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West Nile virus is perhaps the widest distributed arbovirus globally, being maintained in nature by transmission among ornithophagic Culex mosquitoes and a wide-variety of birds. A factor contributing to the success of the WNV invasion is the ability of the virus to successfully overwinter. Experimental studies on the preservation and transmission of West Nile virus by Culex pipiens pallens under simulated overwintering conditions have shown that Cx. p. pallens has a preservative effect on West Nile virus under low-temperature conditions. After the temperature rises, the virus reproduces rapidly and can spread to susceptible animals while the mosquitos feed on blood. This research is of great significance for exploring how West Nile virus enters new areas and how the cycle of transmission is maintained China., Competing Interests: Declaration of Competing Interest There is no conflict of interest of all authors., (Copyright © 2022. Published by Elsevier B.V.)

Mosquito-borne diseases are still threats to public health in the Zhejiang province of China. Surveillance of mosquitoes and the mosquito-borne pathogen is a vital approach for early warning, prevention, and control of the infectious disease. In this study, from 2018 to 2020, a total of 141607 female mosquitoes were caught by means of the light trap method. The main species were Culex pipiens quinquefasciatus/pallens (41.32%), Culex tritaeniorhynchus (47.6%), Aedes albopictus (2.5%), Anopheles sinensis (5.87%), Armigeres subalbatus (2.64%) and other mosquito species (0.07%). Cx. pipiens s.l. were the dominant species in two urban habitats and rural residential areas while Cx. tritaeniorhynchus was the main dominant species in the rural livestock sheds. In terms of seasonal fluctuation, Cx. pipiens s.l fluctuated at a high level from May to October. The peaks of Cx. tritaeniorhynchus, An. sinensis and Ar. subalbatus were in July. In addition, a total of 693 Ae. albopictus were collected with Biogents Mosquitaire CO 2 traps in emergency surveillance of dengue fever (DF) and screened for dengue virus infection. There were three circumstances of collection: The first: the sampling time before mosquito control during the local outbreak of DF in Lucheng of Wenzhou, 2019; The second circumstance: the sampling time after mosquito control during the local outbreak of DF of other cities in 2018-2019; The third circumstance: past DF epidemic areas the sampling time before mosquito control during the local outbreak of DF in Lucheng, Wenzhou, Zhejiang, 2019. The pools formed by mosquitoes collected in these three circumstances were 3 (6.1%), 35 (71.5%), and 11 (22.4%) respectively. Of the 49 pools tested, only one in the first circumstance was positive. The full-length dengue virus sequence of ZJWZ/2019 was obtained by sequencing and uploaded to the NCBI as number OK448162. Full-length nucleotide and amino acid homology analyses showed that ZJWZ2019 and Wenzhou DF serum isolates ZJWZ-62/2019 (MW582816) and ZJWZ-18/2019 (MW582815) had the highest homology. The analysis of full genome and E gene phylogenetic trees showed that ZJWZ2019 belonged to serotype 1, genotype I, lineage II, which was evolutionarily related to OK159963/Cambodia/2019. It implies that ZJWZ2019 originated in Cambodia. This study showed the species composition, seasonal dynamics of mosquitoes in different habitats in Zhejiang province and confirmed the role of Ae. albopictus in the transmission cycle of in outbreak of DF in the Lucheng district of Wenzhou in 2019, suggesting the importance of monitoring of vector Aedes infected dengue virus in the prevention and control of DF., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Liu, Wang, Hou, Wu, Zhang, Xing, Gao, Li, Guo, Jiang, Gong and Zhao.)

A new duck Tembusu-related flavivirus, Baiyangdian virus (BYDV), caused duck egg-drop syndrome in China. The rapid spread, unknown transmission routes, and zoonotic nature, raise serious concern about BYDV as a potential threat to human health. The study provides the first evaluation on the vector competence of Culex and Aedes mosquitoes to transmit BYDV in China. The results show that Culex tritaeniorhynchus , Culex pipiens pallens , Culex pipiens quinquefasciatus , and Aedes albopictus can become infected with BYD-1 virus (BYDV-1) on different days after oral infection. Although the viral copies in Ae. albopictus was higher than that in Cx. p. quinquefasciatus at 13 days postinfection (χ 2  = 10.385, p  = 0.016), there was no significant differences between infection rates of four mosquito species (χ 2  = 3.98, p  = 0.137). In transmission experiment, healthy ducks were infected after being bitten by virus-positive mosquitoes and BYDV-1 disseminated to and replicated in the duck brains. These findings verified the potential role of Cx. p. quinquefasciatus and Cx. tritaeniorhynchus as vectors of BYDV-1. BYDV-1 was also detected in salivary gland of Cx. p. pallens , which indicated that this virus could be transmitted by mosquitoes. These results provide evidence for the role of Culex mosquitoes in the transmission cycles involving BYDV-1 and avian hosts in China.

