Vector competence of northern and southern European Culex pipiens pipiens mosquitoes for West Nile virus across a gradient of temperatures

In Europe, West Nile virus (WNV) outbreaks have been limited to southern and central European countries. However, competent mosquito vectors and susceptible bird hosts are present in northern Europe. Differences in temperature and vector competence of mosquito populations may explain the absence of WNV outbreaks in northern Europe. The aim of the present study was to directly compare vector competence of northern and southern European Culex pipiens (Cx. p.) pipiens mosquitoes for WNV across a gradient of temperatures. WNV infection and transmission rates were determined for two Cx. p. pipiens populations originating from The Netherlands and Italy, respectively. Mosquitoes were orally exposed by providing an infectious bloodmeal, or by injecting WNV (lineage 2) in the thorax, followed by 14‐day incubation at 18, 23, or 28 °C. No differences in infection or transmission rates were found between the Cx. p. pipiens populations with both infection methods, but WNV transmission rates were significantly higher at temperatures above 18 °C. The absence of WNV outbreaks in northern Europe cannot be explained by differences in vector competence between Cx. p. pipiens populations originating from northern and southern Europe. This study suggests that low temperature is a key limiting factor for WNV transmission.     

Unraveling microbe-mediated interactions between mosquito larvae in a laboratory microcosm

Interspecies interactions have important impacts on communities and when multiple trophic levels are involved, effects can be complex and indirect. For mosquitoes, interactions experienced as larvae affect adult attributes such as survivorship, reproductive output, and longevity, factors that can affect their ability to vector disease. We examined how larvae of two ecologically distinct mosquito species, Aedes japonicus japonicus and Culex quinquefasciatus, interact at different temperatures (17 and 27 °C) and at different relative densities. We also quantified abundances of bacteria and protozoan flagellates to uncover how changes in the microbial community affect the outcome of the two mosquitoes’ interaction. At 17 °C, survival and size of both mosquito species were not affected by the other’s presence. Cx. quinquefasciatus was strongly affected by intraspecific, but not interspecific, competition at both temperatures. At 27 °C, Ae. j. japonicus larvae experienced 100 % mortality in treatments by themselves and treatments where Cx. quinquefasciatus was abundant, surviving only in the presence of low densities of Cx. quinquefasciatus. Both the total bacteria count and counts of a protozoan flagellate identified as Spumella spp. decreased with increasing numbers of Cx. quinquefasciatus. We postulate that at 27 °C, the survival of Ae. j. japonicus depends on the interaction between Cx. quinquefasciatus and the microbial community. This study demonstrates that one mosquito species may alter the microbial community in ways that indirectly influence another mosquito species’ larval survival, and by extension adult abundance and potential disease transmission.     

First record of West Nile Virus detection inside wild mosquitoes in Khartoum capital of Sudan using PCR

This study aimed to explore the presence of West Nile Virus (WNV) inside four species of mosquitoes: Culex univittatus (Theobald), Culex quinquefasciatus (Say) Aedes vittatus (Bigot) and Aedes vexans (Meigen). Adult wild mosquitoes were collected from different sites: Soba West, Hellat Kuku, Shambat, and Khartoum North Central Live Stock Market (KCLM). Surveys were carried out at Khartoum State during two phases: pre to the rainy season and post to the rainy season. Mosquito specimens were identified using classical keys then preserved at −80 °C freezer for two weeks till the virus examination using polymerase chain reaction (PCR) were carried out. WNV has been detected inside the three species of mosquitoes: A. vexans, C. univittatus, and C. quinquefasciatus. The species were collected from Hellat Kuku, (Shambat and Hellat Kuku), and (Shambat and KCLM) respectively. Two species of mosquitoes were positive for the virus: C. quinquefasciatus and C. univittatus. Positive results for the virus during the first phase of the study; males of C. quinquefasciatus and C. univittatus collected during the second phase of the study were also tested for the existence of the virus and they were positive. For our knowledge this study represents first record of WNV inside wild mosquitoes in Sudan. PCR technique provided reliable information because specific primer-probe sets were used for the detection of the virus. Extra studies are required to incriminate these species of mosquitoes as potential vectors of WNV.     

Genetic diversity and population structure of Culex modestus across Europe: does recent appearance in the United Kingdom reveal a tendency for geographical spread?

