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The Asian longhorned tick (Haemaphysalis longicornis Neumann), native to East Asia, was first reported in the United States in 2017 and is now established in at least 17 states. Haemaphysalis longicornis feeds on birds in its range outside of the United States, and migratory birds disperse this tick and tick-borne pathogens. However, early studies in the United States did not find H. longicornis on migrating passerine birds. The transport of the parthenogenetic H. longicornis on birds has the potential to greatly expand its range. We report the first discovery of H. longicornis on migratory passerine birds in the Americas.

The Asian longhorned tick, Haemaphysalis longicornis Neumann, is a species native to eastern Asia that has recently been discovered in the United States. In its native range, H. longicornis transmits pathogens that cause disease in humans and livestock. It is currently unknown whether H. longicornis will act as a vector in the United States. Understanding its seasonal activity patterns will be important in identifying which times of the year represent greatest potential risk to humans and livestock should this species become a threat to animal or public health. A study site was established in Yonkers, NY near the residence associated with the first reported human bite from H. longicornis in the United States. Ticks were collected once each week from July 2018 to November 2019. Haemaphysalis longicornis larvae were most active from August to November, nymphs from April to July, and adult females from June to September. This pattern of activity suggests that H. longicornis is capable of completing a generation within a single year and matches the patterns observed in its other ranges in the northern hemisphere. The data presented here contribute to a growing database for H. longicornis phenology in the northeastern United States. Potential implications of the short life cycle for the tick’s vectorial capacity are discussed.

Lyme disease is a serious infectious disease having a restricted worldwide distribution for which there is no vaccine available for human use.This study was designed to determine common reactive antigens involved in Borrelia burgdorferi (Bb) infection that are recognized in mammalian sera that may be useful for vaccine development.Blood samples were collected from patients with documented Lyme disease, and from rabbits and mice experimentally infected with either tick-transmitted or culture-grown Borrelia burgdorferi. All samples were then processed for sera. For performing the Western blots, sonicated Bb organisms (whole cell lysates) and protein ladders were separated by protein gel electrophoresis. Immune reactivities of the electrophoresed proteins with the serum samples were then probed with anti-HRP IgG reagent.Rabbit, mouse and human sera consistently reacted with the 41 kDa band of Bb which corresponded to the flagellin protein - the major protein component of this organism's periplasmic flagella, also known as axial filaments or fibrils. Various other Bb antigens of wide molecular weight ranges were also recognized by rabbit and human sera, and less frequently with mouse sera.The strong immune response to the 41 kDa flagellin protein by the different mammalian species suggests the utility of a possible vaccine targeting this protein, although other proteins may also be appropriate, for preventing Lyme disease following a bite from an infected tick.

Doxycycline is recommended for persons with Ixodes scapularis tick bites in certain geographic areas, if the tick had fed for at least 36 hours. Based on the scutal index, over 40% of I. scapularis tick bites from patients seen at the Lyme Disease Diagnostic Center did not warrant antibiotic prophylaxis.

Genetic diversity of Anaplasma phagocytophilum was assessed in specimens from 16 infected patients and 16 infected Ixodes scapularis ticks. A region immediately downstream of the 16S rRNA gene, which included the gene encoding SdhC, was sequenced. For the A. phagocytophilum strains from patients no sequence differences were detected in this region. In contrast, significantly fewer ticks had a sequence encoding SdhC that was identical to that of the human strains (11/16 vs. 16/16, p = 0.04). This variation is consistent with the premise that not all A. phagocytophilum strains present in nature are able to cause clinical illness in humans. A strain referred to as A. phagocytophilumVariant-1 that is regarded as non-pathogenic for humans was previously described using a different typing method. Data from the current study suggest that both typing methods are identifying the same non-pathogenic strains.

Summary Borrelia burgdorferi sensu lato (s.l.), transmitted by Ixodes spp. ticks, is the causative agent of Lyme disease. Although Ixodes spp. ticks are distributed in both Northern and Southern Hemispheres, evidence for the presence of B. burgdorferi s.l. in South America apart from Uruguay is lacking. We now report the presence of culturable spirochetes with flat-wave morphology and borrelial DNA in ende- mic Ixodes stilesi ticks collected in Chile from environmental vegetation and long-tailed rice rats (Oligoryzomys longicaudatus). Cultured spirochetes and borrelial DNA in ticks were characterized by multilocus sequence typing and by sequencing five other loci (16S and 23S ribosomal genes, 5S-23S intergenic spacer, flaB, ospC). Phylogenetic analysis placed this spirochete as a new genospecies within the Lyme borreliosis group. Its plasmid profi- le determined by polymerase chain reaction and pulsed-field gel electrophoresis differed from that of B. burgdorferi B31A3. We propose naming this new South American member of the Lyme borreliosis group B. chilensis VA1 in honor of its country of origin.

Ticks expend energy while host-seeking and must consume blood to advance to the next life stage. The energy required for activity is derived from the tick's lipid reserves, a valuable resource that sustains the tick until it finds the next host and can take another bloodmeal. The amount of lipid reserves in an unfed tick has been proposed as an index of tick biological age. Two different methods for aging nymphal blacklegged ticks, Ixodes scapularis Say, were analyzed in this study. To study lipid usage, colony-raised nymphs were held in lab-controlled chambers at 21.0 °C and ≥95% relative humidity, with a photoperiod of 12:12 (L:D) h. Samples of ticks were frozen at -80 °C every 2-3 wk, starting at 12-wk postmolt and continuing until 38-wk postmolt. Lipid reserves were determined indirectly through measurements of "physiological age" that estimate the energy a tick has based on the evaluation of morphometric size ratios of the tick scutum and alloscutum, and quantified directly through chloroform extractions of lipid from individual ticks. Morphometric age ratios and lipid amounts were compared to determine if morphometric measurements accurately estimated a tick's physiological state. Although the morphometric age ratio did correlate significantly with total tick lipid content, the predictive value of the ratio was not reliable; chloroform extraction results showed that lipid amounts declined steadily through the study and more accurately characterized the physiological condition of nymphal I. scapularis. The study of physiological aging of blacklegged ticks may lead to a better understanding of how changing environmental conditions affect tick longevity.