The Japanese encephalitis virus (JEV), a mosquito-borne zoonotic pathogen, is the major cause of viral encephalitis worldwide. An investigation of mosquito species diversity, JEV infection rate, and seasonal population fluctuations of Culex tritaeniorhynchus in Huaihua County, Hunan Province, China, revealed the distribution of vector mosquito populations and genotypes and molecular characteristics of current, common JEV strains in this region. Research on mosquito species diversity in different habitats in Huaihua revealed that local community composition was relatively simple, including five species from four genera (two Culex spp., one Anopheles sp., one Aedes sp., and one Armigeres sp.). Cx. tritaeniorhynchus was clearly the dominant species comprising 94.2-98.6% of all specimens and was always the most common species captured in paddy fields, pigpens, and human dwellings. The seasonal abundance of Cx. tritaeniorhynchus was relatively even, with a single seasonal peak in late August. Two Huaihua JEV strains isolated from the mosquito and pig were highly congruent. The genetic affinities were determined by analyzing capsid/premembrane (C/PrM) and envelope (E) gene variation. The results showed that they were of genotype III and most closely related to the live, attenuated vaccine strains SAl4-14-2 and SA14 and JaGAr01. The Huaihua E protein shares high similarity (mosquito 98.8% and pig 97.6%) at the nucleotide level with the SA14-14-2 vaccine. Although we found that the E gene sequences of the Huaihua JEV mosquito strain and pig strain have 11 and 15 amino acid site substitutions compared with the SAl4-14-2 vaccine, key sites that associated with JEV's antigenic activity and neurovirulence were unchanged. The SA14-14-2 vaccine should therefore be effective in preventing JEV infection in the Huaihua region.

Mosquitoes are hematophagous insects that carry-on and transmit many human viruses. However, little information is available regarding the common mechanisms underlying the infection of mosquitoes by these viruses. In this study, we reveal that the hematophagous nature of mosquitoes contributes to arboviral infection after a blood meal, which suppresses antiviral innate immunity by activating the GABAergic pathway. dsRNA-mediated interruption of the GABA signaling and blockage of the GABA A receptor by the specific inhibitors both significantly impaired arbovirus replication. Consistently, inoculation of GABA enhanced arboviral infection, indicating that GABA signaling facilitates the arboviral infection of mosquitoes. The ingestion of blood by mosquitoes resulted in robust GABA production from glutamic acid derived from blood protein digestion. The oral introduction of glutamic acid increased virus acquisition by mosquitoes via activation of the GABAergic system. Our study reveals that blood meals enhance arbovirus replication in mosquitoes through activation of the GABAergic system.

Information regarding the infection rate and genotype shifts for Japanese encephalitis virus (JEV) are important for JE vaccine application. In Jiangsu province, China, which is one of the provinces with a high prevalence of JE, JEV infection in swine and mosquitoes in certain cities has only been investigated in 2008-2009. Lianyungang City has one of the highest numbers of JE cases in Jiangsu province, and it has a high risk of JEV invasion via migrant birds. JEV infection in vectors in Lianyungang City, which has urban and rural parts, has not been investigated. In 2015-2016, we collected mosquitoes in cowsheds with ultraviolet light traps and detected JEV by reverse transcription-polymerase chain reaction (RT-PCR) method in Culex tritaeniorhynchus in Xintan village, Xuzhuang village, and Xiaogaozhuang village in Lianyungang City, China. The proportion of positive pools, which is calculated by the number of infected pools to the total number of pools tested in these villages, were 16.67%, 20.00%, and 4.17%, respectively, and the minimum infection rates, which is calculated as the ratio of the number of positive pools to the total number of mosquitoes tested, were 3.33‰, 4.00‰, and 0.83‰, respectively. Four JEV strains from positive samples were coded as LYG-1, LYG-2, LYG-3, and LYG-4, and the complete E genes were sequenced. Furthermore, the complete genome of LYG-3 was sequenced. The phylogenetic analysis indicated that all the four JEV strains belonged to genotype I-b. This is the first report of genotype I JEV strain in Jiangsu province. The high JEV infection rate in Culex tritaeniorhynchus indicated a high risk of JE reemergence in Lianyungang. The detected JEV strains may have similar antigenicity to that of SA14-14-2 according to molecular characters. These findings suggest that the vaccine can still be effective in Lianyungang.

