Host preferences of Aedes trivittatus (Diptera: Culicidae) in central Iowa.

The vertebrate host preferences of Aedes trivittatus mosquitoes were studied to gain an insight into the possible hosts of trivittatus (TVT) virus (California encephalitis group) in Iowa. Engorged mosquitoes were collected with a Malaise trap and Dry Ice-baited CDC miniature light traps. The origin of mosquito blood meals was determined by the capillary tube precipitin test. Of 600 A. trivittatus blood meals tested, 409 (68.2%) reacted positively with anti-rabbit serum. The incidences of mosquitoes feeding on other vertebrate species ranged from 0.2% to 2.5%. The vertebrate host preferences of A. trivittatus suggest a close association between this mosquito species and the eastern cottontail rabbit (Sylvilagus floridanus) in central Iowa. Furthermore, the results indicate that the eastern cottontail rabbit may be an important host for TVT virus in Iowa.     

Host feeding patterns of Connecticut mosquitoes (Diptera: Culicidae).

Blood-engorged Coquillettidia perturbans, Psorophora ferox, Culex, Culiseta, and Aedes mosquitoes were collected principally by sweep net from salt marsh and woodland habitats in Connecticut. Of the 570 mosquitoes tested, precipitin tests identified the origins of 517 blood meals and revealed distinct host feeding patterns. Aedes mosquitoes fed chiefly on mammals; A. abserratus, A. cantator, and A. vexans showed selectivity for cattle and (or) horses. A. cantator also obtained blood from avian hosts and, in some instances, showed mixed passerine-mammal blood meals. These findings increase the vector potential of this salt marsh mosquito for eastern equine encephalomyelitis virus. Feedings on deer by A. abserratus suggest potential involvement of this mosquito in the transmission of certain subtypes of California encephalitis. Culex-pipiens, C. restuans, Culiseta melanura, and Cs. morsitans dyari acquired blood almost exclusively from passeriform birds.     

Molecular identification of blood-meal sources in Culiseta melanura and Culiseta morsitans from an endemic focus of eastern equine encephalitis virus in New York

Eastern equine encephalitis (EEE) virus perpetuates in an enzootic cycle involving ornithophilic mosquito vectors, principally Culiseta melanura (Coquillett) and avian amplification hosts. To better understand the role of Cs. melanura and Culiseta morsitans (Theobald) in the epizootiology of EEE virus, we collected blood-fed mosquitoes between 31 May and 15 October 2004 at two sites associated with an EEE virus focus in central New York and identified the source of vertebrate blood by nucleotide sequencing of polymerase chain reaction (PCR) products of the cytochrome b gene. Analysis of 484 Cs. melanura and 122 Cs. morsitans revealed that 94.2% and 86.9%, respectively, acquired blood solely from avian hosts. Blood meals derived exclusively from mammals were detected in 0.8% of Cs. melanura and 1.6% of Cs. morsitans. Individual mosquitoes containing mixed-blood meals from both avian and mammalian hosts were also detected in 5.0% of Cs. melanura and 11.5% of Cs. morsitans. Wood thrush constituted the most common vertebrate host for Cs. melanura (23.6%) and Cs. morsitans (30.9%), followed by American robin, song sparrow, ovenbird, red-eyed vireo, and common yellowthroat. Mammalian-derived blood meals were identified as white-tailed deer, horse, domestic cat, and eastern pipistrelle bat. There were three isolations of EEE virus from Cs. melanura and one from Cs. morsitans. These results suggest that wood thrush and a few other passerine birds may play key roles in supporting EEE virus transmission in the northeast and possibly throughout the geographic range of EEE in North America. The frequency of mammalian feedings also suggests that Cs. melanura and Cs. morsitans may play a role in the transmission of EEE virus to equines, in addition to maintaining enzootic transmission among avian hosts. We report the first isolation of arboviruses from mosquito vectors concomitant with the identifications of their blood meal sources.     

