Mutations and intron polymorphisms in voltage-gated sodium channel genes of different geographic populations of Culex pipiens pallens/Culex pipiens quinquefasciatus in China.

BACKGROUND: Culex pipiens pallens and Culex pipiens quinquefasciatus are the dominant species of Culex mosquitoes in China and important disease vectors. Long-term use of insecticides can cause mutations in the voltage-gated sodium channel (vgsc) gene of mosquitoes, but little is known about the current status and evolutionary origins of vgsc gene in different geographic populations. Therefore, this study aimed to determine the current status of vgsc genes in Cx. p. pallens and Cx. p. quinquefasciatus in China and to investigate the evolutionary inheritance of neighboring downstream introns of the vgsc gene to determine the impact of insecticides on long-term evolution. METHODS: Sampling was conducted from July to September 2021 in representative habitats of 22 provincial-level administrative divisions in China. Genomic DNA was extracted from 1308 mosquitoes, the IIS6 fragment of the vgsc gene on the nerve cell membrane was amplified using polymerase chain reaction, and the sequence was used to evaluate allele frequency and knockdown resistance (kdr) frequency. MEGA 11 was used to construct neighbor-joining (NJ) tree. PopART was used to build a TCS network. RESULTS: There were 6 alleles and 6 genotypes at the L1014 locus, which included the wild-type alleles TTA/L and CTA/L and the mutant alleles TTT/F, TTC/F, TCT/S and TCA/S. The geographic populations with a kdr frequency less than 20.00% were mainly concentrated in the regions north of 38° N, and the geographic populations with a kdr frequency greater than 80.00% were concentrated in the regions south of 30° N. kdr frequency increased with decreasing latitude. And within the same latitude, the frequency of kdr in large cities is relatively high. Mutations were correlated with the number of introns. The mutant allele TCA/S has only one intron, the mutant allele TTT/F has three introns, and the wild-type allele TTA/L has 17 introns. CONCLUSIONS: Cx. p. pallens and Cx. p. quinquefasciatus have developed resistance to insecticides in most regions of China. The neighboring downstream introns of the vgsc gene gradually decreased to one intron with the mutation of the vgsc gene. Mutations may originate from multiple mutation events rather than from a single origin, and populations lacking mutations may be genetically isolated.

Mosquitoes in urban green spaces and cemeteries in northern Spain.

BACKGROUND: Mosquitoes inhabiting urban green spaces and cemeteries in Europe represent a crucial facet of public health concern and contribute to the ecological balance. As urbanization intensifies, these areas increasingly serve as vital habitats for various mosquito species, fostering breeding grounds and increasing the risk of disease transmission. METHODS: A study was conducted in the three main cities (inland, coastal, and estuarine) of the Basque Country, northern Spain, to investigate the species composition, abundance, dynamic populations, larval habitats, and host preferences of mosquitoes in urban green spaces and cemeteries. CDC traps and dipping were used to collect mosquitoes for 2 years (2019-2020). RESULTS: A total of 21 mosquito species were identified, with Culex pipiens s.l. being the most abundant and widespread. The three ecological forms of Cx. pipiens were found, and Cx. pipiens pipiens was the most common in both green areas and cemeteries. Morphological identification together with molecular tools identified 65 COI sequences with high homology. The highest species richness was found in the inland city, followed by the coastal city and the estuarine city. Mosquito abundance was significantly higher in green areas compared to cemeteries and in the coastal and estuarine cities compared to the inland city. The investigation of larval breeding sites highlighted the dominance of Cx. pipiens s.l., particularly in semi-artificial ponds, diverse water-holding containers (tyres and buckets) and drainage systems in green areas; in cemeteries, most of the larvae were found in flowerpots and funerary urns. Seasonal activity exhibited variable peaks in mosquito abundance in the different cities, with a notable increase in July or August. Additionally, blood meal analysis revealed that Cx. pipiens s.l. fed on several common urban avian species. CONCLUSIONS: Studies on mosquitoes are essential to understand their role in disease transmission and to design targeted and sustainable management strategies to mitigate the associated risks.

