The influence of ecological factors on mosquito abundance and occurrence in Galápagos

We sampled mosquitoes across 18 sites established at different elevations and stretching from the north to the south of Isla Santa Cruz, Galápagos. Two commonly occurring species, Ae. taeniorhynchus and Cx. quinquefasciatus, were collected along with environmental variables characteristic of the trapping sites to assess their influence on mosquito abundance and occurrence in the dry season of 2015. We captured Ae. taeniorhynchus at 14 out of 18 sites and Cx. quinquefasciatus at low and high elevation sites on Santa Cruz. We utilized two generalized linear models; the first assessed the influence of environmental variables on abundances of Ae. taeniorhynchus and the second assessed the influence of these variables on the presence of Cx. quinquefasciatus. Populations of both mosquito species declined with elevation. Rainfall data were limited, as we sampled during the dry season of 2015. Distance to mangroves and maximum humidity were significant in influencing the abundance of Ae. taeniorhynchus, while maximum humidity was found to significantly influence the presence of Cx. quinquefasciatus. Both species occurred in sites where temperature, precipitation, and humidity should allow for mosquito development as well as parasitic development of the protozoan parasites that cause avian malaria. Further research involving year‐round sampling of mosquitoes and accompanying meteorological data as well as experimental studies on vector competence are required to understand disease dynamics of parasites such as avian malaria in Galápagos.     

Piperonyl butoxide synergizes the larvicidal activity of Origanum vulgare essential oil and its major constituents against the larvae of Aedes albopictus and Culex pipiens quinquefasciatus

Mosquitoes bite human beings and transmit many diseases, such as malaria, dengue fever, and Zika virus. Vector control of mosquitoes is an effective strategy for reducing the spread of disease. However, extensive use of insecticides (e.g. pyrethroids and organophosphorus) has caused resistance in mosquitoes, which weakens the effectiveness of mosquito control. Phytochemicals have been considered an alternative approach to mosquito control. Essential oil (EO) was obtained from the leaves and flowers of Origanum vulgare, and its synergistic activity with piperonyl butoxide (PBO) was tested against Aedes albopictus and Culex pipiens quinquefasciatus larvae. Thirty-seven compounds were identified, among which carvacrol and thymol were two major constituents (30.73 % and 18.81 %, respectively). O. vulgare EO had a significant toxic effect against fourth-stage larvae of Cx. p. quinquefasciatus and Ae. albopictus, with LC₅₀ values of 17.51 and 75.90 mg/L. Carvacrol and thymol also each appeared to be more effective against Cx. p. quinquefasciatus (LC₅₀ = 19.30 and 11.56 mg/L, respectively) than Ae. albopictus (LC₅₀ = 26.62 and 26.66 mg/L, respectively). PBO interacted synergistically with O. vulgare EO, carvacrol and thymol with 2.60–6.26 times as much of the active compound needed without PBO as against Cx. p. quinquefasciatus and Ae. albopictus larvae. Overall, our results contribute to the development of new natural mosquito insecticides.     

Aquatic thermal conditions predict the presence of native and invasive rock pool Aedes (Diptera: Culicidae) in the southern Appalachians,U.S.A.

The native rock pool mosquito, Aedes atropalpus (Coquillett), and the invasive Aedes japonicus (Theobald) have been found in many types of artificial and natural containers throughout North America. Little is known about the ecology of these two species in habitats where they co‐occur, although multiple investigators have reported the decline of the native species concurrent with the introduction and spread of the invasive species. Here we report the results of riverine rock pool collections (n=503) in the southern Appalachian Mountains between 2009‐2015. Surface water temperatures strongly predicted the presence of each species across a broad range of observed temperatures (11‐39.8° C). For every unit of increase in temperature (°C) the odds of collecting Ae. atropalpus larvae increased by 0.34 while the odds of collecting Ae. japonicus larvae decreased by 0.28. No Ae. japonicus larvae or pupae were collected at temperatures greater than 36° C; however, immature Ae. atropalpus were found in rock pools with temperatures up to 39.8° C. In contrast, Ae. japonicus were highly abundant in cooler rock pools (<17° C) where Ae. atropalpus were infrequent or absent. Our findings suggest that in spite of the successful invasion by Ae. japonicus, Ae. atropalpus remains well established in the southern Appalachian Mountains. Given the strong correlation of temperature with the presence of the two species and the contrasting absence of each species at observed temperature extremes, the role of thermal conditions should be carefully explored in the context of other ecological factors likely influencing the range and abundance of these mosquitoes.     