Ixodes scapularis Say, the blacklegged tick, vectors Borrelia burgdorferi Johnson et al. 1984, the bacterium that causes Lyme disease, the most important vector-borne disease in the United States. Efforts to reduce I. scapularis populations are shifting toward the development of biological control methods. Currently, only a few entomopathogenic fungal species are considered virulent to ticks. We hypothesized that these species may not represent the most abundant local taxa that would be pathogenic to ticks in situ. To identify potential entomopathogenic fungi at a study site in Westchester County, New York, we sampled soils and ticks, extracted and amplified the internal transcribed spacer region of nuclear ribosomal DNA (nrDNA), and compared sequences with those in GenBank. Over three sampling periods from June 2007 to May 2008, 70 fungal taxa were isolated and identified from soils (48 taxa) and ticks (27 taxa; 5 taxa were found both in soil and on ticks) collected in this study, encompassing species in 25 different genera. In laboratory bioassays, 15 fungal taxa were found to be significantly virulent, although none of these were previously considered common pathogens of I. scapularis. Two species, Hypocrea lixii Patouillard 1891 and Penicillium soppii K. M. Zalessky 1927, were tested in field trials by spraying suspensions on forested plots. Mean tick mortality was 71% after treatment with H. lixii, 58% after treatment with P. soppii, and 32% in the control plots. The complete diversity of entomopathogenic fungal species at this site is yet to be defined, but, in general, such fungi appear to be more common in forest habitats where I. scapularis resides than previously thought. Examination of intact fungal communities can provide information that serves as the foundation for site-specific biocontrol programs.

We compared spatial and temporal patterns of resource use by feral and abandoned domestic dogs (Canis familiaris) on the Navajo reservation in Arizona and New Mexico. Community dumps provide locally abundant food resources utilized both by feral dogs and dogs abandoned at the dump site. Although population parameters were much the same for feral and abandoned dogs, the use of space varied distinctly and reflected behavioral differences in the way each population responded to the absence of human control, the need to acquire food, and the developmental state of pups. Temporal use of resources by feral dogs varied seasonally with the age of pups in one pack, but not in a second pack. Priority of access to local resources may be influenced by aggressive interactions among dogs at a dump. Barking may serve to warn dogs already present at a dump that competitors have arrived.

As part of the Northeast Area-wide Tick Control Project (NEATCP), meta-analyses were performed using pooled data on the extent of tick-vector control achieved through seven concurrent studies, conducted within five states, using U.S. Department of Agriculture "4-Poster" devices to deliver targeted-acaricide to white-tailed deer. Although reductions in the abundance of all life-stages of Ixodes scapularis were the measured outcomes, this study focused on metrics associated with I. scapularis nymphal tick densities as this measure has consistently proven to directly correlate with human risk of acquiring Lyme disease. Since independent tick sampling schemes were undertaken at each of the five environmentally distinct study locations, a meta-analytic approach permitted estimation of a single true control-effect size for each treatment year of the NEATCP. The control-effect is expressed as the annual percent I. scapularis nymphal control most consistent with meta-analysis data for each treatment year. Our meta-analyses indicate that by the sixth treatment year, the NEATCP effectively reduced the relative density of I. scapularis nymphs by 71% on the 5.14 km(2) treatment sites, corresponding to a 71% lower relative entomologic risk index for acquiring Lyme disease.

Entomopathogenic fungi are commonly found in forested soils that provide tick habitat, and many species are pathogenic to Ixodes scapularis Say, the blacklegged tick. As a first step to developing effective biocontrol strategies, the objective of this study was to determine the best methods to isolate entomopathogenic fungal species from field-collected samples of soils and ticks from an Eastern deciduous forest where I. scapularis is common. Several methods were assessed: (1) soils, leaf litter, and ticks were plated on two types of media; (2) soils were assayed for entomopathogenic fungi using the Galleria bait method; (3) DNA from internal transcribed spacer (ITS) regions of the nuclear ribosomal repeat was extracted from pure cultures obtained from soils, Galleria, and ticks and was amplified and sequenced; and (4) DNA was extracted directly from ticks, amplified, and sequenced. We conclude that (1) ticks encounter potentially entomopathogenic fungi more often in soil than in leaf litter, (2) many species of potentially entomopathogenic fungi found in the soil can readily be cultured, (3) the Galleria bait method is a sufficiently efficient method for isolation of these fungi from soils, and (4) although DNA extraction from ticks was not possible in this study because of small sample size, DNA extraction from fungi isolated from soils and from ticks was successful and provided clean sequences in 100 and 73% of samples, respectively. A combination of the above methods is clearly necessary for optimal characterization of entomopathogenic fungi associated with ticks in the environment.