Combining transinfected Wolbachia and a genetic sexing strain to control Aedes albopictus in laboratory-controlled conditions.

The global expansion of Aedes albopictus has stimulated the development of environmentally friendly methods aiming to control disease transmission through the suppression of natural vector populations. Sterile male release programmes are currently being deployed worldwide, and are challenged by the availability of an efficient sex separation which can be achieved mechanically at the pupal stage and/or by artificial intelligence at the adult stage, or through genetic sexing, which allows separating males and females at an early development stage. In this study, we combined the genetic sexing strain previously established based on the linkage of dieldrin resistance to the male locus with a Wolbachia transinfected line. For this, we introduced either the wPip-I or the wPip-IV strain from Culex pipiens in an asymbiotic Wolbachia-free Ae. albopictus line. We then measured the penetrance of cytoplasmic incompatibility and life-history traits of both transinfected lines, selected the wPip-IV line and combined it with the genetic sexing strain. Population suppression experiments demonstrated a 90% reduction in population size and a 50% decrease in hatching rate. Presented results showed that such a combination has a high potential in terms of vector control but also highlighted associated fitness costs, which should be reduced before large-scale field assay.

Oxidative stress induction by essential oil from Piper alatipetiolatum (Piperaceae) triggers lethality in the larvae of Culex quinquefasciatus (Diptera: Culicidae).

Culex quinquefasciatus is the main vector of lymphatic filariasis in Brazil, which present resistance to commercial insecticides. Nowadays, essential oils (EOs) exhibiting larvicidal activity, such as those derived from Piper alatipetiolatum, provide a promising alternative for vector control, including Culex species. This study aimed to investigate the larvicidal activity and the oxidative stress indicators of the EO from P. alatipetiolatum in Cx. quinquefasciatus larvae. The EO was extracted from P. alatipetiolatum leaves using the hydrodistillation method, resulting in a yield of 7.2 ± 0.1%, analysed by gas chromatography coupled with spectrometry and gas chromatography coupled with flame ionization detector (GC-MS and GC-FID), and evaluated against Cx. quinquefasciatus larvae. Reactive Oxygen and Nitrogen Species (RONS), Catalase (CAT), glutathione-S-transferase (GST), acetylcholinesterase (AChE), and Thiol levels were used as oxidative stress indicators. Analysis by CG-MS and CG-FID revealed that the main compound in the EO was the oxygenated sesquiterpene ishwarone, constituting 78.6% of the composition. Furthermore, the EO exhibited larvicidal activity, ranging from 26 to 100%, with an LC50 of 4.53 µg/mL and LC90 of 15.37 µg/mL. This activity was accompanied by a significant increase in RONS production, alterations in CAT, GST, AChE activity, and thiol levels compared to the control groups (p < 0.05). To the best of our knowledge, this is the first report describing the larvicidal activity and oxidative stress induced by the EO from P. alatipetiolatum against Cx. quinquefasciatus larvae. Therefore, we propose that this EO shows promise as larvicidal agent for the effective control of Cx. quinquefasciatus larvae.

[Comparison of the microbiota diversity between autogenous and anautogenous Culex pipiens pallens].