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.     

Invasive Process and Repeated Cross-Sectional Surveys of the Mosquito Aedes japonicus japonicus Establishment in Belgium

When accidentally introduced in a new location, a species does not necessarily readily become invasive, but it usually needs several years to adapt to its new environment. In 2009, a national mosquito survey (MODIRISK) reported the introduction and possible establishment of an invasive mosquito species, Aedes j. japonicus, in Belgium. First collected in 2002 in the village of Natoye from a second-hand tire company, then sampled in 2003 and 2004, the presence of adults and larvae was confirmed in 2007 and 2008. A repeated cross-sectional survey of Ae. j. japonicus was then conducted in 2009 in Natoye to study the phenology of the species on two different sites using three kinds of traps: Mosquito Magnet Liberty Plus traps, BG sentinel traps and CDC Gravid traps. An analysis of the blood meals was done on females to assess the epidemiological risks. Five species of mosquitos were caught using the different kind of traps: Culex pipiens, Cx torrentium, Anopheles claviger, Aedes geniculatus and Ae. j. japonicus, Cx pipiens being the most abundant. The CDC gravid traps gave the best results. Surprisingly Ae. j. japonicus was only found on one site although both sites seem similar and are only distant of 2.5 km. Its population peak was reached in July. Most of the engorged mosquitoes tested acquired blood meals from humans (60%). No avian blood meals were unambiguously identified. Larvae were also collected, mostly from tires but also from buckets and from one tree hole. Only one larva was found in a puddle at 100 m of the tire storage. A first local treatment of Ae. j. japonicus larvae population was done in May 2012 using Bacillus thuringiensis subsp. israelensis (Bti) and was followed by preventive actions and public information. A monitoring is also presently implemented.     

Larvicidal and mosquito repellent action of peppermint (Mentha piperita) oil

Oil of Mentha piperita L. (Peppermint oil), a widely used essential oil, was evaluated for larvicidal activity against different mosquito species: Aedes aegypti, Anopheles stephensi and Culex quinquefasciatus by exposing IIIrd instar larvae of mosquitoes in enamel trays 6 × 4 inch² size filled to a depth of 3 inch with water. Of the three species tested Cx. quinquefasciatus was most susceptible followed by Ae. aegypti and An. stephensi. Application of oil at 3 ml/m² of water surface area resulted in 100% mortality within 24 h for Cx. quinquefasciatus, 90% for Ae. aegypti and 85% for An. stephensi. For Ae. aegypti 100% mortality was achieved at 3 ml/m² in 48 h or 4 ml/m² in 24 h. For An. stephensi 100% mortality was observed at 4 ml/m² in 72 h. The emergence at 3 ml/m² was also inhibited to a great extent and the few adults which emerged did not ovipost even after taking a blood meal. The oil showed strong repellent action against adult mosquitoes when applied on human skin. Percent protection obtained against An. annularis, An. culicifacies, and Cx. quinquefasciatus was 100%, 92.3% and 84.5%, respectively. The repellent action of Mentha oil was comparable to that of Mylol oil consisting of dibutyl and dimethyl phthalates.     

Knockdown and lethal effects of three mosquito coil formulations against Aedes aegypti and Culex quinquefasciatus under different nutritional conditions

Three mosquito coil formulations, each containing either metofluthrin 0.025% w/w, d-allethrin 0.225% w/w, or esbiothrin 0.10% w/w were evaluated for knockdown and killing properties against laboratory populations of female Aedes aegypti (L.) and Culex quinquefasciatus Say under different nutritional-energy sources of blood, sucrose, and water. The tests were conducted in a 70 cm × 70 cm × 70 cm glass chamber. Mosquito responses were measured by knockdown times during the 20-min exposure period and mortality at 24 h post-exposure. The results showed the metofluthrin coil provided the most rapid knockdown for both test species and regardless of nutritional condition compared with the other two coils. Metofluthrin and d-allethrin were highly effective in killing Ae. aegypti (95–100% mortality), whereas esbiothrin produced 100% mortality to water-fed mosquitoes and 78.3 and 80% mortality for blood- and sucrose-fed specimens, respectively. >85% mortality was observed in sucrose- and water-fed Cx. quinquefasciatus against metofluthrin, while 78.3% blood-fed females survived exposure. This species showed very low mortality with d-allethrin (3.3% to 28.3%), with the highest mortality recorded (71.7%) for water-fed with esbiothrin. Overall, Ae. aegypti was more sensitive to all three coil products than Cx. quinquefasciatus. The mortality between species and nutritional conditions showed significant differences for all comparisons except for sucrose-fed mosquitoes exposed to metofluthrin.     