Blacklegged ticks (Ixodes scapularis) are one of the most important pathogen vectors in the United States, responsible for transmitting Lyme disease and other tick-borne diseases. The structure of a host's microbial community has the potential to affect the ecology and evolution of the host. We employed high-throughput sequencing of the 16S rRNA gene V3-V4 hypervariable regions in the first study to investigate the tick microbiome across all developmental stages (larvae, nymphs, adults). In addition to field-collected life stages, newly hatched lab-reared larvae were studied to determine the baseline microbial community structure and to assess transovarial transmission. We also targeted midguts and salivary glands due to their importance in pathogen maintenance and transmission. Over 100,000 sequences were produced per life stage replicate. Rickettsia was the most abundant bacterial genus across all sample types matching mostly the Ixodes rickettsial endosymbionts, and its proportion decreased as developmental stage progressed, with the exception of adult females that harbored a mean relative abundance of 97.9%. Lab-reared larvae displayed the lowest bacterial diversity, containing almost exclusively Rickettsia. Many of the remaining bacteria included genera associated with soil, water, and plants, suggesting environmental acquisition while off-host. Female organs exhibited significantly different β-diversity than the whole tick from which they were derived. Our results demonstrate clear differences in both α and β diversity among tick developmental stages and between tick organs and the tick as a whole. Furthermore, field-acquired bacteria appear to be very important to the overall internal bacterial community of this tick species, with influence from the host blood meal appearing limited. This article is protected by copyright. All rights reserved.

Background. Blacklegged ticks (Ixodes scapularis) are important disease vectors in the United States, known to transmit a variety of pathogens to humans, including bacteria, protozoa, and viruses. Their importance as a disease vector necessitates reliable and comparable methods for extracting microbial DNA from ticks. Furthermore, to explore the population genetics or genomics of this tick, appropriate DNA extraction techniques are needed for both the vector and its microbes. Although a few studies have investigated different methods of DNA isolation from ticks, they are limited in the number and types of DNA extraction and lack species-specific quantification of DNA yield. Methods. Here we determined the most efficient and consistent method of DNA extraction from two different developmental stages of I. scapularis-nymph and adult-that are the most important for disease transmission. We used various methods of physical disruption of the hard, chitinous exoskeleton, as well as commercial and non-commercial DNA isolation kits. To gauge the effectiveness of these methods, we quantified the DNA yield and confirmed the DNA quality via PCR of both tick and microbial genetic material. Results. DNA extraction using the Thermo GeneJET Genomic DNA Purification Kit resulted in the highest DNA yields and the most consistent PCR amplification when combined with either cutting or bead beating with select matrices across life stages. DNA isolation methods using ammonium hydroxide as well as the MoBio PowerSoil kit also produced strong and successful PCR amplification, but only for females. Discussion. We contrasted a variety of readily available methods of DNA extraction from single individual blacklegged ticks and presented the results through a quantitative and qualitative assessment.

Ticks and whole blood were collected from American black bears (Ursus americanus Pallas) between October 2011 and October 2012 across four counties in northwestern New Jersey, an area where blacklegged ticks (Ixodes scapularis Say) and their associated tick-borne pathogens are prevalent. Adult American dog ticks (Dermacentor variabilis Say) were the most frequently collected tick species in late spring, whereas adult and nymphal blacklegged ticks were found in both the late spring and fall months. Additionally, for blacklegged ticks, we determined the quality of bloodmeals that females acquired from black bears compared with bloodmeals from white-tailed deer (Odocoileus virginianus Zimmerman), the most important host for the adult stage of this tick species. Measures of fecundity after feeding on each host species were not significantly different, suggesting that the bloodmeal a female blacklegged tick acquires from a black bear is of similar quality to that obtained from a white-tailed deer. These results establish the American black bear as both a host and quality bloodmeal source to I. scapularis. Thus, black bears may help support blacklegged tick populations in areas where they are both present. In addition, samples of black bear blood were tested for DNA presence of three tick-borne pathogens. Anaplasma phagocytophilum Foggie and Babesia microti Franca were found in 9.2 and 32.3% of blood samples, respectively. All blood samples were quantitative polymerase chain reaction-negative for Borrelia burgdorferi Johnson, Schmid, Hyde, Steigerwalt, & Brenner. Although circulating pathogens were found in blood, the status of black bears as reservoirs for these pathogens remains unknown.

Background. Blacklegged ticks (Ixodes scapularis) are important disease vectors in the United States, known to transmit a variety of pathogens to humans, including bacteria, protozoa, and viruses. Their importance as a disease vector necessitates reliable and comparable methods for extracting microbial DNA from ticks. Furthermore, to explore the population genetics or genomics of this tick, appropriate DNA extraction techniques are needed for both the vector and its microbes. Although a few studies have investigated different methods of DNA isolation from ticks, they are limited in the number and types of DNA extraction and lack species-specific quantification of DNA yield. Methods. Here we determined the most efficient and consistent method of DNA extraction from two different developmental stages of I. scapularis – nymph and adult - that are the most important for disease transmission. We used various methods of physical disruption of the hard, chitinous exoskeleton, as well as commercial and non-commercial DNA isolation kits. To gauge the effectiveness of these methods we quantified the DNA yield and confirmed the DNA quality via PCR of both tick and microbial genetic material. Results. DNA extraction using the Thermo GeneJET Genomic DNA Purification kit resulted in the highest DNA yields and the strongest, most consistent PCR amplification. We also found that physical disruption of the tick exoskeleton was most effective using cross-sectional cutting compared to any type of bead-beating matrices used. Storing ticks at -80°C resulted in considerably higher DNA yields than those from ticks stored in ethanol. Discussion. We contrasted a variety of readily available methods of DNA extraction from single individual blacklegged ticks and presented the results through a quantitative and qualitative assessment.

