Synthesis, insecticidal Activity, and molecular docking analysis of some benzo[h]quinoline derivatives against Culex pipiens L. Larvae.

Some heterocycles bearing a benzo[h]quinoline moiety were synthesized through treating a 3-((2-chlorobenzo[h]quinolin-3-yl)methylene)-5-(p-tolyl)furan-2(3H)-one with four nitrogen nucleophiles comprising ammonium acetate, benzylamine, dodecan-1-amine, and 1,2-diaminoethane. Also, thiation reactions of furanone and pyrrolinone derivatives were investigated. The insecticidal activity of these compounds against mosquito larvae (Culex pipiens L.) was evaluated. All tested compounds exhibited significant larvicidal activity, surpassing that of the conventional insecticide chlorpyrifos. In silico docking analysis revealed that these compounds may act as acetyl cholinesterase (AChE) inhibitors, potentially explaining their larvicidal effect. Additionally, interactions with other neuroreceptors, such as nicotinic acetylcholine receptor and sodium channel voltage-gated alpha subunit were also predicted. The results obtained from this study reflected the potential of benzo[h]quinoline derivatives as promising candidates for developing more effective and sustainable mosquito control strategies. The ADME (absorption, distribution, metabolism, and excretion) analyses displayed their desirable drug-likeness and oral bioavailability properties.

Next generation sequencing improves the resolution of detecting mixed host blood meal sources in field collected arboviral mosquito vectors.

Accurate knowledge of blood meal hosts of different mosquito species is critical for identifying potential vectors and establishing the risk of pathogen transmission. We compared the performance of Miseq next generation sequencing approach relative to conventional Sanger sequencing approach in identification of mosquito blood meals using genetic markers targeting the 12S rRNA and cytochrome oxidase I (COI) genes. We analysed the blood meals of three mosquito vector species (Aedes aegypti, Aedes simpsoni s.l. and Culex pipiens s.l.) collected outdoors, and compared the frequency of single- versus multiple-blood feeding. Single host blood meals were mostly recovered for Sanger-based sequencing of the mitochondrial 12S rRNA gene, whereas Miseq sequencing employing this marker and the COI marker detected both single and multiple blood meal hosts in individual mosquitoes. Multiple blood meals (two or more hosts) which mostly included humans were detected in 19%-22.7% of Ae. aegypti samples. Most single host blood meals for this mosquito species were from humans (47.7%-57.1%) and dogs (9.1%-19.0%), with livestock, reptile and rodent hosts collectively accounting for 4.7%-28.9% of single host blood meals. The frequency of two or more host blood meals in Ae. simpsoni s.l. was 26.3%-45.5% mostly including humans, while single host blood meals were predominantly from humans (31.8%-47.4%) with representation of rodent, reptile and livestock blood meals (18.2%-68.2%). Single host blood meals from Cx. pipiens s.l. were mostly from humans (27.0%-39.4%) and cows (11.5%-27.36%). Multiple blood meal hosts that mostly included humans occurred in 21.2%-24.4% of Cx. pipiens s.l. samples. Estimated human blood indices ranged from 53%-76% for Ae. aegypti, 32%-82% for Ae. simpsoni s.l. and 26%-61% for Cx. pipiens s.l. and were consistently lower for Sanger-based sequencing approach compared to Miseq-based sequencing approach. These findings demonstrate that Miseq sequencing approach is superior to Sanger sequencing approach as it can reliably identify mixed host blood meals in a single mosquito, improving our ability to understand the transmission dynamics of mosquito-borne pathogens.

Blood meal sources of mosquitoes (Diptera: Culicidae) in domestic and open green environments from two urbanisations of temperate Argentina.

