The current decline of the European Honey Bee (Apis Mellifera) has been linked to the increasing use of neonicotinoid pesticides within agriculture. Whilst the toxicity of these pesticides to Apis has long been established, the possibility of low dosages inducing molecular stress has not yet been fully explored. Of particular interest is the action of these nicotine derivatives on the nicotinic acetylcholine receptor, and its association with the DNA methyltransferase family (Dnmts).
An experimental group of three hives were exposed to sugar water contaminated with a low concentration of imidacloprid (2µg/l). From these hives, 12 third instar larvae were selected. A corresponding number of larvae were also selected from three control hives, for a total of 12 samples. Using quantitative reverse transcription-PCR, known Dnmt transcripts were detected and amplified from these larval samples. Specially designed oligonucleotide primers were used containing gene specific sequences that linked to universal DNA sequences, ensuring that PCR amplification products were of predetermined sizes. Products of this amplification were resolved by capillary electrophoresis and detected by fluorescence spectrophotometry. Simultaneously, the transcriptomes of 3 larval samples each from the control and experimental groups were generated using SOLiD platform sequencing. The statistical package EdgeR was then utilised to identify differential candidates of known honey bee microRNA’s.
Statistical analysis utilising a one-way Analysis of Variance (ANOVA) found no significant differences in the expression levels of known Dnmt transcripts between control and experimental groups. However, comparisons of sequenced control and experimental transcriptomes identified a number of differential microRNA candidates, most notably miR-9/14