An important variable in determining the vectorial capacity of mosquito species for arthropod-borne infections is the degree of contact of the vector and the vertebrate reservoir. This parameter can be estimated by examining the host-feeding habits of vectors. Serological and polymerase chain reaction based methods have been used to study the host-feedings patterns of 21 mosquito species from New York, New Jersey, and Tennessee, 19 of which previously have been found infected with West Nile virus. Mammalophilic mosquito species in New Jersey and New York fed primarily upon white-tailed deer, while those from Memphis, Tennessee, fed mainly upon domestic dogs. A total of 24 different avian host species were detected among the avian-derived blood meals. American Robin, Northern Cardinal, Northern Mockingbird, Tufted Titmouse, and Brown-headed Cowbird were common avian hosts, while blood meals derived from the American Crow were relatively rare. Although the majority of common host species were potentially among the most abundant birds at each location, the proportion of blood meals from the most commonly fed upon avian species was greater than was predicted based upon the likely abundance of these species alone. These findings suggest that vector species for West Nile virus may preferentially feed upon certain avian hosts.

Pathogen prevalence within blacklegged ticks (Ixodes scapularis Say, 1821) tends to vary across sites and geographic regions, but the underlying causes of this variation are not well understood. Efforts to understand the ecology of Lyme disease have led to the proposition that sites with higher host diversity will result in lower disease risk due to an increase in the abundance of inefficient reservoir species relative to the abundance of species that are highly competent reservoirs. Although the Lyme disease transmission cycle is often cited as a model for this "dilution effect hypothesis", little empirical evidence exists to support that claim. Here we tested the dilution effect hypothesis for two pathogens transmitted by the blacklegged tick along an urban-to-rural gradient in the northeastern United States using landscape fragmentation as a proxy for host biodiversity. Percent impervious surface and habitat fragment size around each site were determined to assess the effect of landscape fragmentation on nymphal blacklegged tick infection with Borrelia burgdorferi and Anaplasma phagocytophilum. Our results do not support the dilution effect hypothesis for either pathogen and are in agreement with the few studies to date that have tested this idea using either a landscape proxy or direct measures of host biodiversity.

Mosquitoes and wild birds were collected from three sites near locations in the New York City metropolitan area where single, West Nile (WN) virus-positive dead birds were found early in the 2000 transmission season. The mosquitoes were tested for the presence of infectious virus with a Vero cell culture assay and for WN viral RNA by using reverse transcriptase-polymerase chain reaction (RT-PCR) protocols. Serum samples from wild birds were tested for the presence of neutralizing antibodies against WN virus. Infectious WN virus and WN viral RNA were found in Culex species adult mosquitoes from each of the three sites, and a seropositive hatch-year house sparrow (Passer domesticus) was found in one of the three sites. Molecular techniques used to identify the species in the positive mosquito pools found that most of the pools contained a combination of Culex pipiens and Cx. restuans. The minimum infection rate in Culex species mosquitoes from the sites ranged from 0.2 to 6.0 per 1,000 specimens tested. The results demonstrated that, at least early in the transmission season, detection of a WN virus-positive dead bird indicates a local WN virus transmission cycle. This information is valuable in focusing subsequent surveillance and vector management programs. In addition, the RT-PCR procedure for detecting WN viral RNA in mosquito pools detected more positive pools than did the Vero cell plaque assay.

Understanding the role that nymphal and female ticks, Ixodes scapularis, have in the epidemiology of Lyme disease is essential to the development of successful prevention programs. In this study, the authors sought to evaluate the seasonal and annual relations between tick densities and patientsor = 16 years of age diagnosed with erythema migrans (EM), the rash associated with early Lyme disease. Ticks were collected weekly by drag sampling throughout most of the year from 1991 to 1996 in Westchester County, New York. The number of EM cases was based on patients diagnosed at the Westchester County Medical Center using Centers for Disease Control and Prevention (CDC) criteria. No patients with EM were diagnosed from January through April, when only adult ticks were active. Correlation analysis between monthly tick densities and EM incidence was significant for nymphs (r = 0.87, p0.01), but not for adult ticks (r = -0.57, p0.05). There was a strong, although not significant, correlation between peak annual number of patients with EM and peak nymphal tick abundance (r = 0.76, p = 0.08). These data indicate that bites from adult I. scapularis only rarely result in Lyme disease, and that annual nymphal tick abundance determines exposure. This suggests that annual fluctuations in Lyme disease case numbers are largely due to natural changes in tick abundance and, therefore, that control of nymphal I. scapularis should be a major component of Lyme disease prevention efforts.

Effects of deer exclusion on abundance of immature Ixodes scapularis Say parasitizing small and medium-sized mammals, and the role mammals have in introducing ticks to exclosure areas, were examined at two sites in Westchester County, New York. In total, 686 mammal captures representing nine species were obtained, with white-footed mice, Peromyscus leucopus Rafinesque, composing > 80% of all captures, followed by raccoons, Procyon lotor (L.); opossums, Didelphis virginiana (Kerr); and striped skunks, Mephitis mephitis Schreber. At the Near Archives site, 14% of 81 individual mice, 46% of raccoons, and 33% of opossums captured were found to cross the exclosure fence, as did 12% of 50 mice and 38% of raccoons at the Hudson Pines site. Skunks apparently did not cross the fence at either site. Mice captured exclusively outside the Near Archives exclosure hosted significantly more larvae than mice captured inside only, with fence crossers hosting an intermediate number of larvae. At Hudson Pines, numbers of larvae on fence-crossing mice and those captured solely inside the exclosure were equivalent, with tick loads on mice captured outside the exclosure significantly greater. The number of larvae per raccoon did not differ significantly with capture location (inside, outside, or both sides of exclosure fence) at either site. Densities of host-seeking larvae and nymphs were significantly higher outside the exclosure than inside at Near Archives, though not at Hudson Pines. Differences in tick density inside and outside exclosures declined with each successive developmental stage so that adult density inside exclosures tended to converge with that outside at both sites. Although deer exclosures can have a significant impact on nymphal I. scapularis abundance, they may not reduce the risk of encountering infected adults. Mice, raccoons, and opossums have a role in introducing potentially infective ticks to areas where deer have been excluded, though the level of immigration of ticks into the area will likely depend on the density of ticks outside the exclosure.