Understanding the blood-feeding patterns of mosquitoes is essential for evaluating their potential as disease vectors, especially in urban areas where mosquitoes coexist with humans, domestic animals and wildlife. This study aimed to bridge a substantial gap in regional knowledge by identifying the blood meal sources of field-collected mosquitoes in domestic and open green environments from two urbanisations of temperate Argentina, the Área Metropolitana de Buenos Aires (AMBA) and Tandil, using molecular techniques. Female mosquitoes were collected from November 2019 to March 2020 and April-May 2021. A bipartite network analysis was performed for each environment and urbanisation. A total of 103 blood meals from Aedes (2 species) and Culex (7 species) were identified. Among these, five mammal and 18 bird species were recognised as hosts. Aedes mosquitoes exclusively fed on mammals, while Culex mosquitoes exhibited a broader host range including both birds and mammals. In AMBA, the open green environments were composed by more mosquito species than the domestic environments, while both presented similar numbers of vertebrate species. In contrast, in open green environments from Tandil only blood-fed Aedes albifasciatus were collected. For open green environments of AMBA and domestic environments of Tandil, results suggested some degree of host selection. For the three main vectors of diseases in the region, Aedes aegypti, Ae. albifasciatus and Culex pipiens molestus, we present the first molecular evidence of human blood meals in South America. Epidemiological significance of the present findings is discussed.

Chemical composition of four essential oils and their adulticidal, repellence, and field oviposition deterrence activities against Culex pipiens L. (Diptera: Culicidae).

Effective mosquito repellents can limit the transmission of vector-borne diseases to humans. Consequently, there is an urgent need to develop mosquito control strategies that prioritize eco-friendly and cost-effective repellents. Essential oils (EOs) have enormous potential for mosquito repellency. Here, cinnamon, basil, eucalyptus, and peppermint EOs were investigated for adulticide and repellency properties against Culex pipiens as well on the oviposition behavior of gravid females from laboratory (lab test) and field (field test) populations. Cinnamon oil was an effective oviposition deterrent regardless of the population and had high adulticidal activity with toxicity index of 75.00% at 24 h of exposure, relative to deltamethrin. In addition, it exhibited effective repellency at 98.01% and 71.22% at 6.67 and 1.71 µl/cm2, respectively. Peppermint oil had the least adulticidal activity with toxicity index of 6.2% at 24 h, and it resulted in low repellency at 70.90% and 50.64% at 6.67 and 1.71 µl/cm2, respectively. On average, basil and eucalyptus oils showed some adulticidal efficiency, repellency, and oviposition deterrent activity. For all treatments, the oviposition deterrent index values of gravid females from natural populations (field test) were lower than those from lab-reared (lab test) females. Different ratios of monoterpenoids, phenylpropanoids, and fatty acids in the EOs tested likely account for the activity variations observed. Our results suggest cinnamon, basil, eucalyptus, and peppermint EOs, which are widely available, economical, and eco-friendly, with good potential for mosquito control strategies.

Exploring the range of Culex mosquitoes in Western Argentinean Patagonia, unveiling the presence of Culex pipiens bioform pipiens in South America.

Culicids in Argentinean Patagonia are characterized by low species diversity and adaptation to extreme environmental conditions, yet few studies have been conducted in the region. To further assess the occurrence of Culicidae in Western Patagonia, and in particular the presence of Culex pipiens bioforms at the southernmost extent of their distribution, immature and adult specimens were collected aboveground across various land uses located in shrubland, steppe, and deciduous forest between 38.96 and 46.55°S. Mosquitoes were reported at 35 of the 105 inspected sites. Five species from the genus Culex were identified, all of which were present in the steppe and the forest, while only Cx. apicinus and members of the Cx. pipiens complex were collected in the shrubland. Within the latter, a total of 150 specimens were molecularly identified by PCR amplification of Ace-2 and CQ11 loci. The first-to-date occurrence of bioform pipiens in South America is reported, along with the first records of Cx. quinquefasciatus signatures in Patagonia. In addition, the distribution of Cx. acharistus and Cx. dolosus as south as Santa Cruz province is expanded, and the first record of Cx. eduardoi in Río Negro province is provided. Immature specimens of Cx. pipiens were conspicuous in human-made aquatic habitats (both containers and in the ground), while Cx. acharistus was more prominent in artificial containers and Cx. eduardoi was mainly in ground habitats, either natural or human-made. These findings provide valuable insights into the distribution and ecological roles of these mosquito species in a region of extreme environmental conditions.

