Search

Background: Mosquito biting frequency and how bites are distributed among different people can have significant epidemiologic effects. An improved understanding of mosquito vector-human interactions would refine knowledge of the entomological processes supporting pathogen transmission and could reveal targets for minimizing risk and breaking pathogen transmission cycles., Methodology and Principal Findings: We used human DNA blood meal profiling of the dengue virus (DENV) vector, Aedes aegypti, to quantify its contact with human hosts and to infer epidemiologic implications of its blood feeding behavior. We determined the number of different people bitten, biting frequency by host age, size, mosquito age, and the number of times each person was bitten. Of 3,677 engorged mosquitoes collected and 1,186 complete DNA profiles, only 420 meals matched people from the study area, indicating that Ae. aegypti feed on people moving transiently through communities to conduct daily business. 10-13% of engorged mosquitoes fed on more than one person. No biting rate differences were detected between high- and low-dengue transmission seasons. We estimate that 43-46% of engorged mosquitoes bit more than one person within each gonotrophic cycle. Most multiple meals were from residents of the mosquito collection house or neighbors. People ≤ 25 years old were bitten less often than older people. Some hosts were fed on frequently, with three hosts bitten nine times. Interaction networks for mosquitoes and humans revealed biologically significant blood feeding hotspots, including community marketplaces., Conclusion and Significance: High multiple-feeding rates and feeding on community visitors are likely important features in the efficient transmission and rapid spread of DENV. These results help explain why reducing vector populations alone is difficult for dengue prevention and support the argument for additional studies of mosquito feeding behavior, which when integrated with a greater understanding of human behavior will refine estimates of risk and strategies for dengue control.

We conducted a study to determine the effect of container size and location on oviposition site selection by Ae. aegypti in large outdoor field enclosures (10 x 10 x 4 m high). There was a strong positive relationship between increasing container diameter, container volume, and water surface area with egg numbers over both high (rainy, July) and low (cool-dry, January) dengue transmission seasons. Location of containers (indoors versus immediately outdoors and underneath houses) did not influence the number of eggs deposited for containers 5-32 cm in diameter in either season. No trends based on container color (black, brown, or grey) were observed. A slight trend with a greater numbers of eggs laid outdoors in the largest containers (42 cm diameter) during the dry season was observed. Three separate models were run using the mixed model procedure in SAS for each container attribute. Controlling for season, time, and date, the most important container attribute predicting total egg numbers was container volume (total capacity) explaining 88% of the variation, followed by water surface area (85%), and container diameter opening (83%). Oviposition peaked in the afternoon at 1600 hrs and 2000 hrs in the dry and rainy seasons, respectively. Few eggs were laid overnight (2000 hrs-0600 hrs). Our results indicate that physical attributes of oviposition sites, such as size, light-dark contrasts, and specular reflectance from water surfaces, play a significant role in oviposition site selection.

The assumption that mosquito survival remains constant and that it is independent of age was tested with free-ranging Aedes aegypti (L.) (Diptera: Culicidae) by using a "synthetic cohort" mark-release-recapture method. Mosquito age cohorts (1-2, 3-4, and 19-20 d) were released simultaneously in homes in rural Thai villages during dry and rainy seasons (2002). Significant age-dependent effects were detected during the dry and rainy seasons. More young than older mosquitoes were recaptured (1-4 versus 5-20 d). The best predictive fit for recaptures was obtained using a quadratic function of age. Our results provide the first field-based evidence for age-dependent Ae. aegypti mortality in which the death rate increases with advancing age and highlight the need for research on ecological and epidemiological aspects of this process.

Background: Millions of people are infested by head lice every year. However, louse transfer between hosts is not well-understood. Our goals were to determine: (1) which stages were most likely to disperse and why, (2) the likelihood of fomites transmission, and (3) if host blood gender affects louse development., Methods: Various life stages of lice at differing densities were permitted to cross over a 15-cm hair bridge placed between two artificial blood-feeding arenas. Louse transfer caused by hot air movements, combing, toweling, and passive transfer to fabric was investigated. The ability of lice to oviposit on different foreign substrates and the hatching potential of eggs intermittently incubated for 8 h/night on a host were likewise investigated. Louse in vitro development following feeding on human female or male donor blood was compared., Results: Adult lice were the most likely to disperse. Neither population density nor hunger significantly affected dispersal tendencies. Lice were dislodged by air movement, combs and towels, and passively transferred to fabric within 5 min. Females oviposited on a variety of substrates and 59% of eggs incubated for 8 h/night hatched after 14-16 days. There was no survivorship difference between lice artificially fed on female vs. male blood., Conclusions: Adult lice are the most mobile, indicating that they are most likely to initiate new infestations. Although head-to-head contact may be the primary route of transmission, less direct routes involving fomites may play a role and need further evaluation. Blood-borne factors do not appear to cause any gender-biased host preference.

Knowledge of mosquito dispersal is critical for vector-borne disease control and prevention strategies and for understanding population structure and pathogen dissemination. We determined Aedes aegypti flight range and dispersal patterns from 21 mark-release-recapture experiments conducted over 11 years (1991-2002) in Puerto Rico and Thailand. Dispersal was compared by release location, sex, age, season, and village. For all experiments, the majority of mosquitoes were collected from their release house or adjacent house. Inter-village movement was detected rarely, with a few mosquitoes moving a maximum of 512 meters from one Thai village to the next. Average dispersal distances were similar for males and females and females released indoors versus outdoors. The movement of Ae. aegypti was not influenced by season or age, but differed by village. Results demonstrate that adult Ae. aegypti disperse relatively short distances, suggesting that people rather than mosquitoes are the primary mode of dengue virus dissemination within and among communities.