Borrelia burgdorferi sensu lato (s.l.), transmitted by Ixodes spp. ticks, is the causative agent of Lyme disease. Although Ixodes spp. ticks are distributed in both Northern and Southern Hemispheres, evidence for the presence of B. burgdorferi s.l. in South America apart from Uruguay is lacking. We now report the presence of culturable spirochetes with flat-wave morphology and borrelial DNA in endemic Ixodes stilesi ticks collected in Chile from environmental vegetation and long-tailed rice rats (Oligoryzomys longicaudatus). Cultured spirochetes and borrelial DNA in ticks were characterized by multilocus sequence typing and by sequencing five other loci (16S and 23S ribosomal genes, 5S-23S intergenic spacer, flaB, ospC). Phylogenetic analysis placed this spirochete as a new genospecies within the Lyme borreliosis group. Its plasmid profile determined by polymerase chain reaction and pulsed-field gel electrophoresis differed from that of B. burgdorferi B31A3. We propose naming this new South American member of the Lyme borreliosis group B. chilensis VA1 in honor of its country of origin.

DNA sequences of the second internal transcribed spacer (ITS2) of the nuclear ribosomal DNA were determined for six species of the genus Salvelinus: S. alpinus (Arctic char), S. malma (Dolly Varden), S. confluentus (bull trout), S. leucomaenis (Japanese char), S. fontinalis (brook trout), and S. namaycush (lake trout), and for Hucho perryi (huchen). The ITS2 sequence data (approximately 375 base pairs (bp)) were combined with previously determined sequence data for the ITS1 (approximately 575 bp), giving a total of 981 bp of aligned sequence for each species. Phylogenetic analysis of the aligned sequences was done using maximum parsimony, maximum likelihood, and distance matrix methods with H. perryi as an outgroup. The results were consistent with previous work based on comparisons of morphologies, allozymes, and karyotypes. Comparison of these results with those based on mitochondrial DNA sequences suggests that hybridization may have occurred between S. confluentus and S. alpinus or S. malma.

A prior study in mice has shown that the timely application of topical antibiotics to the skin at the tick bite site could eradicate Borrelia burgdorferi infection. That study, however, did not evaluate antibiotic preparations that are considered suitable for use in humans. In this murine study, topical application of 2% erythromycin and 3% tetracycline preparations that are acceptable for use in humans was found to be ineffective in eliminating B. burgdorferi from the tick bite site or in preventing dissemination to other tissues. Reasons for the discrepant findings are discussed.

The effect of deer exclosures upon the numbers of immature Ixodes scapularis Say, the vector of Lyme disease in the eastern United States, was examined at five sites in Westchester County, NY. Study areas ranged in size from 6 to 101 ha where deer had been excluded for a period of 25 yr by > 2.4-m-high fencing that surrounded each site. A total area of 40,506 m2 was drag-sampled during the study to measure tick abundance. Nymphal densities (ticks per 1,000 m2) averaged 4.6 (range, 1.3-9.6) inside exclosures and 27.7 (range, 7.3-79.4) outside. Larval densities averaged 36.7 (range, 1.2-132.1) inside exclosures and 354.4 (range, 7.5-914.5) outside. Comparisons between exclosure sites and outside areas immediately adjacent to the exclosure fence, where deer had unrestricted access, revealed that exclosures had 83% fewer host-seeking nymphs and 90% fewer host-seeking larvae. Tick numbers inside exclosures did not always decline with increasing distance from the fence. There was no significant difference in the rate of Borrelia burgdorferi Johnson, Schmid, Hyde, Steigerwalt & Brenner infection for host-seeking ticks collected inside (20%, n = 50) exclosures compared with ticks collected outside (26%, n = 50) exclosures. Deer fencing may provide a means of significantly reducing the abundance of I. scapularis and the risk of Lyme disease in relatively large areas without the need to reduce or eliminate the deer population.

The efficacy of topically treating white-tailed deer with an acaricide was evaluated in a Lyme disease-endemic community of southern New York State. Twenty-four 4-Poster feeders were placed in a 5.2 km(2) treatment area in Bedford, NY, while a site in Lewisboro, NY, 4.8 km distant, served as control. Treatment periods ran from 15 September to 15 December each fall from 1997 to 2001, and from 15 March to 15 May each spring from 1998 to 2002. Corn consumption averaged 15,779 kg in fall sessions and 9054 kg in spring sessions, and a mean of 89.6% of deer in the study area showed evidence of using the feeders. Deer densities, estimated by aerial snow counts, averaged 22 and 28 deer per km(2) in Bedford and Lewisboro, respectively, over a 3-year period. Significant reductions in tick numbers on deer captured in the treatment area were noted in fall 1999 compared to deer captured at the control site. Drag sampling for nymphal host-seeking ticks indicated 63.6% control in 2001, which dropped to 54.8% the following year, but reached 80% in 2003. Higher-than-normal acorn production in 2001 that likely caused a drop in deer visitation to the feeders may have reduced efficacy against larval ticks in 2002. The 4-Poster effectively reduced the density of Ixodes scapularis, though the level of control is dependent on environmental factors that affect feeding behavior of white-tailed deer.