In mainland Europe, the mosquito species Culex modestus Ficalbi (1890) is a bridge vector for West Nile virus (WNV) from its natural bird-mosquito cycle to mammals. The present study assessed the genetic diversity of Cx. modestus, as well as related Culex species, using the mitochondrial COI DNA barcoding region and compared this with the population structure across Europe. A haplotype network was mapped to determine genealogical relationships among specimens. The intraspecific genetic diversity within individual Culex species was below 2%, whereas the interspecific genetic divergence varied from 2.99% to 13.74%. In total, 76 haplotypes were identified among 198 sequences. A median-joining network determined from 198 COI sequences identified two major lineages that were separated by at least four mutation steps. A high level of intraspecific genetic diversity was not detected in Cx. modestus in samples submitted from different European populations, which indicates that morphologically identified specimens represent a single species and not a species complex. Therefore, it is deduced that different populations of Cx. modestus will show a similar potential to transmit WNV, lending support to concerns that the population present in southeast England represents a risk of transmission to humans.     

The vector ecology of introduced Culex quinquefasciatus populations,and implications for future risk of West Nile virus emergence in the Galápagos archipelago

Culex quinquefasciatus Say (Diptera: Culicidae), an important vector of West Nile virus (WNV) in the U.S.A., was first detected on the Galápagos Islands (Ecuador) in the 1980s. However, little is known of its ecology, distribution or capacity for arbovirus transmission in the Galápagos. We characterize details of lifecycle (including gonotrophic period), temporal abundance, spatial distribution, vector competence and host‐feeding behaviour. Culex quinquefasciatus was detected on five islands of the Galápagos during 2006–2011. A period of 7–14 days was required for egg–adult emergence; water salinity above 5 ppt was demonstrated to hinder larval development. Blood‐meal analysis indicated feeding on reptiles, birds and mammals. Assessment of WNV vector competency of Galápagos C. quinquefasciatus showed a median infectious dose of 7.41 log₁₀ plaque‐forming units per millilitre and evidence of vertical transmission (minimal filial infection rate of 3.7 per 1000 progeny). The distribution of C. quinquefasciatus across the archipelago could be limited by salt intolerance, and its abundance constrained by high temperatures. Feeding behaviour indicates potential to act as a bridge vector for transmission of pathogens across multiple taxa. Vertical transmission is a potential persistence mechanism for WNV on Galápagos. Together, our results can be used for epidemiological assessments of WNV and target vector control, should this pathogen reach the Galápagos Islands.     

Behavioural response of the house mosquitoes Culex quinquefasciatus and Culex pipiens molestus to avian odours and its reliance on carbon dioxide

How Culex (Diptera: Culicidae) mosquitoes select and discriminate between potential avian hosts is critical for understanding the epidemiology of West Nile virus. Therefore, the present authors studied the behavioural responses of Culex quinquefasciatus (Say) and Culex pipiens molestus (Forsskål) to headspace volatiles of three avian species [chicken and pigeon (sexes analysed separately), and magpie], presented either alone or in combination with 600 p.p.m. carbon dioxide (CO₂). The attraction of Cx. quinquefasciatus to the headspace volatiles of both sexes of chicken, and of female pigeon, in combination with CO₂ was significantly higher than that achieved by the CO₂ and solvent control. Although Cx. p. molestus was attracted to headspace volatiles of chickens and magpies, it was repelled by those of female pigeons when combined with CO₂. An increased effect between the avian volatiles and CO₂ was observed for Cx. quinquefasciatus, whereas the addition of CO₂ had no effect on the attraction of Cx. p. molestus females. The results of this study demonstrate that Cx. quinquefasciatus and Cx. p. molestus are attracted to the odour of potential avian hosts. Future studies aimed at identifying the bioactive volatile compounds in the headspace of chickens may contribute to the potential development of effective surveillance and control tools against Culex species.     

Sympatric occurrence of Culex pipiens (Diptera,Culicidae) biotypes pipiens,molestus and their hybrids in Portugal,Western Europe: feeding patterns and habitat determinants

Culex (Culex) pipiens (Diptera: Culicidae) has two recognized biotypes, pipiens and molestus, which differ in physiology and behaviour; this difference may influence vectorial capacity for West Nile virus (WNV). Our goal was first to determine the presence of Cx. pipiens populations in 31 locations in Portugal and to subsequently analyse their host‐feeding preferences and habitat determinants. Molecular identification of Cx. pipiens forms and their hybrids was performed in 97 females; bloodmeal sources were identified in 59 engorged specimens. Overall, 61.9% of specimens were identified as Cx. pipiens f. pipiens, 20.6% as Cx. pipiens f. molestus, and 17.5% as hybrid forms. Culex pipiens f. pipiens fed preferentially on birds, and Cx. pipiens f. molestus on humans. Hybrid forms fed mostly on birds, but human bloodmeals were common. With reference to habitat, Cx. pipiens f. pipiens and hybrid forms were positively correlated with peri‐urban habitats. Our results confirm the sympatric presence of different Cx. pipiens biotypes in 14 of the 31 locations studied. Peri‐urban areas were a common habitat of all biotypes and may represent zones of hybridization. The feeding preferences and sympatric distribution of the Cx. pipiens biotypes observed in Portugal favour the epizootic circulation of WNV and the occurrence of disease outbreaks of WNV.     

