RNAi-mediated knockdown of arginine kinase genes leads to high mortality and negatively affect reproduction and blood-feeding behavior of Culex pipiens pallens

Abstract

Background Arginine kinase (AK) is one of the crucial enzymes involved in energy metabolism in inverte-brates, and has been proposed as the target for RNA interference (RNAi)-based control of agricultural insect pests. While there is only one AK gene in most insects, two AK genes were identified in Culex pipiens pallens, the primary vector of lymphatic filariasis and epi-demic encephalitis. Methods The full-length cDNA sequences of CpAK1 and CpAK2 genes were obtained by reverse transcription PCR(RT-PCR) and rapid amplification of cDNA ends (RACE). The expression levels of CpAK1 and CpAK2 in different developmental stages and tissues were detected by reverse transcription quantitative PCR (RT-qPCR). The role of CpAK1 and CpAK2 in the reproduction and blood feeding behavior was analyzed using RNA interference (RNAi). Results Full-length cDNAs of CpAK1 and CpAK2 were isolated from Cx. pipiens pallens. Analysis of the expression pattern revealed that the mRNA level of CpAK1 was significantly higher than CpAK2 in all development stages and tissues examined, and the expressions of both CpAK1 and CpAK2 were upregulated in response to blood feeding. The co-knockdown of CpAK1 and CpAK2 mediated by RNAi led to high mortality (74.3%) of adult female mosquitoes and decreased hatchability (59.9%). Remarkably, the blood feeding rate and the engorgement rate of the female mosquitoes were negatively affected by co-injection of dsRNAs targeting CpAK1 and CpAK2. Conclusion CpAK1 and CpAK2 were detected in all developmental stages and tissues, but showed divergence in expression level. RNAi-mediated knockdown of AK genes leads to high mortality and negatively affect blood-feeding behavior of Cx. pipiens pallens, suggesting that AK could be used for the target of RNAi-based mosquito control in the future.