应用多重PCR法鉴定蚊胃血血源的研揪

目的建立多重PCR法鉴定蚊胃血来源。方法依据常见蚊吸血对象(人、牛、猪和犬)的线粒体DNA细胞色素b序列的差异,设计种特异引物,建立多重PCR法,并应用该法检测现场按蚊标本。结果应用多重PCR法共检测249只按蚊,血源来自牛和猪的共91只和63只,未检出吸人血按蚊。立即处死并干燥保存的现场按蚊标本检测成功率最高,为92.50%。结论多重PCR法鉴定蚊胃血血源快速、灵敏,结果客观、可靠。     

Temporal analysis of feeding patterns of Culex erraticus in central Alabama

Host blood meals in seven mosquito species previously shown to be infected with eastern equine encephalitis virus at a site in the Tuskegee National Forest in southcentral Alabama were investigated. Of 1374 blood meals derived from 88 different host species collected over 6 years from these seven mosquito species, 1099 were derived from Culex erraticus. Analysis of the temporal pattern of Cx. erraticus meals using a Runs test revealed that the patterns of feeding upon avian and mammalian hosts from March to September of each year were not randomly distributed over time. Similarly, meals taken from the three most commonly targeted host species (yellow-crowned night heron, great blue heron, and white-tailed deer) were not randomly distributed. A Tukey's two-way analysis of variance test demonstrated that although the temporal pattern of meals taken from avian hosts were consistent over the years, the patterns of meals taken from the individual host species were not consistent from year to year.     

Quantifying mosquito biting patterns on humans by DNA fingerprinting of bloodmeals.

A major debate in infectious disease epidemiology concerns the relative importance of exposure and host factors, such as sex and acquired immunity, in determining observed age patterns of parasitic infection in endemic communities. Nonhomogeneous contact between hosts and vectors is also expected to increase the reproductive rate, and hence transmission, of mosquito-borne infections. Resolution of these questions for human parasitic diseases has been frustrated by the lack of a quantitative tool for quantifying the exposure rate of people in communities. Here, we show that the polymerase chain reaction (PCR) technique for amplifying and fingerprinting human DNA from mosquito bloodmeals can address this problem for mosquito-borne diseases. Analysis of parallel human and mosquito (resting Culex quinquefasciatus) samples from the same households in an urban endemic focus for bancroftian filariasis in South India demonstrates that a 9-locus radioactive short-tandem repeat system is able to identify the source of human DNA within the bloodmeals of nearly 80% of mosquitoes. The results show that a person's exposure rate, and hence the age and sex patterns of exposure to bites in an endemic community, can be successfully quantified by this method. Out of 276 bloodmeal PCR fingerprints, we also found that on average, 27% of the mosquitoes caught resting within individual households had fed on people outside the household. Additionally, 13% of mosquitoes biting within households contained blood from at least 2 people, with the rate of multiple feeding depending on the density of humans in the household. These complex vector feeding behaviors may partly account for the discrepancies in estimates of the infection rates of mosquito-borne diseases calculated parasitologically and entomologically, and they underline the potential of this tool for investigating the transmission dynamics of infection.     

Detection of host virus-reactive antibodies in blood meals of naturally engorged mosquitoes

Although serosurvey in human or animals is a useful and straightforward strategy routinely used for public health, it often faces different types of impediments: ethics, beliefs, limitation by animal owners, hazard of access to wild animals. To survey virus circulation, we applied the enzyme-linked immunosorbent assay (ELISA) technique to detect Dengue and Japanese encephalitis (JE) virus-reactive antibodies in blood meals collected from mosquitoes without regard to the potential of mosquito species to be a virus vector. ELISA was performed on mosquito colonies and wild specimens collected from farms and urban areas. Blood meals from Aedes aegypti freshly fed on naturally infected volunteers showed the same levels of dengue immunoglobulin (Ig)G and IgM as the sera directly collected from volunteers. A significant clearance of antibodies during the digestion process started from 13 hours after blood meal, and a negative baseline was reached after 30 hours. The ELISA test performed on wild mosquitoes showed that 37% of Culex quinquefasciatus mosquitoes that engorged on humans in a dengue urban endemic area tested positive for dengue IgG, and in a JE virus-endemic area, 88% of Culex tritaeniorhynchus mosquitoes that engorged on pigs from a large pig farm tested positive for JE virus antibodies versus 11% in a small farm. The main limitation of the ELISA method is the antibody cross-reactivity among flaviviruses; also, sampling strategy should be adjusted to take into account that the actual host from which the blood meal was taken may not be determined. Nevertheless, ELISA performed on recently (1-2 days) engorged mosquito, or any other hematophagous arthropod species, could potentially be used as a "wild phlebotomist" to monitor the prevalence or emergence of a variety of pathogens, with less of the practical, ethical, or risk limitations due to direct blood collection from humans and wild or domestic animals.     