Background: Pesticide resistance due to sodium channel point mutations has been well documented in many mosquito species., Methods: We tested the resistance of six, wild, Chinese populations of the mosquito Culex pipiens quinquefasciatus to deltamethrin and cyhalothrin. The full length of the sodium channel gene was cloned and sequenced from a pooled sample of mosquitoes from each population., Results: Seven amino acid alterations were found (V250M, R436K, M943V, I973T, L1035F, L1035S and E1901D). Correlation between the frequencies of these mutations and the level of pesticide resistance (LC50) associated with them indicates that those at position L1035 (corresponding to position L1014F in the house fly, Musca domestica; GenBank Accession No.: X96668) are associated with resistance to deltamethrin and cyhalothrin. The frequency of the L1035F mutation was significantly correlated with resistance to deltamethrin (R2 = 0.536, P = 0.049) and cyhalothrin (R2 = 0.626, P = 0.030), and the combined frequency of the L1035F and L1035S mutations was significantly correlated with resistance to both deltamethrin (R2 = 0.661, P = 0.025), and cyhalothrin (R2 = 0.803, P = 0.008). None of the other mutations were correlated with either deltamethrin or cyhalothrin resistance. Interestingly, an HWE test indicated significant linkage between the M943V and I973T mutations (P < 0.01), but further research is required to determine the biological significance of this linkage., Conclusions: Identifying these mutations may be of practical benefit to the development of pesticide resistance management programs.

The distribution of the West Nile virus (WNV) in the organs and tissues of the mosquito Culex pipiens pallens, a potential vector of WNV in China, was investigated up to 14 days after oral infection. The WNV antigen was detected in paraffin-embedded mosquitoes using immunocytochemistry and viral titers of post-infected mosquitoes determined by plaque assay. Viral titers sharply decreased 24 h post-infection, were undetectable for the first few days, then rose over the course of infection. The first midgut infection appeared after one day, and the overall infection rate (based on midgut infection) was 43.9%. Other tissues, including hindgut, foregut, ovarian follicles, Malpighian tubules, and ommatidia, showed weak WNV antigens as early as three days post-infection. Staining in the salivary glands first appeared after seven days, and the salivary gland infection rate on the 14th day was 37.5%. Specimens with no detectable WNV antigens in any tissues, and with positive results confined to the midgut, anterior midgut, and hindgut, were observed on the 14th day. The route of viral dissemination from the midgut, and the relative importance of amplifying tissues in mosquitoes' susceptibility to infection, were evaluated. The results indicate that Cx. p. pallens has the ability to harbor WNV throughout its alimentary system and that midgut epithelial cells may be the initial site of the replication of this virus in this species., (© 2014 The Society for Vector Ecology.)

Acetylcholinesterase resistance has been well documented in many insects, including several mosquito species. We tested the resistance of five wild, Chinese strains of the mosquito Culex pipiens quinquefasciatus to two kinds of pesticides, dichlorvos and propoxur. An acetylcholinesterase gene (ace1) was cloned and sequenced from a pooled sample of mosquitoes from these five strains and the amino acids of five positions were found to vary (V185M, G247S, A328S, A391T, and T682A). Analysis of the correlation between mutation frequencies and resistance levels (LC50) suggests that two point mutations, G247S (r2 = 0.732, P = 0.065) and A328S (r2 = 0.891, P = 0.016), are associated with resistance to propoxur but not to dichlorvos. Although the V185M mutation was not associated with either dichlorvos or propoxur resistance, its RS genotype frequency was correlated with propoxur resistance (r2 = 0.815, P = 0.036). And the HWE test showed the A328S mutation is linked with V185M, also with G247S mutation. This suggested that these three mutations may contribute synergistically to propoxur resistance. The T682A mutation was negatively correlated with propoxur (r2 = 0.788, P = 0.045) resistance. Knowledge of these mutations may help design strategies for managing pesticide resistance in wild mosquito populations.

Two strains of the Western equine encephalitis virus (WEEV) were first detected and isolated in China in 2001. The maintenance and transmission cycles of WEEV in China are currently not well understood, and the mosquito vectors involved in these cycles are unknown. To understand the ability of the local mosquitoes in China to transmit WEEV, the vector competence of five mosquito species, namely, Culex pipiens pallens Coquillett, Cx. p. quinquefasciatus Say, Aedes (Stegomyia) albopictus Skuse, Ae. (Stegomyia) aegypti Linnaeus, and C. tritaeniorhynchus Giles, for WEEV were evaluated. Infection rates for Cx. p. pallens, Cx. p. quinquefasciatus, Cx. tritaeniorhynchus, Ae. Albopictus, and Ae. aegypti were 46%, 60%, 80%, 37%, and 25%, respectively. Dissemination rates for the same species were 60%, 61%, 75%, 55%, and 50%, respectively. Transmission rates were 41%, 53%, 57%, and 45% for Cx. p. pallens, Cx. p. quinquefasciatus, Ae. Albopictus, and Ae. Aegypti, respectively. Infection rates were significantly different between species, but the difference between dissemination and transmission rates were nonsignificant. These results suggest that several local mosquito species in China are competent laboratory vectors for WEEV.

