The current study examined temporal and spatial distribution patterns of anopheline malaria vectors in a highland site and determined the number of houses to be sampled to achieve the targeted precision level. Adult mosquito sampling was conducted seasonally in May and August 2002 in a 3 by 3-km² area, and in November 2002 and February 2003 in an expanded 4 by 4-km² area in Kakamega District, western Kenya. Anopheles gambiae Giles was the predominant malaria vector species, constituting 84.6% of the specimens, whereas Anopheles funestus Giles constituted 15.4% of the vector populations. An. gambiae abundance increased by six- to eight-fold in the long rainy season over the dry seasons, but An. funestus abundance peaked 3 mo after the long rainy season. For both species, the coefficient of variation was larger than 1, suggesting that the distribution of mosquito adults was aggregated. Mosquito clustering occurred in houses <400 m from a valley bottom. The negative binomial distribution was accepted in one sample period (August 2002) for An. gambiae and in two sampling periods (May and August 2002) for An. funestus. Taylor’s power law analyses indicated that An. gambiae distribution was more aggregated in the wet seasons than in the dry seasons, whereas the degree of aggregation of An. funestus was similar in all four seasons. The minimum number of houses required to estimate anopheline female abundance within the commonly acceptable precision level (0.2) should be 17 houses per km² for An. gambiae and 42 houses per km² for An. funestus. The potential factors causing aggregated anopheline mosquito distribution are discussed.

Determinant aspects of the development cycle of Panstrongylus rufotuberculatus (Champion, 1899) were evaluated under laboratory conditions. Fertility, duration of life cycle, mortality rate, feeding time, defecation time, thermal reaction time to food source, and volume of blood ingested were studied. From an initial cohort of fifth instars, nine adult pairs were formed, the females of which oviposited 2,338 eggs over 231 d. The eclosion rate was 84.4%; female and male progeny lived 223.56 (14.41) and 180.78 (14.41) d, respectively. The average total life cycle of immature stages (138.82 ± 51.42 d) was distributed as egg, 24.22 ± 0.42 d; nymph I, 18.94 ± 5.15 d; nymph II, 15.80 ± 3.29 d; nymph III, 16.55 ± 4.36 d; nymph IV, 25.50 ± 7.32 d; and nymph V, 36.71 ± 2.75 d. Given these values, 2.6 generations per year are possible. Because of its longevity, rapid response to the presence of a host, large volume of blood ingested, and frequent defecation during feeding, this species constitutes a potentially important vector of Chagas’ disease in nonendemic rural areas.

We analyzed triatomine blood feeding patterns to evaluate the role of peridomiciles in Trypanosoma cruzi transmission at the rural village of Tepehuaje de Morelos at Jalisco State, Mexico (1999). A total of 206 bugs were collected in 11 out of 26 households (42.3%). Nymphs predominated in the collections (64.9% of the total). Except for one Triatoma barberi female, a species that belongs to the protracta species complex, all adults were Triatoma longipennis, a species of the phyllosoma complex. Triatomines were exclusively present in peridomestic sites mainly piles of tiles and bricks, and none were found indoors. Overall infection rate was 56.6% and no significant differences (P > 0.05) were observed between nymphs and adults or males and females. Identified blood meals were chicken (29.4%), opossum (20.9%), pig (24.5%), murid (20.9%), dog (3.5%), and armadillo (0.7%). No gut content reacted against anti-human, anti-bovine, anti-rabbit, and anti-cat sera. In contrast to fifth nymphs and adults, 87% of the small nymphs fed on one host, indicating that they are less mobile than other stages. Most fifth nymphs and adults fed on domestic hosts, while small nymphs mainly fed on opossum and murid. Infection blood-meal indexes were around 50% for single meals on opossum and murid, stressing their importance as trypanosome donors. Peridomiciles in Tepehuaje could be regarded as interaction sites among domestic and wild and synanthropic mammals and triatomines, which would facilitate circulation of the same T. cruzi strains between domestic and sylvatic cycles. Stone-made walls and building materials, which hold synanthropic rodents and opossums, should be considered as targets for vector control measures.