As the frequency and level of pediculicide resistance increases throughout the world, the need for novel solutions to control pediculosis has intensified. The development and registration of new pesticides has become so costly that many chemical companies are unwilling to pursue it and health-care providers now face a serious lack of new commercial pediculicides. Many infested people resort to using "home-remedy" approaches that have not been scientifically tested. In this article, we examined the potential value of six purportedly effective "home remedies" (vinegar, isopropyl alcohol, olive oil, mayonnaise, melted butter, and petroleum jelly) to treat head louse infestations and the likelihood of drowning lice by water submersion. Results indicated that only the application of petroleum jelly caused significant louse mortality but no treatment prevented lice from laying eggs. Most home remedy products did little to kill eggs, despite prolonged exposure. Petroleum jelly caused the greatest egg mortality, allowing only 6% to hatch. It was extremely difficult to drown lice, despite extended periods (i.e., 8 hr) of water submersion, suggesting that killing lice by depriving them of oxygen is inefficient. None of the home remedy products we surveyed was an effective means of louse control. This suggests that when treatment failure occurs, an increased amount of time and effort should be focused on alternative chemical pediculicides and/or manual louse removal (i.e., combing) rather than using any of these products.

Temporal changes in the cuticular hydrocarbons of female Anophelesstephensi (Liston) (Diptera: Culicidae) were quantified using gas-liquid chromatography with flame-ionization detection. The ratio of two prominent hydrocarbons, nonacosane (C29) and hentriacontane (C31), was found to change significantly with respect to mosquito age over a period of 15 d. A regression model was developed using this ratio, C29/C31 = 3.96 - 1.63 log (age), and prediction intervals, based on a 12-d developmental interval necessary for females to transmit malaria, were generated using confidence levels for one-sided tests. The model predicted that females that had a C29/C31 ratio of 2.6 or greater were only 10% probable to be old enough to transmit malaria, whereas females with ratios of 1.8 or less were 90% probable.

In previous studies, we developed linear regression models to age-grade female Aedes aegypti L. reared and maintained under controlled laboratory conditions. The models were based on temporal differences between two cuticular hydrocarbons, pentacosane (C25H52) and nonacosane (C29H60), which were extracted from Ae. aegypti legs and analyzed by gas-liquid chromatography. These initial models predicted adult female age up to 165 DD (12-15 calendar d at 28 degrees C). The age of older mosquitoes, however, could not be accurately predicted. In this study, our original regression models were tested using age data obtained from mosquitoes maintained in a field laboratory and those that were marked, released, and recaptured in northwestern Thailand. Our field data led to the development of two new regression models: one for the cool-dry season (February-March) and one for the rainy season (July-August). Both models resulted in better estimates of age than the original model and thus improved our ability to predict the age of Ae. aegypti to 15 calendar d. Females older than 15 d can be identified as such, but their exact age cannot yet be estimated. The new models will be useful for epidemiological studies and evaluating the impact of Ae. aegypti control interventions for disease prevention.

An automated feeding apparatus was developed to maintain the human head louse (Pediculus capitis DeGeer) in vitro. With the use of valves and timers, banked human blood and saline from refrigerated reservoirs were pumped into and flushed out of the system every 7 d. During this rotational interval, bloodmeals were provided to head lice continuously and ad libitum through a stretched Nescofilm-silicone sandwich membrane. Compared with our previous in vitro human head louse-rearing apparatus, greater numbers of lice could be fed simultaneously with minimal human monitoring. Development of second to third instars and third instars to adults was significantly faster when lice were reared in vivo than on either of the in vitro rearing systems; there was no significant difference in the duration of the first instar. Although fecundity and hatch rates were significantly higher for female lice reared in vivo, similar trends have been observed for other membrane-fed arthropods. Body lice (Pediculus humanus L.) and bed bugs (Cimex lectularius [L.]) also completed most of their life cycle on this apparatus. Our automated mass-rearing system has broad applications for maintaining fluid-sucking ectoparasites and will facilitate various toxicological, behavioral, and disease-transmission investigations.

Objective: To differentiate groups of highly conditioned, competitive athletes on the basis of Exercise Orientation Questionnaire (EOQ) scores and self-reported psychiatric symptoms., Method: A total of 99 runners, 36 cyclists, and 55 paddlers were administered the EOQ and a symptom checklist., Results: Analysis of variance and chi-square associated self-loathing subscale (SLSS) scores with self-reported eating disorder (ED) symptoms for the entire group [F(1)=4.83; P<.05] and for females only [F(1)=9.30; P<.001]. The paddler group reported more anxiety/panic symptoms than runners or cyclists (chi(2)=7.91; P<.01) and higher SLSS scores than cyclists [F(2)=6.91; P<.01]. The female-only paddler group reported more anxiety/panic [chi(2)(1)=10.27; P<.001] than the other groups. Although total group scores were comparable to controls, 12% runners, 14% cyclists, and 18% paddlers scored above previously established ED patient mean., Discussion: The paddler group appeared less healthy than runners or cyclists and were more likely to report ED symptoms. These differences could have been related to a dissimilar ethnic composition or particular social stress generated within paddling groups. In this study, SLSS was highly specific in its association with ED symptoms. This is of particular interest because SLSS is based on questions about exercise rather than eating pathology.