Synthesis and evaluation of larvicidal efficacy against C. pipiens of some new heterocyclic compounds emanated from 2-cyano-N'-(2-(2,4-dichlorophenoxy)acetyl)acetohydrazide.

Several infectious diseases are transmitted and spread by mosquitoes, and millions of people die annually from them. The mosquito, Culex pipiens is a responsible for the emergence of various Virus in Egypt. So, we devote our work to evaluate the larvicidal efficacy against C. pipiens of some new heterocyclic compounds containing chlorine motifs. The implementation was emanated from using 2-cyano-N'-(2-(2,4-dichlorophenoxy)acetyl)acetohydrazide (3) as scaffold to synthesize some new heterocyclic compounds. The structures of the synthesized compounds were interpreted scrupulously by spectroscopic and elemental analyses. Thereafter, the larvicidal activity against C. pipiens of thirteen synthesized compounds was estimated. Noteworthy, cyanoacetohydrazide derivative 3 and 3-iminobenzochromene derivative 12 showed a fabulous potent efficacy with LC50 equal to 3.2 and 3.5 ppm against C. pipiens, respectively, and are worth being further evaluated in the field of pest control.

Anti-Culex pipiens activity of different pomegranate cultivars and determination of their bioactive compounds using LC-MS profiling.

INTRODUCTION: Pomegranate (Punica granatum L.) peels are rich in various bioactive compounds. Characterization of these compounds is crucial for the utilization of peel waste in industrial processing. OBJECTIVE: The study aimed (1) to establish and compare the metabolic profiles of the peel of seven pomegranate cultivars and (2) to identify bioactive compounds contributing to the larvicidal activity against the third instar larvae of Culex pipiens. MATERIALS AND METHODS: UPLC-ESI-MS/MS was utilized to analyze peel methanol extracts of different pomegranate cultivars. The larvicidal activity was determined by calculating the larval mortality among the third instar larvae of C. pipiens. Multivariate data analysis was conducted to identify the metabolites that exhibited a larvicidal effect. RESULTS: A total of 24 metabolites, including hydrolyzable tannins, flavonoids, and alkaloids, were tentatively identified in both negative and positive ionization modes. The extract of cultivar 'Black' exhibited the most potent larvicidal effect with LC50 values of 185.15, 156.84, and 138.12 ppm/mL after 24, 48, and 72 h of treatment, respectively. By applying chemometric techniques, the larvicidal activity could be directly correlated to the bioactive compounds punicalagin, quercetin-O-rhamnoside, quercetin-O-pentoside, and galloyl-HHDP-glucose. CONCLUSION: The present study implemented UPLC-ESI-MS/MS and chemometric techniques as potential tools for metabolomics analysis and differentiation between peels of different pomegranate cultivars. In addition, cultivar 'Black' extract could be a promising natural insecticide against mosquitoes since it is rich in bioactive compounds with larvicidal activity.

Evaluation of the evolutionary genetics and population structure of Culex pipiens pallens in Shandong province, China based on knockdown resistance (kdr) mutations and the mtDNA-COI gene.