Lutzomyia longipalpis (Lutz and Neiva) is a species complex of Lutzomyia pseudolongipalpis (Arrivillaga and Feliciangeli) and at least three other as yet undefined siblings. Isozyme and mitochondrial studies of allopatric populations across Central and South America have suggested the presence of four “clades” that have been hypothesized to have arisen mainly because of geographical isolation mechanisms. Parallel studies of sexual behavior as well as cross-mating and genetic analysis, of both allopatric and sympatric populations, suggest at least four sibling species that do not seem to correspond to the defined four “clades.” In an effort to understand this apparent discrepancy, sympatric populations of L. longipalpis from a single South American country, Brazil, are being studied. In Brazil, three putative species can be identified by their male-produced sex pheromones: (S)-9-methylgermacrene-B, 3-methyl-α-himachalene, and a cembrene. We report here that analysis by coupled gas chromatography–mass spectrometry shows that L. longipalpis from Jaíbas, Minas Gerais State, Brazil, occurs as two sympatric sex pheromone chemotypes. One chemotype is the cembrene type previously recorded in a L. longipalpis population from Sobral, Ceará State, Brazil, and the other is a new cembrene isomer not previously observed in L. longipalpis. The finding of this new chemotype strongly suggests that the L. longipalpis species complex in Brazil consists of four members rather than the three previously recognized and confirms previous analysis of genetic variation that had suggested the presence of a complex in Brazil.

We have initiated efforts to determine the molecular basis for the M^D meiotic drive system in the mosquito, Aedes aegypti. The effect of the M^D gene is a highly male-biased sex ratio, but varies depending on the frequency and sensitivity of a susceptible responder m^s allele. The M^D system has potential as a mechanism for driving trangenes for pathogen resistance into natural Ae. aegypti populations. Because all previously existing laboratory strains carrying the M^D gene have been lost, we have selected for a new strain, T37, that carries a strong driver. Matings between T37 males and drive-susceptible m^s females result in progeny with highly biased sex ratios, wherein only ≈14.7% females are produced. We discuss the potential for identifying M^D candidate genes based on comparisons with the well-described Drosophila melanogaster segregation distorter (SD) meiotic drive system and considerations for release of transgenic Ae. aegypti into natural populations where M^D and insensitive mⁱ alleles are likely segregating.

We have optimized a larval immersion microassay (LIM) that offers superior sensitivity, flexibility to accommodate multiple formulations, and a robust capability for rapidly screening many compounds with a minimal requirement of test article for evaluation. Dose–response studies were conducted for representative members from the organophosphate, pyrethroid, pyrazole, carbamate, macrocyclic lactone, and formamidine chemistries against Amblyomma americanum (L.). Time–response experiments revealed that permethrin was the most rapid acting, whereas fipronil had the slowest speed-of-kill against A. americanum. Comparison of drug susceptibility profiles between multiple ixodid ticks suggests that A. americanum is an effective model for predicting compound potency against Boophilus spp. in this bioassay. The LIM is suitable for the identification and characterization of active molecules from small- and medium-sized compound or natural product libraries, and it can be a useful tool to prioritize molecules for further in vivo testing in animal models.

A 3-yr community-based study was conducted on residential properties on Mason’s Island, Mystic, CT, to determine the efficacy of a rodent-targeted acaricide (fipronil) to control immature Ixodes scapularis (Say) on Peromyscus leucopus. Results indicated that modified commercial bait boxes were effective as an acaricide delivery method for reducing nymphal and larval tick infestations on white-footed mice by 68 and 84%, respectively. Passive application of fipronil significantly reduced the infection rate of Borrelia burgdorferi among white-footed mice by 53%. Moreover, the abundance of questing I. scapularis adults on treated properties was reduced by 77% and fewer were infected with spirochetes (31%) compared with untreated sites (47%) after 3 yr of treatment. Likewise, the abundance of host-seeking nymphs was significantly reduced on treated properties by >50%. Finally, infection rates in flagged nymphal ticks for both B. burgdorferi and Anaplasma phagocytophilum were reduced by 67 and 64%, respectively, after only 2 yr of treatment. Results from this 3-yr trial indicate that the use of fipronil passively applied to reservoir animals by bait boxes is an environmentally acceptable means to control ticks, interrupt the natural disease transmission cycle, and reduce the risk of Lyme disease for residents of treated properties.

