House Mosquito Culex Quinquefasciatus Research Articles

Prevention of resistance to spinosad by a combination of spinosad and Bacillus thuringiensis subsp. israelensis in Culex quinquefasciatus (Diptera: Culicidae).

Spinosad consists of spinosyn A and spinosyn D that are produced by the soil-dwelling actinomycete Saccharopolyspora spinosa. It has been used to control a wide variety of arthropod pests of economic importance. Formulations of spinosad have been used to control larval mosquitoes since approximately 2010. However, the target site- and metabolism-based resistance to this neurotoxin has been reported since 2000 in agricultural pests The current studies aim to further evaluate resistance risk and develop resistance prevention tactics in the southern house mosquito Culex quinquefasciatus. The first comparison group involved selection at lethal concentration, 75% (LC75) of technical spinosad for 30 generations, resulting in baseline resistance ratios (RRs) of 51.1-fold at LC50 and 45.4-fold at LC90 in Cx. quinquefasciatus. However, under the same conditions, selection by a combination of spinosad and Bacillus thuringiensis subsp. israelensis (B.t.i.), negated resistance development to spinosad, RRs ranging 1.00-1.75-fold at LC50 and 0.83-1.76-fold at LC90. At the same time, the selected population remained susceptible to the combination throughout the selection process, RRs fluctuating 0.74-1.38-fold at LC50 and 0.63-1.23-fold at LC90. Combination of spinosad and B.t.i. negates resistance development to spinosad, as opposed to spinosad alone in Cx. quinquefasciatus. Moreover, the species tested does not develop resistance to this combination upon repeated exposures, implying the potential for further developing this combination as a viable product for larval mosquito control. © 2024 Society of Chemical Industry.

Dopamine modulates odour learning in mosquitoes

Researchers have long been fascinated with figuring out how mosquitoes choose a host to feed on and how they can change their preferences. One important missing piece of the puzzle may be the way that mosquitoes learn different smells. Gabriella Wolff and colleagues from the University of Washington, USA, had previously found that yellow fever mosquitoes could form memories associated with odours related to their favourite food – blood – but not with odours that had nothing to do with their life history. These mosquitoes could be trained to avoid those odours by associating them with swatting. However, it was still unknown whether other mosquito species could learn to avoid the same odours and whether dopamine, a chemical messenger important for learning, could be responsible for this olfactory learning. To figure this out, the team set out to train four different species of mosquitoes.Wolff and colleagues exposed the mosquitoes to the scents of human skin, chicken feathers and rose oil. Two of the mosquito species feed on humans [the yellow fever mosquito (Aedes aegypti) and the Asian malaria mosquito (Anopheles stephensi)], another, the southern house mosquito (Culex quinquefasciatus), feeds on birds, and the elephant mosquito (Toxorhynchites amboinensis) dines on flowers. The team found that all the mosquito species had the strongest attraction to the scent of the hosts that they preferred to feed on.To test whether multiple mosquito species could learn to recognize the same odours, the team trained female mosquitoes from each species to avoid different scents by exposing them to scent components from human odours (octenol and hexanoic acid) or a flower scent component (linalool), regardless of whether they naturally feed on humans or flowers, and then they mimicked swatting at the insects to make the scent unattractive. The team then investigated each mosquito’s preference by putting them into a maze that provided them with a choice between either a host odour (octenol, hexanoic acid or linalool) or no scent. The team found that the mosquitoes learned to avoid the odours associated with their favourite food, but not other odours. For example, the yellow fever mosquito, which dines on people, learned to avoid octenol and hexanoic acid, but not linalool. Meanwhile, nectar-feeding elephant mosquitoes only learned to avoid linalool, but not octenol and hexanoic acid.To see whether these differences in species learning were due to the effects of dopamine, the researchers investigated where dopamine was localized in the brain of each of these species. Wolff and colleagues found that dopamine was localized in the antennal lobe and mushroom bodies – structures in the brain that are important for learning and memory. However, the dopamine was localized in different areas of these structures from species to species, which could be important for differences in their learning abilities. To see whether dopamine was necessary for this olfactory learning, the team also trained yellow fever mosquitoes and the Asian malaria mosquitoes, which feed on mammals, to avoid an odour associated with their preferred diets, and found that when they blocked the dopamine receptor, they eliminated their ability to learn to avoid the odour. Last, the team found that areas in the antennal lobe that had an increased response to these scents also had a raised level of dopamine.Learning the basis of how mosquitoes smell and the impact on their host preference is vital, as the insects can be responsible for the spread of some terrible diseases. The researchers highlight that being able to change dopamine inputs without having to change brain structure may give mosquitoes an evolutionary advantage and could be a way for mosquitoes to change their host preferences quickly.

