Ross River virus (RR) is a mosquito-borne arbovirus responsible for outbreaks of polyarthritic disease throughout Australia. To better understand human and environmental factors driving such events, 57 historical reports on RR outbreaks between 1896 and 1998 were examined collectively. The magnitude, regularity, seasonality, and locality of outbreaks were found to be wide ranging; however, analysis of climatic and tidal data highlighted that environmental conditions act differently in tropical, arid, and temperate regions. Overall, rainfall seems to be the single most important risk factor, with over 90% of major outbreak locations receiving higher than average rainfall in preceding months. Many temperatures were close to average, particularly in tropical populations; however, in arid regions, below average maximum temperatures predominated, and in southeast temperate regions, above average minimum temperatures predominated. High spring tides preceded coastal outbreaks, both in the presence and absence of rainfall, and the relationship between rainfall and the Southern Oscillation Index and La Niña episodes suggest they may be useful predictive tools, but only in southeast temperate regions. Such heterogeneity predisposing outbreaks supports the notion that there are different RR epidemiologies throughout Australia but also suggests that generic parameters for the prediction and control of outbreaks are of limited use at a local level.

The effectiveness of five mosquito traps at sampling anopheline mosquitoes was compared with landing/biting (L/B) collections in western Thailand. Traps evaluated included a CDC style light trap (CDC LT) with dry ice, the American Biophysics Corporation (ABC) standard light trap (ABC LT) with dry ice and octenol, the ABC counterflow geometry (CFG) trap with dry ice and octenol, the ABC mosquito magnet (MM) trap with octenol, and the Nicosia and Reinhardt Company Mosquito Attractor Device (N&R trap). Mosquito numbers captured in landing-biting collections were 5.2, 7.0, 7.3, 31.1, and 168.8 times greater than those collected in the ABC LT, MM, CDC LT, CFG, and N&R traps, respectively, for Anopheles minimus Theobald, the predominant malaria vector in the region. Similar results were obtained for the secondary malaria vectors Anopheles maculatus Theobald and Anopheles sawadwongporni Rattanarithikul & Green. Only Anopheles kochi Doenitz was collected in significantly greater numbers in the CDC LT, ABC LT, and MM traps compared with L/B collections. Although none of the traps were as effective as L/B collections, the ABC LT, MM, and CDC LT were the best alternatives to human bait for the collection of anopheline malaria vectors in Thailand.

The current study was performed on the Bioko Island (Equatorial Guinea) with the aim of establishing a rapid assessment technique for mapping malaria risk and measuring vector densities. Human bait collection, tent traps, light traps, indoor resting collection, and window exit traps were used to collect Anopheles gambiae s.s. and Anopheles funestus, the two anopheline species involved in malaria transmission in this island. Capture data were used to compare differences in the behavior and vectorial capacity of An. gambiae s.s. and An. funestus. Differences in the two species of mosquitoes were found in relation to the season and trapping methods used. Entomological inoculation rates (EIR) for Plasmodium falciparum were calculated using a polymerase chain reaction (PCR) test with individual anopheline mosquitoes from human bait collections in two villages during the dry and rainy seasons. P. falciparum sporozoites were detected from both dissected heads/thorax and abdomens of both species.