Behavioral responses of nine Aedes aegypti (L.) strains, six from recent field collections and three from the long-established laboratory colonies, were tested under laboratory-controlled conditions by using an excito-repellency test system. All nine strains showed significant behavioral escape responses when exposed to deltamethrin at the standard field dose (0.02 g/m²), regardless of background insecticide susceptibility status (susceptible or tolerant/resistant). Insecticide contact irritancy played a predominate role in overall female mosquito escape responses, whereas noncontact repellency was not observed at levels significantly different from paired noncontact control tests (P > 0.01). Among the six field populations, the Jakarta (Indonesia) Toba (north Sumatra), and Bangkok female mosquitoes showed rapid exit (>78%) during 30 min of direct contact with insecticide-treated surfaces, whereas the other three strains demonstrated only moderate escape responses (32–56%) from the chambers. Moderate escape responses during direct insecticidal contact also were observed in the three laboratory test populations (44–60%). Higher percentage of mortality was observed from laboratory strains (8–33%) that failed to escape compared with nonescape females of field strains (2–16%), possibly a reflection of background deltamethrin susceptibility status. We conclude that contact irritancy is a major behavioral response of Ae. aegypti when exposed directly to deltamethrin and that rapid flight escape from areas exposed to space sprays or surfaces treated with residual pyrethroids could have a significant impact on the effectiveness of adult mosquito control and disease transmission reduction measures.

Ixodes ricinus can transmit several microorganisms, out of which Borrelia burgdorferi and tick-borne encephalitis (TBE) virus are the most important pathogens in humans. A lemon eucalyptus extract (Citriodiol) has been shown to be a natural repellent against mosquitoes, stable flies, and midges. We have investigated whether the use of the extract would reduce the number of attached I. ricinus ticks in humans. A volunteer sample of 111 individuals was recruited among healthy, outdoor active adults living in two highly I. ricinus–infested areas. They were randomly divided into two groups: one group applied the Citriodiol spray daily for 2 wk to the lower extremities, whereas the other group was instructed not to use any type of repellent on any part of the body. After 2 wk, the scheme was reversed. All volunteers filled in a daily diary, giving information about outdoor activities and the number of observed, attached, or not yet attached ticks, including the anatomical location of the ticks found. Forty-two attached ticks were reported during the weeks when the Citriodiol spray was used, and 112 were reported when it was not. The median number of reported attached ticks per person decreased from 1.5 (range, 0–9) to 0.5 (range, 0–2; P < 0.05) during the weeks when the repellent was used. The number of reported attached ticks noted below the waist was 13/42 (31%) during the period when the spray was used and 73/112 (65%) when no spray was used (P < 0.001). No evident redistribution of attached ticks from protected areas to unprotected areas was seen. Citriodiol may become a useful means in reducing the number of tick bites and thereby tick-borne infections, although additional studies are warranted.

Laboratory cultures of the mites Blomia tropicalis (van Bronswijk, Cock & Oshima) and Blomia kulagini (Zakhvatkin) were used to study the population dynamics of the mites and the kinetics of released allergens during the growth cycle. The analysis of extracts obtained after different incubation periods, by means of immunoblotting, and quantification of the major allergen Blo t 5, allowed definition of three different growth phases, demonstrating that mite cultures during the maximum growth (end of exponential growth curve–beginning maximum growth plateau) contain the largest amount of allergenic components as well as the highest Blo t 5 concentration.