OBJECTIVE: To investigate the microbiota composition and diversity between autogenous and anautogenous Culex pipiens pallens, so as to provide insights into unraveling the pathogenesis of autogeny in Cx. pipiens pallens. METHODS: Autogenous and anautogenous adult Cx. pipiens pallens samples were collected at 25 ℃, and the hypervariable regions of the microbial 16S ribosomal RNA (16S rRNA) gene was sequenced on the Illumina NovaSeq 6000 sequencing platform. The microbiota abundance and diversity were evaluated using the alpha diversity index, and the difference in the microbiota structure was examined using the beta diversity index. The microbiota with significant differences in the abundance between autogenous and anautogenous adult Cx. pipiens pallens samples was identified using the linear discriminant analysis effect size (LEfSe). RESULTS: The microbiota in autogenous and anautogenous Cx. pipiens pallens samples belonged to 18 phyla, 28 classes, 70 orders, 113 families, and 170 genera, and the dominant phyla included Proteobacteria, Bacteroidetes, and so on. At the genus level, Wolbachia was a common dominant genus, and the relative abundance was (77.6 ± 11.3)% in autogenous Cx. pipiens pallens samples and (47.5 ± 8.5)% in anautogenous mosquito samples, while Faecalibaculum (0.4% ± 0.1%), Dubosiella (0.5% ± 0.0%) and Massilia (0.5% ± 0.1%) were specific species in autogenous Cx. pipiens pallens samples. Alpha diversity analysis showed that higher Chao1 index and ACE index in autogenous Cx. pipiens pallens samples than in anautogenous samples (both P values > 0.05), and lower Shannon index (P > 0.05) and Simpson index (P < 0.05) in autogenous Cx. pipiens pallens samples than in anautogenous samples. LEfSe analysis showed a total of 48 significantly different taxa between autogenous and anautogenous Cx. pipiens pallens samples (all P values < 0.05). CONCLUSIONS: There is a significant difference in the microbiota diversity between autogenous and anautogenous Cx. pipiens pallens.

The effect of physical barriers under a raised house on mosquito entry: an experimental study in rural Gambia.

BACKGROUND: Anopheles gambiae, the major malaria mosquito in sub-Saharan Africa, feed largely indoors at night. Raising a house off the ground with no barriers underneath reduces mosquito-house entry. This experiment tested whether walling off the space under an elevated hut affects mosquito-hut entry. METHODS: Four inhabited experimental huts, each of which could be moved up and down, were used in rural Gambia. Nightly collections of mosquitoes were made using light traps and temperature and carbon dioxide levels monitored indoors and outdoors using loggers. Each night, a reference hut was kept at ground level and three huts raised 2 m above the ground; with the space under the hut left open, walled with air-permeable walls or solid walls. Treatments were rotated every four nights using a randomized block design. The experiment was conducted for 32 nights. Primary measurements were mosquito numbers and indoor temperature in each hut. RESULTS: A total of 1,259 female Anopheles gambiae sensu lato were collected in the hut at ground level, 655 in the hut with an open ground floor, 981 in the hut with air-permeable walls underneath and 873 in the hut with solid walls underneath. Multivariate analysis, adjusting for confounders, showed that a raised hut open underneath had 53% fewer mosquitoes (95% CI 47-58%), those with air-permeable walls underneath 24% fewer (95% CI 9-36%) and huts with solid walls underneath 31% fewer (95% CI 24-37%) compared with a hut on the ground. Similar results were found for Mansonia spp. and total number of female mosquitoes, but not for Culex mosquitoes where hut entry was unaffected by height or barriers. Indoor temperature and carbon dioxide levels were similar in all huts. CONCLUSION: Raising a house 2 m from the ground reduces the entry of An. gambiae and Mansonia mosquitoes, but not Culex species. The protective effect of height is reduced if the space underneath the hut is walled off.

Reducing the Antigen Prevalence Target Threshold for Stopping and Restarting Mass Drug Administration for Lymphatic Filariasis Elimination: A Model-Based Cost-effectiveness Simulation in Tanzania, India and Haiti.

