Refractoriness of the western fence lizard (Sceloporus occidentalis) to the Lyme disease group spirochete Borrelia bissettii

The western fence lizard, Sceloporus occidentalis, is refractory to experimental infection with Borrelia burgdorferi sensu stricto, one of several Lyme disease spirochetes pathogenic for humans. Another member of the Lyme disease spirochete complex, Borrelia bissettii, is distributed widely throughout North America and a similar, if not identical, spirochete has been implicated as a human pathogen in southern Europe. To determine the susceptibility of S. occidentalis to B. bissettii, 6 na?ve lizards were exposed to the feeding activities of Ixodes pacificus nymphs experimentally infected with this spirochete. None of the lizards developed spirochetemias detectable by polymerase chain reaction for up to 8 wk post-tick feeding, infected nymphs apparently lost their B. bissettii infections within 1-2 wk after engorgement, and xenodiagnostic L. pacificus larvae that co-fed alongside infected nymphs did not acquire and maintain spirochetes. In contrast, 3 of 4 na?ve deer mice (Peromyscus maniculatus) exposed similarly to feeding by 1 or more B. bissettii-infected nymphs developed patent infections within 4 wk. These and previous findings suggest that the complement system of S. occidentalis typically destroys B. burgdorferi sensu lato spirochetes present in tissues of attached and feeding I. pacificus nymphs, thereby potentially reducing the probability of transmission of these bacteria to humans or other animals by the resultant adult ticks.     

Impact of the experimental removal of lizards on Lyme disease risk

The distribution of vector meals in the host community is an important element of understanding and predicting vector-borne disease risk. Lizards (such as the western fence lizard; Sceloporus occidentalis) play a unique role in Lyme disease ecology in the far-western United States. Lizards rather than mammals serve as the blood meal hosts for a large fraction of larval and nymphal western black-legged ticks (Ixodes pacificus--the vector for Lyme disease in that region) but are not competent reservoirs for the pathogen, Borrelia burgdorferi. Prior studies have suggested that the net effect of lizards is to reduce risk of human exposure to Lyme disease, a hypothesis that we tested experimentally. Following experimental removal of lizards, we documented incomplete host switching by larval ticks (5.19%) from lizards to other hosts. Larval tick burdens increased on woodrats, a competent reservoir, but not on deer mice, a less competent pathogen reservoir. However, most larvae failed to find an alternate host. This resulted in significantly lower densities of nymphal ticks the following year. Unexpectedly, the removal of reservoir-incompetent lizards did not cause an increase in nymphal tick infection prevalence. The net result of lizard removal was a decrease in the density of infected nymphal ticks, and therefore a decreased risk to humans of Lyme disease. Our results indicate that an incompetent reservoir for a pathogen may, in fact, increase disease risk through the maintenance of higher vector density and therefore, higher density of infected vectors.     

Spirochetes in mammals and ticks (Acari: Ixodidae) from a focus of Lyme borreliosis in California

In northern California, antibodies to Borrelia burgdorferi were detected in 58 of 73 (79%), and spirochetemias in one of 26 (4%) black-tailed jackrabbits (Lepus californicus californicus), by indirect and direct immunofluorescence, respectively. Five species of ticks (Dermacentor occidentalis, D. parumapertus, Ixodes neotomae, I. pacificus, and Haemaphysalis leporispalustris) were collected from rabbits. Two of these species of ticks were found to contain spirochetes; two of 10 (20%) I. neotomae and two of 174 (1%) H. leporispalustris. A strain of B. burgdorferi was recovered from I. neotomae. One infected H. leporispalustris female passed spirochetes via eggs to about 67% of her progeny. The widespread distribution of the black-tailed jackrabbit, its infestation by at least four ticks (D. occidentalis, D. parumapertus, I. neotomae, and I. pacificus) known to be infected naturally with B. burgdorferi, and the high prevalence of spirochetal antibody in this lagomorph suggest that it might be useful as a sentinel for surveillance of Lyme borreliosis. Spirochetes were detected in 15% of 40 Columbian black-tailed deer (Odocoileus hemionus columbianus) by direct immunofluorescence bound with a Borrelia-specific monoclonal antibody (H9724), but not with a monoclonal antibody (H5332) specific for B. burgdorferi. The geographical overlap of different borreliae in ticks that bite wildlife such as deer may confound spirochetal serosurveys, and underscores the need for more specific serologic tests than those currently available.     

Abundance of Ticks (Acari: Ixodidae) Infesting the Western Fence Lizard, Sceloporus Occidentalis, in Relation to Environmental Factors

We examined the impact of environmental characteristics, such as habitat type, topographic exposure and presence of leaf litter, on the abundance of Ixodes pacificus ticks infesting the western fence lizard (Sceloporus occidentalis) at the University of California Hopland Research and Extension Center (HREC), Mendocino County, California. A total of 383 adult lizards were slip-noosed and examined for tick infestation in April and May 1998. At least 94% of the lizards were infested by ticks and at least 20% of the females and 33% of the males carried > 15 ticks. This intensive utilization of western fence lizards (which do not serve as natural reservoirs for Lyme disease spirochetes) by subadult ticks, is probably the primary reason for the low prevalence of infection with Borrelia burgdorferi in I. pacificus nymphs and adults previously recorded at the HREC. Tick loads were higher on male than female lizards. Also, male lizards were generally more heavily infested in late April than in late May. The prevalence of tick infestation exceeded 88% in all habitat types but males collected in woodland and grass/woodland edges had higher tick loads than those collected in open grassland. Male lizards captured in open, exposed grassland tended to carry heavier tick loads in northern/eastern, as compared to southern/western, exposures, and when leaf litter was present.     