A mass rearing cost calculator for the control of Culex quinquefasciatus in Hawaiʻi using the incompatible insect technique

BackgroundHawaiʻi’s native forest avifauna is experiencing drastic declines due to climate change-induced increases in temperature encroaching on their upper-elevation montane rainforest refugia. Higher temperatures support greater avian malaria infection rates due to greater densities of its primary vector, the southern house mosquito Culex quinquefasciatus, and enhance development of the avian malaria parasite Plasmodium relictum. Here we propose the use of the incompatible insect technique (IIT) or the combined IIT/sterile insect technique (SIT) for the landscape-scale (i.e., area-wide) control of Cx. quinquefasciatus, and have developed a calculator to estimate the costs of IIT and IIT/SIT applications at various sites in Hawaiʻi.MethodsThe overall cost of the infrastructure, personnel, and space necessary to produce incompatible adult males for release is calculated in a unit of ~ 1 million culicid larvae/week. We assessed the rearing costs and need for effective control at various elevations in Hawaiʻi using a 10:1 overflooding ratio at each elevation. The calculator uses a rate describing the number of culicids needed to control wild-type mosquitoes at each site/elevation, in relation to the number of larval rearing units. This rate is a constant from which other costs are quantified. With minor modifications, the calculator described here can be applied to other areas, mosquito species, and similar techniques. To test the robustness of our calculator, the Kauaʻi-specific culicid IIT/SIT infrastructure costs were also compared to costs from Singapore, Mexico, and China using the yearly cost of control per hectare, and purchasing power parity between sites for the cost of 1000 IIT/SIT males.ResultsAs a proof of concept, we have used the calculator to estimate rearing infrastructure costs for an application of IIT in the Alakaʻi Wilderness Reserve on the island of Kauaʻi. Our analysis estimated an initial investment of at least ~ $1.16M with subsequent yearly costs of approximately $376K. Projections of rearing costs for control at lower elevations are ~ 100 times greater than in upper elevation forest bird refugia. These results are relatively comparable to those real-world cost estimates developed for IIT/SIT culicid male production in other countries when inflation and purchasing power parity are considered. We also present supplemental examples of infrastructure costs needed to control Cx. quinquefasciatus in the home range of ʻiʻiwi Drepanis coccinea, and the yellow fever vector Aedes aegypti.ConclusionsOur cost calculator can be used to effectively estimate the mass rearing cost of an IIT/SIT program. Therefore, the linear relationship of rearing infrastructure to costs used in this calculator is useful for developing a conservative cost estimate for IIT/SIT culicid mass rearing infrastructure. These mass rearing cost estimates vary based on the density of the targeted organism at the application site.Graphic

Mass Trapping and Larval Source Management for Mosquito Elimination on Small Maldivian Islands.

