presentation on bt _final

7/31/2019 Presentation on Bt _final

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Commercial Production of Biopesticides with reference

to

Bacillus Thurigiensis


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What are biopesticide?

Biopesticides are certain types of pesticides derived from

such natural materials as animals, plants, bacteria, and

certain minerals. For example, canola oil and baking soda

have pesticidal applications and are consideredbiopesticides.


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Biopesticides fall into three majorclasses

1. Microbial pesticide - consist of a microorganism (e.g., a

bacterium, fungus, virus or protozoan's .

2. Plant-Incorporated-Protectants (PIPs)-pesticidal

substances that plants produce from genetic material

that has been added to the plant.

3. Biochemical pesticides-naturally occurring substances

that control pests by non-toxic mechanisms.


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What are the advantages of usingbiopesticides?

Biopesticides are usually inherently less toxic thanconventional pesticides.

Biopesticides generally affect only the target pestand closely related organisms, in contrast to broadspectrum, conventional pesticides that may affectorganisms as different as birds, insects, andmammals.

Biopesticides often are effective in very smallquantities and often decompose quickly, therebyresulting in lower exposures and largely avoiding the

pollution problems caused by conventional pesticides.


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BACILLUS THURIGIENSIS ASBIOPESTICIDE

The most widely used microbial pesticides are subspecies

and strains ofBacillus thuringiensis, or Bt. Each strain ofthis bacterium produces a different mix of proteins, and

specifically kills one or a few related species of insect

larvae.

some Bt's control moth larvae found on plants, other Bt's

are specific for larvae of flies and mosquitoes. The targetinsect species are determined by whether the particular Bt

produces a protein that can bind to a larval gut receptor,

thereby causing the insect larvae to starve.


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WHAT IS NATURAL BACILLUSTHURIGIENSIS?Gram Positive

Spore Forming Bacteria

During speculation produce

protein crystal (cry) called -endotoxins, that

have insecticidal action

In most strains ofB.thuringiensis, the crygenes arelocated on a plasmid (in other

words, cryis not achromosomal gene in most

strains).
http://en.wikipedia.org/wiki/%CE%94-endotoxinshttp://en.wikipedia.org/wiki/%CE%94-endotoxinshttp://en.wikipedia.org/wiki/%CE%94-endotoxinshttp://en.wikipedia.org/wiki/Insecticidalhttp://en.wikipedia.org/wiki/Plasmidhttp://en.wikipedia.org/wiki/Plasmidhttp://en.wikipedia.org/wiki/Insecticidalhttp://en.wikipedia.org/wiki/%CE%94-endotoxinshttp://en.wikipedia.org/wiki/%CE%94-endotoxinshttp://en.wikipedia.org/wiki/%CE%94-endotoxins


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Cry toxins have specific activities against insect species of

the orders Lepidoptera (moths and butterflies), Diptera (fliesand

mosquitoes), Coleoptera (beetles), Hymenoptera (wasps, be

es, antsand sawflies) and nematodes.

Bthas to be eaten to cause mortality. The Bttoxindissolve in the high pH insect gut and become active. The

toxins then attack the gut cells of the insect, punching

holes in the lining. The Btspores spills out of the gut and

germinate in the insect causing death within a couple days.

Bacillus thurigiensis toxin
http://en.wikipedia.org/wiki/Lepidopterahttp://en.wikipedia.org/wiki/Dipterahttp://en.wikipedia.org/wiki/Coleopterahttp://en.wikipedia.org/wiki/Hymenopterahttp://en.wikipedia.org/wiki/Wasphttp://en.wikipedia.org/wiki/Beehttp://en.wikipedia.org/wiki/Beehttp://en.wikipedia.org/wiki/Anthttp://en.wikipedia.org/wiki/Sawflyhttp://en.wikipedia.org/wiki/Nematodehttp://en.wikipedia.org/wiki/Nematodehttp://en.wikipedia.org/wiki/Sawflyhttp://en.wikipedia.org/wiki/Anthttp://en.wikipedia.org/wiki/Beehttp://en.wikipedia.org/wiki/Beehttp://en.wikipedia.org/wiki/Wasphttp://en.wikipedia.org/wiki/Hymenopterahttp://en.wikipedia.org/wiki/Coleopterahttp://en.wikipedia.org/wiki/Dipterahttp://en.wikipedia.org/wiki/Lepidoptera


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How BT Works?


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1. Insect eats Btcrystals andspores.

2. The toxin binds to specific

receptors in the gut and the

insects stops eating.

3. The crystals cause the gut

wall to break down, allowing

spores and normal gut bacteria

to enter the body.

4. The insect dies as spores and

gut bacteria proliferate in the

body.


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Application of BT Bt is used in agriculture as a liquid applied through overhead

irrigation systems or in a granular form for control of Europeancorn borer. The treatments funnel down the corn whorl to where

the feeding larvae occur.

Bt useful in applications where pesticide drift onto Gardens islikely to occur, such as treating trees and shrubs.