Estimating reaction norms for predictive population parameters,age specific mortality,and mean longevity in temperature‐dependent cohorts of Culex quinquefasciatus Say (Diptera: Culicidae)

Culex quinquefasciatus plays a major role in the transmission of important parasites and viruses throughout the world. Because temperature is an important limiting factor on growth and longevity of all mosquito species, estimating the reaction norms provides very important basic information for understanding both plasticity and individual variations of the population. In the present study, Cx. quinquefasciatus were maintained at five different constant temperatures (15°, 20°, 23°, 27°, and 30°C) for two subsequent generations. Reproductive population parameters in blood‐fed mated females and longevities of virgin and blood‐fed mated adults reared at different temperatures were compared for the two generations. Longevity increased as temperature decreased within a range of 15° to 30°C for the unmated adults, and 15° to 27°C for the mated and blood‐fed adults. Generation times were as long as 124.07 and 106.76 days for two subsequent generations reared at 15°C, and the highest intrinsic rate of increase (r_m) values were estimated at 0.22 and 0.18, respectively, from the cohorts reared at 27°C. For survival rates, reproductive rates (R₀), and r_m values, 30°C was found to be a critical temperature for this species. These cohorts produced the smallest amount of eggs (R₀= 5.06), r_m values decreasing across generations (from 0.11 to 0.06), and the survival rates from egg to adult were found to be insufficient (16.1 and 10.8%). Additionally, the rate of exponential increase with age and age specific mortalities (b) were calculated for the virgin cohorts. Age specific mortality rates increased as temperature decreased. The increase in mortality rates started to accelerate at 27°C and was more pronounced at 30°C, for both females and males. We estimated the coefficients of variation for the b values in which females have smaller coefficients than those of the males at all temperatures.     

Temperature modulates the effects of predation and competition on mosquito larvae

1. Predation risk affects interspecific competition by decreasing foraging activity and relative competitive ability. Predation risk is determined by predators' prey choice and prey responses, both of which can be influenced by temperature. Temperature is especially important for larval prey and can result in a trade‐off between predator‐induced decreases in foraging activity and growth. Interspecific competition must also be examined in relation to intraspecific density‐dependent competition; weaker interspecific competition leads to coexistence of competitors. 2. This study explored how temperature (15 and 25 °C) could affect a focal species, larvae of the mosquito Culex quinquefasciatus, by examining prey choice in a shared predator (mosquitofish; Gambusia holbrooki) and the effects of predation risk on interspecific competition with Limnodynastes peronii tadpoles. Intraspecific density‐dependent competition in C. quinquefasciatus at these temperatures was also examined. 3. At 25 °C, G. holbrooki consumption of both C. quinquefasciatus and L. peronii increased; however, the effects of interspecific competition on mosquito survival did not decrease with L. peronii exposure to predation risk. The relationship between intraspecific density‐dependent competition and interspecific competition was temperature‐dependent, with competitive dominance of L. peronii at 25 °C. Male and female mosquitoes had different temperature‐dependent responses, indicating sex‐specific intrinsic responses to starvation and differential selection pressures. At 25 °C, females were susceptible to interspecific competition by L. peronii, while males were susceptible to intraspecific competition. 4. The use of competitors as biological controls has implications for mosquito disease transmission, and these results suggest that control effectiveness may be modified by climate change.     