Simple SummaryThe globalization of trade and travel, in combination with climate change, have resulted in the geographical expansion of mosquito-borne diseases. Moreover, over-reliance on chemical pesticides to control mosquitoes has resulted in resistance, which threatens the management of disease risk. We show, for the first time, that mosquito traps baited with human odors, in combination with controlling mosquito larvae in breeding sites, resulted in the near elimination of mosquito populations on two small islands, and the elimination of Aedes mosquitoes for 6+ months on a third island, in the Maldives. The levels of control achieved are comparable to current genetic control methods that are far more costly and impractical for implementation on small islands. The approach presented here poses the first alternative in decades to manage mosquito-borne disease risk on small (tropical) islands in an affordable and environmentally friendly manner.Globally, environmental impacts and insecticide resistance are forcing pest control organizations to adopt eco-friendly and insecticide-free alternatives to reduce the risk of mosquito-borne diseases, which affect millions of people, such as dengue, chikungunya or Zika virus. We used, for the first time, a combination of human odor-baited mosquito traps (at 6.0 traps/ha), oviposition traps (7.2 traps/ha) and larval source management (LSM) to practically eliminate populations of the Asian tiger mosquito Aedes albopictus (peak suppression 93.0% (95% CI 91.7–94.4)) and the Southern house mosquito Culex quinquefasciatus (peak suppression 98.3% (95% CI 97.0–99.5)) from a Maldivian island (size: 41.4 ha) within a year and thereafter observed a similar collapse of populations on a second island (size 49.0 ha; trap densities 4.1/ha and 8.2/ha for both trap types, respectively). On a third island (1.6 ha in size), we increased the human odor-baited trap density to 6.3/ha and then to 18.8/ha (combined with LSM but without oviposition traps), after which the Aedes mosquito population was eliminated within 2 months. Such suppression levels eliminate the risk of arboviral disease transmission for local communities and safeguard tourism, a vital economic resource for small island developing states. Terminating intense insecticide use (through fogging) benefits human and environmental health and restores insect biodiversity, coral reefs and marine life in these small and fragile island ecosystems. Moreover, trapping poses a convincing alternative to chemical control and reaches impact levels comparable to contemporary genetic control strategies. This can benefit numerous communities and provide livelihood options in small tropical islands around the world where mosquitoes pose both a nuisance and disease threat.

Mosquito odorant receptor sensitive to natural spatial repellents and inhibitory compounds

Previously, we have identified an odorant receptor (OR) from the southern house mosquito Culex quinquefasciatus, CquiOR32, which responded to both odorants (agonists) and inhibitory compounds (antagonists). CquiOR32/CquiOrco-expressing oocytes responded to methyl salicylate and other odorants with inward (regular) currents but gave currents in the reverse direction when challenged with eucalyptol and other inhibitors. To determine whether hitherto unknown ORs show this intrareceptor inhibition, we have now examined two other receptors in the same cluster, CquiOR27 and CquiOR28. We cloned and tested four variants of CquiOR28, but none of the 250 compounds in our panel of odorants, including an Orco ligand candidate (OLC12), elicited inward or upward deflections of the current traces. By contrast, CquiOR27/CquiOrco-expressing oocytes gave robust, dose-dependent inward currents when challenged with γ-octalactone and other odorants. On the other hand, octylamine and other phenolic compounds elicited dose-dependent currents in the reverse direction. When stimulatory and inhibitory compounds were presented in binary mixtures, γ-octalactone-elicited inward currents were attenuated in a dose-dependent manner according to the concentration of octylamine. As part of our chemical ecology approach, we tested the repellency activity of the most potent ligands in the surface landing and feeding assay and a newly reported hand-in cage assay. Protection elicited by γ-octalactone did not differ significantly from that of DEET at the same dose. In the hand-in cage assay, a cream formulation of γ-octalactone showed 97.0 ± 1.3% protection, with 47.6 ± 8.3% and 1.4 ± 0.7% landings per trial in the hands covered with a control and γ-octalactone cream, respectively (N = 8, p = 0.0078, Wilcoxon matched-pairs signed-rank test).

Comparative Efficacy of Selected Biorational Insecticides against Larvae of Southern House Mosquito Culex quinquefasciatus Say (Diptera: Culicidae)

Comparative Efficacy of Selected Biorational Insecticides against Larvae of Southern House Mosquito Culex quinquefasciatus Say (Diptera: Culicidae)

Identification and characterisation of mosquitoes from different locations in Qatar in 2017-2019.

Mosquito-borne infections have considerable consequences for public health. The mere presence of a single case of vector-borne disease (VBD) introduces a risk to the local community particularly when associated with the compatible vector, host, and suitable environmental factors. Presently, there is no well-established vector control and surveillance programme in Qatar; therefore, the likelihood of VBDs spreading is undetermined. As a result, there is a pressing need to address this gap and enable successful management of VBDs. This study presents the results of three consecutive field surveys conducted between 2017 and 2019 with the aim of defining the types and distribution of mosquitoes that are of public health importance in Qatar. The results of the adult mosquito trappings show that the southern house mosquito Culex quinquefasciatus is the most widespread and abundant mosquito species, followed by Cx. perexiguus, both species representing a risk of West Nile virus transmission. All sampling methods show that the malaria vector Anopheles stephensi is widespread including in urbanised areas, suggesting a risk of local malaria transmission. The wetland mosquito Aedes caspius is also widespread, representing a risk of Rift Valley fever virus transmission. The dengue vector Ae. aegypti was not detected and can be considered neither widespread nor abundant, suggesting a minimal risk for local transmission of dengue, chikungunya and Zika viruses. Interestingly, the study detected Culiseta longiareolata for the first time in Qatar. Regular field studies are needed to further address the knowledge gaps in terms of distribution, ecology, and biting habits of different mosquito species currently present in Qatar to accurately assess the risk of mosquito-borne diseases.