This study evaluates the rickettsial presence in Amblyomma ticks from eight areas of the Amazon forest in Rondônia, Brazil. The following tick species (number in parentheses) were examined: Amblyomma ovale Koch (121), Amblyomma cajennense (F.) (41), Amblyomma naponense (Packard) (36), Amblyomma scalpturatum Neumann (35), Amblyomma oblongoguttatum Koch (30), Amblyomma incisum Neumann (27), Amblyomma rotundatum Koch (16), Amblyomma coelebs Neumann (10), and Amblyomma humerale Koch (6). Ticks were examined individually or in pools (2–10 ticks) by polymerase chain reaction (PCR) targeting the gltA gene. The PCR-determined minimal infection rate for each tick species was A. ovale 28%, A. cajennense 27%, A. naponense 0%, A. scalpturatum 11%, A. oblongoguttatum 3%, A. incisum 0%, A. rotundatum 87%, A. coelebs 10%, and A. humerale 50%. Partial sequences of the gltA gene of Rickettsia from A. ovale, A. scalpturatum, A. oblongoguttatum, A. rotundatum, and A. humerale were 99.9% (349/350) identical to Rickettsia bellii. DNA sequences of PCR products from A. cajennense and A. coelebs were 100% (350/350) identical to Rickettsia amblyommii. R. bellii organisms were isolated in Vero cells from A. scalpturatum, A. ovale, A. rotundatum, and A. oblongoguttatum, but only one of the isolates, cultured from A. scalpturatum, was established in continuous cell culture passage. R. amblyommii was isolated from A. cajennense and was successfully established in continuous passage in cell culture. R. amblyommii infection of Vero cells was analyzed by transmission electron microscopy. This study adds South America to the known geographic distribution of R. amblyommii and reports rickettsiae in six Amblyomma species for the first time.

The persistence of Escherichia coli in artificially fed larvae was examined for up to 48 h after ingestion by house flies, Musca domestica L., and stable flies, Stomoxys calcitrans (L.). The rate of change in the E. coli load was similar for both species for up to 5 h after ingestion. Up to 48 h after ingestion, abundance of E. coli declined in immature house flies but remained constant in immature stable flies. When different E. coli concentrations were fed to larvae, the abundance of E. coli increased in stable fly larvae regardless of the initial concentration. The E. coli load in house fly larvae increased when larvae were fed a low concentration of bacteria, but it declined when larvae were fed a high concentration of bacteria. Survival of house fly and stable fly larvae averaged 62 and 25%, respectively, when reared on pure E. coli cultures. These observations suggest that house fly larvae digest E. coli and use it as a food source but stable fly larvae do not.

From 1986 to 1990, we conducted our second longitudinal study in the central (upstate) New York (CNY) area on the wild avian hosts of eastern equine encephalomyelitis (EEE) virus. Field-collecting methods mirrored a study conducted from 1978 to 1980 at the same endemic focus. Over the 5-yr study period, we captured 6,296 birds representing 99 species and took 4,174 blood samples from representatives of 83 species. Gray catbirds, song sparrows, and veerys were the three dominant species captured and bled, accounting for 40 and 55% of birds captured and bled. Blood clots were assayed for virus and sera tested for hemagglutination-inhibition (HI) antibodies to EEE and Highlands J virus. Virus isolations from birds defined two epiornitics of EEE virus in 1988 and 1990, and an epiornitic of HJ virus in 1986. Infected birds responded with the production of HI antibodies with titers indicative of recent infection (HI ≥ 1:160), and titers of sera positive during the epiornitics were significantly higher than positive sera during nonepiornitics. The 1990 EEE epiornitic extended from mid-July to the end of September, providing data to compare infection rates among species, habitats, and combinations of species with habitats. Few significant differences were found. The HJ epiornitic was only the second time this virus has occurred in CNY. Song sparrows were identified as the primary amplifying avian host of both viruses, although our capture and serological data would suggest a role for gray catbirds as the species most likely involved in yearly virus reintroduction. However, the cryptic nature of enzootic virus maintenance remains unresolved for the CNY virus foci. The appearances of HJ and EEE viruses were not epidemiologically linked, and there were no virus isolations from adults returning on site or virus isolations without concurrent isolations from mosquito vectors. Whether EEE and/or HJ virus are consistently present in or sporadically introduced into the inland foci of CNY area still has not been determined.

The lone star tick, Amblyomma americanum (L.), has increased in abundance in several regions of the northeastern United States, including areas of Long Island, NY. Adult and nymphal stage A. americanum collected from several sites on Long Island were evaluated for infection with Ehrlichia chaffeensis, the causative agent of human monocytic ehrlichiosis (HME), by using a nested polymerase chain reaction assay. Fifty-nine (12.5%) of 473 adults and eight of 113 pools of five nymphs each (estimated minimum prevalence of infection 1.4%) contained DNA of E. chaffeensis. These data, coupled with the documented expansion of lone star tick populations in the northeastern United States, confirm that E. chaffeensis is endemic to many areas of Long Island and that HME should be considered among the differential diagnoses of the many distinct tick-borne diseases that occur in this region.

