1. The study aimed to isolate and characterize phosphate solubilizing bacteria (PSB) from heavy metal contaminated soil near cement industries in Srinagar. Soil samples were collected and PSB were isolated using serial dilution and spread plate technique on Pikovskaya media. 2. Isolated PSB were tested for their ability to solubilize phosphate and resistance to heavy metals. Those showing resistance to heavy metals were further evaluated for various plant growth promoting activities. 3. The expected outcomes were isolation of PSB and heavy metal resistant PSB from contaminated soil. Evaluation of heavy metal resistant PSB for their plant growth promoting potential.

1. Characterization of Heavy Metal Resistant Phosphate Solubilizing Bacteria
Isolated from Nearby Contaminated Soil of Cement Industry
METHODOLOGY SEMINAR
PRESENTED BY : SAKIDAH MANZOOR
ENROLLMENT NO. : 21072119014
MSc. Microbiology, 4th Sem 2021
SUPERVISED BY
DR.ULFAT NAZIR
P.G Proramme In Microbiology
Centre Of Research For Development (CORD)
University Of Kashmir, Hazratbal, Srinagar,190006

3. Presentation
Outline

4. INTRODUCTION
Phosphorus is one of the major essential macronutrient which is involved
in plant metabolism and is applied to soil in the form phosphate fertilizer
and large portion of soluble inorganic phosphate is immobilized rapidly
and become unavailable to plant (Motsim et., al 2019; Balemi et al., 2012;
Razaq et al., 2017; Sanz-Sáez et al.,2017; Malhotra et.,2018).
Phosphate Solubilizing Microorganisms(PSMs) are diverse group of
microorganisms including bacteria and fungi to convert insoluble form of
Phosphorus into soluble forms by a process called Phosphate
Solubilization (Leandro et al., 2023).
Important Genera of bacterial Phosphate Solubilizers include Bacillus,
Pseudomonas, Micrococcus and Flavobacterium, while fungal genera
include Aspergillus, Pencillium, Fusarium and Sclerotium (MS et al.,
2009; Seenivasagan et al., 2021).

5. INTRODUCTION (Cont.)
Phosphate-solubilizing bacteria play an essential role in P-cycling and
promoting plant growth. By using PSMs, they can reduce the need for
synthetic phosphorus fertilizers,which can be costly and have negative
environmental impacts when overused (Zhonghua et al., 2022).
The accumulation of heavy metals by agricultural activities in soil poses a
risk to the environmental and human health. In soil, heavy metals cause
toxicological effects on soil microbes, which may lead to a decrease in
their numbers and activities (DalCorso et al., 2013)
Environmental pollution by heavy metals has become a serious problem
in the world. The problem of heavy metals pollution is becoming more
and more serious with increasing industrialization and disturbance of
natural biogeochemical cycles (Khan et al., 2010).

6. INTRODUCTION (Cont.)
Cement factories are known to emit a variety of pollutants, including
particulate matter, gases, and various heavy metals. Cement dust can
indeed contain heavy metals, including Cr, Ni, Co, Pb and Hg which are
known to be harmful to the biotic environment (Akpambang et al.,
2022).
Heavy metals cannot be degraded, biological processes can still be used
for remediation and management of heavy metal-contaminated sites
through various techniques via Biosorption, Phytoremediation,
Biostimulation, Bioaugmentation (Lebeau et al., 2008).
Microorganisms like Heavy Metal Resistant PSBs have evolved different
strategies and mechanisms to survive in heavy metal-contaminated
environments. Heavy Metal Resistant PSB includes, Pseudomonas spp.,
Bacillus spp., Enterobacter spp., Arthrobacter spp., Stenotrophomonas
maltophilia, Acinetobacter spp. (Fakhar et al., 2022).

7. INTRODUCTION (Cont.)
Phosphate-solubilizing bacteria (PSB) do not directly degrade heavy
metals but mitigate the adverse effects of heavy metal toxicity, enhancing
plant growth and health through several mechanisms Phytostimulation,
Enhancing Metal Tolerance, Inducing Metal Sequestration,
Bioremediation and Improving Soil Structure (Mohamed et al., 2017).
The Heavy metal resistance of these bacteria may vary depending on the
specific heavy metal and its concentration in the environment.
Additionally, the efficiency of these bacteria in phosphate solubilization
and heavy metal resistance can be influenced by soil pH, temperature, and
other environmental factors (Paul et al., 2015).
However, it's essential to carefully select and evaluate the most suitable
strains for specific soil conditions and targeted heavy metal contaminants
to ensure successful applications (Priya et al., 2023).

8. OBJECTIVES
OBJECTIVE 1
Isolation and
Screening of
Phosphate
Solubilizing
Bacteria.
OBJECTIVE 2
Assay of Bacterial
strains for Heavy
Metal
Resistance.
OBJECTIVE 3
Evaluation of
Heavy Metal
Resistant Bacteria
for Plant Growth
Promoting
activities.
Keeping all these considerations in view the present investigation is proposed to be undertaken
with the following objectives:

9. METHODOLOGY
Isolation of Phosphate Solubilizing Bacteria
S o i l S a m p l i n g
Serial Dilution (10⁻⁶ to 10⁻⁷)
Spread Plate Technique
Phosphate Solublizing Bacteria (PSB) will be isolated from nearby Heavy Metal
contaminated soil of different Cement Industries in Srinagar in sterilized polythene bags
aseptically with proper labels to prevent moisture loss.
Serial Dilution technique of the soil sample will be employed as it will reduce the number
of bacteria in the sample.
 An aliquots from decimal dilutions will be spreaded by Spread Plate technique on the
respective media, Pikovskaya media (PVK).
 Incubation(28±2°C) upto 5 days will show the Zone of Clearance (Halo zone).
1

10. METHODOLOGY (Cont.)
P u r i f i c a t i o n
Screening of Phosphate Solubilizing Bacteria for
P- Solubilization
Identification and Characterization through Morphological
and Biochemical Parameters
To isolate desire colony, there will be Purification of cultures and shall be
maintained in slants containing the respective medium.
2
The isolates obtained shall be morphologically and biochemically characterized as
per the procedures outlined by Cappuccino and Sherman, 1992

11. Identification
of PSB

12. METHODOLOGY (Cont.)
Assay of Bacterial Strains for Heavy Metal Resistance
Assay of Minimum Inhibitory Concentration
3
The Resistance of bacterial isolates will be determined by observing Minimum
Inhibitory Concentration (MIC) against Heavy Metals on agar plates.
Evaluation of Heavy Metal Resistant Bacteria for Plant
Growth Promoting activities.
4

13. METHODOLOGY (Cont.)
H e a v y M e t a l R e s i s t a n t b a c t e r i a w i l l b e s c re e n e d f o r d i f f e re n t P l a n t G ro w t h P ro m o t i n g
a c t i v i t i e s :

14. POSSIBLE OUTCOME
I will be expected to isolate Phosphate Solubilizing Bacteria.
I will be expected to isolate Heavy Metal Resistant Phosphate Solubilizing
Bacteria.
Evaluating the Heavy Metal Resistant Bacteria for Plant Growth Promoting
Bacteria activities.

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