The Function Odorant Receptors In Culex Pipiens Quinquefasciatus

Culex pipiens quinquefasciatus is widely distributed in tropical and subtropical regions of the world, and also is the most common mosquito species in southern China. It is not only an important vector of lymphatic filariasis and epidemic encephalitis B, but a potential vector of West Nile virus and western equine encephalitis in our country as well. In nature, the mosquito complete the reproductive cycle by biting birds, mammals and humans. In order to find hosts for a blood meal, mosquito need sense host odors in the olfactory system with odorant receptor (OR), transfer olfactory signal to central nervous system and locate a host to complete a blood meal. Mosquito behavior in response to host ordors, such as being attracted and being repelled, are decided by nerve impulses caused by the specific binding of odorant receptors and host odors. Since the odorant receptor is a key factor of mosquito olfaction, better understanding its functions will help develop new strategies for mosquito control by blocking or interfering chemical communication between mosquitoes and hosts, and also may provide insight into the development of new repellents or attractants.The main research results of this study are as follows:1. Expression of odorant receptors genes in different developmental stages and after blooding feeding in Cx. pipiens quinquefasciatusThe expression of eleven odorant receptor genes (CquiOR5, CquiOR15, CquiOR22, CquiOR20, CquiOR54, CquiOR71, CquiOR78, CquiOR85, CquiOR104, CquiOR117 and CquiOR118) in different stages and after blood feeding were investigated in Cx.pipiens quinquefasciatus mosquitoes by real-time PCR. CquiOR117 showed the highest expression in Cx. pipiens quinquefasciatus various developmental stages as well as in the adult head; CquiOR54 showed the lowest expression in stages from egg to pupa and one week old male mosquitoes after emergence; CquiOR78 was specifically expressed in the male mosquito; the expressions of CquiOR118, CquiOR15, CquiOR71 and CquiOR54 were significantly different in blood meal and non blood meal mosquitoes.2. Identification of CquiOR117 in Cx. pipiens quinquefasciatusA successful microinjection method was established. Briefly,3-5 days old mosquitoes after eclosion were selected, starved for 4h, frozen at-30℃ for 4 min, placed in the ice box, then injected 0.7-0.8μL dsRNA solution into the side of the mosquito thorax using a microinjection system. The detection of CquiOR117 expression levels of mosquitoes after being injectied CquiOR117dsRNA by real-time PCR showed that the expressions of CquiOR117 decreased by 19.96%,71.51%,72.99%,70.30%,86.36% and 85.41%1-6 days post injection, respectively. It is found that, mosquitoes had less successful blood meal after CquiOR117 was interfered, however, there was correlation between the gene expression levels and blood meal decrease。3. Cloning and comparison of odorant receptor (OR) genes in Cx. pipiens complex mosquitoesTotal 19 full-length OR genes were cloned in the head of Cx pipiens quinquefasciatus, Culex pipiens molestus and Culex pipiens pallens, and sequences were visually checked and aligned using DNAstar. The results showed that all the sequences we cloned had mutation sites, and the CquiORll had the largest number of mutation sites. In the Neighbor-Joining tree CquiOR01, CquiOR39, CquiOR06, CquiOR13, CquiOR83, CquiOR66 in Cx.pipiens molestus form a segregated cluster, and CquiOR42, CquiOR48, CquiOR65 and CquiOR14 in Cx.pipiens pallens also form a segregated cluster, and only CquiOR04 in Cx.pipiens quinquefasciatus form a segregated cluster. The structural analysis of the encoded protein was done through the TMHMM Server v.2.0. The results showed that Cx. pipiens molestus had mutation sites in the transmembrane helices regions of CquiOR04、CquiOR05 and CquiOR06, besides Cx. pipiens pallens and Cx.pipiens quinquefasciatus both had mutation sites in the transmembrane helices regions of CquiOR42 and CquiOR65.