One of the major obstacles for studies of the biology, ecology, and behavior of Neotropical vectors of human Plasmodium has been the lack of reliable and efficient means of identifying many species. Although the subgenus Nyssorhynchus includes most species responsible for human transmission in South America, there are no polymerase chain reaction (PCR)-based techniques for identifying members of this taxon. We describe the first multiplex PCR for identifying four species in the subgenus Nyssorhynchus that are vectors of human Plasmodium spp. Four species specific primers, together with a universal primer that anneals to the 5.8S rDNA region, produce amplicons of the internal transcribed spacer two with base pair sizes of 131, 308, 371, and 441 for An. triannulatus, An. trinkae, An. strodei, and An. rangeli, respectively.

Ross River virus is a common mosquito-borne arbovirus responsible for outbreaks of polyarthritic disease throughout Australia. To better understand climatic factors preceding outbreaks, we compared seasonal and monthly rainfall and temperature trends in outbreak and nonoutbreak years at four epidemic-prone locations. Our analyses showed that rainfall in outbreak years tended to be above average and higher than rainfall in nonoutbreak years. Overall temperatures were warmer during outbreak years. However, there were a number of distinct deviations in temperature, which seem to play a role in either promoting or inhibiting outbreaks. These preliminary findings show that climatic differences occur between outbreak and nonoutbreak years; however, seasonal and monthly trends differed across geo-climatic regions of the country. More detailed research is imperative if we are to optimize the surveillance and control of epidemic polyarthritic disease in Australia.

Large-scale longitudinal cohort studies are necessary to characterize temporal and geographic variation in Aedes aegypti (L.) (Diptera: Culicidae) production patterns and to develop targeted dengue control strategies that will reduce disease. We carried out pupal/demographic surveys in a circuit of ≈6,000 houses, 10 separate times, between January 1999 and August 2002 in the Amazonian city of Iquitos, Peru. We quantified the number of containers positive for Ae. aegypti larvae and/or pupae, containers holding pupae, and the absolute number of pupae by 4-mo sampling circuits and spatially by geographic area by using a geographic information system developed for the city. A total of 289,941 water-holding containers were characterized, of which 7.3% were positive for Ae. aegypti. Temporal and geographic variations were detected for all variables examined, and the relative importance of different container types for production of Ae. aegypti was calculated. Ae. aegypti larvae and pupae were detected in 64 types of containers. Consistent production patterns were observed for the lid status (lids: 32% wet containers, 2% pupal production), container location (outdoor: 43% wet containers, 85% pupal production), and method by which the container was filled with water (rain filled: 15% wet containers, 88.3% pupal production); these patterns were consistent temporally and geographically. We describe a new container category (nontraditional) that includes transient puddles, which were rare but capable of producing large numbers of pupae. Because of high variable pupal counts, four container categories (large tank, medium storage, miscellaneous, and nontraditional) should be targeted in addition to outdoor rain-filled containers that are not covered by a lid. The utility of targeted Ae. aegypti control is discussed, as well as the ability to achieve control objectives based on published but untested threshold values.