BACKGROUND: The Global Programme to Eliminate Lymphatic Filariasis (GPELF) aims to reduce and maintain infection levels through mass drug administration (MDA), but there is evidence of ongoing transmission after MDA in areas where Culex mosquitoes are the main transmission vector, suggesting that a more stringent criterion is required for MDA decision making in these settings. METHODS: We use a transmission model to investigate how a lower prevalence threshold (<1% antigenemia [Ag] prevalence compared with <2% Ag prevalence) for MDA decision making would affect the probability of local elimination, health outcomes, the number of MDA rounds, including restarts, and program costs associated with MDA and surveys across different scenarios. To determine the cost-effectiveness of switching to a lower threshold, we simulated 65% and 80% MDA coverage of the total population for different willingness to pay per disability-adjusted life-year averted for India ($446.07), Tanzania ($389.83), and Haiti ($219.84). RESULTS: Our results suggest that with a lower Ag threshold, there is a small proportion of simulations where extra rounds are required to reach the target, but this also reduces the need to restart MDA later in the program. For 80% coverage, the lower threshold is cost-effective across all baseline prevalences for India, Tanzania, and Haiti. For 65% MDA coverage, the lower threshold is not cost-effective due to additional MDA rounds, although it increases the probability of local elimination. Valuing the benefits of elimination to align with the GPELF goals, we find that a willingness to pay per capita government expenditure of approximately $1000-$4000 for 1% increase in the probability of local elimination would be required to make a lower threshold cost-effective. CONCLUSIONS: Lower Ag thresholds for stopping MDAs generally mean a higher probability of local elimination, reducing long-term costs and health impacts. However, they may also lead to an increased number of MDA rounds required to reach the lower threshold and, therefore, increased short-term costs. Collectively, our analyses highlight that lower target Ag thresholds have the potential to assist programs in achieving lymphatic filariasis goals.

Effect of Mosquitocidal, histopathological alteration and non target effects of Sigesbeckia orientalis L. on Anopheles stephensi Liston, Culex quinquefasciatus say and Aedes aegypti L.

Diseases transmitted by vectors have a significant collision on society and community health, particularly in tropical and subtropical regions, where they can cause large-scale outbreaks shortly after initial transmission. The intent of this investigation was to study the plant extract derived from Sigesbeckia orientalis L. in controlling the immature stages of Anopheles, Culex and Aedes mosquitoes, while also considering its potential toxicity to ecosystems. The immature stages were exposed to different extracts (62.5-500 ppm), and the mortality of larvae and pupae, as well as ovicidal activity, were noted after 24 and 120 h of the experiment. The hexane and ethyl aceate extract of S. orientalis presented 100% ovicidal activity against the eggs of Anopheles, Aedes and Culex at 500 ppm concentration after 5 days of treatment. The hexane and ethylacetate extracts presented strong larvicidal activity with LC50 values of 215.7, 332.0, 197.4 and 212.6, 694.9 and 201.7 ppm against treated mosquitoes at 24 h, respectively. The same extract also presented promising pupicidal activity. The LC50 values of hexane extract were 219.6, 353.6, 194.2 and LC50 values of ethyl acetate were 257.6, 387.8 and 259.07 ppm against early stage pupae of three vector mosquitoes, respectively. The extracts from S. orientalis had strong inhibitory activity against growth and development of mosquitoes. SI/PSF values showed that the extracts of S. orientalis did not harm Poecilia reticulata, Diplonychus indicus (Water bug), Gambusia affinis and dragon fly nymph at tested concentrations. Furthermore, examinations of histopathology and growth disruption revealed significant damage to the midgut cells in the treated larvae. The formulations utilizing hexane and ethyl acetate extracts exhibited potent activity without posing any toxicity towards non-target organisms. This study clearly indicated that hexane and ethylacetate extracts showed promising results against treated mosquitoes. The present study documents the first report of the extracts from S. orientalis and they can be further assessed to identify compounds for application purposes.

Year-round Observations of Adult Culex Mosquitoes in Storm Sewer Pipes in the Northwestern Chicago Suburbs.

Over the course of three years, 200 ft to 0.75 mi (60 m to 1.2 km) sections of 3 larger (>6 ft [1.8 m] diam) belowground storm sewer conveyance pipes in the northwestern Chicago suburbs were inspected for the presence of adult mosquitoes. Culex mosquitoes were by far the most common (555 of 556 [99.8%] total mosquitoes) collected within pipes during all four meteorological seasons (i.e. during months of October, January, May, August). These observations support prior work elsewhere, suggesting storm sewer pipes are consistent sites of refuge for adult Culex mosquitoes.

Evaluation of the mosquitocidal efficacy of fluralaner, a potential candidate for drug based vector control.