In this study, we measured the phenology of Ixodes ricinus ticks and their infection with Borrelia burgdorferi sensu lato (sl) simultaneously along an altitudinal gradient to assess the impact of climate on the phenology of ticks and on their infection with B. burgdorferi sl. From 1999 to 2001, free-living I. ricinus ticks were collected monthly by flagging vegetation at three different altitudes (620, 740, and 900 m above sea level) on the slope of a mountain in Chaumont (Neuchâtel, Switzerland). I. ricinus ticks were examined for the presence of B. burgdorferi sl by using direct fluorescent antibody assay and isolation of spirochetes. Borrelia species were characterized by polymerase chain reaction followed by restriction fragment-length polymorphism. Tick density and tick phenology varied with altitude. Although the peak tick density decreased and the onset of ticks was delayed with altitude, the phenology was much more stable among years at the highest altitudes than at the lowest. The prevalence of B. burgdorferi infection in nymphs and adults decreased with altitude. The prevalence of infection differed significantly among years, and it was significantly higher in adults (30%) than in nymphs (21%). B. burgdorferi infection in adults was positively related with adult density, but this was not observed for nymphs. Five B. burgdorferi sl genospecies were successfully isolated: B. garinii, B. burgdorferi sensu stricto, B. afzelii, B. valaisiana, and B. lusitaniae. Mixed infections were obtained from five of 140 infected ticks. The greatest diversity in Borrelia species was observed at the lowest altitude where all five Borrelia species were present, whereas at the two highest altitudes, B. lusitaniae was not observed.

Parasitic life stages of Amblyomma maculatum Koch were collected from domestic cattle and several species of wild mammals during a 3.5-yr study (May 1998–October 2001) in north-central Oklahoma. Adult ticks were the predominant life stage collected from cattle, white-tailed deer, coyotes, and raccoons, whereas only immature ticks were collected from cotton rats and white-footed mice. The prevalence of adult A. maculatum on white-tailed deer (n = 15) examined in June, July, and August 1998 was 80, 100, 100%, respectively. The prevalence of adult A. maculatum on cattle (n = 84) ranged from 52% in February 1999 to 100% in May 1999. The prevalence of adult A. maculatum on coyotes (n = 16) was 100% in April 1998 and 43% on coyotes (n = 7) examined in January 2001. The prevalence of adult A. maculatum on raccoons (n = 23) examined during May, June, and July 1999 was 13%. No A. maculatum of any life stage were recovered from opossums (n = 7). Nine hundred forty-five rodents were trapped over 294 trap-nights; prevalence of A. maculatum larvae and nymphs on cotton rats (n = 395) was 34 and 15%, respectively, whereas on white-footed mice (n = 517), prevalence was 1.5 and 1.4%, respectively. No A. maculatum were recovered from pack rats (n = 33). There were significant differences (P = 0.0001) in larval infestation prevalence between cotton rats and white-footed mice in the spring, summer, and fall and for nymphs in the spring and summer. Results of A. maculatum parasitism and seasonal occurrence on hosts in this study are compared with previous research conducted in Oklahoma and with collection records of A. maculatum in the Entomology Museum at Oklahoma State University.

Oviposition is a critical stage in the mosquito lifecycle, and may determine population levels, distribution, biting behavior, and pathogen transmission. Knowledge of the oviposition behavior of Culex restuans Theobald has become particularly important with the emergence of West Nile virus (WNV) in North America. Laboratory and field studies have examined some factors that contribute to oviposition choice in Culex spp., but few studies have investigated responses to cues of future competition and breeding habitat availability in the field. We hypothesized that female Cx. restuans mosquitoes avoid laying eggs in habitats containing cues of larval competition, and that increased availability of larval habitat decreases egg density. To test these hypotheses, a series of field experiments were conducted in southeastern Michigan during summer 2002. We found that female mosquitoes prefer nutrient-enriched containers and decrease ovipositing in containers with conspecific larvae. In addition, greater habitat abundance decreased egg clutch density per container, although there was considerable aggregation of egg clutches. These results support our hypotheses and have potentially important implications for pathogen transmission by mosquitoes.