Seasonal surveillance confirms the range expansion of Aedes japonicus japonicus (Theobald) (Diptera: Culicidae) to the Hawaiian Islands of Oahu and Kauai

The Asian bush mosquito, Aedes japonicus japonicus (Theobald) was not known to occur in the Hawaii archipelago until it was identified on the island of Hawaii in 2003. This mosquito species remained undetected on the neighboring islands for 8?years before it was discovered at the Honolulu International Airport on Oahu in 2012. By 2015, four Ae. j. japonicus mosquitoes were collected in the western mountains of Oahu and one was collected in the central mountains of Kauai. The collection of this invasive mosquito species across the neighboring Hawaiian Islands of Oahu and Kauai indicated the need for increased seasonal surveillance on these islands. Following nearly four years of surveillance, Ae. j. japonicus was also confirmed to occur in the eastern mountains of Oahu and in the central mountainous region of Kauai. To expand the knowledge of the spread of invasive mosquitoes species further surveillance is necessary to identify all possible areas where populations of Ae. j. japonicus and other invasive mosquito species occur in Hawaiian archipelago.     

A recent survey of mosquito fauna in Guandong Province, southern China, with a review of past records

Abstract The southern province of Guangdong has long been subject to endemic mosquito‐borne diseases. In recent years, this region of China has experienced rapid, extensive economic development involving environmental change, making much of the scant knowledge of its mosquito fauna obsolete. This paper reviews previous mosquito surveys, some of which may be too old to be of relevance to present‐day conditions, and presents the results of a recent survey of adult and immature mosquitoes. The main vectors of mosquito‐borne diseases endemic to the area, such as dengue and Japanese encephalitis virus, develop in container habitats. A three‐year survey was carried out, between 2004 and 2006, of 4131 breeding containers in residential areas and in open, sparsely populated areas, of which approximately 50% were positive for the presence of mosquitoes, and 10 156 larvae and pupae were collected and identified. Twelve species were found in both residential and sparsely populated areas: Aedes albopictus (Skuse) (Diptera: Culicidae), Ae. lineatopennis (Ludlow), Ae. vexans (Meigen), Tanakaius togoi (Theobald), Culex barraudi Edwards, Cx dispectus Bram, Cx malayi (Leicester), Cx pallidothorax Theobald, Cx quinquefasciatus Say, Cx sitiens Wiedemann, Lutzia fuscanus Wiedemann and Tripteroides bambusa (Yamada). Armigeres subalbatus (Coquillett) was found only in containers in villages, whereas Ae. macfarlanei Edwards, Cx mimeticus Noé, Cx sinensis Theobald, Cx vegans Wiedemann, Cx wilfredi Colless and Mansonia uniformis (Theobald) were found only in non‐ or sparsely populated areas. In residential areas, the rank order of most common species, as measured by the proportion of containers colonized, was Ae. albopictus > Cx quinquefasciatus > Lu. fuscanus, whereas in sparsely populated areas the rank order was Cx quinquefasciatus > Ae. albopictus > Lu. fuscanus. Light traps in non‐ or sparsely populated areas caught 5995 adult mosquitoes of 25 species, some of which are not container breeders. The most common species were: Anopheles sinensis Wiedemann, An. maculatus Theobald, An. minimus Theobald, Ta. togoi, Cx bitaeniorhynchus Giles, Cx malayi, Cx quinquefasciatus, Cx sinensis Theobald, Cx sitiens, Cx tritaeniorhynchus Giles and Lu. fuscanus. It is noteworthy that nine species caught had not been previously recorded in Guangdong Province, highlighting the deficient knowledge of the current composition and distribution of the mosquito fauna of this part of China.     