Four geographically distinct colonies of the human head louse, Pediculus humanus capitis De Geer (Anoplura: Pediculidae) were reared on a live host and exhibited significantly different life history patterns. Florida head lice exhibited approximately 10% slower development and approximately 15% reduced longevity relative to California or Ecuador head lice. Fecundity (4.9 +/- 0.2 eggs/female/d) and fertility (76.4 +/- 2.9% mean hatching rate) declined over the lifetime of female lice, especially when separated from males (i.e., unmated recently). All four colonies (above plus one from Panama) were similar in their ability to tolerate starvation, although older stages tended to die sooner. An in vitro feeding apparatus was developed to rear head lice. Teneral first instar lice were placed on human hair tufts on the upper side of membrane-covered feeders, which were immersed bottom-side down within a vessel containing warmed human blood. Relative to lice reared on a human host, in vitro-reared lice required a significantly longer time (10-20%) to molt and survived a significantly shorter time as adults (30-50%); the addition of antibiotics did not adversely affect louse development. Teneral first instars were more likely than any other stage to feed through the membrane. Lice spent a significantly greater proportion of time searching in the in vitro apparatus than on a host, but the proportion of time spent feeding did not differ. This research is the first to demonstrate that head lice can be reared successfully in vitro through a complete life cycle.

We used polymerase chain reaction-based DNA profiling to construct allelic profiles for residents and visitors of 22 houses in Florida, Puerto Rico, and human DNA from blood meals in Aedes aegypti that were collected in those homes. Complete profiles were obtained for < or = 2 days after blood ingestion. Eighteen percent of the meals came from two different people. There was no evidence of meals from > or = 2 people. Eighty percent of the meal sources were identified, > 70% were taken from residents of the collection house, and > 90% were from residents of the study community. Across the community, feeding was non-random with a bias towards young adults and males. Three people accounted for 56% of the meals. Our results confirm that multiple feeding on different people is an important component in the role of Ae. aegypti in dengue virus transmission and help explain the spatial distribution of dengue cases in a previous epidemic in Florida, Puerto Rico.

Methods for the estimation and comparison of survival rates are considered when data arises from a release of individuals followed by a sequence of recaptures, with recaptured individuals removed from the population. It is shown that commonly used methods based on linear regression of the log of recapture numbers versus time can lead to substantial errors if individuals are removed from the population. A general nonlinear regression approach is proposed combined with bootstrap techniques for obtaining confidence intervals and tests of hypotheses. Simulations demonstrate that these techniques perform well using data from an Aedes aegypit L. mark-release-recapture study in Thailand.

Phenotypic expression of trans-sialidase (TS) by Trypanosoma cruzi (Silvio strain) is restricted to a subpopulation (20-30%) of broad trypomastigotes (TS+ parasites), while the larger subpopulation of slender trypomastigotes lacks TS expression (TS- parasites). Rhodnius prolixus nymphs were infected with fractionated T. cruzi parasite populations (Silvio strain). Bugs ingesting TS- parasites produced significantly higher parasite loads than bugs ingesting either TS+ parasites or unfractionated populations. The addition of 1.0 microg of TS monoclonal antibody (TCN-2)/ml of blood meal to TS+ populations increased parasite populations to levels comparable to those of TS- populations. In contrast, the addition of exogenous TS to TS- parasite blood meals significantly reduced parasite loads to levels comparable to bugs ingesting TS+ parasites. These results suggest that T. cruzi trypomastigote polymorphism may serve to enhance survival in different host environments.

Behavioral implications of Trypanosoma cruzi Chagas infection in Rhodnius prolixus Stål were observed. Feeding and defecation behaviors of infected versus uninfected insects were assessed on an artificial membrane-feeding system and on live guinea pigs. Based on a defecation index, fifth instars were the most efficient vectors, followed by adult females, fourth instars, and adult males. Bugs fasted for longer periods (5-6 mo) took smaller blood meals but defecated significantly earlier than bugs fasted for shorter periods (2-3 mo). Multiple blood feeding, degree of fasting, life stage, T cruzi infection, and gender affected the vector potential of R. prolixus. Our data indicate that T. cruzi and R. prolixus have not coevolved to facilitate the transmission of T. cruzi, which suggests that this parasite-host relationship may be relatively recent.

The study employed an innovative exercise-based instrument to identify individuals at risk for eating disorder (ED). The Exercise Orientation Questionnaire (EOQ), a reliable and valid instrument, was used to compare 80 ED patients, 74 obese patients, 99 elite runners, and 214 controls from a previous study. Runners resembled ED patients in scoring high on total EOQ and exercise intensity but differed in that the athletes scored low on self-loathing (SLSS), a sub-scale of the EOQ. SLSS scores clearly distinguished clinical (ED, obese) from non-clinical groups, while other sub-scales (self-control, weight loss, and identity) did not. The study suggests that it is feasible to detect ED risk through assessment of exercise attitudes and behaviors. The SLSS subscale of the EOQ can differentiate clinical from non-clinical groups and anorexic patients from healthy individuals with body mass index within the anorexic range. The SLSS could provide a theoretical bridge between compulsive athleticism and ED.

We observed the movement, attraction, and resting behavior of laboratory Rhodnius prolixus Stål within a house environment, simulated by a Styrofoam box. Movement of R. prolixus into various refuge types (fiberpots or white-, cream-, or black-colored modified Gomez-Nunez boxes) at different strata was assessed. Greater numbers of fifth instars sought a refuge than third instars. Fasted nymphs sought a refuge more than fed nymphs. Our simulated house environment showed potential as a laboratory arena to observe the resting preferences, dispersal, and response of kissing bugs to potential attractants. Our data indicated that fiberpot refuges may serve as an effective and inexpensive monitoring device to detect low density kissing bug infestations.