Vector control is a key intervention against mosquito borne diseases. However, conventional methods have several limitations and alternate strategies are in urgent need. Vector control with endectocides such as ivermectin is emerging as a novel strategy. The short half-life of ivermectin is a limiting factor for its application as a mass therapy tool for vector control. Isoxazoline compounds like fluralaner, a class of veterinary acaricides with long half-life hold promise as an alternative. However, information about their mosquitocidal effect is limited. We explored the efficacy of fluralaner against laboratory reared vector mosquitoes-Aedes aegypti, Anopheles stephensi, and, Culex quinquefasciatus. 24 h post-blood feeding, fluralaner showed a significant mosquitocidal effect with LC50 values in the range of 24.04-49.82 ng/mL for the three different mosquito species tested. Effects on life history characteristics (fecundity, egg hatch success, etc.) were also observed and significant effects were noted at drug concentrations of 20, 25 and 45 ng/mL for Ae. aegypti, An. stephensi, and, Cx. quinquefasciatus respectively. At higher drug concentration of 250 ng/mL, significant mortality was observed within 1-2 h of post blood feeding. Potent mosquitocidal effect coupled with its long half-life makes fluralaner an excellent candidate for drug based vector control strategies.

Evaluation of vector susceptibility in Aedes aegypti and Culex pipiens pallens to Tibet orbivirus.

Mosquito-borne viruses cause various infectious diseases in humans and animals. Tibet orbivirus (TIBOV), a newly identified arbovirus, efficiently replicates in different types of vertebrate and mosquito cells, with its neutralizing antibodies detected in cattle and goats. However, despite being isolated from Culicoides midges, Anopheles, and Culex mosquitoes, there has been a notable absence of systematic studies on its vector competence. Thus, in this study, Aedes aegypti and Culex pipiens pallens were reared in the laboratory to measure vector susceptibility through blood-feeding infection. Furthermore, RNA sequencing was used to examine the overall alterations in the Ae. aegypti transcriptome following TIBOV infection. The results revealed that Ae. aegypti exhibited a high susceptibility to TIBOV compared to Cx. p. pallens. Effective replication of the virus in Ae. aegypti midguts occurred when the blood-feeding titer of TIBOV exceeded 105 plaque-forming units mL-1. Nevertheless, only a few TIBOV RNA-positive samples were detected in the saliva of Ae. aegypti and Cx. p. pallens, suggesting that these mosquito species may not be the primary vectors for TIBOV. Moreover, at 2 dpi of TIBOV, numerous antimicrobial peptides downstream of the Toll and Imd signaling pathways were significantly downregulated in Ae. aegypti, indicating that TIBOV suppressed mosquitos' defense to survive in the vector at an early stage. Subsequently, the stress-activated protein kinase JNK, a crucial component of the MAPK signaling pathway, exhibited significant upregulation. Certain genes were also enriched in the MAPK signaling pathway in TIBOV-infected Ae. aegypti at 7 dpi.IMPORTANCETibet orbivirus (TIBOV) is an understudied arbovirus of the genus Orbivirus. Our study is the first-ever attempt to assess the vector susceptibility of this virus in two important mosquito vectors, Aedes aegypti and Culex pipiens pallens. Additionally, we present transcriptome data detailing the interaction between TIBOV and the immune system of Ae. aegypti, which expands the knowledge about orbivirus infection and its interaction with mosquitoes.

Nanoparticles-mediated entomotoxicology: lessons from biologica.

Nanotechnology has grown in importance in medicine, manufacturing, and consumer products. Nanoparticles (NPs) are also widely used in the field of insect pest management, where they show a variety of toxicological effects on insects. As a result, the primary goal of this review is to compile and evaluate available information on effects of NPs on insects, by use of a timely, bibliometric analysis. We also discussed the manufacturing capacity of NPs from insect tissues and the toxic effects of NPs on insects. To do so, we searched the Web of Science database for literature from 1995 to 2023 and ran bibliometric analyses with CiteSpace© and Bibliometrix©. The analyses covered 614 journals and identified 1763 relevant documents. We found that accumulation of NPs was one of the top trending topics. China, India, and USA had the most published papers. The most overall reported models of insects were those of Aedes aegypti (yellow fever mosquito), Culex quinquefasciatus (southern house mosquito), Bombyx mori (silk moth), and Anopheles stephensi (Asian malaria mosquito). The application and methods of fabrication of NPs using insect tissues, as well as the mechanism of toxicity of NPs on insects, were also reported. A uniform legal framework is required to allow nanotechnology to fully realize its potential while minimizing harm to living organisms and reducing the release of toxic metalloid nanoparticles into the environment.