Evaluation of collection methods for Culex quinquefasciatus,Aedes aegypti,and Aedes simpsoni in northeastern Tanzania

In East Africa, significant morbidity and mortality are caused by infections spread by Culex quinquefasciatus and Aedes aegypti. Sticky traps have been shown to be effective tools for sampling populations of Aedes mosquitoes and have been found to catch Cx. quinquefasciatus. Thus, they could potentially be used to sample populations of this species. This study compared Sticky ovitraps (SO) and MosquiTraps (MQT) with the CDC Gravid trap (CDC‐GT) for collection of Culex and Aedes mosquito populations in Tanzania. A follow‐up experiment was carried out using traps set for a 24‐h period to accommodate the oviposition habits of Aedes aegypti and Ae. simpsoni s.l. mosquitoes. The results showed that the CDC‐GT caught significantly more Cx. quinquefasciatus and Ae. aegypti than the SO or MQT, but there was no significant difference in the number of mosquitoes caught between the two sticky traps or of Ae. simpsoni s.l. caught among the three trap types. The results suggest that CDC‐GTs are the most appropriate in sampling of Cx. quinquefasciatus. Although CDC‐GTs collected more Ae. aegypti than the sticky traps, the simplicity and cost benefit of sticky traps facilitates large scale studies. All three trap types should be considered for monitoring Aedes mosquitoes.     

Expression of Binary Toxin Genes in the Mosquito-Colonizable Bacteria, <Emphasis Type="Italic">Bacillus cereus</Emphasis>, Leads to High Toxicity Against <Emphasis Type="Italic">Culex quinquefasciatus</Emphasis> Larvae

Two B. cereus strains, Ae10 and Cx5, isolated from mosquito larval guts, were transformed with a recombinant plasmid, pBS373, harboring binary toxin genes from Bacillus sphaericus 2297. Immunoblotting analysis clearly revealed the production and presence of the 51-kDa toxin protein in both strains. Two recombinant B. cereus strains Ae10 and Cx5 showed very high toxicity against C. quinquefasciatus larvae. Since both strains have a close relationship with the mosquito larvae in the native environment and are capable of recolonizing in the guts of mosquito larvae, these strains can be considered promising new hosts for an effective delivery of mosquito-larvicidal toxins.     

Microbial synthesis of spherical nanosilver and nanogold for mosquito control

Biosynthesis of metal nanoparticles using microorganisms is an important area of research in nanobiotechnology, which is an emerging eco-friendly science of well-defined sizes, shapes and controlled monodispersity. The present study proposed a green process for the extracellular production of silver (Ag) and gold (Au) nanoparticles (NPs) using the soil fungi Chrysosporium keratinophilum and Verticillium lecanii. The synthesized NPs were formed fairly uniform with spherical shape determined by Transmission Electron Microscope (TEM) and confirmed by Scanning Electron Microscope (SEM). Elemental analysis on single particle was carried by EDX analysis. The results were further supported by UV-vis spectrophotometry. In addition, we have also investigated the effect of synthesized AgNPs and AuNPs against the larvae and pupae of Anopheles stephensi, Culex quinquefasciatus and Aedes aegypti. The efficacy test was performed at different concentrations for periods of different lengths by the probit analysis. The larvae and pupae of Cx. quinquefasciatus, An. stephensi and Ae. aegypti were found highly susceptible to the synthesized AgNPs than the AuNPs. The larvae of Cx. quinquefasciatus and Ae. aegypti were found to be more susceptible to the AgNPs and AuNPs synthesized using the C. keratinophilum and V. lecanii compared with the larvae of An. stephensi. The pupae of Ae. aegypti have shown higher mortality against the synthesized AgNPs than the pupa of Cx. quinquefasciatus, while no adverse effects could be observed in the pupa of An. stephensi. By this approach, it is suggested that this rapid synthesis of nanoparticles would be useful for developing a biological process for mosquito control.     