Unexpected anthropophily in the potential secondary malaria vectors Anopheles coustani s.l. and Anopheles squamosus in Macha, Zambia

Anopheles coustani s.l. and Anopheles squamosus are sub-Saharan mosquito species that have been implicated in malaria transmission. Although generally believed to be of negligible importance due to their overwhelmingly zoophilic behavior, An. coustani s.l. and An. squamosus made up a large proportion of the anophelines collected by human landing catches during the 2007-2008 and 2008-2009 rainy seasons in Macha, Zambia. Further, polymerase chain reaction-based blood meal identification showed that the majority of blood meals from these mosquito species caught in human-baited Centers for Disease Control light traps were from human hosts. Although no An. coustani s.l. or An. squamosus were found to be positive for Plasmodium, the demonstrated anthropophilic tendencies of these mosquitoes in southern Zambia suggest their potential as secondary malaria vectors.     

Identification of mosquito blood meals by enzyme-linked immunosorbent assay

A micro enzyme-linked immunosorbent assay for identifying mosquito blood meals is described. Antisera for the assay were prepared in rabbits and guinea pigs and purified by precipitation with cross-reacting blood sera. Positive blood source identifications could be made to the generic level using as little as 58 micrograms of ingested blood serum. The blood meals of 37 of 38 laboratory mosquitoes fed on seven host animals were correctly identified, including a mosquito that contained a double feed.     

Physico-chemical signals involved in host localization and in the induction of mosquito bites

Disease vector female mosquitoes respond to physic-chemical signals to localize vertebrate hosts for blood meals. Zoophylic mosquitoes preferentially respond to CO2 and octenol released in the breath and bodily fluids, while anthropophylic mosquitoes respond to lactic acid and a variety of sweat compounds. These compounds are modified by saprophytic microorganisms in the skin sebaceous glands. Other factors present in human dwellings contribute to the integration of microsystems with characteristic odors that have different attraction for mosquitoes, explaining the focalization of malaria transmission in few households in endemic areas. The identification of the chemical attractants and their molecular receptors could be used to complement new methods to attract mosquitoes to traps during epidemiological surveys, to increase their contact with insecticides in control interventions, and for genetic manipulation to divert mosquito bites towards other animal populations. The English version of this paper is available at:http://www.insp.mx/salud/index.html.     

Identification of mosquito avian-derived blood meals by polymerase chain reaction-heteroduplex analysis

A polymerase chain reaction (PCR) heteroduplex assay (HDA) was developed to identify avian derived mosquito blood meals to the species level. The assay used primers amplifying a fragment of the cytochrome B gene from vertebrate but not invertebrate species. In Culex tarsalis fed on quail, PCR products derived from the quail cytochrome B gene were detected seven days post-engorgement. In an analysis of wild-caught mosquitoes, 85% of blood-fed mosquitoes produced detectable PCR products. Heteroduplex patterns obtained from bird-derived PCR products were found to permit the unambiguous identification of all species examined. No intraspecific variation in HDA patterns was found. The PCR-HDA was used to characterize blood meals in wild caught Cx. tarsalis. Of the 67 blood meals analyzed, 60% were derived from avian sources. Of the avian blood meals, 65% were derived from a single host, the common grackle.     

Heterogeneity and Changes in Inequality of Malaria Risk after Introduction of Insecticide-Treated Bed Nets in Macha,Zambia

In 2007, the first free mass distribution of insecticide-treated bed nets (ITNs) occurred in southern Zambia. To determine the effect of ITNs on heterogeneity in biting rates, human DNA from Anopheles arabiensis blood meals was genotyped to determine the number of hosts that had contributed to the blood meals. The multiple feeding rate decreased from 18.9% pre-ITN to 9.1% post-ITN, suggesting that mosquito biting had focused onto a smaller fraction of the population. Pre-ITN, 20% of persons in a household provided 40% of blood meals, which increased to 59% post-ITN. To measure heterogeneity over a larger scale, mosquitoes were collected in 90 households in two village areas. Of these households, 25% contributed 78.1% of An. arabiensis, and households with high frequencies of An. arabiensis were significantly spatially clustered. The results indicate that substantial heterogeneity in malaria risk exists at local and household levels, and household-level heterogeneity may be influenced by interventions, such as ITNs.     