Avian host community structure and prevalence of West Nile virus in Chicago,Illinois

Vertebrate host diversity has been postulated to mediate prevalence of zoonotic, vector-borne diseases, such that as diversity increases, transmission dampens. This “dilution effect” is thought to be caused by distribution of infective bites to incompetent reservoir hosts. We quantified avian species richness, avian seroprevalence for antibodies to West Nile virus (WNV), and infection of WNV in Culex mosquitoes, in the Chicago metropolitan area, Illinois, USA, a region of historically high WNV activity. Results indicated high overall avian seroprevalence and variation in seroprevalence across host species; however, there was no negative correlation between avian richness and Culex infection rate or between richness and infection status in individual birds. Bird species with high seroprevalence, especially northern cardinals and mourning doves, may be important sentinels for WNV in Chicago, since they were common and widespread among all study sites. Overall, our results suggest no net effect of increasing species richness to West Nile virus transmission in Chicago. Other intrinsic and extrinsic factors, such as variation in mosquito host preference, reservoir host competence, temperature, and precipitation, may be more important than host diversity for driving interannual variation in WNV transmission. These results from a fine-scale study call into question the generality of a dilution effect for WNV at coarser spatial scales.     

Microsatellite loci from the northern house mosquito (Culex pipiens), a principal vector of West Nile virus in North America

Microsatellites were isolated and characterized in the northern house mosquito, Culex pipiens, a widespread pest species and important vector of diseases such as West Nile virus. An enrichment protocol yielded 150 positive clones. We designed primers to amplify 17 unique (GT)_n microsatellites, eight of which amplified cleanly and were polymorphic. A survey of 29 individuals showed that these loci are highly variable with the number of alleles ranging from seven to 19 and expected heterozygosity ranging from 0.66 to 0.93. These markers will be useful for studies of population structure and intraspecific variation in epidemiological characteristics of Cx. pipiens.     

Mosquito and West Nile virus surveillance in northeast Montana,U.S.A., 2005 and 2006

Mosquito and West Nile virus (WNV) surveillance was conducted on a national wildlife refuge in northeast Montana in 2005 and 2006, during which outbreaks of WNV in a colony of American white pelicans (Pelecanus erythrorhynchos Gmelin) (Pelecaniformes: Pelecanidae) resulted in juvenile mortality rates of ～ 31%. Both years, floodwater species Ochlerotatus dorsalis (Meigen) (Diptera: Culicidae), Aedes vexans (Meigen) (Diptera: Culicidae) and Ochlerotatus flavescens (Muller) (Diptera: Culicidae) comprised 78% of the total collection and heightened host‐seeking activity was observed from mid‐June to mid‐July. Culex tarsalis Coquillett (Diptera: Culicidae) was most active from mid‐July to mid‐August and comprised 18% of the collection in 2005 and 20% in 2006. However, fewer than 10% of the Cx. tarsalis females collected in 2006 were obtained adjacent to the pelicans' nesting grounds. Minimum infection rates per 1000 Cx. tarsalis tested for WNV were 1.36 in 2005 and 1.41 in 2006. All pools in which WNV was detected in 2006 were composed of females collected 10 km from the nesting grounds. Substantial juvenile pelican mortality in 2006 despite reductions in the population of the primary vector and in mosquito infection rates near the colony suggests that the methods used to detect the introduction of WNV were too coarse and that amplification of the virus within the colony may reflect causes other than mosquito infection.     

Inhibitory effect of Ocotea quixos (Lam.) Kosterm. and Piper aduncum L. essential oils from Ecuador on West Nile virus infection

West Nile virus (WNV) is a mosquito-borne flavivirus responsible of neuroinvasive manifestations. Natural products are well-known for their biological activities and pharmaceutical application. In this study, the inhibitory effects of essential oils (EOs) of Ocotea quixos (Lam.) Kosterm. and Piper aduncum L. on WNV replication were investigated. WNV was incubated with EOs before adsorption on Vero cells, viral replication was carried out in the absence or presence of EO. Cells were exposed to EO before the adsorption of untreated-virus. GC-MS and GC-FID were used for chemical characterization of EOs. Cell protection from infection was observed for both EOs. P. aduncum EO was characterized by dillapiole as main compound (48.21%) and O. quixos EO by 1,8-cineole (39.15%). Further investigations, such as the study of molecular and cellular mechanisms of action and in vivo evaluation, should be performed on these essential oils to derive new potential drugs against WNV.     

Evaluation of factors for rapid development of Culex quinquefasciatus in belowground stormwater treatment devices

Water samples from 11 belowground stormwater treatment Best Management Practices (BMPs) were evaluated for their capacity to support rapid development of the West Nile virus (WNV) mosquito vector, Culex quinquefasciatus. The observed minimum development time from egg to pupa ranged from six to over 30 days. Concentrations of potential food resources (total suspended solids and the particulate organic matter in water samples) were significantly correlated to development times. In addition, the rate of immature mosquito development was both site‐dependent and variable in time, suggesting that factors favorable to rapid development were strongly influenced by watershed characteristics and seasonal changes in temperature. Measured temperatures in belowground BMPs suggest that these structures may remain amenable to WNV virus activity longer each year than sites aboveground.     

