The completion of the Anopheles gambiae Giles genome sequencing project is a milestone toward developing more effective strategies in reducing the impact of malaria and other vector borne diseases. The successes in developing transgenic approaches using mosquitoes have provided another essential new tool for further progress in basic vector genetics and the goal of disease control. The use of transgenic approaches to develop refractory mosquitoes is also possible. The ability to use genome sequence to identify genes, and transgenic approaches to construct refractory mosquitoes, has provided the opportunity that with the future development of an appropriate genetic drive system, refractory transgenes can be released into vector populations leading to nontransmitting mosquitoes. An. gambiae populations incapable of transmitting malaria. This compelling strategy will be very difficult to achieve and will require a broad substantial research program for success. The fundamental information that is required on genome structure, gene function and environmental effects on genetic expression are largely unknown. The ability to predict gene effects on phenotype is rudimentary, particularly in natural populations. As a result, the release of a refractory transgene into natural mosquito populations is imprecise and there is little ability to predict unintended consequences. The new genetic tools at hand provide opportunities to address an array of important issues, many of which can have immediate impact on the effectiveness of a host of strategies to control vector borne disease. Transgenic release approaches represent only one strategy that should be pursued. A balanced research program is required.

In a Central Michigan wetland setting, with abundant Aedes vexans (Meigen), Anopheles walkeri (Theobald), and Coquillettidea perturbans (Walker), electric fan-generated wind strongly reduced mosquito catches in Centers for Disease Control and Prevention light traps releasing carbon dioxide at 650 or 1,950 ml/min. This relationship was negatively logarithmic over the range of velocities tested (0 m/s to 3.7 m/s; 0–8.3 mph) and closely matched the theoretical dilution function predicted to occur with increasing wind flows over a constantly releasing point source of volatile chemical. Tripling the carbon dioxide release rate consistently doubled the number of mosquitoes caught across the range of wind velocities tested. Neither applied wind velocity nor carbon dioxide release rate was correlated with mosquito body mass across the range of wind velocities tested. Collectively, these data support the conceptual model that applied wind diminishes mosquito catches primarily by diluting attractants rather than by exceeding mosquito flight capability. Previously published estimates of maximum air speeds of which mosquitoes are capable may now need to be reassessed if they were based upon shut-down of catches in traps baited with chemical attractants. Addition of DEET vapor significantly reduced the number of mosquitoes caught relative to equivalently moving air without repellent. We recommend that fan-generated wind should be pursued as a practical means of protecting humans or pets from mosquitoes in the backyard setting.

The sand fly, Lutzomyia longipalpis (Lutz & Neiva) reputedly is a complex of cryptic species; however, there is currently no consensus as to the number of species in the complex or their geographic distributions. We conducted phylogenetic analyses of 31 populations from throughout the species range, using seven isozyme loci and genes in the mitochondrial genome. Analyses of these two independent sets of markers were largely concordant and revealed four distinct clades that support the existence of four species. The four clades have distinct geographic ranges: (1) Brazil (Species A = Lu. longipalpis sensu stricto), (2) Laran (Species B = Lu. pseudolongipalpis), (3) cis-Andean (Species C), and (4) trans-Andean (Species D). The cis-Andean clade may be subdivided further into two groups, one in Colombia and one in northwestern Venezuela, but their taxonomic status remains unresolved. Knowledge that Lu. longipalpis is a complex of species may ultimately shed light on anomalies in the epidemiology of visceral leishmaniasis in the New World.

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 ≈10% slower development and ≈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.

Studies on seasonal anopheline fauna variation were performed in two distinct settlements in the State of Rondônia, Brazil: one at the Madeira River banks (Portuchuelo) with stable native Amazonian population; the other at an inland lumber-extracting farm (Urupá) in dry land, in which adults are mostly migrants. During a 6-yr period (1994–2000), 8,638 adult anophelines were collected: 2,684 in Urupá and 5,954 in Portuchuelo. Anopheles darlingi represented >95% of total mosquitoes caught. Dissection of 4,424 A. darlingi females yielded a very low sporozoite infection index below 0.1%. Oocysts were found in both localities in ∼0.1% of dissected mosquitoes. Determination of the hour biting rates disclosed seasonal variations in both localities. However, in Portuchuelo, mosquito density peaked at the acme of the rainy season, whereas at Urupá it peaked in the dry season. The increase in mosquito density and incidence of malaria cases were coincident. The high mosquito densities observed in the riverine settlement of Portochuelo sector B, which permits evaluation in >10,000 mosquitoes’ bites/person/year, could explain, in spite of the low mosquito’s infection index, the previously described development of natural immunity in the local population that is not observed in the dry land agroindustrial settlement of Urupá.