We evaluated the effects of tick control by acaricide self-treatment of white-tailed deer on the infection prevalence and entomologic risk for three Ixodes scapularis-borne bacteria in host-seeking ticks. Ticks were collected from vegetation in areas treated with the "4-Poster" device and from control areas over a 6-year period in five geographically diverse study locations in the Northeastern United States and tested for infection with two known agents of human disease, Borrelia burgdorferi and Anaplasma phagocytophilum, and for a novel relapsing fever-group spirochete related to Borrelia miyamotoi. Overall, 38.2% of adults and 12.5% of nymphs were infected with B. burgdorferi; 8.5% of adults and 4.2% of nymphs were infected with A. phagocytophilum; and 1.9% of adults and 0.8% of nymphs were infected with B. miyamotoi. In most cases, treatment with the 4-Poster device was not associated with changes in the prevalence of infection with any of these three microorganisms among nymphal or adult ticks. However, the density of nymphs infected with B. burgdorferi, and consequently the entomologic risk for Lyme disease, was reduced overall by 68% in treated areas compared to control areas among the five study sites at the end of the study. The frequency of bacterial coinfections in ticks was generally equal to the product of the proportion of ticks infected with a single bacterium, indicating that enzootic maintenance of these pathogens is independent. We conclude that controlling ticks on deer by self-application of acaricide results in an overall decrease in the human risk for exposure to these three bacterial agents, which is due solely to a reduction in tick density.

From 1997 to 2002, the U.S. Department of Agriculture's Northeast Area-wide Tick Control Project used acaricide-treated 4-Poster Deer Treatment Bait Stations in five eastern states to control ticks feeding on white-tailed deer. The objectives of this host-targeted technology were to reduce free-living blacklegged (Ixodes scapularis Say) and lone star (Amblyomma americanum [L.]) tick populations and thereby to reduce the risk of tick-borne disease. During 2002 to 2004, treatments were suspended, and tick population recovery rates were assayed. Subsequently, the major factors that influenced variations in efficacy were extrapolated to better understand and improve this technology. Treatments resulted in significant reductions in free-living populations of nymphal blacklegged ticks at six of the seven sites, and lone star ticks were significantly reduced at all three sites where they were present. During the study, maximal significant (p < or = 0.05) efficacies against nymphal blacklegged and lone star ticks at individual sites ranged from 60.0 to 81.7 and 90.9 to 99.5%, respectively. The major environmental factor that reduced efficacy was the occurrence of heavy acorn masts, which provided an alternative food resource for deer. Although the 4-Poster technology requires 1 or more years to show efficacy, this host-targeted intervention was demonstrated to be an efficacious, economical, safe, and environment-friendly alternative to area-wide spraying of acaricide to control free-living populations of these tick species.

The ability of an animal to discriminate between kin and non-kin (kin recognition) has been the subject of numerous recent investigations. Grafen ( Anim. Behav. , 1990, 39 , 42–54) recently reported that the evidence in support of kin recognition is weak and the data illustrating a preference for kin to associate in the laboratory may be more consistently explained as species recognition. It is suggested here, however, that in many cases it may be impossible to distinguish between species recognition and kin recognition, but in some cases, kin recognition seems apparent. It is also emphasized that very little is known about the adaptive value of kin recognition.

Publisher Summary A list of health challenges for the twenty-first century included sanitation and hygiene, vaccination, antibiotics and other antimicrobials, technological advances in detecting and monitoring diseases, serologic testing, viral isolation and tissue culture, and molecular techniques that can help diagnose and track the transmission of new threats. The proliferation of suburban development and fragmented landscapes in the northeastern and midwestern US has generated an ideal habitat for people emigrating from the cities, for commensal animal species such as deer that proliferate in the presence of humans, and for the parasites those animals host. This mixture of large numbers of vectors and people brings with it an increased potential for diseases such as Lyme disease, babesiosis, and human granulocytic anaplasmosis. Cases of Lyme disease are on the rise throughout the nation, notably from regions that have experienced similar alterations of landscape, have rapidly growing deer herds, and are becoming increasingly suburbanized. Much of this progress comes as a result of the interest in global warming, a critically important phenomenon that cannot be ignored any longer. Climate constrains the range of many infectious diseases, and weather affects the timing and intensity of outbreaks.

The spatial distribution of larval Ixodes dammini Spielman, Clifford, Piesman, & Corwin was examined at two sites in Westchester County, N.Y., an area endemic for Lyme disease. Larval abundance peaked in early August and was marked by a highly clumped distribution reflecting high concentrations of newly hatched larvae. Subsequent sampling indicated an increasingly uniform distribution of larvae as measured by the variance/mean ratio and Lloyd's index of patchiness. An experiment using ticks marked with fluorescent powder to illustrate the degree of active dispersal (tick disperses by its own movement) indicated little lateral movement; only 3 (4%) of the 77 marked larvae recovered during the experiment were found beyond 3 m from the release point. The recovery of unfed larvae (12 larvae [0.6% of total] were recovered from raccoons, Procyon lotor (L.); 9 [3.4%] from mice, Peromyscus leucopus Rafinesque; and 32 [9.0%] from opossums, Didelphis virginiana (Kerr)) from captive hosts in the laboratory suggests that passive transport of unfed larvae may be a mechanism of dispersal.

Summary Ixodes scapularis ticks play an important role in the transmission of a wide variety of pathogens between various mammalian species, including humans. Pathogens transmitted by ticks include Borrelia, Anaplasma and Babesia. Although ticks may harbour both pathogenic and non-pathogenic microflora, little is known about how the diversity of the microflora within ticks may influence the transmission of pathogens. To begin addressing this question, we examined the composition of bacterial communities present in Ixodes scapularis collected from Westchester and Dutchess Counties, New York State, at different developmental and nutritional stages. Genetic fingerprints of bacterial populations were generated by temporal temperature gradient gel electrophoresis (TTGE) separation of individual polymerase chain reaction (PCR)-amplified 16S rRNA gene fragments, followed by DNA sequence analysis for bacterial identification. The fingerprints of the TTGE bands were grouped into five clusters. The most abundant DNA sequence found in all the samples was Rickettsia, followed by Pseudomonas and Borrelia. Ralstonia, Anaplasma, Enterobacterias, Moraxella, Rhodococcus and uncultured proteobacterium were present as well. We also determined the prevalence of Anaplasma phagocytophilum and Borrelia burgdorferi by PCR and DNA sequence analysis. Statistical analyses indicated significant variations in the bacterial communities depending on tick developmental stage and degree of engorgement. We suggest that these two elements affect microbial diversity within the tick and may in turn influence pathogen transmission to humans and animals after tick bite.