Mosquitoes that do not oviposit their entire batch of eggs in one location, but choose instead to oviposit a few eggs in several different sites exhibit skip oviposition. Although skip oviposition may ensure the greater distribution of progeny from an individual female and reduce sibling competition, it also uses greater maternal energy reserves, may increase the risk of adult female mortality and does not reduce other competition. To test the hypothesis that female Aedes aegypti (L.) do not oviposit all of their eggs at one time and place, we examined females collected with an aspirator resting inside houses in Thailand. Females (n = 384) were dissected and the stage and number of developing oocytes were enumerated. If skip oviposition is a common behavior among female Ae. aegypti, one would expect to find significantly fewer mature eggs (Christophers' stage V) in gravid females than developing oocytes in half-gravid females (Christophers' stage IIIa-IVb). No significant differences were found between ovaries containing immature or mature oocytes, even when the data were stratified by female body size and when immature oocytes that potentially could undergo resorption (Christophers' stages IIIa-IIIb) were excluded from the analysis. We found no evidence that skip oviposition over several days is a common behavior among domestic populations of Ae. aegypti at the end of the rainy season in this region of Thailand.

It generally is assumed that the daily probability of survival of wild adult mosquitoes is independent of age. To test this assumption we conducted mark-release-recapture studies in Puerto Rico and Thailand to determine if estimated daily survival rates between two different age cohorts of the dengue vector Aedes aegypti (L.) were the same. Survivorship was estimated with nonlinear regression analysis using bootstrapping to obtain estimates of errors. Initial recapture success of the younger cohort was greater than the older cohort at both locations. Our analysis revealed a significantly greater survival rate for the younger cohort of females in Puerto Rico, and no significant differences between age cohorts in Thailand. For comparison, a traditional approach for analyzing these type of data, linear regression of log-transformed captures over time (exponential model), was used to calculate the probability of daily survival based on slopes of linear regression lines for recaptured mosquitoes. With this method, the estimated daily survival rate of older females (13-23 d old) was significantly greater than survival of younger ones (3-13 d old) in Puerto Rico and Thailand. In addition, short-range movement of mosquitoes was observed in Puerto Rico; maximum dispersal distance detected was 79 m. Survival rates of adult Ae. aegypti may be age-dependent and nonlinear regression analysis is a sensitive approach for comparing patterns of mosquito survival based on mark, single release, multiple recapture data.

Adult female Aedes aegypti (L.), the vector of dengue and yellow fever viruses, have an affinity for feeding on human blood and a tendency to forego feeding on sugar. This observation challenges two tenets of mosquito biology: (1) mosquitoes imbibe plant carbohydrates for synthesis of energy reserves and blood for reproduction and (2) egg production is reduced when mosquitoes feed on human blood compared with blood from other species. Sub-optimal amounts of the amino acid isoleucine in human blood (particularly free isoleucine in plasma) are thought to be responsible for lowered egg production when human blood is ingested. We tested the hypothesis that feeding on human blood is associated with a selective advantage for Ae. aegypti and is an underlying reason for this mosquito's intimate and epidemiologically important relationship with human beings. Our five experiments examined the effects of different isoleucine concentrations on accumulated energy reserves, frequency of host contact, survival, and egg production. When mosquitoes imbibed blood meals over a 7- to 10-d period and were not fed sugar, increased isoleucine concentration decreased energy reserves and did not increase egg production. Aedes aegypti took smaller but more frequent blood meals when feeding on a low-isoleucine human host daily compared with a high-isoleucine mouse host. Previous reports that isoleucine enhances egg production were confirmed only when females were fed sugar, an unusual behavior for most domestic Ae. aegypti populations. Females fed human blood and water had greater age-specific survival (l(x)), reproductive output (m(x)), and cumulative net replacement (R0) than cohorts fed human blood plus sugar or isoleucine-rich mouse blood with or without access to sugar. The unique isoleucine concentration of human blood is associated with Ae. aegypti's unusual propensity to feed preferentially and frequently on humans--a behavior that increases this mosquito's fitness, synthesis of energy reserves, and contact with human hosts, making it an especially effective disseminator of human pathogens.

The effect of eastern equine encephalomyelitis (EEE) virus on the survivorship of Aedes albopictus (Skuse), Anopheles quadrimaculatus Say, and Coquillettidia perturbans (Walker) was determined experimentally. Female mosquitoes were allowed to feed on EEE viremic chicks, and survival rates were compared for infected and uninfected mosquitoes. Additionally, the survival of female Cq. perturbans and An. quadrimaculatus intrathoracically (i.t.) inoculated with EEE was compared with controls receiving diluent inoculations. Infection with EEE significantly reduced survival in Cq. perturbans compared with uninfected individuals in per os infection experiments. I.t. infections of Cq. perturbans did not reduce survival when compared with diluent inoculated groups. In contrast, infection with EEE did not affect the survival of Ae. albopictus after per os infection or An. quadrimaculatus after either i.t. or per os infections.

Geographic information system (GIS) technology and remote sensing were used to identify landscape features determining risk of eastern equine encephalomyelitis virus (EEE) transmission as defined by the abundance of Culiseta melanura (the enzootic vector) and 6 putative epidemic-epizootic vectors in Massachusetts. Landsat Thematic Mapper data combined with aerial videography data were used to generate a map of landscape elements at epidemic-epizootic foci in southeastern Massachusetts. Geographic information system technology was used to determine the proportion of landscape elements surrounding 15 human and horse case sites where abundance data were collected for Culiseta melanura, Aedes canadensis, Aedes vexans, Culex salinarius, Coquillettidia perturbans, Anopheles quadrimaculatus, and Anopheles punctipennis. The relationships between vector abundance and landscape proportions were analyzed using stepwise linear regression. Stepwise regression indicated wetlands as the most important major class element, which accounted for up to 72.5% of the observed variation in the host-seeking populations of Ae. canadensis, Ae. vexans, and Cs. melanura. Moreover, stepwise linear regression demonstrated deciduous wetlands to be the specific wetland category contributing to the major class models. This approach of utilizing GIS technology and remote sensing in combination with street mapping can be employed to identify deciduous wetlands in neighborhoods at risk for EEE transmission and to plan more efficient schedules of pesticide applications targeting adults.