To control mosquito larvae, formulations containing

the israelensisstrain are placed into the standing water of

mosquito breeding sites.

Use of Bt (israelensis) for control of fungus gnat larvae involvesdrenching the soil.


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Insects Controlled by Bt

Cabbage worm (cabbage looper, imported cabbageworm, diamondback moth,etc.).

Tomato and tobacco hornworm

Vegetable insects

European corn borer (granular formulations have given good control of firstgeneration corn borers).

Alfalfa caterpillar, alfalfa webworm

Field and forage crop insects

Leafroller.

Achemon sphinx.

Fruit crop insects

Tent caterpillar.

Pine budworm

Western spruce budworm.

Fruit crop insects


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Israelensisstrains (Vectobac, Mosquito Dunks, Gnatrol,Bactimos, etc.)

Insects Controlled by Bt continue..

Mosquito.

Black fly.

Fungus gnat.

San diego/tenebrionisstrains (Trident, M-One, M-Trak, Foil,Novodor, etc.)

Colorado potato beetle.

Elm leaf beetle.

Cottonwood leaf beetle.


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Disadvantages

Bt is susceptible to degradation by sunlight

The highly specific activity of Bt insecticides might limit their

use on Crops where problems with several pests occur,

including nonsusceptible insects (aphids, grasshoppers, etc.).

Since Bt does not kill rapidly, users may incorrectly assume

that it is ineffective a day or two after treatment.

Bt-based products tend to have a shorter shelf life than other

insecticides.


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Advantages

The specific activity of Bt generally is considered highly

beneficial Perhaps the major advantage is that Bt isessentially nontoxic to people, pets and wildlife.

This high margin of safety recommends its use on food

Crops or in other sensitive sites where pesticide use can

cause adverse effects.

Perhaps the major advantage is that Bt is essentially

nontoxic to people, pets and wildlife.


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Since 1996, a wide range of crop plants have been geneticallyengineered to contain the delta-endotoxin gene from Bacillusthuringiensis.

These "Bt crops" are now available commercially in the USA. They

include "Bt corn", "Bt potato", "Bt cotton" and "Bt soybean". Such plants

have been genetically engineered to express part of the active Cry toxin in

their tissues, so they kill insects that feed on the crops.

In some respects, this is an important technological and practical

development, because it ensures that only those insects that attack the

crop will be exposed to Bt toxins - there is no risk to other types of insect.

However, there is also a "downside", because the target insects are

perpetually exposed to toxins and this creates a very strong selection

pressure for the development of resistance to the toxins. Various crop-

management strategies are being developed to try to minimise this risk.

Plants genetically engineered with the Btgene


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.

Biopesticide Production from Bacillusthuringiensis: An Environmentally

Friendly AlternativeNinfa M. Rosas Garca*

Received: October 29, 2008; Accepted: November 26, 2008; Revised:November 28, 2008

Laboratorio de Biotecnologa Ambiental. Centro de Biotecnologa

Genmica-IPN. Blvd. del Maestro s/n. Reynosa,Tamp. CP 88710 Mxico


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Introduction

The bacterium Bacillus thuringiensis was discovered by

Shigetane Ishiwata in 1901.

The bacterium was isolated from diseased larvae of

Anagasta kuehniella, and this finding led to theestablishmentofB. thuringiensis as microbial insecticide.

The first record of its application to control insects was in

Hungary at the end of 1920, and in Yugoslavia at the

beginning of 1930s, it was applied to control the European

corn borer.

During the following two decades, several field tests were

conducted to evaluate its effectiveness against lepidopterans,

both in Europe and in the United States.


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results favored the development of formulations against on

this pathogen. Subsequently, the first commercial product was

produced in 1938 by Libec I France.

serious environmental and health issues began to be

recognized by the presence of chemical residues in food,

water, and air during 1950.

To counteract this contamination, attention and efforts weredirected to the use of biological control agents including insect

pathogens.

As an entomopathogenic organism, B.thuringiensis fulfills allthese requirements.


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MODE OF ACTION

The C-terminal extension found in the long protoxins is

necessary for toxicity and is believed to play a role in theformation of the crystal within the bacterium.

During proteolytic activation, peptides from the N terminus and C

terminus are cleaved from the full protein.

During proteolytic activation, peptides from the N terminus and

C terminus are cleaved from the full protein.

Activated toxin binds to receptors located on the apical

microvillus membranes of epithelial midgut cells.


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For Cry1A toxins, at least four different binding sites have been

described in different lepidopteran insects:

1. a cadherin-like protein(CADR),2. a glycosylphosphatidyl-inositol (GPI)-anchored

aminopeptidase-N (APN),

3. a GPI-anchored alkaline

4. phosphatase (ALP) and a 270 kDa glycoconjugate .

