Transcriptomic analysis to elucidate the response of honeybees (Hymenoptera: Apidae) to amitraz treatment

Abstract

Amitraz is an acaricide that is widely used in apiculture. Several studies have reported that in honeybees (Apis mellifera Linnaeus; Hymenoptera: Apidae), amitraz affects learning, memory, behavior, immunity, and various other physiological processes. Despite this, few studies have explored the molecular mechanisms underlying the action of amitraz on honeybees. Here, we investigated the transcriptome of honeybees after exposure to 9.4 mg/L amitraz for 10 d, a subchronic dose. Overall, 279 differentially expressed genes (DEGs) were identified (237 upregulated, 42 downregulated). Several, including Pla2, LOC725381, LOC413324, LOC724386, LOC100577456, LOC551785, and P4504c3, were validated by quantitative PCR. According to gene ontology, DEGs were mainly involved in metabolism, biosynthesis, and translation. Kyoto Encyclopedia of Genes and Genomes pathway analyses revealed that amitraz treatment affected the relaxin signaling pathway, platelet activation, and protein digestion and absorption.

Fig 1. Survival of forager bees subjected to chronic exposure to field-realistic concentrations of amitraz after 10 d.

Fig 2. Average number of genes specifically expressed in amitraz and control libraries.

Shown as the number of genes expressed in each class.

Fig 3. Volcano plot of differentially expressed genes in honeybees exposed to 9.4 mg/L of amitraz for 10 days.

Genes with an adjusted P value of <0.05 (FDR correction method) were considered to be differentially expressed. Red: upregulated genes in amitraz-treated bees; green: downregulated genes in amitraz-treated bees; blue: no significant difference.

Fig 4. Gene ontology enrichment analysis of differentially expressed genes (DEGs) in honeybees exposed to 9.4mg/L amitraz for 10 days.

Green bars: DEGs enriched for biological process; orange bars: DEGs enriched for cellular components; purple bars: DEGs enriched for molecular function. * indicates that GO terms were significantly enriched by DEGs (corrected P values of <0.05, FDR correction method).

Fig 5. Real-time quantitative PCR and RNA-seq analysis of LOC725381, CYP4C3, LOC41332, Pla2, LOC724386, LOC100577456, and LOC551385 genes expression multiple in honeybees exposed to 9.4 mg/L of amitraz for 10 days.

Reference gene were RPS5 and β-actin. Values represent means ± SEM. * indicates a significant difference in comparison with controls (P < 0.05) and ** indicates a statistically significant difference in comparison with controls (P < 0.01).