Synopsis 17 April Final

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Evaluation of biocontrol potential of an indigenous bacterialisolate: Optimization and characterization studies

Synopsis Seminar

Sunita DeviS-2010-40-04

Department of MicrobiologyCollege of Basic Sciences

CSK Himachal Pradesh Krishi VishvavidyalayaPalampur-176062



INTRODUCTION

Among the greatest hazards in the crop production, diseasescaused by phytopathogens are one of the major threats tosustainable food production.

For combating phytopathogens, many successful measuresincluding chemical control have been developedThese chemicals have resulted in a variety of harmful andundesirable effects on man, wildlife and ecosystem.

Biological control seems to be a potent means of reducing thedamage caused by phytopathogens which can be achieved bymaking use of various types of microbes including bacteria,fungi and viruses.



W hy use biological control?

Biological control agents are ± Expensive ± Labor intensive ±

Host specificChemical pesticides are:

± cost-effective ± easy to apply ±

Broad spectrum



Contd.W ILL:

Chemical pesticides ± Implicated in ecological, environmental, and human health problems ± Require yearly treatments ± Broad spectrum

Toxic to both beneficial and pathogenic speciesBUT:

Biological control agents ± Non-toxic to human ± Not a water contaminant concern ± Once colonized may last for years ± Host specific

Only effect one or few species



Organic farming is becoming popular due to its eco-friendlynature and quality of the produce. The application of

buttermilk is one of the organic inputs being used in theprocess of organic farming to control some of the diseases.The microbial diversity of this organic input has been studiedin the dept. of Microbiology, where various types of microbes

possessing some of the biocontrol properties have beenisolated.

One of such microbes which is gram positive, spore formerand exhibiting fairly a good amount of biocontrol activityagainst some of the fungal pathogens has also been isolated.This particular indigenous microbe needs further systematicevaluation of its biocontrol potential with regards to the typeof metabolites involved in biocontrol activity.



OBJECTIVES:

To examine the antagonistic activity of indigenous bacterial isolate against differentphytopathogenic fungi and nematode

To optimize different parameters to augmentproduction of biocontrol agent(s)To purify and characterize the biocontrolcompounds produced by the aboveindigenous isolate



Technical programme of work



The following plant pathogenic bacterium and fungi will be usedin the present investigation:

Plant pathogenic bacterium:

Plant pathogenic fungi: Rhizoctonia solaniSclerotium rolfsii

Fusarium oxysporumColletorichum truncatum Alternaria brassica



A gainst phytopathogenic fungi

The bacterial isolate will be tested for its antagonistic activity against

five fungal pathogens, viz., Rhizoctonia solani , Sclerotium rolfsii,Fusarium oxysporum , colleotricium tructam and Alternaria brassicaby dual culture technique (Ganesan and Gnanamanickam, 1987).

Each fungal pathogen will be grown on a PD A plate till it covers thewhole surface of the agar. W ith the help of a sterile cork borer, a discof fungal growth will be taken and placed at the center of a fresh PD A

plate.24 hour old culture of bacterial isolate will be then streaked on one

side of the fungal disc.The plates will be kept for incubation at 30°C for 2-7 days.

Inhibition of growth of fungal pathogen will be then recorded withrespect to control plate, inoculated with the fungal pathogen only.



Against nematode pathogen

The culture filtrate and the whole cell culture of thebacterial isolate will be examined for their ability toimmobilize or kill root knot nematode, M eloidogyneincognita , under laboratory condition (Jayakumar et al .2002).



100 ml of 24 hour old bacterial suspension

25 ml will becentrifuged at 8000 rpm

for 10 minutes

Supernatant will befiltered

Filterate will be used forbioassay against root knot

nematode

Remaining broth culturewill be used as whole

cell culture for bioassay.



Bioassay

The egg masses of root knot nematode, M eloidogyne incognita will be made to hatch in to juveniles .Populationof juveniles present in water suspension will be counted under stereomicroscope and used for bioassay test

Three ml of the nematode suspension containingabout 50 juveniles will be taken in a petri plate to

which 10 ml of culture filtrate will be added andkept for 72 hours for incubation.

At 24 hours, 48 hours and 72 hours of incubation, observations for immobilized and killednematodes will be recorded in each plate.

The revival test will also be conducted by transferring the immobilized nematodes into

fresh water to confirm mortality in both the sets of plates

In another set of platescontaining nematode

juveniles, 10 ml of whole cell culture will be

added.

The platescontainingnematode

suspension andplain broth willserve as control



The following parameters will be optimize to augmentproduction of biocontrol agent(s) using Responsesurface methodology:

Carbon sourcesNitrogen sources

PH

AerationThe partial identification of biocontrol agent(s) will be

done using suitable solvent systems by TLC and HPLCbased detection methods.

MALDI/ LC/ MS/ GC-MS or NMR based study will bedone to elucidate the structure of biocontrol agent(s)

.



Facilities A vailable



Collaboration with Other

Department/Institute/University



Synopsis 17 April Final

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