Rodent trapping and drag sampling in Van Cortlandt Park, New York City, yielded all stages of Ixodes scapularis, the deer tick vector of Lyme disease and human granulocytic ehrlichiosis (HGE). Polymerase chain reaction analyses of the ticks showed Borrelia burgdorferi and the Ehrlichia sp. that causes HGE.

The stringent response is a global bacterial response to nutritional stress mediated by (p)ppGpp. We previously found that both noninfectious Borrelia burgdorferi strain B31 and infectious B. burgdorferi strain N40 produced large amounts of (p)ppGpp during growth in BSK-H medium and suggested that the stringent response was triggered in B. burgdorferi under these conditions. Here we report that (p)ppGpp levels in B. burgdorferi growing in BSK-II or BSK-H medium are not further increased by nutrient limitation or by serine hydroxamate-induced inhibition of protein synthesis and that the presence of (p)ppGpp during growth of N40 in BSK-H medium is not associated with decreased 16S rRNA synthesis. Decreased 16S rRNA synthesis was associated with the decreased growth rate of N40 seen during coculture with tick cells, which are growth conditions that were previously shown to decrease (p)ppGpp levels. One-half as much of the mRNA of the gene encoding the Rel protein of B. burgdorferi ( rel Bbu ) was produced by B31 as by N40 during in vitro growth (2 ± 0.5 and 4 ± 0.8 fg of rel Bbu mRNA/ng of total Borrelia RNA, respectively). Although the amounts of N40 rel Bbu mRNA were identical during growth in vitro and in rat peritoneal chambers, they were markedly decreased during growth in nymphal ticks. In contrast to the lack of change in rel Bbu mRNA levels, larger amounts of a 78-kDa protein that was cross-reactive with antibodies to Bacillus subtilis Rel Bsu were detected in immunoblots of N40 lysates after growth in rat peritoneal chambers than after growth in vitro. Differences in the level of production of (p)ppGpp between B31 and N40 could not be explained by differences in rel Bbu promoters since identical transcriptional start sites 309 nucleotides upstream from the B31 and N40 rel Bbu ATG start codon and identical σ 70 -like promoters were identified by primer extension and sequencing analysis. rel Bbu complemented an Escherichia coli CF1693 relA spoT double mutant for growth on M9 minimal medium, and the transformed cells produced rel Bbu mRNA. These results indicate that rel Bbu is functional and that its transcription and translation and production of (p)ppGpp are affected by environmental conditions in strains N40 and B31. They also suggest that in B. burgdorferi , an organism with few rRNA operons that grows slowly, the role of (p)ppGpp may differ from the classic role played by this molecule in E. coli and that (p)ppGpp may not be responsible for growth rate control.

The seasonal occurrence and abundance of a newly introduced mosquito in the United States, Ochlerotatus (Finlaya) japonicus (Theobald), are reported for Westchester and Putnam Counties in southern New York State. Adult mosquitoes were sampled at 39 sites distributed throughout the two counties. CDC (Centers forDisease Control and Prevention) light traps (2,107 trap nights) and gravid traps (1,813 trap nights) were used to collect a total of 44,428 mosquitoes from May through October 2000. Oc. japonicus was found at 97.4% of sites sampled and accounted for 9.8% of all specimens collected. Oc. japonicus was recovered from 1.5% (n = 326) of the light trap collections and 18.1% (n = 4,026) of the gravid trap collections. Although gravid traps collected significantly more specimens than light traps, the seasonal activity patterns measured by each trap type were congruent. In all, 30 mosquito species were collected. Unlike other Ochlerotatus or Aedes species, Oc. japonicus was collected throughout the study, indicating a broad seasonal activity period. There was significant regional variation in Oc. japonicus abundance, with higher trap densities occurring in the northern-most trapping sites. This study demonstrates that Oc. Japonicus is established in southern New York State.

Ixodes scapularis , the tick vector of Lyme disease and human granulocytic ehrlichiosis (HGE), is prevalent in much of southern New York state. The distribution of this species has increased, as have reported cases of both Lyme disease and HGE. The unreliability of case reports, however, demonstrates the need for tick and pathogen surveillance in order to accurately define areas of high risk. In this study, a total of 89,550 m 2 at 34 study sites was drag sampled in 1995 and a total of 51,540 m 2 at 40 sites was sampled in 1996 to determine tick and pathogen distribution in southern New York state. I. scapularis was collected from 90% of the sites sampled, and regionally, a 2.5-fold increase in nymphal abundance occurred from 1995 to 1996. I. scapularis individuals from all sites were infected with Borrelia burgdorferi in 1995, while an examination of ticks for both B. burgdorferi and the agent of HGE in 1996 confirmed that these organisms were present in all counties; the average coinfection rate was 1.9%. No correlation was found between estimated risk and reported cases of Lyme disease. The geographic disparity of risk observed among sites in this study underscores the need for vector and pathogen surveillance on a regional level. An entomologic risk index can help identify sites for targeted tick control efforts.