BACKGROUND: Mosquitoes are important vectors for a range of diseases, contributing to high rates of morbidity and mortality in the human population. Culex pipiens pallens is dominant species of Culex mosquito in northern China and a major vector for both West Nile virus and Bancroftian filariasis. Insecticide application were largely applied to control the mosquito-mediated spread of these diseases, contributing to increasing rates of resistance in the mosquito population. The voltage-gated sodium channel (Vgsc) gene is the target site of pyrethroids, and mutations in this gene cause knockdown resistance (kdr). While these kdr mutations are known to be critical to pyrethroid resistance, their evolutionary origins remain poorly understood. Clarifying the origins of these mutations is potential to guide further vector control and disease prevention efforts. Accordingly, the present study was designed to study the evolutionary genetics of kdr mutations and their association with the population structure of Cx. p. pallens in Shandong province, China. METHODS: Adult Culex females were collected from Shandong province and subjected to morphological identification under a dissection microscope. Genomic DNA were extracted from the collected mosquitoes, the Vgsc gene were amplified via PCR and sequenced to assess kdr allele frequencies, intron polymorphisms, and kdr codon evolution. In addition, population genetic diversity and related population characteristics were assessed by amplifying and sequencing the mitochondrial cytochrome C oxidase I (COI) gene. RESULTS: Totally, 263 Cx. p. pallens specimens were used for DNA barcoding and sequencing analyses to assess kdr allele frequencies in nine Culex populations. The kdr codon L1014 in the Vgsc gene identified two non-synonymous mutations (L1014F and L1014S) in the analyzed population. These mutations were present in the eastern hilly area and west plain region of Shandong Province. However, only L1014F mutation was detected in the southern mountainous area and Dongying city of Shandong Province, where the mutation frequency was low. Compared to other cities, population in Qingdao revealed significant genetic differentiation. Spatial kdr mutation patterns are likely attributable to some combination of prolonged insecticide-mediated selection coupled with the genetic isolation of these mosquito populations. CONCLUSIONS: These data suggest that multiple kdr alleles associated with insecticide resistance are present within the Cx. p. pallens populations of Shandong Province, China. The geographical distributions of kdr mutations in this province are likely that the result of prolonged and extensive insecticide application in agricultural contexts together with frequent mosquito population migrations. In contrast, the low-frequency kdr mutation detected in central Shandong Province populations may originate from the limited selection pressure in this area and the relative genetic isolation. Overall, the study compares the genetic patterns revealed by a functional gene with a neutral marker and demonstrates the combined impact of demographic and selection factors on population structure.

The diapausing mosquito Culex pipiens exhibits reduced levels of H3K27me2 in the fat body.

Culex pipiens, the northern house mosquito, is a major vector of West Nile virus. To survive the severe winter, adult mosquitoes enter a diapause programme. Extended lifespan and an increase in lipid storage are key indicators of diapause. Post-translational modifications to histone proteins impact the expression of genes and have been linked to the lifespan and energy utilisation of numerous insects. Here, we investigated the potential contribution of epigenetic alterations in initiating diapause in this mosquito species. Multiple sequence alignment of H3 sequences from other insect species demonstrates a high conservation of the H3 histone in Cx. pipiens throughout evolution. We then compared the levels of histone methylation in the ovaries and fat body tissues of diapausing and non-diapausing Cx. pipiens using western blots. Our data indicate that histone methylation levels in the ovaries of Cx. pipiens do not change during diapause. In contrast, H3K27me2 levels decrease more than twofold in the fat body of diapausing mosquitoes relative to non-diapausing counterparts. H3K27 methylation plays a crucial role in chromosome activation and inactivation during development in many insect species. This is predominantly governed by polycomb repressor complex 2. Intriguingly, a previous ChIP-seq study demonstrated that the transcription factor FOXO (Forkhead box O) targets the genes that comprise this complex. In addition, H3K27me2 exhibits dynamic abundance throughout the diapause programme in Cx. pipiens, suggesting its potential role in the initial activation of the diapause programme. This study expands our understanding of the relationship between alterations in epigenetic regulation and diapause.

Diversity oriented synthesis and SAR studies of new quinazolinones and related compounds as insecticidal agents against Culex pipiens L. Larvae and associated predator.