The principal vector for the pathogens of Lyme disease, human granulocytic ehrlichiosis, and human babesiosis is the tick Ixodes scapularis Say. A chalcid wasp, Ixodiphagus hookeri, in the family Encyrtidae parasitizes populations of the tick on several islands or other geographically isolated sites in New England with high densities of these ticks and white-tailed deer (Odocoileus virginianus), the principal host for adult I. scapularis. Deer densities were reduced at a forested tract in Bridgeport and the Bluff Point Coastal Reserve in Groton, Connecticut, from levels exceeding 90 animals per km² in 1992 (Bridgeport) and 1994 (Bluff Point) to 17 and 10 animals per km², respectively, by fall 2001. Tick densities declined with sustained reductions in the population of white-tailed deer. Similarly, prevalence of tick parasitism by Ixodes hookeri declined at both sites from 30 to 25% to <1.0% and was significantly correlated with previous year’s deer density at both sites (r_s = 0.933 and r_s = 0.867, P ≤ 0.0001) and with nymphal tick densities at Bridgeport (r_s = 0.867, P ≤ 0.0001), but was not as well correlated with tick densities in Groton. The virtual disappearance of I. hookeri in this study corresponds with a lack of I. hookeri in mainland I. scapularis at comparable deer and tick densities, suggesting that there is a threshold deer density of ≈10–20/km², with corresponding tick densities necessary for I. hookeri to successfully parasitize I. scapularis.

Kairomones produced by humans provide female anthropophilic mosquitoes with vital cues used in host-seeking for a blood meal. These chemicals are emanated primarily by the skin and provide the mosquitoes a means to orient themselves to humans at a relatively close range. Chemical studies of these emanations have provided new ideas for the formulation of attractant blends. We report mosquito attraction responses for three binary blends and their separate components. The blends are comprised of L-lactic acid plus either acetone, dichloromethane, or dimethyl disulfide. At the emission rates used in our bioassays, these blends synergistically attract laboratory-reared female Aedes aegypti. Carbon dioxide is not a necessary component to yield high levels of attraction with these blends. It is postulated that at least one of these synergistic blends (L-lactic acid and acetone) produces mosquito attraction behavior similar to L-lactic acid and CO₂.

The influence of indoor microclimate on survival of female Anopheles gambiae sensu stricto Giles (Diptera: culicidae) mosquitoes fed on different nutrition sources was evaluated in a semifield experimental hut exposed to ambient climate in western Kenya. Cages of mosquitoes (n ≈ 50 per cage) were placed in nine positions within the hut combining three different sides and three different heights. At each height and side, mosquitoes were offered either human blood (once every 2 d), glucose (6% wt:vol) or a combination of the two diets over three experiments so that each cage position received one diet source. The effect of diet on survival was significant with mean survival times of 14 d for mosquitoes fed blood alone, 29 d for sugar alone and 33 d for blood plus sugar. Sugar availability decreased the odds of mortality ≈85% compared with the blood group. Micro heterogeneities of temperature but not relative humidity also influenced survival although to a much lesser extent. The side but not height within the hut at which mosquitoes were placed, influenced survival but could not be explained by either temperature or relative humidity differences. The potential influence of seemingly minor heterogeneities of indoor microclimate upon vector longevity and vectorial capacity may merit further investigation. Also, the availability of sugar was shown to be a potentially crucial determinant of vectorial capacity. Compared with blood alone, the availability of sugar served to increase survival potential of vectors beyond ages at which they are old enough to transmit malaria.