Timing of oviposition and larval eclosion for fall- and spring-fed female blacklegged tick, Ixodes scapularis Say, and effect of temperature on those events in southern New York state, where Lyme disease is endemic, were determined in the field. We also examined seasonal pattern of larval host-seeking activity, as well as rate of survival for overwintering, unfed larvae. Totals of 9 and 8 replete female I. scapularis were released in fall 1988 and spring 1989, respectively. For the 10 females that oviposited, egg laying began in mid-to-late May, regardless of when ticks had fed. Likewise, larval eclosion was synchronized in July for all egg masses. Microhabitat temperatures experienced by females were not reliable indicators of female reproductive success, in terms of either presence or absence of eggs, or numbers of larvae recovered. However, average body size was significantly greater for females that oviposited successfully than for those that did not. The percentage of unfed larvae surviving the 8-mo overwintering period from late August to early May ranged from 1.9 to 31.4, with a mean survival rate of 10.4%. Concurrent drag sampling through the year indicated a bimodal pattern of larval abundance marked by a relatively small early peak of activity in late May that extended into early July. Because the first peak of larval activity occurred during the period of oviposition, 8 wk before onset of larval eclosion, it appears that bimodality represents activity of 2 consecutive cohorts, with the early peak composed of overwintering survivors. These data imply variation in timing of reproduction and larval activity from 1 geographic area to another. Efforts to devise a comprehensive scheme for the life cycle of I. scapularis must consider that population differences may exist.

Changes in the abundance of immature Ixodes scapularis Say, the primary vector of Lyme disease, were assessed over a 7-yr period in Westchester County, New York, where the disease is endemic. In total, 4,000 m2 were drag sampled at each of 6 study sites in June and July of 1984 and 1991. The abundance of nymphs collected from all sites was 2.6-fold greater in 1991 than in 1984. Four of the 6 sites had significantly more nymphs in 1991, with increases ranging from 1.1- to 46.5-fold. The abundance of larvae collected from all sites was 34.2-fold higher in 1991 than in 1984. Five of the sites sampled had significantly more larvae in 1991, with increases ranging from 2.7- to 681-fold. These data demonstrate that populations of immature I. scapularis in Westchester County have, overall, increased during the 7-yr period from 1984 to 1991. However, the magnitude of increase is not uniform from site to site.

To the Editor: Human granulocytic ehrlichiosis (HGE), first described in 1994 (1), is the second-most common tick-borne disease in the United States; Lyme disease is the most prevalent. Both diseases are transmitted by blacklegged ticks (Ixodes scapularis Say) that are abundant in southern New York state (2). Factors that influence the risk for infection, particularly the role of wildlife in transmitting the etiologic agent, Anaplasma (formerly Ehrlichia) phagocytophilum (3,4), to vector ticks are not well understood. In the absence of transovarial transmission (5,6), acquisition of A. phagocytophilum by its vector must result either from feeding on the blood of reservoir-competent hosts or by cofeeding of uninfected ticks in close proximity to infected ticks (7).

Surveillance programs that identify areas where both the vector (Ixodes dammini) and etiologic agent (Borrelia burgdorferi) are present may identify the risk of Lyme disease and its spread earlier and more accurately than do programs relying on any single method, particularly human case reports. Hunter-killed deer ( n = 1,204) from 22 counties in Maryland, Massachusetts, New Hampshire, New York, and Pennsylvania were examined in fall 1989 and all ectoparasites were identified. The following spring, canine sera ( n = 884) were obtained from these sites, which included known endemic areas and those where Lyme disease is uncommon, and tested by enzyme-linked immunosorbent assay. Regression analysis of canine seroprevalence versus tick prevalence on deer from the same counties demonstrate a significant positive linear relationship. Sites were designated as low-, moderate-, or high-risk counties based upon their position on the regression curve. The geographic distribution of the sites correlated well with the distribution of known Lyme disease endemic and nonendemic areas. Locations were also identified where Lyme disease may be emerging. The positive relationship between measures of vector and pathogen abundance determined in this study permits public health workers to identify endemic and potentially endemic areas independently of human case reports.

A 2-yr evaluation of a commercial product designed to reduce the risk of Lyme disease by delivering permethrin-treated cotton to white-footed mice (Peromyscus leucopus) was conducted at three sites in Westchester County, N.Y., an area where Lyme disease is endemic. We examined the numbers of host-seeking nymphal Ixodes dammini Spielman, Clifford, Piesman & Corwin, the numbers of larval I. dammini infesting mice, and the proportion of nymphs infected with Borrelia burgdorferi, the causative agent of Lyme disease. The density of nymphs collected by drag sampling did not differ between treatment and control areas in the first year for either a woodland or recreational site. In the second year, no significant decrease in the density of nymphs was observed in the treatment areas compared with the control areas. Numbers of larvae on 67 mice captured on treatment and control grids did not differ for woodland and recreational sites, but significantly fewer larvae were found on mice captured at the residential treatment site. The percentage of host-seeking ticks infected with spirochetes did not differ between treatment and control sites in any habitat for either year. The use of permethrin-treated cotton did not measurably reduce the numbers of I. dammini or the proportion of ticks infected with spirochetes at our study sites.

To the Editor: Human granulocytic ehrlichiosis, a newly discovered tick-borne infection originally described in the upper Midwest,1 has recently been reported in 29 patients from Westchester County, New York.2 We used the polymerase chain reaction (PCR) to examine 173 host-seeking ticks (Ixodes scapularis) collected from vegetation in Westchester County for the presence of the agent of human granulocytic ehrlichiosis, as well as the agent of Lyme disease, Borrelia burgdorferi. After extracting DNA from dissected tissues, we used the 23S ribosomal DNA (rDNA) primers of Schwartz et al.3 for B. burgdorferi and the 16S rDNA primers of Pancholi . . .