Cross Resistance in S-Methoprene-Resistant Culex quinquefasciatus (Diptera: Culicidae).

The juvenile hormone analog S-methoprene is the only synthetic biopesticide that is registered with the United States Environmental Protection Agency to control arthropods of economic importance in public health, livestock, pets, urban, and stored products. The high activity, relative target specificity, and benign environmental profile of S-methoprene have been well documented. While the risk of resistance in mosquitoes to S-methoprene is generally low, there is a lack of information regarding cross resistance in S-methoprene-resistant mosquitoes to other pesticides. In this paper, a population of the southern house mosquito Culex quinquefasciatus Say from southern California acquired low levels of resistance to S-methoprene in the field, where the resistance ratios ranged 7.0- to 8.8-fold as compared with a laboratory reference colony. After 30 generations of laboratory selections by S-methoprene when resistance was elevated to 57.4- to 168.3-fold relative to an unselected population, various levels of cross resistance to other commonly used pesticides were revealed in the selected population. Cross resistance to the microbial mosquito larvicide Lysinibacillus sphaericus (Meyer & Neide) (Bacillales: Bacillaceae) was the most profound, amounting to 77.50- to 220.50-fold. The mechanism and potential management tactics toward cross resistance are discussed to preserve the unique value of this synthetic biopesticide.

Fostering real-time climate adaptation: Analyzing past, current, and forecast temperature to understand the dynamic risk to Hawaiian honeycreepers from avian malaria

Various vector control options are increasingly being considered to safeguard forest birds in their natural habitats from avian malaria transmission. However, vector control options require localized deployment that is not logistically, ethically, ecologically, nor economically viable everywhere and all the time. Based on thermal tolerances of the sporogonic stages of avian malaria (Plasmodium relictum) parasite and its vector, the southern house mosquito (Culex quinquefasciatus), we examined the long-term weather trends for three high value, forest bird refuges (Alakai Wilderness Preserve on Kaua’i, Hanawi Natural Area Reserve on Maui, and Hakalau Forest National Wildlife Refuge on Hawai’i Island) to understand the temporal and site-specific differences of temperature-driven suitability for localized avian malaria transmission.On average, Alakai had mean ambient temperatures suitable for both the vector’s immature stage development and parasite sporogonic development most of the time (85.3%), indicating that observed variability in vector abundance or disease transmission may be driven by other factors. At higher elevation sites like Hakalau and Hanawi, current mean ambient temperatures suitable only for vector development prevail (91.7% and 96.6%, respectively), while mean ambient temperatures for both vector and parasite sporogonic development seldom occur (4.4% and 0% respectively). Our results not only show differences in the temperature suitability for transmission across elevation, but also different levels of vulnerability to avian malaria transmission with any additional projected increase in temperature. For instance, under a conservative warming scenario of 1.0 °C, the joint temperature suitability of parasite and vector development increases at higher elevation sites such as Hakalau (+35.8%) and Hanawi (+15.4%). While mean ambient temperatures suitable for both vector and parasite development already occur most of the time at Alakai, the occurrence also increases (+8.4%) as well under this conservative warming scenario.By linking current site-specific weather data to real-time weather forecasts, we developed a real-time avian malaria warning system to assist managers in identifying conditions when vector control is most needed at these three selected study sites. This online tool determines when conditions are likely to be suitable for local development of P. relictum and C. quinquefasciatus at Alakai, Hanawi, and Hakalau. This tool illustrates how managers can incorporate climate and current weather patterns into decision making without having to consider the uncertainties of long-term climatic and ecological projections.

Three-dimensional structure of the antennal lobe in the Southern house mosquito Culex quinquefasciatus.

