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Identification and characterisation of new aphid killing bacteria for use as biological pest control agents

Paliwal, D. (2017) Identification and characterisation of new aphid killing bacteria for use as biological pest control agents. PhD thesis, University of Reading

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Abstract/Summary

Insecticide resistance and on-going legislation changes on the use of insecticides are likely to reduce their availability for use in agriculture; hence there is an urgent need to develop effective biological controls against these plant pests. Aphids are major insect pests of the agricultural and horticultural sectors. Recent work screened a range of phylloplane-residing bacteria for their ability to kill aphids and used alternative insect targets to determine host specificity. Tests with five other aphid genera indicated the bacteria were also able to kill them. However, the bacteria were generally not effective against non-aphid species including Lepidoptera. I aimed to characterise potential aphid killing pathogens and investigate the killing mechanism. An artificial feeding system with a liquid diet was used to devise a high-throughput screening system to identify pathogenic bacteria against the Green Peach Aphid Myzus persicae (“wild type” insecticide susceptible clones plus insecticide resistant clones). Six bacterial strains were pathogenic to all insecticide susceptible and resistant clones although variation in susceptibility was observed. No single bacterial strain was identified that was consistently more toxic to insecticide resistance clones than susceptible clones, suggesting there was no penalty in resistant clones that makes such clones less fit to bacterial challenge. Pseudomonas poae, which was the most pathogenic to nearly all of aphid clones, was selected for further in-depth analyses. Plant colonisation assays showed that the bacterium could effectively grow and persist on three different plant species. Foliar spray of P. poae did not show any hypersensitive (HR) response and populations (log 5-6) remained stable over three weeks of infestation. Additionally, application of the bacterium to plants before aphid colonisation led to a 68 %, 57 %, 69 % reduction in aphid populations on pre-infested peppers, Arabidopsis and sugar beet plants, respectively. Olfactometer analysis showed that bacterial colonisation of leaves had a deterrent effect on aphids that was not evident for leaves or bacteria alone. Genome analysis of the bacterium revealed three different insecticidal toxins, stress response genes and other pathogenicity-related effector proteins which reflect potential toxicity towards aphids. RNA-Seq was used to examine changes in aphid and bacterial gene expression after 38 h of infection. The altered transcript profiles of the aphid revealed 193 differentially expressed genes and limited gene expression of lysosomal and detoxification genes. 1325 genes were differentially expressed in bacteria, which mainly includes iron acquisition and stress response genes, and putative toxins. Single and combinational deletion, and complementation, of different toxins was conducted. In vitro killing analysis indicated all toxins contributed to aphid killing, with a particularly strong effect seen for one, AprX. Together, these data are being used to understand the molecular basis of aphid mortality to bacterial infection with the aim of utilising them as effective biocontrol agents.

Item Type:Thesis (PhD)
Thesis Supervisor:Jackson, R. and Bass, C.
Thesis/Report Department:School of Biological Sciences
Identification Number/DOI:
Divisions:Faculty of Life Sciences > School of Biological Sciences
ID Code:78229

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