Abstract
BackgroundBed bugs (Cimex lectularius) are hematophagous nocturnal parasites of humans that have attained high impact status due to their worldwide resurgence. The sudden and rampant resurgence of C. lectularius has been attributed to numerous factors including frequent international travel, narrower pest management practices, and insecticide resistance.ResultsWe performed a next-generation RNA sequencing (RNA-Seq) experiment to find differentially expressed genes between pesticide-resistant (PR) and pesticide-susceptible (PS) strains of C. lectularius. A reference transcriptome database of 51,492 expressed sequence tags (ESTs) was created by combining the databases derived from de novo assembled mRNA-Seq tags (30,404 ESTs) and our previous 454 pyrosequenced database (21,088 ESTs). The two-way GLMseq analysis revealed ~15,000 highly significant differentially expressed ESTs between the PR and PS strains. Among the top 5,000 differentially expressed ESTs, 109 putative defense genes (cuticular proteins, cytochrome P450s, antioxidant genes, ABC transporters, glutathione S-transferases, carboxylesterases and acetyl cholinesterase) involved in penetration resistance and metabolic resistance were identified. Tissue and development-specific expression of P450 CYP3 clan members showed high mRNA levels in the cuticle, Malpighian tubules, and midgut; and in early instar nymphs, respectively. Lastly, molecular modeling and docking of a candidate cytochrome P450 (CYP397A1V2) revealed the flexibility of the deduced protein to metabolize a broad range of insecticide substrates including DDT, deltamethrin, permethrin, and imidacloprid.ConclusionsWe developed significant molecular resources for C. lectularius putatively involved in metabolic resistance as well as those participating in other modes of insecticide resistance. RNA-Seq profiles of PR strains combined with tissue-specific profiles and molecular docking revealed multi-level insecticide resistance in C. lectularius. Future research that is targeted towards RNA interference (RNAi) on the identified metabolic targets such as cytochrome P450s and cuticular proteins could lay the foundation for a better understanding of the genetic basis of insecticide resistance in C. lectularius.
Highlights
Bed bugs (Cimex lectularius) are hematophagous nocturnal parasites of humans that have attained high impact status due to their worldwide resurgence
Other possible reasons for read variation include different percentages of poly A+ in different samples, differential efficiency of conversion of RNA to cDNA, size or GC composition variation among libraries. All these factors may contribute to slight variations in the estimation of the library concentration that is applied onto the flow-cell, resulting in differences in cluster densities
Detoxification enrichment We found enriched ATP binding and glutathione peroxidase-associated Gene Ontology (GO) terms in the up-regulated cluster of differentially expressed expressed sequence tags (ESTs) suggesting the involvement of other detoxification genes such as ATP binding cassette (ABC) transporters and quenchers of reactive oxygen species (ROS) (Figure 4A), which is in agreement with other insect studies [34,35]
Summary
Bed bugs (Cimex lectularius) are hematophagous nocturnal parasites of humans that have attained high impact status due to their worldwide resurgence. The sudden and rampant resurgence of C. lectularius has been attributed to numerous factors including frequent international travel, narrower pest management practices, and insecticide resistance. Cimex lectularius (the bed bug), a hematophagous ectoparasite of humans, is spreading at alarming rates across the globe [1,2,3,4,5,6]. The bites of these nocturnal blood feeders result in cutaneous manifestations, urticarial reactions, and occasionally anaphylaxis [7]. The lack of effective pest management tools for this blood-feeding insect has led to its successful establishment around the globe [8]
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