The Southern house mosquito Culex quinquefasciatus relies on its olfactory system to locate the human hosts for blood meals, by which several deadly diseases are transmitted. Olfactory sensory neurons (OSNs) housed in the sensilla on the olfactory appendages send their axons into the antennal lobes (ALs), the primary olfactory center in the brain, where the OSNs expressing the same olfactory receptors converge upon the same spherical structures known as glomeruli in the AL. The structure of the antennal lobe, that is, the spatial organization of the glomeruli, governs the insect's odor identification and discrimination. Drosophila studies have demonstrated the specific connections between receptors and glomeruli based on the 3D structure of the antennal lobe, deepening our understanding of the relationships between glomerular activities and behaviors, but as yet the structure of the Cx. quinquefasciatus antennal lobe remains unknown. We therefore constructed a 3D model of the Cx. quinquefasciatus antennal lobe using nc82 antibody staining, identifying 62 and 44 glomeruli in the female and male mosquito antennal lobe, respectively, with a significant sexual dimorphism in terms of the antennal lobe volume and glomerulus number. These results demonstrate the structural basis of mosquito odor coding and provide a platform for future studies of the mosquito olfactory signal processing mechanism.

The role of native and introduced birds in transmission of avian malaria in Hawaii.

The introduction of nonnative species and reductions in native biodiversity have resulted in substantial changes in vector and host communities globally, but the consequences for pathogen transmission are poorly understood. In lowland Hawaii, bird communities are composed of primarily introduced species, with scattered populations of abundant native species. We examined the influence of avian host community composition, specifically the role of native and introduced species, as well as host diversity, on the prevalence of avian malaria (Plasmodium relictum) in the southern house mosquito (Culex quinquefasciatus). We also explored the reciprocal effect of malaria transmission on native host populationsand demography. Avian malaria infection prevalence in mosquitoes increased with the density and relative abundance of native birds, as well as host community competence, but was uncorrelated with host diversity. Avian malaria transmission was estimated to reduce population growth rates of Hawai'i 'amakihi (Chlorodrepanis virens) by 7-14%, but mortality from malaria could not explain gaps in this species' distribution at our sites. Our results suggest that, in Hawaii, native host species increase pathogen transmission to mosquitoes, but introduced species can also support malaria transmission alone. The increase in pathogen transmission with native bird abundance leads to additional disease mortality in native birds, further increasing disease impacts in an ecological feedback cycle. In addition, vector abundance was higher at sites without native birds and this overwhelmed the effects of host community composition on transmission such that infected mosquito abundance was highest at sites without native birds. Higher disease risk at these sites due to higher vector abundance could inhibit recolonization and recovery of native species to these areas. More broadly, this work shows how differences in host competence for a pathogen among native and introduced taxa can influence transmission and highlights the need to examine this question in other systems to determine the generality of this result.

Effects of predator cues and pesticide resistance on the toxicity of a (bio)pesticide mixture.

Populations of target species are typically exposed to pesticide mixtures and natural stressors such as predator cues, and are increasingly developing resistance to single pesticides. Nevertheless, we have poor knowledge whether natural stressors and the presence of pesticide resistance shape mixture toxicity. We tested the single and combined effects of the pesticide chlorpyrifos and the biopesticide Bacillus thuringiensis israelensis (Bti) on the survival of the Southern house mosquito (Culex quinquefasciatus, Say) and whether these effects were magnified by synthetic predator cues of Notonecta water bugs and differed between a chlorpyrifos-resistant (Ace-1R) and non-resistant (S-Lab) strain. Single exposure to Bti caused mortality in both strains (S-Lab ∼27%, Ace-1R ∼41%) and single exposure to chlorpyrifos caused only mortality in the S-Lab strain (∼33%), while predator cues did not induce mortality. The chlorpyrifos-resistant strain was 1.5-fold more sensitive to Bti, indicating a cost of resistance. The interaction types between chlorpyrifos and Bti (additive), between chlorpyrifos and predator cues (additive), and between Bti and predator cues (synergistic) were consistent in both strains. Despite predator cues making Bti approximately 8% more lethal, they did not change the additive interaction between Bti and chlorpyrifos in their mixture in either strain. These results indicate that the resistance against chlorpyrifos was not partly lifted when chlorpyrifos exposure was combined with Bti and predator cues. Identifying the interaction type within pesticide mixtures and how this depends on natural stressors is important to select control strategies that give a disadvantage to resistant individuals compared to non-resistant individuals. © 2019 Society of Chemical Industry.