The ongoing study reports the synthesis, spectroscopic analyses and larvicidal efficacy of novel series of quinazolinone derivatives and related compounds. The structures of the products were confirmed relied on their analytical and spectral data (IR, 1H NMR, and 13C NMR). The spectral documentation promoted the successful isolation of the desirable compounds. The insecticidal activities of the synthesized compounds were assessed against laboratory and field strains of Culex pipiens larvae and a predator from the same ecological niche, Cybister tripunctatus. The results revealed that most of the tested compounds showed high potencies against lab strain of C. pipiens larvae with low resistance ratios in filed strain. In particular, compounds 15, 6 and 16 showed low LC50 values, 0.094, 0.106, 0.129 (µg/mL), respectively against lab strain of C. pipiens larvae. The present study also explored the toxicity of tested compounds against field strain of non-target C. tripunctatus. Most of tested compounds were safer than temephos, especially 15 and 6 with SI/PSF values 96.746 and 83.167, respectively. Structure-activity relationship (SAR) was discussed the effect of substituents insertion on the derivatives activities. Quinazolinone derivatives and related compounds are promising compounds in the mosquito control programs and further studies are recommended to develop more effective derivatives and reveal their mode of action.

The first molecular confirmation of Culex pipiens complex as potential natural vectors of Dirofilaria immitis in Myanmar.

Dirofilariosis, known as one of the most widespread vector-borne zoonotic diseases, is caused by several different species of the nematodes of the genus Dirofilaria, which can be transmitted by Culex, Anopheles and Aedes mosquito vectors. In order to identify key vector mosquitoes of filarial parasites in Myanmar, mosquitoes were collected during three different seasons (summer, rainy and winter) in three townships in Nay Pyi Taw area, Myanmar. DNA extraction and polymerase chain reaction (PCR) analyses were conducted for 185 mosquito pools, with each pool containing 1-10 mosquitoes. Dirofilaria immitis was detected in 20 pools of Culex pipiens complex mosquitoes. The minimum infection rate of mosquitoes was found to be 16.33. The small subunit ribosomal RNA (12S rDNA) gene targeted PCR revealed that the sequences obtained were completely identical to the sequences of D. immitis derived from dogs in China, Brazil and France. The sequences obtained from mitochondrial cytochrome oxidase subunit I (COI) gene PCR exhibited 100% identity with the sequences of D. immitis derived from dogs in Bangladesh, Iran, Japan and Thailand, as well as humans in Iran and Thailand, and mosquitoes in Germany and Hungary. The findings of this study demonstrated that the mosquito species of Cx. pipiens complex are potential mosquito vectors for dirofilariosis in Myanmar.

Impact of larvae of the Asian tiger mosquito Aedes albopictus on larvae of the Culex pipiens complex from Germany in laboratory co-breeding studies.

Since the first detection of the Asian tiger mosquito Aedes albopictus (Diptera: Culicidae) eggs in Germany in 2007, several populations of this species have established in Germany. Although colloquially Ae. albopictus is called an 'invasive species', it is not considered 'invasive' and therefore to be controlled according to the European Union (EU) Environmental and Nature Protection Act since evidence of displacement of native species is missing. To test the competitive potential of Ae. albopictus towards mosquito species native to Germany, laboratory experiments were conducted with larvae of this species and indigenous Cx. pipiens complex species/biotypes. First instar larvae of Ae. albopictus and of one of the native taxa were exposed to different temperatures and fed with different food sources. The ratio of individuals developing into adults as well as the time the larvae needed for development were taken as a measure of competitive outcome. In addition, the size of emerging adults was compared between control and experimental groups. Regarding developmental time, no significant differences were found between treatments and controls while significant differences were found regarding developmental rate and average wing size of individuals. Because no evidence of competitive repression of the native species was found, Ae. albopictus cannot be included in the EU list of invasive species.

Experimental infection of Aedes (Stegomyia) albopictus and Culex pipiens mosquitoes with Bluetongue virus.