We sampled Anopheles funestus Giles mosquitoes from 10 sites in Kenya to determine (1) the different members of this group present at these sites and (2) the population genetic structure of these species based on chromosomal inversion karyotypes with particular reference to the role of the Eastern arm of the Great Rift Valley and geographical distance as barriers to gene flow. Four members of this species group, namely An. funestus s.s., An. parensis, An. leesoni, and An. rivulorum, were found. An. funestus was found almost exclusively inside human dwellings while An. rivulorum was found almost exclusively outdoors. An. parensis was found both indoors and outdoors in comparable proportions. Earlier attempts to collect this group of mosquitoes from two of the sites yielded no mosquitoes, suggesting that numbers of mosquitoes change during the year, sometime reaching zero or very near zero. Significant levels of differentiation were observed among some An. funestus s.s. populations studied. Results suggest that the Rift Valley acts as a barrier to gene flow for An. funestus, but that levels of differentiation are not associated with the geographical distance between populations. The results are discussed in the light of the fact that inversions may have some adaptive significance to differences in environmental conditions.

Several strains of Bacillus sphaericus (Neide) (Bsph), a spore-forming bacterium that produces potent mosquitocidal toxins, have been widely used against mosquito larvae globally. In some locations in the field and in the laboratory, moderate to high levels of resistance in Culex pipiens complex mosquito larvae to Bsph have been reported. Practical strategies for dealing with the resistance problem and more importantly to delay or possibly prevent development of resistance to Bsph in the first place are urgently needed. From published reports, we deduced that Bsph-resistant mosquito larvae are completely susceptible to Bacillus thuringiensis ssp. isreaelensis (Bti) and that there is no appearance of cross-resistance to Bti. On the basis of this information, we hypothesized that Bti offers a good potential for the management of resistance to Bsph, especially when it comes to delaying or preventing emergence of resistance. We conducted laboratory studies to see whether rotation of Bsph and Bti or mixture of both delays or prevents resistance development in Bsph-susceptible California Cx. quinquefasciatus. Development of resistance using Bsph alone increased after F₁₅ and fluctuated slightly on further selections. Rotation of Bsph and Bti resulted in much higher level and rapid emergence of resistance to Bsph. However, selection with mixtures of Bti and Bsph for 36 generations showed no emergence of resistance. On the basis of this information, we believe mixtures have a good potential in delaying or preventing Bsph resistance.

Nitenpyram (Capstar) is a fast acting, orally administered flea treatment that is absorbed into the blood of the host animal and is readily available for uptake by feeding fleas. We examined the efficacy of a single dose of nitenpyram against adult cat fleas, Ctenocephalides felis (Bouché), over several days. We recorded adult flea mortality and flea egg production on treated and untreated cats. Nitenpyram provided 100% kill of all fleas on the host at the time of treatment and for up to 24 h after treatment. Between 24 and 48 h after treatment, there was a 98.6% reduction in adult flea numbers. From 48 to 72 h, there was a 5% reduction in adult fleas. There was a 97% reduction and 95.2% reduction in the number of flea eggs collected from treated versus untreated animals during the first 48 h and from 48 to 72 h, respectively. In addition, we quantified three distinct behavioral responses of infested adult cats treated with nitenpyram to determine the extent of any immediate, overt behavioral responses in treated animals. A significant increase in scratching, biting, licking, and twitching occurred for 5 h. The biting and licking continued for 7 h after treatment. Administration of nitenpyram provides an effective mechanism to eliminate adult fleas from hosts for up to 48 h after treatment.

Infestation parameters are presented for 227 ticks, all Ixodes angustus Neumann, collected from individual mammals (n = 531) in southeastern and south-central Alaska from 1996 to 1999. This tick was recovered from 12 of the 19 mammal species examined, with four species of shrews (Sorex spp.), two species of voles [Clethrionomys gapperi (Vigors) and Clethrionomys rutilus (Pallas)], one species of mouse [Peromyscus keeni (Rhoads)], and the red squirrel [Tamiasciurus hudsonicus (Erxleben)] being the most frequently parasitized hosts. Larvae (n = 146) of I. angustus were collected most often, followed by nymphs (n = 50), females (n = 30), and a single male. The leptinid mammal-nest beetle Leptinus occidentamericanus Peck (1male, 5 females) was also recovered from five individual small mammals; three of these were C. rutilus.