Bloodmeal Host Selection of Culex quinquefasciatus (Diptera: Culicidae) in Las Vegas, Nevada, United States.

St. Louis encephalitis virus (SLEV) and West Nile virus (WNV) have recently emerged in the southwestern United States. Surveillance for arboviruses in Las Vegas, NV, detected a surge of SLEV activity in the southern house mosquito (Culex quinquefasciatus Say) during 2016. To identify candidate avian amplifiers, we assessed the identification, viral infection, and immune status of vertebrate hosts for 195 blood-engorged Cx. quinquefasciatus mosquitoes collected in August and September 2016. Bloodmeals were identified from 164 engorged abdomens, representing 19 species of birds and three species of mammals. No SLEV or WNV viremia was detected, but one mosquito tested positive for Culex flavivirus. House finch (Haemorhous mexicanus) (Muller) was the most common bloodmeal, followed by domestic chicken (Gallus gallus) (Linnaeus), American robin (Turdus migratorius) L., house sparrow (Passer domesticus) (L.), great-tailed grackle (Quiscalus mexicanus) (Gmelin), northern mockingbird (Mimus polyglottos) (L.) and mourning dove (Zenaida macroura) (L.). SLEV-reactive antibodies were detected in six identified bloodmeals and WNV-reactive antibodies were detected in 33. House sparrow and house finch were the most likely hosts to show previous exposure to SLEV and WNV, respectively. Over-utilization by Cx. quinquefasciatus for bloodmeal hosts was observed primarily among robin, finch and sparrow, all species that roost communally. House finch stands out as a candidate important amplifier for both SLEV and WNV because of its preference by mosquito vectors, and high competence for closely related virus strains. While implicated in previous outbreaks as an important mosquito vector, Cx. quinquefasciatus feeds infrequently on mammals in Las Vegas, indicating a low risk for bridge transmission to humans.

Environmental and social-demographic predictors of the southern house mosquito Culex quinquefasciatus in New Orleans, Louisiana

BackgroundUnderstanding the major predictors of disease vectors such as mosquitoes can guide the development of effective and timely strategies for mitigating vector-borne disease outbreaks. This study examined the influence of selected environmental, weather and sociodemographic factors on the spatial and temporal distribution of the southern house mosquito Culex quinquefasciatus Say in New Orleans, Louisiana, USA.MethodsAdult mosquitoes were collected over a 4-year period (2006, 2008, 2009 and 2010) using CDC gravid traps. Socio-demographic predictors were obtained from the United States Census Bureau, 2005–2009 American Community Survey and the City of New Orleans Department of Code Enforcement. Linear mixed effects models and ERDAS image processing software were used for statistical analysis and image processing.ResultsOnly two of the 22 predictors examined were significant predictors of Cx. quinquefasciatus abundance. Mean temperature during the week of mosquito collection was positively associated with Cx. quinquefasciatus abundance while developed high intensity areas were negatively associated with Cx. quinquefasciatus abundance.ConclusionThe findings of this study illustrate the power and utility of integrating biophysical and sociodemographic data using GIS analysis to identify the biophysical and sociodemographic processes that increase the risk of vector mosquito abundance. This knowledge can inform development of accurate predictive models that ensure timely implementation of mosquito control interventions.

Oviposition preference and offspring performance in container breeding mosquitoes: evaluating the effects of organic compounds and laboratory colonisation.