Two western equine encephalomyelitis virus (WEEV) strains have been isolated in China. Our previous studies have verified that the mosquito Culex pipiens pallens Coquillett (Diptera: Culicidae) infected with WEEV was capable of transmitting this arbovirus, but it was not clear how the sequential multiplication and spread of virus occurred within the mosquito. In this study, we observed the distribution of WEEV antigen in orally-infected Cx. p. pallens by immunohistochemistry in order to better understand the initial infection, dissemination, and transmission of WEEV in the potential vector. Orally-infected WEEV dissemination varied within the different tissues of Cx. p. pallens, with virus antigen consistently observed in the salivary glands, foregut, midgut epithelial cells, Malpighian tubules, hindgut, and ovarian follicles of some individuals after various days of extrinsic incubation. We suggest that Cx. p. pallens, the potential vector of WEEV, has the ability to harbor the virus through the alimentary system, and the midgut epithelial cell may be the initial site of WEEV replication after ingestion of a viremic blood meal., (© 2010 The Society for Vector Ecology.)

A new duck Tembusu-related flavivirus, Baiyangdian virus (BYDV), caused duck egg-drop syndrome in China. The rapid spread, unknown transmission routes, and zoonotic nature, raise serious concern about BYDV as a potential threat to human health. The study provides the first evaluation on the vector competence of Culex and Aedes mosquitoes to transmit BYDV in China. The results show that Culex tritaeniorhynchus, Culex pipiens pallens, Culex pipiens quinquefasciatus, and Aedes albopictus can become infected with BYD-1 virus (BYDV-1) on different days after oral infection. Although the viral copies in Ae. albopictus was higher than that in Cx. p. quinquefasciatus at 13 days postinfection (χ2 = 10.385, p = 0.016), there was no significant differences between infection rates of four mosquito species (χ2 = 3.98, p = 0.137). In transmission experiment, healthy ducks were infected after being bitten by virus-positive mosquitoes and BYDV-1 disseminated to and replicated in the duck brains. These findings verified the potential role of Cx. p. quinquefasciatus and Cx. tritaeniorhynchus as vectors of BYDV-1. BYDV-1 was also detected in salivary gland of Cx. p. pallens, which indicated that this virus could be transmitted by mosquitoes. These results provide evidence for the role of Culex mosquitoes in the transmission cycles involving BYDV-1 and avian hosts in China. [ABSTRACT FROM AUTHOR]

The Japanese encephalitis virus (JEV), a mosquito-borne zoonotic pathogen, is the major cause of viral encephalitis worldwide. An investigation of mosquito species diversity, JEV infection rate, and seasonal population fluctuations of Culex tritaeniorhynchus in Huaihua County, Hunan Province, China, revealed the distribution of vector mosquito populations and genotypes and molecular characteristics of current, common JEV strains in this region. Research on mosquito species diversity in different habitats in Huaihua revealed that local community composition was relatively simple, including five species from four genera (two Culex spp., one Anopheles sp., one Aedes sp., and one Armigeres sp.). Cx. tritaeniorhynchus was clearly the dominant species comprising 94.2–98.6% of all specimens and was always the most common species captured in paddy fields, pigpens, and human dwellings. The seasonal abundance of Cx. tritaeniorhynchus was relatively even, with a single seasonal peak in late August. Two Huaihua JEV strains isolated from the mosquito and pig were highly congruent. The genetic affinities were determined by analyzing capsid/premembrane (C/PrM) and envelope (E) gene variation. The results showed that they were of genotype III and most closely related to the live, attenuated vaccine strains SAl4-14-2 and SA14 and JaGAr01. The Huaihua E protein shares high similarity (mosquito 98.8% and pig 97.6%) at the nucleotide level with the SA14-14-2 vaccine. Although we found that the E gene sequences of the Huaihua JEV mosquito strain and pig strain have 11 and 15 amino acid site substitutions compared with the SAl4-14-2 vaccine, key sites that associated with JEV's antigenic activity and neurovirulence were unchanged. The SA14-14-2 vaccine should therefore be effective in preventing JEV infection in the Huaihua region. [ABSTRACT FROM AUTHOR]

The complete sequence of the mitochondrial genome of theCulex tritaeniorhynchus(Diptera: Culicidae) is presented using traditional Sanger sequencing. Its mitogenome are 15,123 bp in length, consisting of 13 protein-coding genes, 2 ribosomal RNA genes (rRNA) and 22 transfer RNA (tRNA) genes. The percent of A + T of the mitogenome is 77.73%. Most mitochondrial genes are encoded on the heavy strand, except forNAD5,NaD4,NAD4L,NAD1, two rRNA and nine tRNA genes, which are encoded on the light strand. The results of phylogenetic analyzes showed that theCx. tritaeniorhynchushas close relationship with the species of genusCulex. [ABSTRACT FROM PUBLISHER]