Bluetongue disease (BT), caused by Bluetongue virus (BTV), infects wild and domestic ruminants, causing severe economic damage in the cattle and sheep industry. Proven vectors of BTV are biting midges belonging to the Culicoides genus, but other arthropods are considered potential vectors, such as ticks, mosquitoes, wingless flies, and sand flies. The present study represents the first attempt to evaluate the vectorial capacity of Culex pipiens and Aedes albopictus for BTV. Mosquitoes were artificially fed with blood containing BTV serotype 1. Infection, dissemination and transmission rates were evaluated at 0, 3, 7, 14 and 21 days after an infected blood meal. Viral RNA was only detected up to 3 days post infection in the bodies of both species. This study indicates that the two Italian populations of Cx. pipiens and Ae. albopictus are not susceptible to BTV infection.

Mapping the abundance of endemic mosquito-borne diseases vectors in southern Quebec.

BACKGROUND: Climate change is increasing the dispersion of mosquitoes and the spread of viruses of which some mosquitoes are the main vectors. In Quebec, the surveillance and management of endemic mosquito-borne diseases, such as West Nile virus or Eastern equine encephalitis, could be improved by mapping the areas of risk supporting vector populations. However, there is currently no active tool tailored to Quebec that can predict mosquito population abundances, and we propose, with this work, to help fill this gap. METHODS: Four species of mosquitos were studied in this project for the period from 2003 to 2016 for the southern part of the province of Quebec: Aedes vexans (VEX), Coquillettidia perturbans (CQP), Culex pipiens-restuans group (CPR) and Ochlerotatus stimulans group (SMG) species. We used a negative binomial regression approach, including a spatial component, to model the abundances of each species or species group as a function of meteorological and land-cover variables. We tested several sets of variables combination, regional and local scale variables for landcover and different lag period for the day of capture for weather variables, to finally select one best model for each species. RESULTS: Models selected showed the importance of the spatial component, independently of the environmental variables, at the larger spatial scale. In these models, the most important land-cover predictors that favored CQP and VEX were 'forest', and 'agriculture' (for VEX only). Land-cover 'urban' had negative impact on SMG and CQP. The weather conditions on the trapping day and previous weather conditions summarized over 30 or 90 days were preferred over a shorter period of seven days, suggesting current and long-term previous weather conditions effects on mosquito abundance. CONCLUSIONS: The strength of the spatial component highlights the difficulties in modelling the abundance of mosquito species and the model selection shows the importance of selecting the right environmental predictors, especially when choosing the temporal and spatial scale of these variables. Climate and landscape variables were important for each species or species group, suggesting it is possible to consider their use in predicting long-term spatial variationsin the abundance of mosquitoes potentially harmful to public health in southern Quebec.

Co-application of entomopathogenic fungi with chemical insecticides against Culex pipiens.

Culex pipiens (Diptera: Culicidae) is a vector of many human and animal diseases. Its control is regarded as a preventative approach that is focused on effectively managing such diseases. In this context, dose response assays of two insecticides, bendiocarb and diflubenzuron were performed with two entomopathogenic fungi, Beauveria bassiana and Metarhizium anisopliae against 3rd instar C. pipiens larvae. The most effective agents, combination experiments as well as enzymatic activities of phenoloxidase (PO) and chitinase (CHI) were also assessed. The results showed that diflubenzuron was more effective at low concentrations (LC50: 0.001 ppm) than bendiocarb (LC50: 0.174 ppm), whereas M. anisopliae was more effective (LC50: 5.2 × 105 conidia/mL) than B. bassiana (LC50: 7.5 × 107 conidia/mL). Synergistic interactions were observed when diflubenzuron was applied at 2- and 4-days post- exposure to M. anisopliae, with the highest degree of synergism observed when diflubenzuron was applied 2 days post-fungal exposure (χ2 = 5.77). In contrast, additive interactions were recorded with all other insecticide-fungal combinations. PO activities significantly (p ≤ 0.05) increased during 24 h after a single diflubenzuron treatment as well as when diflubenzuron was applied prior to M. anisopliae, whereas suppressed after 24 h when M. anisopliae applied prior to diflubenzuron as well as after 48 h from single and combined treatments. CHI activity increased 24 h after both single and combined treatments, the activity remained elevated 48 h after a single diflubenzuron treatment and when diflubenzuron was applied after M. anisopliae. Histological study of the cuticle by transmission electron microscopy revealed abnormalities following single and combined treatments. Germination of the conidia and production of the mycelium that colonizes the lysing cuticle was obvious when diflubenzuron was applied 48 h after M. anisopliae exposure. Overall, these results demonstrate that M. anisopliae is compatible with diflubenzuron at lower concentrations and that combined applications can improve C. pipiens management.