Aspiration collections of adult Aedes aegypti (L.) were made weekly from inside and outside of houses for 3 yr in a rural Thai village (n = 9,637 females and n = 11,988 males) and for 2 yr in a residential section of San Juan, Puerto Rico (n = 5,941 females and n = 6,739 males). In Thailand, temperature and rainfall fell into distinct seasonal categories, but only temperature was correlated with fluctuations in female abundance. Average weekly temperature 6 wk before mosquitoes were collected and minimum weekly temperature during the week of collection provided the highest correlations with female abundance. Accounting for annual variation significantly improved Thai models of temperature and mosquito abundance. In Puerto Rico, temperature, but not rainfall, could be categorized into seasonal patterns. Neither was correlated with changes in female abundance. At both sites the vast majority of females were collected inside houses and most contained a blood meal. Most teneral females were collected outside. Wing length--an indicator of female size--and parity, egg development or engorgement status were not correlated, indicating that feeding success and survival were not influenced by female size. At both sites, females fed almost exclusively on human hosts (> or = 96%), a pattern that did not change seasonally. In Puerto Rico more nonhuman blood meals were detected in mosquitoes collected outside than inside houses; no such difference was detected in Thailand. Gut contents of dissected females indicated that females in the Thai population had a younger age distribution and fed more frequently on blood than did Ae. aegypti in Puerto Rico. Our results indicated that aspects of this species' biology can vary significantly from one location to another and 1 yr to the next.

We used a histologic technique to study multiple blood feeding in a single gonotrophic cycle by engorged Aedes aegypti (L.) that were collected weekly for 2 yr from houses in a rural village in Thailand (n = 1,891) and a residential section of San Juan, Puerto Rico (n = 1,675). Overall, mosquitoes from Thailand contained significantly more multiple meals (n = 1,300, 42% double meals, 5% triple meals) than mosquitoes collected in Puerto Rico (n = 1,156, 32% double meals, 2% triple meals). The portion of specimens for which frequency of feeding could not be determined was 31% at both sites. We estimated that on average Ae. aegypti take 0.76 and 0.63 human blood meals per day in Thailand and Puerto Rico, respectively. However, frequency of multiple feeding varied among houses and, in Puerto Rico, the neighborhoods from which mosquitoes were collected. In Thailand 65% of the mosquitoes fed twice on the same day, whereas in Puerto Rico 57% took multiple meals separated by > or = 1 d. At both sites, the majority of engorged specimens were collected inside houses (Thailand 86%, Puerto Rico 95%). The number of blood meals detected was independent of where mosquitoes were collected (inside versus outside of the house) at both sites and the time of day collections were made in Puerto Rico. Feeding rates were slightly higher for mosquitoes collected in the afternoon in Thailand. Temperatures were significantly higher and mosquitoes significantly smaller in Thailand than in Puerto Rico. At both sites female size was negatively associated with temperature. Rates of multiple feeding were associated positively with temperature and negatively with mosquito size in Thailand, but not in Puerto Rico. Multiple feeding during a single gonotrophic cycle is a regular part of Ae. aegypti biology, can vary geographically and under different climate conditions, and may be associated with variation in patterns of dengue virus transmission.

Putative epidemic/epizootic eastern equine encephalomyelitis virus (EEE) vector populations were compared at 15 recent (1982-90) human and horse case sites in Bristol and Plymouth counties in southeastern Massachusetts. Carbon dioxide-baited American Biophysics Corporation light traps were used for trapping adult mosquitoes to estimate biting risk in these foci of known transmission. Population data suggest that Coquillettidia perturbans, Aedes canadensis, and Culex salinarius are more likely vectors of EEE in Massachusetts than Aedes vexans, Anopheles punctipennis, and Anopheles quadrimaculatus.

Gas chromatography with flame-ionization detection was used to measure the time-associated, quantitative changes in the cuticular hydrocarbons of female Aedes aegypti (L.). Cohorts of unstressed Ae. aegypti, Rockefeller strain, were reared and held at 3 constant temperatures (24, 28, and 30 degrees C). Five females from each cohort were taken at 33 degree-day (DD) intervals from 0 to 231 DD (using 17 degrees C as the threshold temperature). Quantitative changes over time of cuticular hydrocarbons associated with gas chromatographic peaks 1 and 5 were identified as having promise for age grading. The relative abundance of peak 1 (pentacosane) decreased linearly from 0 to 132 DD, whereas peak 5 (nonacosane) increased linearly over the same period. Suboptimal larval conditions (crowded and starved), which resulted in physiological stress (decreased size), had negligible effect on the relative abundance of pentacosane and nonacosane. Additionally, the rate of change in the relative abundance of pentacosane and nonacosane were the same for both a recently colonized Chachoengsao (Thailand) strain of Ae. aegypti compared with the long-colonized Rockefeller (Caribbean) strain over a 0-99 DD interval. Two linear regression models, one based on the relative abundance of pentacosane and the other on the logit transformation of these values, were developed for aging female Ae. aegypti. A blind study using laboratory-reared mosquitoes and a mark-release-recapture experiment using field mosquitoes validated these age-grading models and produced promising results for aging females up to 132 DD (19, 12, and 10 calendar days at 24, 28 and 30 degrees C, respectively). Therefore the regression models, based on the relative abundance of these 2 cuticular hydrocarbons, appeared to be a useful approach for age-grading Ae. aegypti up to at least 12 d of age regardless of environmental conditions (temperature and stress) and population history (origin and colonization time).