Metagenomic sequencing reveals viral diversity of mosquitoes from Shandong Province, China.

Mosquitoes carry a large number of known and unknown viruses, some of which could cause serious diseases in humans or animals. Metagenomic sequencing for mosquito viromes is crucial for understanding the evolutionary history of viruses and preventing emerging mosquito-borne diseases. We collected 1,598 mosquitoes belonging to four species from five counties in Shandong Province, China in 2021. They were grouped by species and sampling locations and subjected to metagenomic next-generation sequencing for the analysis of the viromes. A total of 233,317,352 sequencing reads were classified into 30 viral families and an unclassified group. Comparative analysis showed that mosquitoes in Shandong Province generally possessed host-specific virome. We detected mosquito-borne viruses including Japanese encephalitis virus, Getah virus, and Kadipiro virus in Culex tritaeniorhynchus and Anopheles sinensis samples. Phylogenetic analysis showed that these pathogenic viruses may have existed in mosquitoes in Shandong Province for a long time. Meanwhile, we identified 22 novel viruses belonging to seven families and the genus Negevirus. Our study comprehensively described the viromes of several common mosquito species in Shandong Province, China, and demonstrated the major role of host species in shaping mosquito viromes. Furthermore, the metagenomic data provided valuable epidemiological information on multiple mosquito-borne viruses, highlighting the potential risk of infection transmission. IMPORTANCE: Mosquitoes are known as the source of various pathogens for humans and animals. Culex tritaeniorhynchus, Armigeres subalbatus, and Anopheles sinensis have been found to transmit the Getah virus, which has recently caused increasing infections in China. Cx. tritaeniorhynchus and Culex pipiens are the main vectors of Japanese encephalitis virus and have caused epidemics of Japanese encephalitis in China in past decades. These mosquitoes are widely present in Shandong Province, China, leading to a great threat to public health and the breeding industry. This study provided a comprehensive insight into the viromes of several common mosquito species in Shandong Province, China. The metagenomic sequencing data revealed the risks of multiple pathogenic mosquito-borne viruses, including Japanese encephalitis virus, Getah virus, and Kadipiro virus, which are of great importance for preventing emerging viral epidemics.

Structural and functional comparisons of salivary α-glucosidases from the mosquito vectors Aedes aegypti, Anopheles gambiae, and Culex quinquefasciatus.

Mosquito vectors of medical importance both blood and sugar feed, and their saliva contains bioactive molecules that aid in both processes. Although it has been shown that the salivary glands of several mosquito species exhibit α-glucosidase activities, the specific enzymes responsible for sugar digestion remain understudied. We therefore expressed and purified three recombinant salivary α-glucosidases from the mosquito vectors Aedes aegypti, Anopheles gambiae, and Culex quinquefasciatus and compared their functions and structures. We found that all three enzymes were expressed in the salivary glands of their respective vectors and were secreted into the saliva. The proteins, as well as mosquito salivary gland extracts, exhibited α-glucosidase activity, and the recombinant enzymes displayed preference for sucrose compared to p-nitrophenyl-α-D-glucopyranoside. Finally, we solved the crystal structure of the Ae. aegypti α-glucosidase bound to two calcium ions at a 2.3 Ångstrom resolution. Molecular docking suggested that the Ae. aegypti α-glucosidase preferred di- or polysaccharides compared to monosaccharides, consistent with enzymatic activity assays. Comparing structural models between the three species revealed a high degree of similarity, suggesting similar functional properties. We conclude that the α-glucosidases studied herein are important enzymes for sugar digestion in three mosquito species.