The maculatum agent, Rickettsia parkeri (a member of the spotted fever group rickettsiae), was inoculated into a colony of the lone star tick, Amblyomma americanum, and followed for two tick generations. In addition, guinea pigs were exposed to the agent by direct injection and by feeding infected ticks on them. Eighty (53%) of 150 nymphal A. americanum that were inoculated with suspensions of R. parkeri were positive by hemolymph test and fluorescent antibody test for rickettsial infection when examined as adults. One-month survival of R. parkeri-infected ticks was similar to that of control (noninfected) ticks. Transstadial and transovarial transmission of R. parkeri in the laboratory was demonstrated in A. americanum. When guinea pigs were exposed to the maculatum agent by either direct injection of Vero cell-grown R. parkeri, injection of homogenates of infected ticks, or by feeding of infected ticks, they developed mild fevers and occasional scrotal reactions. These data indicate that R. parkeri can remain viable in lone star ticks for two generations and suggest that guinea pigs may become infected, displaying mild clinical signs.

During 1998–1999, Ixodes ricinus (L.) populations were investigated in three different biotopes (deciduous, mixed, coniferous forest) situated in popular recreational areas in Poznań, Poland. In total, 1,123 questing ticks (1,002 nymphs, 69 males, 52 females) were collected by flagging vegetation. Additionally, in 1998 between May and September small rodents were trapped and inspected for feeding ticks. Altogether, 213 rodents of three species: Apodemus agrarius Pall., A. flavicollis Melchior, Clethrionomys glareolus Schreber were captured. Of 323 engorged ticks, 304 were larvae and 19 nymphs. All ticks collected from vegetation, as well as from rodents were examined for the presence of Borrelia burgdorferi Johnson, Schmid, Hyde, Steigenwalt & Brenner s.l. spirochetes by indirect immunofluorescence assay (IFA) using PAB 1B29. The seasonal pattern of activity of questing I. ricinus was always bimodal (May/June and August/September). The most abundant tick population occurred in the deciduous forest. The total infection rate in questing ticks was 16.2%. Differences in mean infection prevalence of host-seeking ticks between three biotopes each year were not significant. On average more larvae parasitized on the genus of Apodemus than on C. glareolus. 17.8% of larvae and 31.6% of nymphs fed on rodents harbored spirochetes. The three rodent species contributed to a different degree in to transmission of the pathogen to subadult stages. ≈27% of larvae infested on A. agrarius, 22% on C. glareolus, and only 4.2% on A. flavicollis contained spirochetes. The results suggest that the prevalence of A. agrarius and C. glareolus in disturbed urban forests used for leisure activities seems to be crucial for the maintenance of B. burgdorferi s.l. in I. ricinus populations.

Virus dissemination and associated pathology were examined in Aedes albopictus after intrathoracic inoculation of Sindbis virus (SIN), the prototypic Alphavirus. At 10 days postinfection, virus RNA was detected in all three-body segments of the insect. Colocalization of virus antigen with structural pathology was observed in mosquito salivary glands and midgut-associated visceral muscles, representing yet another example of arbovirus-associated pathology in a mosquito host. SIN antigen and gross pathology were detected in lateral lobes, but not the medial lobe of salivary glands, whereas virus antigen, vacuolated cytoplasm, and myofilament misalignment were detected in the visceral muscles at the midgut exterior surface. Early in the midgut infection, virus antigen was localized in small foci on the organ surface that progressed to a grate work-like banding pattern that eventually cleared. Both the salivary glands and the midgut are essential to insect survival and reproduction. Additionally, these organs provide a pathway for virus transmission in nature. Although SIN infection may not shorten the mosquito life span, persistent coexistence could permit survival of both host and microbe as well as contribute to alterations in insect behavior.

There is a consensus that malaria is a growing problem in African highlands. This is surprising because many parts of the highlands were considered too cold to support transmission. In this report, we examined how transmission of Plasmodium falciparum in six villages changed along an altitude transect in the Usambara Mountains, Tanzania, from 300 m to 1700 m. Routine entomological collections were made using spray catches and light traps for 15 mo. Direct estimates of entomological inoculation rates and indirect estimates of vectorial capacity suggested a >1000-fold reduction in transmission intensity between the holoendemic lowland and the hypoendemic highland plateau. Lowland transmission was perennial with a significant peak in the cool season after the long rains in May, when vectors densities were high. In the highlands, low temperatures prevented parasite development in mosquitoes during the cool season rains, and highland transmission was therefore limited to the warm dry season when vector densities were low. The primary effect of increasing altitude was a log-linear reduction in vector abundance and, to a lesser extent, a reduction in the proportion of infective mosquitoes. Highland malaria transmission was maintained at extraordinarily low vector densities. We discuss herein the implications of these findings for modeling malaria and suggest that process-based models of malaria transmission risk should be improved by considering the direct effect of temperature on vector densities. Our findings suggest that variation in the short rains in November and changes in agricultural practices are likely to be important generators of epidemics in the Usambaras.