Unequivocal identification of phlebotomine sand flies is of crucial importance in epidemiological studies of leishmaniasis, because certain species may act as vectors, depending on behavior and physiology. For Lutzomyia whitmani, a major vector of American human cutaneous leishmaniasis in Brazil, an increasing number of studies have suggested the existence of a species complex. In the present work, we evaluated the genetic variability of L. whitmani populations from four Brazilian foci of that disease: Corte de Pedra, Ilhéus, Martinho Campos, and Serra de Baturité. Computational analysis of 85 characters, generated by RAPD-polymerase chain reaction, demonstrated high intrapopulational variability. Those characters led to sex discrimination in three of the populations, with the exception of Martinho Campos individuals, in which sex distinction was not complete. One and two interpopulational phenograms were obtained for females and males, respectively. A higher similarity was observed among the specimens from Ilhéus, Corte de Pedra, and Serra de Baturité, whereas the Martinho Campos population remained external to that cluster. These results, which are in partial accordance with a previous morphometric survey of L. whitmani from the same regions, provide additional evidence to support the existence of at least two spatial clusters of biogeographical populations of L. whitmani in Brazil.

Amitraz, a formamidine acaricide, plays an important role in the control of the southern cattle tick, Boophilus microplus (Canestrini), and other tick species that infest cattle, dogs, and wild animals. Although resistance to amitraz in B. microplus was previously reported in several countries, the actual measurement of the level of amitraz resistance in ticks has been difficult to determine due to the lack of a proper bioassay technique. We conducted a survey, by using a newly reported technique that was a modification of the standard Food and Agriculture Organization larval packet test, to measure the levels of resistance to amitraz in 15 strains of B. microplus from four major cattle-producing states in Mexico. Low-order resistance (1.68- to 4.58-fold) was detected in 11 of those strains. Our laboratory selection using amitraz on larvae of the Santa Luiza strain, which originated from Brazil, achieved a resistance ratio of 153.93 at F₆, indicating the potential for high resistance to this acaricide in B. microplus. Both triphenylphosphate and piperonyl butoxide significantly synergized amitraz toxicity in both resistant and susceptible tick strains. Diethyl maleate synergized amitraz toxicity in one resistant strain but had no effect on the susceptible strain and had minor antagonistic effects on two other resistant strains. Target site insensitivity, instead of metabolic detoxification mechanisms, might be responsible for amitraz resistance observed in the Santa Luiza strain and possibly in other amitraz resistant B. microplus ticks from Mexico. The Santa Luiza strain also demonstrated high resistance to pyrethroids and moderate resistance to organophosphates. Multiple resistance shown in this strain and other B. microplus strains from Mexico poses a significant challenge to the management of B. microplus resistance to acaricides in Mexico.

The infectivity of Plasmodium-infected humans in western Thailand was estimated by feeding laboratory-reared Anopheles dirus Peyton and Harrison mosquitoes on venous blood placed in a membrane-feeding apparatus. Between May 2000 and November 2001, a total of 6,494 blood films collected during an active malaria surveillance program were checked by microscopy for the presence of Plasmodium parasites: 3.3, 4.5, and 0.1% of slides were P. falciparum- (Pf), P. vivax- (Pv), and P. malariae (Pm)-positive. Venous blood was collected from 70, 52, 6, and 4 individuals infected with Pf, Pv, Pm, and mixed Pf/Pv, respectively, with 167 uninfected individuals serving as negative controls. Only 10% (7/70), 13% (7/52), and 0% (0/6) of membrane feeds conducted on Pf-, Pv-, and Pm-infected blood yielded infected mosquitoes. One percent (2/167) of microscope-negative samples infected mosquitoes; however, both samples were subsequently determined to be Pf-positive by polymerase chain reaction. Gametocytes were observed in only 29% (4/14) of the infectious samples. All infections resulted in low oocyst loads (average of 1.2 oocysts per positive mosquito). Only 4.5% (10/222) of mosquitoes fed on the seven infectious Pf samples developed oocysts, whereas 2.9% (9/311) of mosquitoes fed on the seven infectious Pv samples developed oocysts. The probability of a mosquito becoming infected with Pf or Pv after a blood meal on a member of the human population in Kong Mong Tha was estimated to be 1 in 6,700 and 1 in 5,700, respectively. The implications toward malaria transmission in western Thailand are discussed.