Application of one step RT-PCR assay for detection of flavivirus RNA in mosquitoes

The conditions of one step RT-PCR method for detection of virus RNA in field-collected mosquitoes, and preservation period of infected mosquitoes for one step RT-PCR were examined. We compared several virus RNA extraction methods with artificially contaminated mosquito pools with dengue virus (DV), Japanese encephalitis virus (JEV), and yellow fever virus (YFV) with a known amount of plaque forming unit (PFU) to establish the condition of one step RT-PCR. In this study, most effective RNA extraction method was ISOGEN-LS extraction combined with supernatant of centrifuged mosquito homogenates. Detection limit of one step RT-PCR using flavivirus universal primer in ten mosquitoes/tube (pool) was 10 PFU of DV, JEV and YFV, 1 PFU of each viruses using species-specific primer respectively, in one hundred mosquitoes/tube, 100 PFU/tube using universal primer pairs, 10 PFU/tube using species-specific primer pairs respectively. Dengue virus infected single mosquito was mixed with 99 un-infected mosquitoes, and tested by one step RT-PCR. We could detect single infected mosquito in pools containing 99 un-infected mosquitoes. Aedes aegypti and Aedes albopictus were inoculated intrathoracically with a mouse-adapted strain of dengue-1 virus and were kept up to 30 days at different temperature. Then examined by one step RT-PCR to determine the appropriate mosquito handling method and the condition of transportation. Positive result was obtained up to 30 days after the mosquito died naturally. These results suggested that we could detect flavivirus RNA tested not only from live mosquitoes but also dead mosquitoes as well, and could apply one step RT-PCR as a rapid, specific, and highly sensitive tool for flavivirus surveillance.     

Nutritional stress affects mosquito survival and vector competence for West Nile virus

Most anautogenous female mosquitoes ingest plant carbohydrates for flight energy and survival, and they imbibe vertebrate blood for egg development. We evaluated the effect of different sucrose meals following a blood meal containing West Nile virus (WNV) on Culex pipiens pipiens survival, nutritional status, and susceptibility to viral infection and transmission. Ten days after blood feeding, no mosquitoes survived on distilled water, 55% survived on 2% sucrose, 61% on 10 and 20% sucrose meals, and over 70% survived on 40% sucrose. There was a positive correlation between sucrose meal concentration and detectable sugars, glycogen, and lipid in whole-body homogenates. Average sugar values increased from 0 microg per starved mosquito (range 0-1.0 microg) to an average of 392 microg per mosquito fed on 40% sucrose (85-1088 microg). Average glycogen values increased from 0 microg (0-5.7 microg) to an average of 620 microg (118-1421 microg). Average lipid values were identical for mosquitoes in the starved and 2% sucrose series (38 microg) and increased to 172 microg per mosquito fed on 40% sucrose (92-266 microg). Mosquitoes in all sucrose series were equally susceptible to WNV infection (p > 0.5), but mosquitoes with lower nutrient reserves as a result of lower sucrose meals were more likely to orally transmit virus (p < 0.05). We discuss how mosquito nutritional status influences probability of daily survival, susceptibility to infection, and vectorial capacity. We conclude that maintaining C. p. pipiens on standard 10% sucrose is justified in light of these results.     

