RNA-Seq and molecular docking reveal multi-level pesticide resistance in the bed bug.
BACKGROUND
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.


RESULTS
We 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.


CONCLUSIONS
We 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.
Department of Entomology, The Ohio State University, Ohio Agricultural and Research Development Center, Wooster, OH 44691, USA.
Praveen
P
Mamidala
Asela J
AJ
Wijeratne
Saranga
S
Wijeratne
Karl
K
Kornacker
Babu
B
Sudhamalla
Loren J
LJ
Rivera-Vega
Andrew
A
Hoelmer
Tea
T
Meulia
Susan C
SC
Jones
Omprakash
O
Mittapalli
D000818:Animals; D001511:Bedbugs / Q000235:genetics*; D001665:Binding Sites; D020134:Catalytic Domain; D003198:Computer Simulation; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D030541:Databases, Genetic; D004351:Drug Resistance / Q000235:genetics*; D020224:Expressed Sequence Tags; D019476:Insect Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007306:Insecticides / Q000737:chemistry*; D008969:Molecular Sequence Data; D009154:Mutation; D017423:Sequence Analysis, RNA; D059467:Transcriptome
D019476:Insect Proteins; D007306:Insecticides; D003577:Cytochrome P-450 Enzyme System; C423312:flavonoid 6-hydroxylase