Prevalence and abundance of Ixodes pacificus immatures (Acari: Ixodidae) infesting western fence lizards (Sceloporus occidentalis) in northern California: temporal trends and environmental correlates.

The prevalence and abundance of immature Ixodes pacificus ticks on western fence lizards (Sceloporus occidentalis) were examined in relation to time of year, host attributes (i.e., age, gender, and presence or absence of blood parasites), and 5 environmental characteristics, including topographic exposure and ground cover substrate, over a 2-year period in northern California. Lizards were infested with subadult ticks from early March until late July or early August, with peak median numbers of larvae and nymphs recorded in late April and early May of both years. Peak larval and nymphal abundances differed between years. The overall ratio of larvae to nymphs on adult male lizards was low, ranging from 0.80 in 1999 to 2.41 in 2000. Such intensive feeding of nymphs versus larvae on these lizards, which are reservoir-incompetent for Borrelia burgdorferi spirochetes, may explain previous observations of decreasing spirochetal infection prevalence from the nymphal to adult stage in northwestern California. Adult male lizards were more likely to be infested with nymphs and harbored greater abundances of larvae and nymphs than adult females. Lizards uninfected with blood parasites had more nymphs than infected lizards. The measured environmental characteristics could explain only a small percentage of the total variation observed in larval prevalence (22%) and in larval and nymphal abundance (12 and 3%, respectively).     

Columbian black-tailed deer (Odocoileus hemionus columbianus) as hosts for Borrelia spp. in northern California

The prevalence of infection of Columbian black-tailed deer (Odocoileus hemionus columbianus) with Borrelia spp. was evaluated in an area of northwestern California (USA) where Lyme disease is endemic and the relapsing-fever group spirochete Borrelia coriaceae is enzootic, and in a far-removed comparison area having a disparate climate and lower density of vector ticks. Blood samples collected from both deer herds in 1987, 1988, and from 2000-02 were assayed for borrelial infection with microscopic and molecular methods. Serum specimens from two (5%) of 39 deer from the Dye Creek Preserve in Tehama County versus 13 (20%) of 64 animals from the Hopland Research and Extension Center (HREC) in Mendocino County, California were polymerase chain reaction (PCR) test positive for B. burgdorferi sensu lato. DNA sequencing analyses revealed that eight animals were infected with B. bissettii, six with three unclassified genotypes, and one with B. burgdorferi sensu stricto. One serum sample (2%) from HREC was positive for a relapsing-fever group spirochete that had a 16S rRNA sequence homology of 99% with the C053 type strain of B. coriaceae. Spirochetes undetermined to geno-species were detected in thick-blood drops prepared from three (8%) of 36 deer from the HREC by direct immunofluorescence. Adults of the hippoboscid flies Lipoptena depressa (n=73) and Neolipoptena ferrisi (n=24), the Pacific Coast tick (Dermacentor occidentalis) (n=22), and the western black-legged tick (Ixodes pacificus) (n=1) that had been removed from deer from both study areas in 2002 were PCR test negative for borreliae. The occurrence of diverse borreliae in deer from northern California confounds and, consequently, reduces the utility of borrelial serosurveys for detecting specific genospecies, unless they are complemented by more specific assays (e.g., immunoblotting, PCR/sequencing analysis).     

The roles of birds, lizards, and rodents as hosts for the western black-legged tick Ixodes pacificus.