We previously described methods for age-grading female Aedes aegypti (L.) by gas chromatographic (GC) analysis of whole-body cuticular hydrocarbon patterns. Two regression models were developed that were based on the age-dependent, relative abundance of 2 cuticular hydrocarbons, pentacosane (GC peak 1) and nonacosane (GC peak 5). We have refined this method so that only the legs are required to age individual females. Two new regression models were developed that also use the relative abundance of a 3rd cuticular hydrocarbon, octacosane (GC peak 4). These models improve the overall accuracy of the cuticular hydrocarbon method for aging female mosquitoes, especially for older females from 132 to 165 degree-days (DD) of age (12-15 calendar days at 28 degrees C). The correlation coefficients (R2) for the best-fitted linear regression models for aging females from 0 to 132 and 0-165 DD were 0.80 and 0.81, respectively (P < 0.001 in all cases). The use of leg cuticular hydrocarbons for estimating the age of female Ae. aegypti has a significant advantage over whole-body extracts as indicated by the decreased variability associated with the relative abundance of pentacosane and the expanded range over which the models were able to predict age accurately by the addition of the relative abundance of octacosane.

Laboratory experiments suggest that utilization of blood rather than natural sugar sources for energetic needs affords female Aedes aegypti a reproductive advantage over conspecifics that use sugar. To test this hypothesis under field conditions, we carried out a mark-release-recapture study in Florida, PR. Adult females (F1) reared from field-collected eggs were provided with a diet of human blood alone or human blood plus a 20% honey solution before their release. Backpack aspirators were used to collect mosquitoes from release houses for 5 consecutive days beginning the 2nd day after release. Survival was estimated from the slope of the regression line of the log-transformed daily number of recaptures for each treatment group. To compare fecundity of the treatment groups, each recaptured female was dissected, ovaries were removed, oocytes counted, and Christophers' stages of oocyte development scored. Recapture rates were 30% for the blood-only group and 23% for blood plus honey group. The daily survival rate of the blood-only group (55%) was not statistically different from that of the blood plus honey group (69%) (t = 0.32, P > 0.05). By analysis of variance, fecundity (average number of stage III-V oocytes) was significantly higher in the females fed human blood alone (n = 103, 109 oocytes/female) than in the group fed on blood and honey (n = 50, 95 oocytes/female) (P = 0.0007). The observed gonotrophic cycle length of the recaptured females ranged from 3 to 7 days. Results from our field study are consistent with laboratory life-table experiments that suggest feeding exclusively on human blood provides a reproductive advantage for female A. aegypti.

Survival of adult Aedes aegypti (L.) was studied in Thailand (1995) and Puerto Rico (1996) during periods of high and low dengue virus transmission. Resting males and females were collected inside houses by aspiration. Females were separated into different cages by their degree of engorgement and ovarian development. Teneral adults were obtained from pupae collected from natural breeding sites. All mosquitoes were given access to water, held at ambient temperature in the shade, and their survival monitored daily. We calculated median survival for each stage to estimate when mosquitoes had to feed again or die. No differences in survival between seasons were observed in Thailand. In Puerto Rico, except for wild males, survival was longer in the cool/dry season than in the hot/rainy season, indicating that mosquitoes may need to feed more frequently during the high than low dengue transmission season. During both study periods and at both sites, blood-engorged females survived as long or longer than mosquitoes in other gonotrophic or developmental stages. Except in Puerto Rico during the cool season, when females had a relatively high probability of surviving 3-4 d without feeding, females needed to feed approximately every other day to avoid death caused by starvation. Our results indicate that in some regions, there are seasonal differences in the length of time female Ae. aegypti can survive without feeding, females with a blood meal can survive for a longer time than those without blood, and teneral males can live longer without food than teneral females.

We examined the amount of glycogen, lipid, and sugar in small and large female Ae. aegypti (L.) that were fed different concentrations of sucrose. Replicate groups of laboratory-reared teneral females (< or = 1 d old) were allowed to feed ad libidum on a 0, 5, 10, 15, or 20% sucrose solution, and total glycogen, lipids, and sugars were assayed at 4, 12, 24, 48, and 72 h after exposure to sugar. Mosquitoes fed sugar increased nutrient levels with time. The pattern of accumulation was significantly different between small and large mosquitoes. Large mosquitoes accumulated larger amounts of glycogen than small ones. Accumulation of lipid increased sooner in small (4 h) than large (48 h) Ae. aegypti. A significant size x time interaction for the amount of sugar per female indicated that small mosquitoes may need to feed more than once during a 72-h period to compensate for low energy reserves at emergence. We conclude that the pattern of energy utilization from sugar meals by Ae. aegypti is influenced by the amount of energy reserves that they have before feeding and not by the concentration of sugar on which they feed.

We examined the utilization of sugar and human blood as nutrient sources for small and large female Aedes aegypti (L.) when they were fed blood 2 or 5 d after emergence. Laboratory-reared mosquitoes were fed human blood alone or sugar plus human blood and assayed at 4, 12, 24, and 48 h after the blood meal. Starved and well-fed mosquitoes were obtained by holding teneral females (< or = 1 d old) with 0, 5, 10, and 15% sucrose solutions ad libidum from emergence. Both small and large mosquitoes increased their glycogen and sugar levels significantly by feeding on blood only or on blood plus sugar when they imbibed a human blood meal on day 2 after emergence. Mosquitoes only fed blood on day 2 had the highest lipid levels of any treatment group. Both size classes and all feeding regimes failed to increase the total amount of glycogen, lipid, or sugar when they fed on blood 5 d after emergence. We conclude that there is an energetic advantage to Ae. aegypti when they feed on blood early in adult life (< or = day 2 after emergence).