Burrows within black-tailed prairie dog (Cynomys ludovicianus) colonies on the Rocky Mountain Arsenal National Wildlife Refuge, Colorado, were dusted with deltamethrin insecticide to reduce flea (Insecta: Siphonaptera) abundance. Flea populations were monitored pre- and posttreatment by combing prairie dogs and collecting fleas from burrows. A single application of deltamethrin significantly reduced populations of the plague vector Oropsylla hirsuta, and other flea species on prairie dogs and in prairie dog burrows for at least 84 d. A plague epizootic on the Rocky Mountain Arsenal National Wildlife Refuge caused high mortality of prairie dogs on some untreated colonies, but did not appear to affect nearby colonies dusted with deltamethrin.

In this study, we have developed molecular methods for the identification of reservoir hosts of sylvatic tick-borne zoonoses. The methods are based on the analysis of the blood meal remnant in the tick gut and include detection of pathogens and identification of the host origin of the blood meal. For host identification, a universal primer pair was used to amplify part of the vertebrate 18S rRNA gene followed by reverse line blot hybridization using subgroup-specific probes. Analyses of DNA from whole blood of vertebrates identified the correct subgroup of a broad range of vertebrate species (e.g., Ruminantia, Leporidea, Canidae, Murinae, Arvicolinae, Insectivora, Galliformes, Passeriformes) using probes based on the 18S rDNA sequences. Host DNA in the remnants of larval blood meals was detected in the gut of Ixodes ricinus nymphs maintained under natural conditions up to 9 mo after molting. For pathogen identification, a multiplex polymerase chain reaction was used that targeted parts of the 18S rRNA gene of piroplasm protozoa, the 16S rRNA gene of bacteria, and the intergenic spacer of the Borrelia burgdorferi genospecies complex. The utility of both methods was demonstrated under laboratory conditions by detecting Babesia microti (Franca) and gerbil DNA in 3-mo-old I. ricinus nymphs that had fed on B. microti-infected gerbils as larvae, and under field conditions by analyzing unfed ticks that were collected in a forest. The field study showed that the majority of ticks had fed on ruminants or birds and few on rodents, which is in accord with our knowledge of the fauna in this forest. Few pathogens were detected but the discovery of Borrelia valaisiana and B. burgdorferi s.s. in ticks that had fed on deer and Borrelia afzelii in a tick that had fed on a bird raises questions about the mode of transmission of these spirochetes and possibly about their host specificity.

Host-seeking ticks often remain on clothing of persons returning home from work or recreation in tick habitats, and can pose at least a temporary risk to people and pets in these homes. Laundering clothing has been one of the recommendations to reduce tick exposure. Host-seeking lone star tick, Amblyomma americanum (L.), and blacklegged tick, Ixodes scapularis Say, nymphs confined in polyester mesh packets, were included with laundry in cold, warm, and hot wash cycles of an automatic clothes washer. Ticks were also placed with washed clothing and subjected to drying in an automatic clothes dryer set on high heat and on air only (unheated). Most nymphs (≥90%) of both species survived the cold and warm washes, and 95% of A. americanum nymphs survived the hot wash. At the time of their removal from the washer, I. scapularis nymphs were clearly affected by the hot wash, but 65% were considered alive 20–24 h later. Large percentages of nymphs of both species survived hot washes in which two other detergents (a powder containing a nonchlorine bleach and a liquid) were used. All ticks were killed by the 1 h cycle at high heat in the clothes dryer, but with unheated air some nymphs of both species survived the 1 h cycle in the dryer. Given the laundering recommendations of clothing manufacturers and variation in the use automatic clothes washers, laundry washed in automatic washers should not be considered free of living ticks.