Mosquitoes and West Nile virus along a river corridor from prairie to montane habitats in eastern Colorado

We conducted studies on mosquitoes and West Nile virus (WNV) along a riparian corridor following the South Platte River and Big Thompson River in northeastern Colorado and extending from an elevation of 1,215 m in the prairie landscape of the eastern Colorado plains to 1,840 m in low montane areas at the eastern edge of the Rocky Mountains in the central part of the state. Mosquito collection during June‐September 2007 in 20 sites along this riparian corridor yielded a total of 199,833 identifiable mosquitoes of 17 species. The most commonly collected mosquitoes were, in descending order: Aedes vexans, Culex tarsalis, Ae. dorsalis, Ae. trivittatus, Ae. melanimon, Cx. pipiens, and Culiseta inornata. Species richness was higher in the plains than in foothills‐montane areas, and abundances of several individual species, including the WNV vectors Cx. tarsalis and Cx. pipiens and the nuisance‐biter and potential secondary WNV vector Ae. vexans, decreased dramatically from the plains (1,215‐1,487 m) to foothills‐montane areas (1,524‐1,840 m). Ae. vexans and Cx. tarsalis had a striking pattern of uniformly high abundances between 1,200‐1,450 m followed by a gradual decrease in abundance above 1,450 m to reach very low numbers above 1,550 m. Culex species were commonly infected with WNV in the plains portion of the riparian corridor in 2007, with 14 of 16 sites yielding WNV‐infected Cx. tarsalis and infection rates for Cx. tarsalis females exceeding 2.0 per 1,000 individuals in ten of the sites. The Vector Index for abundance of WNV‐infected Cx. tarsalis females during June‐September exceeded 0.5 in six plains sites along the South Platte River but was uniformly low (0–0.1) in plains, foothills and montane sites above 1,500 m along the Big Thompson River. A population genetic analysis of Cx. tarsalis revealed that all collections from the ≈190 km riparian transect in northeastern Colorado were genetically uniform but that these collections were genetically distinct from collections from Delta County on the western slope of the Continental Divide. This suggests that major waterways in the Great Plains serve as important dispersal corridors for Cx. tarsalis but that the Continental Divide is a formidable barrier to this WNV vector.     

Butterflies of the South Pacific

ABSTRACT

Larvae of the invasive mosquito Culex quinquefasciatus Say are morphologically similar to those of the native Culex pervigilans Bergroth, yet distinguishing these species can be hampered by morphological variations in Cx. quinquefasciatus. We present detail about the extent of these variations in an urban population of Cx. quinquefasciatus in Auckland. To aid in identification of this exotic species, we provide images of key diagnostic characters and some observed exceptions to these. Details regarding variation in diagnostic characters for < 3rd instar and 3rd/4th instar larvae are given. Of the nine characters used for identification, three were highly consistent (dorsal papillae, mantle plate, pecten teeth); each observed in > 90% of larvae, although these characters were not always visible. Other characters were less reliable, for instance, the expected position of seta 1a-S in relation to the pecten teeth was observed in < 10% of larvae. Further exploration of regional morphological variation in both Cx. quinquefasciatus and Cx. pervigilans is recommended, ideally with associated molecular characterisation.     

The Effect of West Nile Virus Perceptions and Knowledge on Protective Behavior and Mosquito Breeding in Residential Yards in Upstate New York

A knowledge, attitudes, and practices (KAP) questionnaire combined with entomological surveys of residential mosquito-breeding sites were conducted in two Upstate New York neighborhoods. We tested the hypothesis that “correct” West Nile virus (WNV) knowledge and perceptions correspond with the use of practices that prevent mosquitoes from breeding and biting. Our results demonstrate that perceptions of WNV relate to the number of positive containers in yards and the use of mosquito preventive measures. In contrast, WNV knowledge was not related. Culex pipiens and Cx. restuans were common species found breeding in containers. Aedes japonicus was the most abundant species in 77% of positive containers (buckets, flower pots, and birdbaths). This new, invasive mosquito together with the Culex species identified in this study represent significant potential as vectors of WNV and other arboviruses affecting human and animal health. We conclude that more training and education programs should focus on WNV control strategies and recognizing mosquito breeding in residential yards. This is the first study to directly investigate the relationship between KAP and breeding of WNV vectors in residential yards.