After binding, toxin adopts a conformation allowing its insertion

into the cell membrane. Subsequently, oligomerization occurs,

and this oligomer forms a ion channel induced by an increase

in cationic permeability within the functional receptorscontained on the brush borders membranes

This causing disruption of membrane transport and cell lysis,

and leading to insect death

Photograph sho s lepidopteran lar ae s sceptible to B th ringiensis to ic


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Photograph shows lepidopteran larvae susceptible to B.thuringiensis toxicactivity.

A) Nine day-old, healthy larvae,

notexposed to B. thuringiensis

B) Nine-day old larvae exposed to

sublethal concentration ofB. thuringiensis.Larvae exhibit a smallersize due to theirpoor feeding.

C) Larvae exposed to lethalconcentrationofB. thuringiensis.


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BACILLUS THURINGIENSIS-BASEFORMULATIONS

The active ingredient in commercial formulations is the

sporecrystal complex.

It is more effective to use and cheaper to obtain than the

crystals alone, which are frequently used in experimental tests.

A great variety of ingredients have been employed to prepare

formulations, including liquid or solid carriers, surfactants,

coadjuvants, fluidity agents, adherents, dispersants,stabilizers, moisturizers, attractants, and protective agents

among others


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An interesting and recent example of these kind of inert

ingredients is the superabsorbent starch graft copolymer,

which combined with a B. thuringiensisstrain among manyother pesticides constitutes a novel formulation that could be

applied in an agricultural environment.

The biological activity of one particular bioinsecticide was

enhanced when the antibiotic zwittermycin was added. Thiscombination was also successful in pest control.


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Bacillus thuringiensis-based products are classifiedaccordingto their formulation.

first-generation products- All products containing a blend ofspores and crystals from a native strain.

Second generation products -based on spores and crystals

from a

B. thuringiensis strain bearing artificially introduced genescoding for delta-endotoxins from several strains, in order to

increase the activity spectrum against other insect pests.

Third generation products- Formulations containing deadrecombinant Pseudomonas fluorescens cells transformed withgenes coding for deltaendotoxins


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Types of Bacillus thuringiensisFormulations andTheir Applications for

Insect Pest Control

Form

ulatio

n Emulsions Encapsulations

Wettable powders

Granules

Powders

Briquettes

Appl

ication Agriculture and forestry

Agriculture and forestry

Gardens and agriculture

Agriculture and forestry

Forestry

Aquatic systems


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Most Common Commercial Bacillusthuringiensis-based Bioinsecticides

Company CommercialName

ActiveIngredient

Target Pest


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CHIMERIC CRYSTAL PROTEINS

In recent years, hybrid delta-endotoxins have arisen as

proteins with potential for enhanced toxic activity orimproved properties.

Recent advances in molecular methodologies have allowed

gene fusions and chimeric protein construction.

This construction can include alteration of amino acid

sequences, fusion of portions of two or more proteins

together into a single recombinant protein, or alteration of

the genetic sequences encoding for proteins withcommercial application.


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There is a great amount of scientific research on the

bacterium B. thuringiensis, involving aspects ranging fromitsmolecular biology to its activity in a bioinsecticide.

The development of formulations with biodegradable

ingredients is a favored approach for the reduction of chemicalinsecticide use, which can threaten the environment.

CURRENT & FUTURE DEVELOPMENTS


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REFERENCES[1] Aizawa K. Shigetane Ishiwata: His discovery of sottokin (Bacillus thuringiensis) in1901 and subsequent investigations in Japan. Proceedings of a CentennialSymposium CommemoratingIshiwatas Discovery of Bacillus thuringiensis. Japan2001.[2] Lord JC. From Metchnikoff to Monsanto and beyond: The path of microbial

control. J Invertebr Pathol 2005; 89 (1): 19-29.

[3] Cern JA. Productos comerciales nativos y recombinantes a base de Bacillusthuringiensis. Bioinsecticidas: Fundamentos yaplicaciones de Bacillus thuringiensis

en el control integrado deplagas. In: Caballero P, Ferr J. Eds, Phytoma-Espaa2001; 153-168.

[4] Aronson A, Beckman W, Dunn P. Bacillus thuringiensis andrelated insectpathogens. Microbiol Rev 1986; 50 (1): 1-24.

[5] Nester EW, Thomashow LS, Metz M, Gordon M. 100 years ofBacillusthuringiensis: A critical scientific assessment. AmericanAcademy of Microbiology.

Washington DC 2002.[6] King E. Control biolgico de insectos y caros plaga. Avances recientes en labiotecnologa en Bacillus thuringiensis. In: Galn-Wong LJ, Rodrguez-Padilla C,Luna-Olvera HA, Eds. Universidad Autnoma de Nuevo Len 1996; 13-19.

[7] Margalith Y, Ben-Dov E. Biological control by Bacillus thuringiensis subp.israelensis. Insect pest management, techniquesfor environmental protection. In:

Rechcigl JE, Rechcigl NA, Eds.Lewis Publishers 2000; 243-301.[8] Aizawa K. Selection and utilization ofBacillus thuringiensisstrains for microbial


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