Landscape level associations between birds, mosquitoes and microclimates: possible consequences for disease transmission?

BACKGROUND: Mosquito-borne diseases are on the rise. While climatic factors have been linked to disease occurrences, they do not explain the non-random spatial distribution in disease outbreaks. Landscape-related factors, such as vegetation structure, likely play a crucial but hitherto unquantified role. METHODS: We explored how three critically important factors that are associated with mosquito-borne disease outbreaks: microclimate, mosquito abundance and bird communities, vary at the landscape scale. We compared the co-occurrence of these three factors in two contrasting habitat types (forest versus grassland) across five rural locations in the central part of the Netherlands between June and September 2021. RESULTS: Our results show that forest patches provide a more sheltered microclimate, and a higher overall abundance of birds. When accounting for differences in landscape characteristics, we also observed that the number of mosquitoes was higher in isolated forest patches. CONCLUSIONS: Our findings indicate that, at the landscape scale, variation in tree cover coincides with suitable microclimate and high Culex pipiens and bird abundance. Overall, these factors can help understand the non-random spatial distribution of mosquito-borne disease outbreaks.

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.

Short Communication: Mosquito Histone 2A Protein Facilitate Japanese Encephalitis Virus Infection in the Mosquito.

Japanese encephalitis virus is mainly prevalent in the tropical and subtropical regions of Asia and Oceania. Through immunoprecipitation-mass spectrometry analysis using monoclonal antibodies targeting JEV E protein, we found that mosquito Histone 2A protein could bind to JEV particles. The binding of H2A and JEV was detected in the salivary gland and supernatant of mosquito cells. Furthermore, RNA interference experiments in vitro and in vivo confirmed that H2A protein promotes JEV infection in mosquitoes. In summary, we found that mosquito H2A is a factor that supports JEV infection and can potentially facilitate cross-species transmission of JEV.

Effects of soil on the development, survival, and oviposition of Culex quinquefasciatus (Diptera: Culicidae) mosquitoes.

BACKGROUND: Water quality is known to influence the development and survival of larval mosquitoes, which affects mosquito-borne pathogen transmission as a function of the number of mosquitoes that reach adulthood and blood feed. Although water properties are known to affect mosquito development, few studies have investigated the link among soil properties, water quality, and mosquito development. Given the large number of ground-breeding mosquito species, this linkage is a potentially important factor to consider in mosquito ecology. In this study, we explored the effects of different soils on multiple life history parameters of the ground-breeding mosquito species Culex quinquefasciatus (Diptera: Culicidae). METHODS: Cx. quinquefasciatus larvae were reared in water combined with different soil substrates (sandy, silt, or clay loam textures) at increasing soil to water volume ratios, with and without the addition of organic matter (fish food). Gravid mosquitoes were offered different soil-water extracts to investigate soil effects on oviposition preference. RESULTS: Without the addition of organic matter, larval survival and development differed significantly among waters with different soil textures and volumes of substrate. Mosquitoes in water with clay loam soil survived longer and developed further than mosquitoes in other soil waters. Larvae survived for longer periods of time with increased volumes of soil substrate. Adding organic matter reduced the differences in larval survival time, development, and pupation among soil-water extracts. Adult female mosquitoes oviposited more frequently in water with clay loam soil, but the addition of organic matter reduced the soil effects on oviposition preference. CONCLUSIONS: This study suggests soil composition affects larval mosquito survival and development, as well as the oviposition preference of gravid females. Future studies could differentiate abiotic and biotic soil features that affect mosquitoes and incorporate soil variation at the landscape scale into models to predict mosquito population dynamics and mosquito-borne pathogen transmission.

Synthesis of eco-friendly layered double hydroxide and nanoemulsion for jasmine and peppermint oils and their larvicidal activities against Culex pipiens Linnaeus.