We examined the accumulation of glycogen, lipid, and sugar obtained from a human blood meal for egg development and body energy reserves by small and large female Aedes aegypti (L.). Small and large mosquitoes were fed a single meal of human blood on day 2 after emergence. Mosquitoes were collected at 4, 12, 24, 48, and 72 h after blood feeding and ovaries and body remnants were separated by dissection and then assayed. Large mosquitoes had greater reserves than small mosquitoes. Mosquitoes deposited similar proportions of lipid reserves obtained from blood meals in their ovaries regardless of body size. Small mosquitoes deposited a significantly higher proportion of their glycogen in ovaries than large mosquitoes. The pattern of energy accumulation and use indicates that to avoid starvation, mosquitoes fed a single human blood meal will need to feed again before ovipositing, and that multiple feeding may be more important for small than for large Ae. aegypti.

Male and female Aedes aegypti fed a 10% sucrose solution and/or blood were tested to determine the duration of fructose detection in their bodies and the volume of sugar solution they ingested. The limit of detection of fructose by the cold anthrone test was investigated in a series of experiments. Results were applied to the interpretation of sugar feeding by Ae. aegypti collected inside houses in Puerto Rico during times of low (cool season) and high (hot season) dengue transmission in 1996. We conclude that, under our experimental conditions, the cold anthrone test can detect a 10% sucrose solution in male and female Ae. aegypti up to approximately 4 days after ingestion, even the smallest volumes of 10% sucrose solution ingested by experimental mosquitoes are detectable, the test is sensitive enough to detect 0.6 microgram of fructose, and the cutoff point for defining positive fructose values in field-collected females should be based on blood-engorged specimens. We confirmed that female Ae. aegypti collected from natural resting sites inside houses in Puerto Rico seldom, compared to males, contain detectable amounts of fructose.

Life table studies were performed in 1996 with Aedes aegypti (L.) during the low (cool/dry) and high (hot/rainy) dengue virus transmission seasons in Puerto Rico. Mated adult females from field-collected pupae were placed individually in cages and divided into 2 treatment groups: one was fed only human blood and the other human blood plus a 10% sucrose solution. Survival and number of eggs laid were recorded daily for each female. During both seasons, age specific survivorship was higher for the blood plus sugar group, groups fed only human blood had higher reproductive outputs (mx), and net replacement rates (Ro) for blood only groups were higher than for those fed blood plus sugar. Intrinsic rates of growth (r) were the same for both treatments during the low (cool/dry) transmission season, but higher for the blood-only treatment during the high (hot/rainy) transmission season. Our results indicate that feeding on only human blood provides an evolutionary advantage to Ae. aegypti females in Puerto Rico. These results are similar to those from an earlier study carried out with Ae. aegypti in Thailand; the advantage of feeding on human blood does not seem to be restricted to a particular geographic region. We also found that the benefits associated with human feeding persist through epidemiologically different times of the year. We conclude that feeding on human blood is reproductively beneficial for Ae. aegypti, which may increase their contact with human hosts, and therefore may influence their vectorial capacity for dengue viruses through frequent feeding on blood.

Marked Aedes aegypti (L.) (5-6 d old) were released inside 2 groups of 5 houses (100 females per house) in a residential community in Florida, PR, to compare behavior of gravid females at sites where oviposition containers were absent to sites where containers were abundant (i.e., 2 tires and 10 ovipots were added to each yard). Two sequential releases were made so that both groups of houses were evaluated with oviposition containers removed and added. Mosquitoes resting inside the 10 release houses plus 20 additional neighboring houses were collected with backpack aspirators for 4 consecutive days, beginning 2 d after release. Because 172 of the 185 recaptured females (93%) were collected in the same houses in which they had been released, dispersal patterns were not directly comparable. However, the recapture rate in houses with containers added (13%) was significantly higher (P < 0.0001) than in houses with containers removed (6%). No difference was observed in the mean number of potential oviposition containers among the nonrelease houses at the 2 sites (3.9 versus 3.8 aquatic containers per house in the prerelease survey). Our results are consistent with the hypothesis that availability of oviposition sites is inversely correlated with the potential for female Ae. aegypti to disperse. These results have important implications because campaigns to reduce Ae. aegypti larval sites during dengue epidemics could have the undesirable effect of inducing the dispersal of infected adult female mosquitoes.

To determine which of 11 trapping methods best sampled populations of 6 potential epidemic vectors of eastern equine encephalomyelitis (EEE) virus, we compared New Jersey (NJ) light trap, CDC light trap, CDC with octenol, CDC with CO2, CDC with CO2 plus octenol, American Biophysics light trap with flickering light (AB flicker), AB steady light, AB flicker with octenol, AB flicker with CO2, AB flicker with CO2 plus octenol, and 3 resting boxes. There was no significant difference between CDC and AB light traps (P > 0.05). The addition of octenol increased trap catch with both CDC and AB light traps; however, this increase was not statistically significant (P > 0.05). Both brands supplemented with CO2 collected more Aedes canadensis, Coquillettidia perturbans, Culex salinarius, and Anopheles spp. than did unbaited light traps (P < 0.05). The addition of octenol to CO2 increased collections of Anopheles spp. and decreased collections of Aedes spp. and Cq. perturbans at some sites, but these changes were not statistically significant (P > 0.05). Resting boxes were best for sampling bloodfed or parous Anopheles spp. The NJ, AB flicker, and AB steady light traps were not effective for sampling potential vectors of EEE virus.

To estimate human exposure to potential vectors of eastern equine encephalomyelitis (EEE) virus, we compared collections of putative EEE virus vectors from human biting collections with collections from CDC, AB (American Biophysics), and New Jersey light traps and resting boxes in enzootic/epidemic foci of EEE virus in southeastern Massachusetts. Human biting collections caught significantly more host-seeking females than resting boxes or unbaited light traps (P < 0.05). Regression analysis of human biting collections to AB traps supplemented with CO2 could predict 60-70% of the actual human biting risk by Aedes and Coquillettidia perturbans. The AB traps supplemented with CO2 and octenol could predict 65% of Anopheles biting risk. No single sampling method was accurate for predicting human biting risk by Culex salinarius, and no method could predict human biting risk by all potential vectors surveyed.

