Cuticular proteins (CPs) are implicated in insecticide resistance in mosquito populations. Here, we investigated the role of cuticular genes in regulation of insecticide resistance in Culex pipiens pallens. We identified two CpCPRs (CpCPR63 and CpCPR47) that exhibited higher transcript levels in pyrethroid-resistant strains than in susceptible strains. Mosquito mortality was increased after knockdown of CpCPR genes by dsRNA injection. The RNA interference experiment suggested an interaction between CpCPR63 and CpCPR47, as silencing of one gene resulted in decreased expression of the other. These findings revealed that CpCPRs may regulate pyrethroid resistance and could be used as a potential genetic marker to monitor pyrethroid resistance in mosquitoes.
Piwi-interacting RNAs (piRNAs) are a novel class of noncoding single-strand RNAs. They play an important role in the germ cell maintenance, brain development, epigenetic regulation of cancer, and antiviral function. However, little is known about the relationship between the piRNAs and insecticide resistance in mosquitoes. In this study, we reported that piRNA-3878 was related with pyrethroid resistance in Culex pipiens pallens. The expression level of piRNA-3878 was lower in both laboratory and field-collected deltamethrin-resistant (DR) strains. Overexpression of piRNA-3878 increased the susceptibility of the DR strain, while inhibiting the expression of piRNA-3878 in DS strain made the mosquitoes more resistant to deltamethrin. Furthermore, we identified that CpCYP307B1 was the target of piRNA-3878. The mosquito mortality rate was increased after downregulating the expression of CpCYP307B1. These findings revealed that piRNA-3878 could target CpCYP307B1 to regulate pyrethroid resistance.
Piwi-interacting RNAs (piRNAs) are a newly identified class of small noncoding RNAs. They are associated with chromatin organization, messenger RNA stability, and genome structure. Although the overexpression of piRNA-3312 in deltamethrin-susceptible (DS) strain of Culex pipiens pallens (L.) was observed in our previous large-scale transcriptome data, the roles of piRNA in insecticide resistance have not been clearly defined. The aim of the present study was to investigate how piRNA-3312 is involved in insecticide resistance. The lower expression level of piRNA-3312 in deltamethrin-resistant (DR) strain of Cx. pipiens pallens was confirmed by quantitative real time polymerase chain reaction (qRT-PCR). Overexpression of piRNA-3312 in the DR strain made the mosquitoes more sensitive to deltamethrin, whereas inhibiting the expression of piRNA-3312 in the DS strain made the mosquitoes more resistant to deltamethrin. Piwi-interacting RNA-3312 was also found to bind 3' UTR (Untranslated Regions) of gut esterase 1 gene and could induce its degradation. In addition, knockdown of gut esterase 1 gene increased the sensitivity of DR strain to deltamethrin. In conclusion, we found that piRNA-3312 targeted the gut esterase 1 gene to negatively regulate the insecticide resistance. This finding facilitates the understanding of various functions of piRNAs and their association with insecticide resistance.
Carbonic anhydrases (CAs) are metabolic enzymes that regulate the physiological equilibrium in a variety of organisms. In this study, we investigated the function of CA II in Culex pipiens pallens using real-time quantitative polymerase chain reaction and double-stranded RNA injection. CA II transcripts were more abundant in the deltamethrin-susceptible strain than the deltamethrin-resistant strain. The activities of metabolic enzymes increased when the CA II expression was silenced. These findings suggest CA II regulates deltamethrin resistance by altering metabolic enzyme activity, and could serve as a potential genetic marker for monitoring deltamethrin resistance in mosquitoes.
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