We evaluated the performance of the VecTest Malaria Antigen Panel (V-MAP) assay for the detection of Plasmodium falciparum and P. vivax (variants 210 and 247) circumsporozoite protein in anopheline mosquitoes in Thailand. The V-MAP assay is a rapid, one-step procedure using a ‘dipstick’ wicking test strip. The circumsporozoite (CS) ELISA was used as the reference standard. Mosquitoes evaluated in the study included field-collected specimens (n = 930) and laboratory-reared specimens that had been fed on blood collected from patients with and without Plasmodium gametocytes (n = 4,110) or on cultured P. falciparum gametocytes (n = 262). Field-collected mosquitoes were triturated individually or in pools of 2–5 and tested using 613 V-MAP assays. Laboratory-reared specimens were tested individually using 4,372 V-MAP assays. Assay performance depended on the species of Plasmodium and the number of sporozoites used as the cut-off. For P. falciparum, optimal performance was achieved using a cut-off of 150 sporozoites (sensitivity = 100%, specificity = 99.2%, and accuracy = 0.99). For P. vivax variant 210, optimal performance was also achieved using a cut-off of 150 sporozoites (sensitivity = 94.8%, specificity = 94.5%, and accuracy = 0.95). We were unable to develop a standard-curve for the CS-ELISA using P. vivax variant 247 because of a lack of sporozoites; however, using a cut-off of 30 pg P. vivax 247 antigen (mosquitoes with less than this amount of antigen were considered negative), assay performance (sensitivity = 94.3%, specificity = 99.2%, and accuracy = 0.99) was comparable to that achieved for P. falciparum and P. vivax 210. These results clearly demonstrate that the V-MAP assay performs at an acceptable level and offers practical advantages for field workers needing to make rapid surveys of malaria vectors.

We examined the prevalence of antibodies to three mosquito-borne arboviruses in blue jays, Cyanocitta cristata, and Florida scrub-jays, Aphelocoma coerulescens, to identify the effects on host survival, the influence of sex and age on infection, and the temporal patterns of antibody prevalence. Blood samples from 306 blue jays and 219 Florida scrub-jays were collected at Archbold Biological Station (Lake Placid, FL) from April 1994 through December 1995. Sera were analyzed for hemagglutination-inhibition antibody to eastern equine encephalitis (EEE) and St. Louis encephalitis (SLE) viruses, and neutralizing antibodies to EEE, Highlands J (HJ), and SLE viruses. Overall, 31.4% of blue jay samples and 22.1% of scrub-jay samples had antibodies to EEE. Antibodies to HJ were detected in slightly >15% of samples in each jay species, and SLE was detected in <3% of the samples in each jay species. A single EEE virus isolation was made from the blood of an 11-d-old scrub-jay nestling. Survival of adult blue jays seropositive to EEE was significantly lower than that of seronegative birds based on resight rates, but infection did not seem to affect survival of adult or juvenile Florida scrub-jays.

The Lyme disease spirochete, Borrelia burgdorferi Johnson, Schmidt, Hyde, Steigerwalt, and Brenner, was discovered in blacklegged ticks, Ixodes scapularis Say at Turkey Point, Ontario, Canada. We report the first isolation of B. burgdorferi from a vertebrate animal collected on mainland Ontario. During this 2-yr study, spirochetes were isolated from the white-footed mouse, Peromyscus leucopus Rafinesque, and attached I. scapularis larvae. Similarly, isolates of B. burgdorferi were cultured from blacklegged tick adults, and confirmed positive with polymerase chain reaction by targeting OspA and rrf (5S)-rrl (23S) genes. Moreover, all isolates of B. burgdorferi from this area had complementary genetic structure, and the second primer set amplicons confirmed the first primer set amplification products. These findings show an epicenter endemic for B. burgdorferi within an established population of I. scapularis at Turkey Point.