Chicory (Cichorium intybus) and wormwood (Artemisia absinthium) extracts exhibit strong larvicidal activity against mosquito vectors of malaria,dengue fever,and filariasis

Vector-borne diseases transmitted by mosquitoes cause globally important diseases such as malaria, dengue fever, and filariasis. The incidence of these diseases can be reduced through mosquito control programs but these control programs currently rely on synthetic insecticides that can impact the environment, and has selected widespread mosquito resistance. Environment friendly and biodegradable natural insecticides discovered in plants offer an alternative approach to mosquito control. Here, we investigated extracts from root or aerial parts of Chicory (Cichorium intybus) and wormwood (Artemisia absinthium) against the early 4th instar larvae of Anopheles stephensi (malaria vector), Aedes aegypti (dengue fever vector), and Culex quinquefasciatus (filariasis vector). The root and aerial parts extracts of A. absinthium and C. intybus at 200, 100, 50, 25 and 12.5?ppm caused significant mortality of the tested mosquito species. Root extracts exhibited higher larvicidal activity that aerial part extracts. The highest larvicidal activity was recorded in methanol extract of roots of C. intybus with LC50?=?66.16, 18.88 and LC¬90?=?197.56, 107.16?ppm for An. stephensi; LC50?=?78.51, 40.15 and LC90?=?277.31, 231.28?ppm for Ae. aegypti and LC50?=?103.99, 64.56 and LC¬90?=?314.04, 247.54?ppm for Cx. quinquefasciatus. These results reveal potent mosquito larvicidal activity against vectors of malaria, dengue fever, and filariasis is present in extracts of chicory and wormwood.     

Oviposition substrate selection by Florida mosquitoes in response to pathogen‐infected conspecific larvae

The impact of the presence of larval mosquito pathogens with potential for biological control on oviposition choice was evaluated for three mosquito species/pathogen pairs present in Florida. These included Aedes aegypti infected with Edhazardia aedis, Aedes albopictus infected with Vavraia culicis, and Culex quinquefasciatus infected with Culex nigripalpus nucleopolyhedrovirus (CuniNPV). Two‐choice oviposition bioassays were performed on each host and pathogen species with one oviposition cup containing infected larvae and the other cup containing uninfected larvae (control). Both uninfected and E. aedis‐infected female Ae. aegypti laid significantly fewer eggs in oviposition cups containing infected larvae. Uninfected gravid female Ae. albopictus and Cx. quinquefasciatus oviposited equally in cups containing uninfected larvae or containing larvae infected with V. culicis or CuniNPV, respectively. Gravid female Ae. albopictus infected with V. culicis did not display ovarian development and did not lay eggs. The decreased oviposition by gravid Ae. aegypti in containers containing E. aedis‐infected larvae may indicate that the infected larvae produce chemicals deterring oviposition.     

Mosquitoes rely on their gut microbiota for development

Field studies indicate adult mosquitoes (Culicidae) host low diversity communities of bacteria that vary greatly among individuals and species. In contrast, it remains unclear how adult mosquitoes acquire their microbiome, what influences community structure, and whether the microbiome is important for survival. Here, we used pyrosequencing of 16S rRNA to characterize the bacterial communities of three mosquito species reared under identical conditions. Two of these species, Aedes aegypti and Anopheles gambiae, are anautogenous and must blood‐feed to produce eggs, while one, Georgecraigius atropalpus, is autogenous and produces eggs without blood feeding. Each mosquito species contained a low diversity community comprised primarily of aerobic bacteria acquired from the aquatic habitat in which larvae developed. Our results suggested that the communities in Ae. aegypti and An. gambiae larvae share more similarities with one another than with G. atropalpus. Studies with Ae. aegypti also strongly suggested that adults transstadially acquired several members of the larval bacterial community, but only four genera of bacteria present in blood fed females were detected on eggs. Functional assays showed that axenic larvae of each species failed to develop beyond the first instar. Experiments with Ae. aegypti indicated several members of the microbial community and Escherichia coli successfully colonized axenic larvae and rescued development. Overall, our results provide new insights about the acquisition and structure of bacterial communities in mosquitoes. They also indicate that three mosquito species spanning the breadth of the Culicidae depend on their gut microbiome for development.     

DsRed2 transient expression in Culex quinquefasciatus mosquitoes

Culex quinquefasciatus mosquitoes have been successfully genetically modified only once, despite the efforts of several laboratories to transform and establish a stable strain. We have developed a transient gene expression method, in Culex, that delivers plasmid DNA directly to the mosquito haemolymph and additional tissues. We were able to express DsRed2 fluorescent protein in adult Cx. quinquefasciatus mosquitoes by injecting plasmids directly into their thorax. The expression of DsRed2 in adult Cx. quinquefasciatus mosquitoes is an important stepping stone to genetic transformation and the potential use of new control strategies and genetic interactions.     

