Bacillus thuringiensis bt toxin

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Bacillus thuringiensis (bt) toxin

• BS322 Environmental Microbiology

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Bacillus thuringiensis (Bt) toxin action, use
and genetic engineering

• References
• Not many suitable ones in MH, but some
  information can be found on the genetic
  manipulation of plants at 631.51 onwards.
• Dines, A.J. and Mottley, J. (1999). Seeds of
  Change Applications of Biotechnology to
  Agriculture. In Springham, D. (ed) Biotechnology
  The Science and the Business (in MH library - ref
  only). Can also can be found at
  http//homepages.uel.ac.uk/J.Mottley/Newchapt.html
  
• Alternatively try the WWW (NBIAP Monthly News
  Reports are useful - to be found at
  www.nbiap.vt.edu where there is also a searchable
  database).

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Bacillus thuringiensis

• Main biological insect control agent
• Gm ve sporulating bacterium
• Produces crystals of endotoxin in sporulating
  cells
• Specific for certain insect groups, particularly
  Lepidoptera, Diptera and Coleoptera

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Types of Bt toxin
Pseudomonas fluorescens
MVP, M-TRAK
Mycogen Corp.
?
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Factors affecting effectiveness of Bt toxin

• Amount of toxin ingestion
• Processing within the insect gut
• Activity at site of action

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Toxin ingestion

• Effect of larval size
• Early stages more susceptible
• Feeding behaviour
• Indiscriminate feeders eat all parts of plant
• Discriminate feeders - only certain parts of
  plant used for food either all the time or at
  certain stages of life cycle

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Gut processing of toxin

• Requires two processes that may differ in
  different insects and produce different
  responses
• Toxin solubilisation
• Occurs in highly alkaline (pH8-9.5) juices of
  target insects mid gut
• Toxin activation
• By action of proteolytic enzymes

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Activity at site of action

• Toxin binds to specific receptors on surface of
  insect mid-gut epithelial cells, specifically to
  the p-lipids in the brush border membranes of the
  columnar cells
• A pore forms through the cell membrane resulting
  in loss of insects ability to osmoregulate
• Insect dies due to massive water uptake and cell
  bursting

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Strategies for increasing the effectiveness of Bt
insecticides

• Two main approaches
• Genetic modification of Bt plasmids
• Genetic modification of plants to produce Bt

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Genetic modification of Bt plasmids

• Depends on plasmid transfer by conjugation
• Bt strain that shows low activity against a wide
  spectrum of insects chosen as recipient. Some of
  its plasmids are cured by growth at high temp.
• Bt strain with high potency but with maybe narrow
  spectrum of activity chosen as donor
• After conjugation, the strains potency against a
  wide spectrum of insects is increased

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Genetic modification of plants with Bt toxin genes

• Plants are genetically modified with Bt toxin
  genes so that they produce Bt toxin in their
  tissues.
• Insects are then killed when they feed on the
  plant tissues
• This has been successful and widely used
  commercially in most important crops, eg. maize,
  tobacco, tomatoes and cotton

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Reduced use of synthetic chemical insecticides

• Should lead to cheaper products, a less polluted
  environment and safer food
• Conventional insecticides may still be needed,
  especially with large populations
• With small pest populations, farmers may choose
  to use cheaper synthetic alternatives

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Table 2. Comparison of strategies for delivery of
Bt toxin either as conventional sprays or via GM
plants
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POTENTIAL RISKS OF UTILIZING BT PRODUCING CROPS

• Toxicity to the consumer
• Toxicity to non-target insect species
• Build-up of insect resistance to BT

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Toxicity to the consumer

• Consumers in industrialised countries have been
  exposed to residues from BT sprays in their food
  for many years with no obvious toxicity problems,
  but
• Long-term toxicity of large dosages needs
  determining
• Toxin chemical structure may differ when produced
  in plants, eg. active form

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Toxicity to non-target insect species

• Although only insects that feed on BT-producing
  plants are targeted, useful insects may also be
  harmed if the spectrum of activity of the toxins
  is altered. For example, honey bees and (Monarch)
  butterflies when they feed on the nectar or
  pollen of Bt crops
• Only signs of this are in lab experiments under
  highly unnatural conditions with highly abnormal
  doses

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Development of insect resistance to Bt toxin

• Few reports of resistance development in over 40
  years of use and these done in lab
• Contrasts with use of synthetic pesticides in lab
  and field

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Possible causes of insect resistance to Bt toxin

• Could result from
• Reduced solubilization in the gut
• Changes in the gut protease composition/activity
• Decreased sensitivity of the gut receptor sites
• But would these reduce overall fitness of
  resistant insects?

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Build-up of insect resistance to BT sprays

• Little information available on the long-term
  effects of constantly exposing insect populations
  to lethal doses of BT sprays
• Buildup of resistance more likely from marginally
  susceptible insects than extremely susceptible
  ones
• If so, Bt may become obsolete as the most
  important biological pest control agent

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Build-up of insect resistance to GM Bt crops

• Selection pressure through increased use of
  single and active Bt toxin strains in GM Bt
  plants on a very wide scale compared to more
  limited use and multiple strains in sprays
• Unlike with synthetic pesticides, different toxin
  strains have slightly different modes of action
  and often insects resistant to one are not
  resistant to another so need multigenic mutations
  to create resistance

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Strategies for reducing insect resistance in Bt
crops

• All based on providing a non-Bt environment so
  that susceptible insects can survive and multiply
• Rotations
• Bt crops rotated with non-Bt crops to reduce
  build-up of resistant insects

2001
2002
2003
Non-GM
GM
Non-GM
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• Refugia
• Whole fields of non-Bt crops placed alongside
  fields of Bt crops

GM
Non-GM
GM
Non-GM
GM
Non-GM
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• Mosaics
• Bt and non-Bt crops grown in the same field

GM Non-Gm Gm Non-GM GM Non-GM Gm Non-GM GM Non-G
M GM Non-GM Gm Non-GM Gm Non-GM GM Non-GM GM
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Conclusions

• Bt toxin delivery can be achieved in TWO ways
  spray or through the plant food of insects
• Both methods can be improved by GM
• Both methods have their advantages and
  disadvantages
• Use of Bt plants definitely reduces use of
  dangerous synthetic pesticides but may not
  eliminate it altogether
• A risk from increased use may be the development
  of insect resistance (but not yet proven)
• Agricultural strategies can be used to reduce the
  build-up of resistance