Mosquito-borne diseases represent a growing health challenge over time. Numerous potential phytochemicals are target-specific, biodegradable, and eco-friendly. The larvicidal activity of essential oils, a jasmine blend consisting of Jasmine oil and Azores jasmine (AJ) (Jasminum sambac and Jasminum azoricum) and peppermint (PP) Mentha arvensis and their nanoformulations against 2nd and 4th instar larvae of Culex pipiens, was evaluated after subjecting to different concentrations (62.5, 125, 250, 500, 1000, and 2000 ppm). Two forms of phase-different nanodelivery systems of layered double hydroxide LDH and oil/water nanoemulsions were formulated. The synthesized nanoemulsions showed particle sizes of 199 and 333 nm for AJ-NE and PP-NE, with a polydispersity index of 0.249 and 0.198, respectively. Chemical and physiochemical analysis of TEM, SEM, XRD, zeta potential, drug loading capacity, and drug release measurements were done to confirm the synthesis and loading efficiencies of essential oils' active ingredients. At high concentrations of AJ and PP nanoemulsions (2000 ppm), O/W nanoemulsions showed higher larval mortality than both LDH conjugates and crude oils. The mortality rate reached 100% for 2nd and 4th instar larvae. The relative toxicities revealed that PP nanoemulsion (MA-NE) was the most effective larvicide, followed by AJ nanoemulsion (AJ-NE). There was a significant increase in defensive enzymes, phenoloxidase, and α and ß-esterase enzymes in the treated groups. After treatment of L4 with AJ, AJ-NE, PP, and PP-NE, the levels of phenoloxidase were 545.67, 731.00, 700.00, and 799.67 u/mg, respectively, compared with control 669.67 u/mg. The activity levels of α-esterase were 9.71, 10.32, 8.91, and 10.55 mg α-naphthol/min/mg protein, respectively. It could be concluded that the AJ-NE and PP-NE nanoformulations have promising larvicidal activity and could act as safe and effective alternatives to chemical insecticides.

New insight into avian malaria vectors in New Zealand.

BACKGROUND: Mosquitoes (Culicidae) are vectors for most malaria parasites of the Plasmodium species and are required for Plasmodium spp. to complete their life cycle. Despite having 16 species of mosquitoes and the detection of many Plasmodium species in birds, little is known about the role of different mosquito species in the avian malaria life cycle in New Zealand. METHODS: In this study, we used nested polymerase chain reaction (PCR) and real-time PCR to determine Plasmodium spp. prevalence and diversity of mitochondrial cytochrome b gene sequences in wild-caught mosquitoes sampled across ten sites on the North Island of New Zealand during 2012-2014. The mosquitoes were pooled by species and location collected, and the thorax and abdomens were examined separately for Plasmodium spp. DNA. Akaike information criterion (AIC) modeling was used to test whether location, year of sampling, and mosquito species were significant predictors of minimum infection rates (MIR). RESULTS: We collected 788 unengorged mosquitoes of six species, both native and introduced. The most frequently caught mosquito species were the introduced Aedes notoscriptus and the native Culex pervigilans. Plasmodium sp DNA was detected in 37% of matched thorax and abdomen pools. When considered separately, 33% of abdomen and 23% of thorax pools tested positive by nested PCR. The MIR of the positive thorax pools from introduced mosquito species was 1.79% for Ae. notoscriptus and 0% for Cx. quinquefasciatus, while the MIR for the positive thorax pools of native mosquito species was 4.9% for Cx. pervigilans and 0% for Opifex fuscus. For the overall MIR, site and mosquito species were significant predictors of Plasmodium overall MIR. Aedes notoscriptus and Cx. pervigilans were positive for malaria DNA in the thorax samples, indicating that they may play a role as avian malaria vectors. Four different Plasmodium lineages (SYAT05, LINN1, GRW6, and a new lineage of P (Haemamoeba) sp. AENOT11) were identified in the pooled samples. CONCLUSIONS: This is the first detection of avian Plasmodium DNA extracted from thoraxes of native Culex and introduced Aedes mosquito species in New Zealand and therefore the first study providing an indication of potential vectors in this country.