The preference-performance hypothesis predicts that organisms lacking parental care should oviposit in habitats that optimize offspring performance. We investigated preference-performance relationships for the Asian tiger mosquito (Aedes albopictus Skuse) and the southern house mosquito (Culex quinquefasciatus Say) (Diptera:Culicidae), two medically important container-breeding species, in response to an organic chemical blend mimicking decaying plant matter. Additionally, we evaluated the effects of long-term laboratory colonization of Cx. quinquefasciatus by using wild and laboratory strains.Oviposition bioassays were conducted by releasing gravid mosquitoes into field enclosures with automobile tires containing low and high concentrations of the chemical blend, and water controls. The offspring were then reared in water collected from the tires in which they were deposited.Aedes albopictus and wild Cx. quinquefasciatus laid more eggs in the chemical blend than water controls but did not differentiate between the low and high concentrations. Conversely, laboratory Cx. quinquefasciatus only preferred the high concentration to the low concentration. No statistical associations between oviposition preference and larval survival were found, as the chemical blend did not affect survivorship of either species.The oviposition preference for the chemical blend over water controls suggests that both species oviposit in the best available resource environment, but further studies are needed before conclusions regarding preference-performance relationships can be drawn.We found that long-term laboratory colonization affects oviposition behavior in Cx. quinquefasciatus, suggesting that behavioral studies on laboratory strains are not always applicable to wild populations.

Testing the causality between CYP9M10 and pyrethroid resistance using the TALEN and CRISPR/Cas9 technologies.

Recently-emerging genome editing technologies have enabled targeted gene knockout experiments even in non-model insect species. For studies on insecticide resistance, genome editing technologies offer some advantages over the conventional reverse genetic technique, RNA interference, for testing causal relationships between genes of detoxifying enzymes and resistance phenotypes. There were relatively abundant evidences indicating that the overexpression of a cytochrome P450 gene CYP9M10 confers strong pyrethroid resistance in larvae of the southern house mosquito Culex quinquefasciatus. However, reverse genetic verification has not yet been obtained because of the technical difficulty of microinjection into larvae. Here, we tested two genome editing technologies, transcription activator-like effector nucleases (TALEN)s and clustered regularly interspaced short palindromic repeats (CRISPR/Cas9), to disrupt CYP9M10 in a resistant strain of C. quinquefasciatus. Additionally, we developed a novel, effective approach to construct a TALE using the chemical cleavage of phosphorothioate inter-nucleotide linkages in the level 1 assembly. Both TALEN and CRISPR/Cas9 induced frame-shifting mutations in one or all copies of CYP9M10 in a pyrethroid-resistant strain. A line fixed with a completely disrupted CYP9M10 haplotype showed more than 100-fold reduction in pyrethroid resistance in the larval stage.

A single nucleotide change in a core promoter is involved in the progressive overexpression of the duplicated CYP9M10 haplotype lineage in Culex quinquefasciatus

Although the importance of cis-acting mutations on detoxification enzyme genes for insecticide resistance is widely accepted, only a few of them have been determined as concrete mutations present in genomic DNA till date. The overexpression of a cytochrome P450 gene, CYP9M10, is associated with pyrethroid resistance in the southern house mosquito Culex quinquefasciatus. The haplotypes of CYP9M10 exhibiting overexpression (resistant haplotypes) belong to one specific phylogenetic lineage that shares high nucleotide sequence homology and the same insertion of a transposable element. Among the resistant haplotypes, allelic progression involving an additional cis-acting mutation and gene duplication evolved a CYP9M10 haplotype associated with extremely high transcription and strong pyrethroid resistance. Here we show that a single nucleotide substitution G−27A, which is located near the transcription start site of CYP9M10, is involved in the progression of the duplicated haplotype lineage. The deletion of a 7-bp AT-rich sequence that includes nucleotide −27 inhibited the initiation of transcription from the original transcriptional initiation site. The mutation was suspected to reside within a core promoter, TATA-box, of CYP9M10.

A standard cytogenetic map of Culex quinquefasciatus polytene chromosomes in application for fine-scale physical mapping.

