Identification of potential biomarker genes for selecting varroa tolerant honey bees (Apis mellifera) and biochemical characterization of a differentially expressed carboxylesterase gene in response to mite infestation
Previously a large number of differentially expressed genes were identified by a DNA microarray analysis of two contrasting honey bee colonies for tolerance and susceptibility to varroa mite infestation. This study initially analyzed the expression patterns of ten of these genes in detail for a wide range of colonies with a range of phenotypes for susceptibility and tolerance to varroa mite infestation using real time qRT-PCR. Dark eyed stage 4 pupae with and without varroa infestation were sampled for the molecular analysis. The results showed that three out of the ten genes, AmCbE E4, AmApoD and AmCYP6A1 displayed relatively consistent differential expression patterns among the colonies and could be used as potential biomarkers for identifying varroa tolerant colony phenotypes. In general, these biomarker genes exhibited higher expression in tolerant colonies and lower expression in susceptible colonies with varroa mite infestation, compared to non-infested colonies. Tissue expression analysis showed AmCbE E4 was more differentially expressed in the head and AmApoD was differentially expressed in the abdomen, and AmCYP6A1 showed more differential expression in the thorax and abdomen among the honey bees differing in varroa tolerance and susceptibility. Expression of the three genes also responded to miticide treatments in the colonies. The miticide treatments (Apistan®, Apivar®, Thymovar®) could stimulate their expression in tolerant colonies, but not in susceptible colonies. In addition, the infection of deformed wing virus (DWV), another biotic stressor for honey bees primarily vectored by the mite, was also quantitatively evaluated by real time qRT-PCR in the varroa tolerant and susceptible honey bee colonies. The results showed that DWV infections were considerably increased in the susceptible colonies infested by varroa mites or treated with miticides (Apistan®, Apivar®, Thymovar®). AmCbE E4 encoding a putative esterase E4 was identified for its highly differential expression between the susceptible and tolerant bees in response to the mite infestation. Its biochemical function was analyzed by cloning the AmCbE E4 from the head of the dark eyed stage 4 pupae and heterologously expressing it in E. coli. The enzymatic assays revealed that AmCbE E4 could hydrolyze synthetic esterase substrates, α-naphthyl acetate, β-naphthyl acetate and para-nitrophenyl acetate, as well as carbaryl, a carbamate pesticide. This result suggests a defensive function of AmCbE E4 in protecting the varroa tolerant bees from the toxic stresses of carboxylester miticides and ester compounds possibly produced by the Varroa destructor parasitism.
Honey bee, Varroa destructor
Master of Science (M.Sc.)
Food and Bioproduct Sciences