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°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.

Epidemiological evidence suggests that the nymph of the western black-legged tick, Ixodes pacificus Cooley and Kohls, is the primary vector of the Lyme disease spirochete, Borrelia burgdorferi Johnson, Schmid, Hyde, Steigerwalt, and Brenner, to humans in northwestern California. In spring 2002, six different human behaviors were evaluated as potential risk factors for acquiring I. pacificus nymphs in a deciduous woodland in Mendocino County, California. Also, the prevalence of B. burgdorferi sensu lato (s.l.) and the causative agents of human granulocytic (Anaplasma phagocytophilum [Foggie] Dumler, Barbet, Bekker, Dasch, Palmer, Ray, Rikihisa, and Rurangirwa) and monocytic ehrlichioses (Ehrlichia chaffeensis Anderson, Dawson, Jones, and Wilson) was determined in nymphs that had been collected from subjects or by dragging leaf litter. Activities involving a considerable degree of contact with wood resulted in greater acquisition of nymphs than those involving exposure solely to leaf litter. Time-adjusted tick-acquisition rates demonstrated that sitting on logs was the riskiest behavior, followed, in descending rank, by gathering wood, sitting against trees, walking, stirring and sitting on leaf litter, and just sitting on leaf litter. The number of ticks acquired appeared to be unrelated to the type of footwear worn (hiking boots, hiking sandals, or running shoes). Overall, 3.4% (n = 234) of the nymphs were infected with A. phagocytophilum, 3.9% (n = 181) with B. burgdorferi s.l., and none (n = 234) with E. chaffeensis. Of 13 nymphs infected with either A. phagocytophilum or B. burgdorferi s.l., 2 (15.4%) were coinfected with both bacteria, as were 1.3% of 158 nymphs obtained from leaf litter, the first report of coinfection in this life stage of I. pacificus. Four unattached, infected nymphs were removed from subjects, including two acquired while sitting on logs that contained A. phagocytophilum, another with the same bacterium obtained while walking, and one acquired while gathering wood that was infected with B. burgdorferi s.l. Despite the use of extreme personal preventive measures by both subjects, two attached, uninfected nymphs were removed from one of them ≥1–2 d postexposure. The public health implications of these findings are discussed.

The repellents N,N-diethyl-3-methylbenzamide (deet) and racemic 2-methylpiperidinyl-3-cyclohexene-1-carboxamide (AI3-37220) were evaluated using two different laboratory bioassays to determine their relative effectiveness against host-seeking nymphs of the blacklegged tick, Ixodes scapularis Say, and the lone star tick, Amblyomma americanum (L.). In a petri dish bioassay, ticks were released within a ring of repellent on a horizontal filter paper disk. In the second bioassay, ticks were allowed to climb a vertical strip of filter paper whose central portion was treated with a repellent. Deet and AI3-37220 were more effective against I. scapularis than A. americanum nymphs. In the petri dish bioassay, none of the concentrations of deet or AI3-37220 tested confined A. americanum within the treated ring. However, in the vertical bioassay, both species exhibited avoidance of the repellents, and I. scapularis was repelled by much lower concentrations than A. americanum. I. scapularis were repelled by lower concentrations in the vertical bioassay than in the petri dish bioassay. Deet was slightly more effective against I. scapularis than AI3-37220 in both bioassays, but AI3-37220 was significantly more effective than deet against A. americanum in the vertical bioassay.

Knowledge of biogeographic variation in Wolbachia infection rates and inferred susceptibility to infection among different mosquito taxa has fundamental implications for the design and successful application of Wolbachia-based vector-borne disease control strategies. Using a Wolbachia-specific polymerase chain reaction assay, we tested 14 North American mosquito species in five genera (Aedes, Anopheles, Culiseta, Culex, and Ochlerotatus) for Wolbachia infection. Wolbachia infections were only detected in members of the Culex pipiens (L.) species complex.