BackgroundSouthern house mosquito Culex quinquefasciatus belongs to the C. pipiens cryptic species complex, with global distribution and unclear taxonomy. Mosquitoes of the complex can transmit human and animal pathogens, such as filarial worm, West Nile virus and avian malarial Plasmodium. Physical gene mapping is crucial to understanding genome organization, function, and systematic relationships of cryptic species, and is a basis for developing new vector control strategies. However, physical mapping was not established previously for Culex due to the lack of well-structured polytene chromosomes.MethodsInbreeding was used to diminish inversion polymorphism and asynapsis of chromosomal homologs. Identification of larvae of the same developmental stage using the shape of imaginal discs allowed achievement of uniformity in chromosomal banding pattern. This together with high-resolution phase-contrast photography enabled the development of a cytogenetic map. Fluorescent in situ hybridization was used for gene mapping.ResultsA detailed cytogenetic map of C. quinquefasciatus polytene chromosomes was produced. Landmarks for chromosome recognition and cytological boundaries for two inversions were identified. Locations of 23 genes belonging to 16 genomic supercontigs, and 2 cDNA were established. Six supercontigs were oriented and one was found putatively misassembled. The cytogenetic map was linked to the previously developed genetic linkage groups by corresponding positions of 2 genetic markers and 10 supercontigs carrying genetic markers. Polytene chromosomes were numbered according to the genetic linkage groups.ConclusionsThis study developed a new standard cytogenetic photomap of the polytene chromosomes for C. quinquefasciatus and was applied for the fine-scale physical mapping. It allowed us to infer chromosomal position of 1333 of annotated genes belonging to 16 genomic supercontigs and find orientation of 6 of these supercontigs; the new cytogenetic and previously developed genetic linkage maps were integrated based on 12 matches. The map will further assist in finding chromosomal position of the medically important and other genes, contributing into improvement of the genome assembly. Better assembled C. quinquefasciatus genome can serve as a reference for studying other vector species of C. pipiens complex and will help to resolve their taxonomic relationships. This, in turn, will contribute into future development of vector and disease control strategies.Electronic supplementary materialThe online version of this article (doi:10.1186/s13071-015-0912-4) contains supplementary material, which is available to authorized users.

Laboratory selection of resistance to spinosad in Culex quinquefasciatus (Diptera: Culicidae).

A southern house mosquito Culex quinquefasciatus Say colony was established from surviving late instars and pupae from a semifield evaluation on Natular XRG (a granular formulation containing 2.5% spinosad). The initial lethal levels ofNatular XRG against this colony were determined in the laboratory for the first-generation progeny (designated as F1). Selection pressure was applied at LC70-90 levels to 10,000-15,000 late third- and early fourth-instar larvae of each generation with Natular XRG. Susceptibility changes in response to selection were determined every other generation, where a gradual and steady decline in susceptibility occurred from generation F1 to F35, followed by significant decline from generations F37 to F45 For reference purposes, susceptibility of freshly collected wild populations as well as a laboratory colony of the same species was also determined concurrently, which fluctuated within a slightly wider range for the wild populations and a tighter range for the laboratory colony. By comparing with wild populations and laboratory reference colony, tolerance to spinosad was observed from generations up to F9 in the selected population. Resistance levels increased gradually from generation F11 to F35, and elevated significantly from generations F37 to F45, when resistance ratios reached 1,415.3- to 2,229.9-fold at LC50 and 9,613.1- to 17,062.6-fold at LC90. Possible mechanisms of resistance development to spinosad were discussed.

Influence of resource levels, organic compounds and laboratory colonization on interspecific competition between the Asian tiger mosquito Aedes albopictus (Stegomyia albopicta) and the southern house mosquito Culex quinquefasciatus.

The mosquitoes Aedes albopictus (Stegomyia albopicta) (Skuse) and Culex quinquefasciatus (Say) (Diptera: Culicidae) are common inhabitants of tyres and other artificial containers, which constitute important peridomestic mosquito breeding habitats. We tested the hypotheses that interspecific resource competition between the larvae of these species is asymmetrical, that the concentration of chemicals associated with decomposing detritus affects the competitive outcomes of these species, and that wild and colonized strains of Cx. quinquefasciatus are affected differently by competition with Ae. albopictus. We conducted two laboratory competition experiments wherein we measured survivorship and estimated population growth (λ') in both species under multiple mixed-species densities. Under varying resource levels, competition was asymmetrical: Ae. albopictus caused competitive reductions or exclusions of Cx. quinquefasciatus under conditions of limited resources. In a second experiment, which used both wild and colonized strains of Cx. quinquefasciatus, organic chemical compounds associated with decomposing detritus did not affect the competitive outcome. The colonized strain of Cx. quinquefasciatus had greater survivorship and adult mass, and faster development times than the wild strain, but both strains were similarly affected by competition with Ae. albopictus. Competition between these species may have important consequences for vector population dynamics, especially in areas in which tyres and artificial containers constitute the majority of mosquito breeding habitats.