Abstract
BackgroundBacillus thuringiensis var. israelensis (Bti) is a natural larval mosquito pathogen producing pore-forming toxins targeting the midgut of Diptera larvae. It is used worldwide for mosquito control. Resistance mechanisms of an Aedes aegypti laboratory strain selected for 30 generations with field-collected leaf litter containing Bti toxins were investigated in larval midguts at two levels: 1. gene transcription using DNA microarray and RT-qPCR and 2. differential expression of brush border membrane proteins using DIGE (Differential In Gel Electrophoresis).ResultsSeveral Bti Cry toxin receptors including alkaline phosphatases and N-aminopeptidases and toxin-binding V-ATPases exhibited altered expression levels in the resistant strain. The under-expression of putative Bti-receptors is consistent with Bt-resistance mechanisms previously described in Lepidoptera. Four soluble metalloproteinases were found under-transcribed together with a drastic decrease of metalloproteinases activity in the resistant strain, suggesting a role in resistance by decreasing the amount of activated Cry toxins in the larval midgut.ConclusionsBy combining transcriptomic and proteomic approaches, we detected expression changes at nearly each step of the ingestion-to-infection process, providing a short list of genes and proteins potentially involved in Bti-resistance whose implication needs to be validated. Collectively, these results open the way to further functional analyses to better characterize Bti-resistance mechanisms in mosquitoes.
Highlights
Bacillus thuringiensis var. israelensis (Bti) is a natural larval mosquito pathogen producing pore-forming toxins targeting the midgut of Diptera larvae
The toxic crystal is dissolved in the alkaline pH of the midgut, protoxins are activated by digestive proteases to activated-toxins that bind to specific membrane receptors, form pores, disrupt the midgut epithelium, allowing spore penetration and bacterial proliferation in the host tissues [7,8]
Resistance levels to Bti toxins in the LiTOX strain After 30 generations of selection with leaf litter containing Bti, bioassays indicated that the LiTOX strain exhibited a moderate 3.5-fold resistance to commercial Bti mixture VectobacW WG compared to the susceptible strain at the larval stage (Table 1)
Summary
Bacillus thuringiensis var. israelensis (Bti) is a natural larval mosquito pathogen producing pore-forming toxins targeting the midgut of Diptera larvae. Israelensis (Bti) is a natural larval mosquito pathogen producing pore-forming toxins targeting the midgut of Diptera larvae. It is used worldwide for mosquito control. Israelensis (Bti) produces a mosquitocidal toxic crystal during sporulation and represents the best alternative to chemical insecticides for mosquito larval control due to its high potency and specificity [7]. The toxic crystal is dissolved in the alkaline pH of the midgut, protoxins are activated by digestive proteases to activated-toxins that bind to specific membrane receptors, form pores, disrupt the midgut epithelium, allowing spore penetration and bacterial proliferation in the host tissues [7,8]. The receptors for mosquitocidal Bti Cry toxins are similar to the lepidopteran-active Cry toxins which utilize Naminopeptidase, alkaline phosphatase and cadherin proteins as midgut receptors [9]
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