The mechanisms of resistance to the Bacillus thuringiensis crystal toxin Cry1Ac were explored in the Plutella xylostella SERD4 population, which shows polygenic resistance to Cry1Ac with cross-resistance to the pyrethroid deltamethrin that can be partially overcome by co-administering the esterase inhibitor piperonyl butoxide (PBO). The novelty of SERD4 cross-resistance was confirmed by testing other Cry1Ac-resistant populations for resistance to deltamethrin and for synergism of Cry1Ac toxicity by PBO. Esterase-mediated sequestration of Cry1Ac was tested using electrophoretic mobility shift assays, gut esterase activity inhibition assays and ligand blotting. Additionally, isozyme profiles and gut esterase activity were compared between various populations in an attempt to correlate the pattern and activity of esterases with cross-resistance. No correlation was found between esterase isozymes and Cry1Ac resistance and no evidence was found to indicate a direct interaction between CrylAc and esterases. Previous work showed that a maternal effect contributes to Cry1Ac resistance in SERD4. Therefore factors that are known to be maternally inherited in Lepidoptera and which have been implicated in Cry1Ac resistance were screened to investigate possible correlations with resistance. Various immune parameters were screened including both cell-free and haemocyte-mediated responses. No correlation was observed between immune status and Cry1Ac resistance. The composition of the gut microbiota was compared between SERD4 and a susceptible P. xylostella population and eliminated to assess any contribution to Cry1Ac toxicity. The microbiotas of resistant and susceptible larvae were identical. Furthermore, the gut microbiota of a Cry1Ac-susceptible strain that is adapted to artificial diet was not observed to affect Cry1Ac toxicity. This led to further work concerning a recent novel hypothesis that gut bacteria are obligately required for the toxicity of B. thuringiensis and Cry1Ac towards numerous lepidopterans including the tobacco hornworm, Manduca sexta. Techniques were developed to allow the rearing of aseptic M. sexta larvae. The toxicity of B. thuringiensis was not reduced in the absence of gut bacteria. However, B. thuringiensis was less toxic to larvae that had been recently exposed to antibiotics. Antibiotic exposure did not reduce the toxicity of Cry1Ac.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:506940 |
| Date | January 2009 |
| Creators | Johnston, Paul |
| Publisher | University of Sussex |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
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