Phytochemical Investigation of Three Cystoseira Species and Their Larvicidal Activity Supported with In Silico Studies.

Culex pipiens mosquitoes are transmitters of many viruses and are associated with the transmission of many diseases, such as filariasis and avian malaria, that have a high rate of mortality. The current study draws attention to the larvicidal efficacy of three methanolic algal extracts, Cystoseira myrica, C. trinodis, and C. tamariscifolia, against the third larval instar of Cx. pipiens. The UPLC-ESI-MS analysis of three methanol fractions of algal samples led to the tentative characterization of twelve compounds with different percentages among the three samples belonging to phenolics and terpenoids. Probit analysis was used to calculate the lethal concentrations (LC50 and LC90). The highest level of toxicity was attained after treatment with C. myrica extract using a lethal concentration 50 (LC50) of 105.06 ppm, followed by C. trinodis (135.08 ppm), and the lowest level of toxicity was achieved by C. tamariscifolia (138.71 ppm) after 24 h. The elevation of glutathione-S-transferase (GST) and reduction of acetylcholine esterase (AChE) enzymes confirm the larvicidal activity of the three algal extracts. When compared to untreated larvae, all evaluated extracts revealed a significant reduction in protein, lipid, and carbohydrate contents, verifying their larvicidal effectiveness. To further support the observed activity, an in silico study for the identified compounds was carried out on the two tested enzymes. Results showed that the identified compounds and the tested enzymes had excellent binding affinities for each other. Overall, the current work suggests that the three algal extractions are a prospective source for the development of innovative, environmentally friendly larvicides.

Physiological and developmental dysfunctions in the dengue vector Culex pipiens (Diptera: Culicidae) immature stages following treatment with zinc oxide nanoparticles.

The medical value of mosquitoes attracted researchers worldwide to search for a valuable way to control such serious insects. The continuous development of resistance against chemical insecticides pushed toward looking for novel and promising compounds against mosquitoes. In this study, the toxicity and physio-developmental effects of 10-30 nm spherical zinc oxide nanoparticles (ZnONPs) in aqueous suspension was addressed against the first larval instar of Culex pipiens mosquito. The calculated value of LC50 was about 0.892 g/L while the sub lethal concentration LC20 recorded about 0.246 g/L. Larvae treated with ZnONPs suffered reduced growth rate, longer developmental period and malformations in the breathing tube. Furthermore, the treated larvae showed clear abnormal appearance of the gastric caeca and midgut epithelia under transmission electron microscope (TEM). These abnormalities appeared as condensation of the nuclear chromatin, abnormal shape or absence of microvilli, highly increased amount of smooth endoplasmic reticulum in the cytoplasm and appearance of numerous vacuoles. Additionally, ZnONPs interfered with several biochemical pathways such as induction of oxidative stress which appeared in the form of increased levels of hydrogen peroxide and inability to activate the detoxifying enzymes alkaline phosphatase (ALP), catalase and glutathione peroxidase (GPX). On the contrary, the activity of the antioxidant enzyme superoxide dismutase (SOD) increased in treated larvae. Furthermore, LC20 and LC50 of ZnONPs inhibited the growth rate of the larval gut fauna in vitro. These results clearly show that ZnONPs target several tissues leading to serious alteration in the physiological and developmental processes in C. pipiens mosquito larvae.

Inference of selection pressures that drive insecticide resistance in Anopheles and Culex mosquitoes in Korea.

