14 Effect of different microbial inoculants on enzyme activities in soil after
harvest of summer soybean
.
B Kranti Kumar and Syed Ismail (2017) Influence of different microbial inoculants on biological health of soil and economics
in Soybean crop grown on Vertisol, Journal of Pharmacognosy and Phytochemistry, 6(3): 198-201
Tr. No. Treatment Enzyme activities in soil after harvest of
Soybean(μg g-1)
Alkaline
phosphatase
Acid
Phosphatase
Dehydrogenas
e
T1 RDF+ Rhizobium 99.6 44.6 37.03
T6 T1+Trichoderma viride 118.0 53.5 46.1
S.E. ± 2.29 1.35 0.58
C.D. at 5 % 7.04 4.13 1.78
C.V. % 3.44 4.47 2.42

conclusion
Microorganism population, microbial activity is high in rhizospheric soil
than that of bulk soil due ideal soil environment and food and
Rhizospheric soil also act as source of nutrient for plant and
microorganism.
Biofertilizers being essential components of organic farming play a vital
role in maintaining long term soil fertility and sustainability by fixing
atmospheric nitrogen, mobilizing fixed macro and micro nutrients in the
soil into forms available to plants.
Plant growth promoting rhizobacteria help plants tolerate drought stress.
Some of the physiological mechanisms that have been proposed include
alterations in root architecture, root surface area which results in
improved water and nutrient uptake, with positive effects on the overall
plant growth.
1
2
3

conclusion
Nutrient- solubilizing rhizobacteria (NSR) have emerged as one of the
alternatives to application of chemical inputs for needs of fertilizers.
Their use in agriculture in preference to chemical fertilizers offers
economic and ecological benefits by way of soil health and fertility to
farmers.
5
Rhizosphere soil having high availability of nutrients than the bulk soil
due to organic acid produced during secretion by root, microorganism
and decomposition of organic matter and maintain physical, chemical
and biological environment of soil which play an important role in
mobilization N, P , Fe, Mn, Zn and Cu etc
4

conclusion
Soil biota and soil fertility are key interacting factors in determining
the complementarity effects in plant and the plant species richness–
community biomass relationship
Nutrient-solubilizing rhizobacteria (NSR) have emerged as one of the
alternatives to application of chemical inputs for needs of fertilizers.
Their use in agriculture in preference to chemical fertilizers offers
economic and ecological benefits by way of soil health and fertility to
farmers.
4
6
Rhizosphere soil having high avilabity of nutrients than the bulk soil
due to organic acid produced during secretion by root, microorganism
and decomposition of organic matter and maintain physical, chemical
and biological environment of soil which play an important role in
mobilization N, P , Fe, Mn, Zn and Cu etc
5

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