Literature on arthropod-borne diseases has traditionally supported the notion that mosquito vectors maintain a feeding duality that includes vertebrate blood meals for egg development and sugar meals from plants for the synthesis of flight and survival energy reserves. Aedes aegypti was found to deviate from that feeding pattern by obtaining a reproductive advantage when feeding only on human blood. Female mosquitoes fed human blood alone had a greater net replacement rate and intrinsic rate of growth during all phases of their reproductive life than conspecifics fed human blood plus sucrose. Feeding frequently on human hosts during each gonotrophic cycle is necessary to avoid death due to starvation and increases exponentially the spread of Ae. aegypti-borne disease. Our results help explain why Ae. aegypti is such an unusually efficient vector of human disease; frequent biting of humans results in a high reproductive rate for vectors as well as the viruses they transmit.

Frequency of sugar feeding and blood feeding can have an impact on the infection by and transmission of malaria parasites. Data presented here indicate that frequent blood feeding has a deleterious effect on infection by malaria parasites in Aedes aegypti. In addition, mosquitoes that do not blood feed, but instead feed on sugar alone after an infected blood meal, have a higher rate of parasite transmission than mosquitoes fed additional blood meals.

We tested susceptibility to per os infection and potential salivary transmission for eastern equine encephalomyelitis (EEE) virus in Aedes canadensis (Theobald), Aedes vexans (Meigen), Anopheles quadrimaculatus (Say), Anopheles punctipennis (Say), Coquillettidia perturbans (Walker), and Culex salinarius (Coquillett). Culiseta melanura (Coquillett), the documented enzootic vector of EEE virus, was our control. Based on these estimates of laboratory vector competence and other behavioral and ecological components of vectorial capacity, we ranked these 6 species from the most to least probable epidemic vectors: Cx. salinarius, An. quadrimaculatus, Ae. canadensis, Cq. perturbans, Ae. vexans, and An. punctipennis.

The numbers of blacklegged tick, Ixodes scapularis Say, per hectare that engorged as larvae and nymphs on white-footed mice, Peromyscus leucopus (Rafinesque), were estimated at 3 sites in coastal Massachusetts. Estimates of numbers of engorged ticks dropping per day from trapped mice were multiplied by mark-recapture estimates of the density (numbers per hectare) of white-footed mice to estimate numbers of ticks engorging on mice per hectare. In 1991, estimates of seasonal totals per hectare of I. scapularis nymphs completing blood meals on P.leucopus ranged from 825 +/- 68 at Nonamesset to 6,654 +/- 770 at Menemsha. This absolute sampling method (the drop-off method) was compared with drag sampling for nymphal ticks. Per hectare estimation of engorgement rates overcomes a shortcoming associated with drag sampling because site-to-site comparisons can be made even where vegetation structure differs markedly among sites. The drop-off method may be used to quantify the contribution of any host to the engorged tick population, provided sufficient such hosts can be collected on a weekly basis.

Ten-day-old, parous Aedes aegypti (L.) had a higher threshold of blood meal size for the inhibition of continued host-seeking than did 5 d-old, nulliparous females of the same size. Older females of a smaller-bodied cohort had a higher threshold of blood meal size for the initiation of egg development than did younger females. In contrast, older larger-sized females had a lower blood meal-size threshold for egg development. The blood of paired avian hosts was marked with rubidium and cesium to study the effect of age and body size on the frequency of multiple blood feeding. Chronologically old females had a higher frequency of multiple blood feeding than younger females. In the laboratory, the frequency of multiple blood feeding among larger-sized females was higher than among small-sized females.

Daily survivorship and fecundity of Aedes aegypti mosquitoes were monitored for cohorts provided with five nutritional treatments: water, 2% sucrose, blood from a live chicken plus water, blood plus 2% sucrose, and blood alone. The median mortality time (LT50) for these females was 6, 54, 16, 12, and 29 d, respectively. There was no significant difference in the number of eggs laid by females in any of the treatments containing a host. Females maintained on blood alone laid as many or more eggs during their lifetime as females with access to sugar who had a greater life expectancy. Males maintained on sugar alone survived significantly longer than those in any of the other treatments. Large- and small-bodied, sugar-starved Ae. aegypti females that were marked, released, and recaptured in a Thai village survived as well as the replicate cohorts that received sugar or sugar plus blood during the 36 +/- 12 h period from emergence until their release. These results indicate that Ae. aegypti females live longer in the laboratory if they are provided a source of carbohydrate, but the increased survival associated with sugar feeding does not increase reproductive success.

Anopheles maculatus Theobald sensu lato is a species complex now consisting of eight sibling species; An. maculatus is still represented by two cytologically distinct forms; i.e., the widely distributed sensu strictu or B, and E from southern Thailand and adjacent areas in northern Malaysia. Cuticular lipid profiles in conjunction with principal component analysis was used to separate An. maculatus form E from sensu stricto form B in a preliminary survey of the An. maculatus complex at five locations spanning peninsular Malaysia. The relative rank orders, from the areas of the five gas chromatographic peaks used to determine lipid differences for specimens from peninsular Malaysia, matched well with those from cytogenetically identified colony specimens of An. maculatus forms B and E. The two-dimensional principal component pattern of specimens identified as form E was highly clumped, which indicated that very similar cuticular lipids were present within this putative malaria vector. Both forms coexisted in peninsular Malaysia, but form E may be dominant except in the south.