三带喙库蚊和致倦库蚊经口感染乙型脑炎病毒疫苗SA14-14-2株的研究

目的通过实验，确定被SA14—14—2乙脑减毒活疫苗免疫的宿主动物和人在被媒介蚊虫叮咬后，是否存在感染和传播的可能。方法建立三带喙库蚊和致倦库蚊的实验室种群，用乙型脑炎SA14—14—2疫苗株病毒和乙脑野毒株经口感染两种库蚊，感染后不同时间取一定数量的蚊，研磨制成悬液，应用空斑试验方法检测感染后不同时间蚊体内的病毒存在情况和病毒滴度。结果在用6．0610gPFU／ml SA14～14—2病毒经口感染的两种库蚊中，没有检测到病毒空斑，即没发现蚊虫的感染；在用较高的6．1810gPFU／ml病毒经口感染的三带喙库蚊和致倦库蚊中，分别有一组蚊虫出现低滴度感染，空斑形成单位分别是1．24和1．11log10PFU／ml；在用野毒株经口感染的两种库蚊，共计19组蚊虫中，有14组发生感染，空斑形成单位在3．18～4．7910g10PFU／ml。结论作为乙脑主要传播媒介的三带喙库蚊和致倦库蚊对SA14—14—2疫苗株病毒的经口感染和病毒在体内的复制严重受限，当叮咬接种疫苗的人后，不具备发生乙脑病毒感染和传播的能力。     

Avian host preference by vectors of eastern equine encephalomyelitis virus

An important variable in the amplification and escape from the enzootic cycle of the arboviral encephalitides is the degree of contact between avian hosts and mosquito vectors. To analyze this interaction in detail, blood-fed mosquitoes that were confirmed vectors of eastern equine encephalomyelitis (EEE) virus were collected in 2002 from an enzootic site in central Alabama during the time this virus was actively transmitted. Avian-derived blood meals were identified to the species level of the host, and the proportion derived from each species was compared with the overall composition of the avifauna at the study site. The EEE vector mosquito species fed significantly more on some bird species and less on other species than expected given the overall abundance, biomass, or surface area of the local avifauna. When viewed collectively, these data suggest that these mosquitoes are selectively targeting particular avian species.     

Blood meal size as a factor affecting continued host-seeking by Aedes aegypti (L.)

The effect of ingested blood on the host-seeking response of two strains of Aedes aegypti was examined. Using an olfactometer, females fed partial blood meals were scored for host-seeking behavior within 1 h, and their blood meal sizes were measured chemically immediately afterwards. The suppression of host-seeking within 1 h after a blood meal appears to be caused by abdominal distention from ingested blood. Mosquitoes of either strain were attracted to a host when the blood meal size was less than 2.5 microliter; above this threshold there was a sharp decline in the tendency to respond. Small mosquitoes resulting from a low larval diet had a lower threshold, and were more likely to cease host-seeking after a small blood meal. Multiple feeding within a single gonotrophic cycle may result if mosquitoes take small blood meals which are insufficient to terminate host-seeking. Partial meals and reduced feeding success of mosquitoes can result from defensive host behavior, which in the laboratory rat was shown to increase at high mosquito densities.     

Host Selection of Potential West Nile Virus Vectors in Puerto Barrios,Guatemala, 2007

The selection of vertebrate hosts by Culex mosquitoes relative to West Nile virus (WNV) transmission in neotropical countries such as Guatemala is not described. This study determined the feeding patterns of Cx. quinquefasciatus and Cx. nigripalpus and estimated the relative contribution of two common and frequently infected wild bird species, Turdus grayi and Quiscalus mexicanus, to WNV transmission. Engorged mosquitoes were collected from rural and urban habitats after the dry and wet seasons in the Department of Izabal in 2007. Host selection by Cx. nigripalpus varied significantly between urban and rural habitats. Both Cx. quinquefasciatus and Cx. nigripalpus fed predominantly on chickens and other domestic animals. Blood meals from wild birds were rare, accounting for 1.1% of blood meals identified from Cx. quinquefasciatus and 6.5% of blood meals from Cx. nigripalpus. Transmission of WNV by these two mosquito species may be dampened by extensive feeding on reservoir-incompetent hosts.     

Blood meals of mosquito vectors of filariasis in Guizhou Province, China

The blood meals of mosquito vectors of W. bancrofti and B. malayi were determined by performing CIEP of the eluant of crushed mosquitoes in filter paper against rabbit anti-human, cow and pig sera. The mosquitoes were collected from houses, cowsheds and pigpens in two counties in Guizhou Province. It was shown that all three species fed on blood from humans, cows and pigs with different preference. While An sinensis fed more on cows, Cx. fatigans and An. lesteri fed on the hosts that were nearby, i.e., Cx. fatigans caught from households fed more on humans, and those collected from cowsheds or pigpens fed more on cows and pigs, respectively.