Pyrethroids are primarily used for mosquito control in Korea. However, high frequencies of mutations conferring resistance to not only pyrethroids but also to other insecticides have been found in mosquito populations. This study aimed to examine the hypothesis that insecticides used outside of public health may play a role in selection. Briefly, the resistance mutation frequencies to three insecticide groups (pyrethroids, organophosphates, and cyclodienes) were estimated in two representative groups of mosquito species (Anopheles Hyrcanus Group and Culex pipiens complex). The relationship between these frequencies and the land-use status of the collection sites was investigated through multiple regression analysis. In the Anopheles Hyrcanus Group, the frequencies of both ace1 (organophosphate resistance) and rdl (cyclodiene resistance) mutations were positively correlated with 'proximity to golf course', possibly be due to the insecticides used for turf maintenance. They also showed positive correlations with field area and rice paddy area, respectively, suggesting the role of agricultural insecticides in the selection of these resistance traits. For the Cx. pipiens complex, the kdr (pyrethroid resistance), ace1, and rdl mutations were positively correlated with the residential area, field, and rice paddy, respectively. Therefore, pyrethroids used for public health could serve as a direct source of resistance selection pressure against kdr, whereas non-public health insecticides may pose primary selection pressure against the ace1 and rdl traits. The current findings suggest that the insecticides used in agriculture and the golf industry play a significant role in mosquito selection, despite variations in the extent of indirect selection pressure according to the mosquito groups and insecticide classes.

Potential for urban warming to postpone overwintering dormancy of temperate mosquitoes.

Cities are generally hotter than surrounding rural areas due to the Urban Heat Island (UHI) effect. These increases in temperature advance plant and animal phenology, development, and reproduction in the spring. However, research determining how increased temperatures affect the seasonal physiology of animals in the fall has been limited. The Northern house mosquito, Culex pipiens, is abundant in cities and transmits several pathogens including West Nile virus. Females of this species enter a state of developmental arrest, or reproductive diapause, in response to short days and low temperatures during autumn. Diapausing females halt reproduction and blood-feeding, and instead accumulate fat and seek sheltered overwintering sites. We found that exposure to increased temperatures in the lab that mimic the UHI effect induced ovarian development and blood-feeding, and that females exposed to these temperatures were as fecund as non-diapausing mosquitoes. We also found that females exposed to higher temperatures had lower survival rates in winter-like conditions, despite having accumulated equivalent lipid reserves relative to their diapausing congeners. These data suggest that urban warming may inhibit diapause initiation in the autumn, thereby extending the active biting season of temperate mosquitoes.

Transcriptome profiling reveals sex-specific gene expressions in pupal and adult stages of the mosquito Culex pipiens.

Understanding the development process of male and female mosquitoes provides important basic information for sterile insect release programmes and is important for improving other vector control strategies. However, little is known about the molecular mechanisms that distinguish male from female-specific developmental processes in this species. We used IlluminaRNA-seq to identify sex-specific genes during pupal and adult stages. One hundred and forty-seven genes were expressed only in pupal males, 56 genes were expressed in adult males and another 82 genes were commonly expressed in both male samples. In addition, 26 genes were expressed only in the pupal females, 163 genes were found in the adult females and only one gene was expressed in both female samples. A further quantitative real-time PCR validation of selected genes from the RNA sequencing (RNA-seq) analysis confirmed upregulation of those genes in a sex-specific manner, including: fibrinogen and fibronectin, a zinc finger protein, phospholipase A(2) and a serine protein for female pupae; venom allergen 3, a perlecan, testis-specific serine/threonine-protein kinase 1, testis-specific serine/threonine-protein kinase 6 and cytochrome c-2 for male pupae; a salivary protein, D7 protein precursor, trypsin 7 precursor, D7 protein and nanos for female adults; and tetraspanin F139, cytosol aminopeptidase, testis-specific serine/threonine-protein kinase 1, a testis-specific serine/threonine-protein kinase 6 and a C-type lectin for male adults. These findings provide insight into the development and physiology of Culex mosquitoes, which will help in the development of more effective control methods for these disease vectors.