Many mosquito control agencies use carbon dioxide-baited traps as surveillance tools for adult vector populations. However, decisions regarding the number and location of trap sites and the frequency of collections are often based on logistical issues, and not on the bionomics or spatial distribution of the target species. Therefore, with the aim of providing practical information for adult mosquito surveillance programs, we used an array of 81 carbon dioxide- and octenol-baited lights traps to obtain weekly samples of adult mosquitoes in Redland Shire in southeastern Queensland, Australia. The spatial patterns of four different mosquito species were examined, and positive spatial autocorrelation in trap counts was evident for Ochlerotatus vigilax (Skuse), Coquillettidia linealis (Skuse), and Culex annulirostris Skuse, but not for the container species Ochlerotatus notoscriptus (Skuse). Of the three species that exhibited spatially correlated trap counts, the autocorrelation was greatest in Oc. vigilax at a lag distance of 0–1.5 km, with Moran’s I values of 0.30–0.64. Moran’s I indices were also positive and statistically significant (P < 0.05) at lag distances of 1.5–3.0 and 3.0–4.5 km on each of the 15 sampling occasions. However, at 3.0–4.5 km the Moran’s I values were low, which indicated only weak spatial autocorrelation in trap counts. Universal kriging was used to estimate the numbers of each species at unsampled locations throughout the study area, and leave-one-out cross validation analyses indicated that this was a robust method for Cq. linealis and Oc. vigilax. In contrast, trap counts for the container-breeding species Oc. notoscriptus were randomly distributed and the interpolated counts were not reliable. Comparisons of weekly contour maps of adult mosquito counts indicated a consistent spatial pattern for Oc. vigilax and Cq. linealis. Particular geographic areas had consistently high or low numbers of vectors, and these patterns were stable from year to year. Definition of geographic areas with consistently high or low numbers of vectors may allow control activities to be focused in areas with the greatest risk of arbovirus transmission.

We present a population model to understand the effects of temperature and rainfall on the population dynamics of the southern house mosquito, Culex quinquefasciatus Say, along an elevational gradient in Hawaii. We use a novel approach to model the effects of temperature on population growth by dynamically incorporating developmental rate into the transition matrix, by using physiological ages of immatures instead of chronological age or stages. We also model the effects of rainfall on survival of immatures as the cumulative number of days below a certain rain threshold. Finally, we incorporate density dependence into the model as competition between immatures within breeding sites. Our model predicts the upper altitudinal distributions of Cx. quinquefasciatus on the Big Island of Hawaii for self-sustaining mosquito and migrating summer sink populations at 1,475 and 1,715 m above sea level, respectively. Our model predicts that mosquitoes at lower elevations can grow under a broader range of rainfall parameters than middle and high elevation populations. Density dependence in conjunction with the seasonal forcing imposed by temperature and rain creates cycles in the dynamics of the population that peak in the summer and early fall. The model provides a reasonable fit to the available data on mosquito abundance for the east side of Mauna Loa, Hawaii. The predictions of our model indicate the importance of abiotic conditions on mosquito dynamics and have important implications for the management of diseases transmitted by Cx. quinquefasciatus in Hawaii and elsewhere.

Laboratory mice, Mus musculus (L.), BALB/c strain, were assessed for their ability to develop resistance to repeated infestation by Amblyomma americanum (L.) ticks. Mice were infested five consecutive times with A. americanum nymphs. No decrease in tick viability was seen after five infestations, suggesting that BALB/c mice do not develop immune-mediated resistance to A. americanum. In contrast, tick viability was significantly reduced in the second infestation of a New Zealand White rabbit, a laboratory animal known to develop resistance to A. americanum.

Wolbachia symbionts are obligate intracellular bacteria that cause host reproductive alterations in many arthropods and filarial nematodes. We identified Wolbachia symbionts in the cliff swallow bug (Oeciacus vicarious Horvath) and the human bed bug (Cimex lectularius L.) (Hemiptera: Cimicidae) by polymerase chain reaction (PCR) amplification and sequencing using Wolbachia-specific 16S rDNA and FtsZ primers. Phylogenetic analyses using Bayesian, maximum likelihood, and maximum parsimony algorithms indicated, with strong support, that (1) Wolbachia infections in these two cimicid hosts form a monophyletic group, and (2) the Wolbachia strains detected belong to the F clade, previously associated with termites, weevils, and filarial nematodes.

The argasid tick Carios (Ornithodoros) kelleyi Cooley & Kohls is a common ectoparasite of bats and has been found in massive numbers in homes with associated bat colonies in eastern Iowa. This tick feeds nearly exclusively on bats in nature. Several inhabitants of infested homes complained of “bug bites” at night while asleep that may have resulted in erythematous, edematous, urticaric skin lesions and constitutional signs and symptoms. We provide laboratory evidence that a single, engorged C. kelleyi nymph contained host blood from a human female. The clinical implications of our findings are intriguing but unclear.