Mosquito-only flaviviruses,isolated from <Emphasis Type="Italic">Aedes albopictus</Emphasis> in Slovenia: results of a pilot mosquito monitoring program

Autochthonous transmissions of mosquito-borne diseases were described in close proximity to Slovenia where the knowledge of mosquito species and the viruses they transmit remains very scarce. Therefore we aimed to assess the burden mosquitoes bring to the local community and to test mosquitoes for the presence of the selected zoonotic arboviruses, i. e. Dengue (DENV), West Nile (WNV) and Chikungunya (CHIKV). A pilot mosquito monitoring was conducted in 2012 within two municipalities based on adult mosquito trapping. Altogether 2007 adult mosquitoes were caught. Females of Aedes albopictus and Culex pipiens represented the majority of the total mosquito catch. In sites with more households and nearby greenery, catches of Ae. albopictus were more abundant. Highest numbers of Cx. pipiens females were caught close to a nature reserve, where many bird species stop during their migrations. Trapping results are consistent with both species’ feeding behaviour. Female mosquitoes were grouped into 58 pools according to species, location and date of catch. Real time RT-PCRs for WNV, DENV and CHIKV were performed, but no arboviral genomes were detected in any of the tested mosquito pools. However, amplicons were obtained in three pools when using generic RT-PCR markers for flaviviruses. The viruses detected were propagated and isolated in mosquito cell line C3/36 and identified with sequencing as mosquito-only flaviviruses (MOFs). The important contributions of this study are a successful first isolation of MOFs and confirmation of the presence of the invasive mosquito Ae. albopictus in Slovenia.     

Host range of the parasite Strelkovimermis spiculatus (Nematoda: Mermithidae) in Argentina mosquitoes

Strelkovimermis spiculatus ( Poinar and Camino 1986 ) is a nematode parasite of mosquitoes isolated from the Neotropical region. We investigated the host range of this parasite in mosquito populations for a better understanding of the dynamics of temporary breeding sites. Five grassy‐pool habitats filled by rainwater were sampled from the summer 2007 through the fall 2008. Eight mosquito species were collected: Anopheles albitarsis, Culex chidesteri, Culex dolosus, Culex maxi, Aedes albifasciatus, Psorophora ciliata, Psorophora cyanescens, and Psorophora albigenu. Six of these species were parasitized: Cx. chidesteri, Cx. dolosus, Cx. maxi, Ae. albifasciatus, Ps. ciliata, and Ps. cyanescens. The occurrence of this mermithid in natural mosquito populations was increased from the end of winter to the end of the spring. Prevalence ranged from 11% to 100%. High levels of infections were registered only in Ae. albifasciatus larvae, the most abundant mosquito species (95%), followed by Cx. dolosus (2.7%). Strelkovimermis spiculatus completed its development in all infected mosquito larvae. The presence of S. spiculatus in six natural mosquito populations increases the number of susceptible species to 24.     

Exposure of Culex quinquefasciatus to the oomycete Pythium insidiosum: A protocol for in vitro studies

Pythium insidiosum causes pythiosis, an infection that affects different species of mammals, including humans, and inhabits marshy ecosystems of tropical, subtropical, and temperate regions worldwide. Therefore, this study proposes a protocol to expose Culex quinquefasciatus to P. insidiosum zoospores. Cx. quinquefasciatus immatures (eggs, larvae, and pupae) were exposed to zoospores (8x10³ zoospores/mL) of the oomycete for 24 h. The exposure of Cx. quinquefasciatus to the zoospores from L1 to the emergence of adults was evaluated, and P. insidiosum detection was performed by microbiological culture, polymerase chain reaction, and histopathological analysis of stage 4 larvae. The protocol used to produce Cx. quinquefasciatus colonies and adapted for this study proved viable for research on the interaction between P. insidiosum and this Culicidae species. Moreover, P. insidiosum presence was evident in all larval stages of the mosquito, although the presence of the oomycete was not detected in the eggs, pupae, and adults. This study is a pioneer in the development of a protocol to evaluate Cx. quinquefasciatus exposure to P. insidiosum zoospores, and under experimental conditions, P. insidiosum can establish itself in Cx. quinquefasciatus larval stages. The developed protocol is expected to serve as a basis for developing studies to evaluate the interactions of P. insidiosum with these mosquitoes and shed more light on the participation of culicids in expanding the ecological niche of P. insidiosum.