We compared the infestation by ixodid ticks of lizards, rodents, and birds collected simultaneously within areas representing common habitat types in Mendocino County, CA. Lizards were infested only by Ixodes pacificus Cooley and Kohls, birds by I. pacificus and Haemaphysalis leporispalustris (Packard), and rodents by I. pacificus, I. spinipalpis Hadwen and Nuttall, I. woodi Bishopp, Dermacentor occidentalis Marx, and D. variabilis (Say). Infestation by I. pacificus larvae and nymphs of lizards (Sceloporus occidentalis Baird and Girard; Elgaria spp.) and western gray squirrels (Sciurus griseus Ord) (means of 9-35 larvae and 5-6 nymphs per animal) was several times greater than for Neotoma fuscipes Baird woodrats, Peromyscus spp. mice, and birds (means of 0.9-3.5 larvae and 0-0.3 nymphs). Overall, Borrelia-refractory lizards accounted for 84% of I. pacificus larvae and 91% of nymphs collected from animals in dense woodlands. Bird species frequently utilizing tick-questing substrates such as leaf litter (guild I birds) were more heavily infested by I. pacificus subadults (5.2 larvae and 1.0 nymphs per bird) than guild IV birds with minimal perceived contact with tick-questing substrates (0.08 larvae and 0.06 nymphs per bird). Notably, guild I birds carried similar larval loads and at least 20-fold higher nymphal loads relative to woodrats and mice. Only guild IV birds carried as few I. pacificus nymphs as did these rodents. The ratios of larvae to nymphs suggest that, relative to birds, lizards, and squirrels (infested by 1.3-6.0 larvae per nymph), nocturnally active ground-dwelling rodents such as woodrats and mice are underutilized by the nymphal stage (69 to >100 larvae per nymph). The western gray squirrel and guild I-II birds (e.g., the dark-eyed junco, Junco hyemalis [L.]) were the only potential reservoirs of Borrelia burgdorferi Johnson, Schmid, Hyde, Steigerwalt, and Brenner (the causative agent of Lyme disease in North America) that were frequently infested with both I. pacificus larvae and nymphs and commonly utilized dense woodland habitats.     

Feeding of Ticks on Animals for Transmission and Xenodiagnosis in Lyme Disease Research

Transmission of the etiologic agent of Lyme disease, Borrelia burgdorferi, occurs by the attachment and blood feeding of Ixodes species ticks on mammalian hosts. In nature, this zoonotic bacterial pathogen may use a variety of reservoir hosts, but the white-footed mouse (Peromyscus leucopus) is the primary reservoir for larval and nymphal ticks in North America. Humans are incidental hosts most frequently infected with B. burgdorferi by the bite of ticks in the nymphal stage. B. burgdorferi adapts to its hosts throughout the enzootic cycle, so the ability to explore the functions of these spirochetes and their effects on mammalian hosts requires the use of tick feeding. In addition, the technique of xenodiagnosis (using the natural vector for detection and recovery of an infectious agent) has been useful in studies of cryptic infection. In order to obtain nymphal ticks that harbor B. burgdorferi, ticks are fed live spirochetes in culture through capillary tubes. Two animal models, mice and nonhuman primates, are most commonly used for Lyme disease studies involving tick feeding. We demonstrate the methods by which these ticks can be fed upon, and recovered from animals for either infection or xenodiagnosis.     

Habitat-related variation in infestation of lizards and rodents with Ixodes ticks in dense woodlands in Mendocino County, California

During the spring and early summer of 2002, we examined the relative importance of Borrelia-refractory lizards (Sceloporus occidentalis, Elgaria spp.) versus potential Borrelia burgdorferi sensu lato (s.l.)-reservoirs (rodents) as hosts for Ixodes pacificus immatures in 14 woodland areas (six oak, five mixed oak/Douglas fir, and three redwood/tanoak areas) distributed throughout Mendocino County, California. Lizards were estimated to serve as hosts for 93-98% of all larvae and > or =99.6% of all nymphs infesting lizards or rodents in oak woodlands and oak/Douglas fir sites in the southern part of the county. In redwood/tanoak woodlands and oak/Douglas fir sites in northern Mendocino County, the contribution of rodents to larval feedings reached 36-69% but lizards still accounted for 94-100% of nymphal bloodmeals. From late April to mid-June, I. pacificus larvae were recovered from 95 to 96% of lizards and dusky-footed woodrats (Neotoma fuscipes) and from 59% of Peromyscus spp. mice. In contrast, 99% of lizards but few woodrats (15%) and none of the mice were infested by nymphs. Comparisons of tick loads for 19 lizard-Peromyscus spp. mouse pairings, where the lizard and mouse were captured within 10m of each other, revealed that the lizards harbored 36 times more larvae and >190 times more nymphs than the mice. In oak woodlands, loads of I. pacificus larvae decreased from late April/early May to late June for S. occidentalis lizards but increased for Peromyscus spp. mice. We conclude that the relative utilization of Borrelia-refractory lizards, as compared to rodents, by I. pacificus larvae was far higher in dry oak woodlands than in moister habitats such as redwood/tanoak and oak/Douglas fir woodlands in northern Mendocino County. Non-lizard-infesting potential enzootic vectors of B. burgdorferi s.l. (I. angustus and I. spinipalpis) were recorded from rodents in three of six oak woodland areas, two of five oak/Douglas fir woodland areas, and two of three redwood/tanoak woodland areas.     

Birds and their ticks in northwestern California: minimal contribution to Borrelia burgdorferi enzootiology

Birds and their attendant ticks were surveyed for infection with the Lyme disease spirochete Borrelia burgdorferi, in chaparral and woodland-grass habitats in northwestern California from March to July, 1998 to 1999. In total, 234 birds were captured and recaptured (15%); nearly 2.5 times more birds were captured in chaparral than in woodland-grass. Overall, 34 species representing 15 families were collected during this study; of these, 24 species were caught in chaparral, 19 in woodland-grass, and 9 in both vegetational types. The most frequently captured birds were sage sparrows (Amphispiza belli) in chaparral, and American robins (Turdus migratorius) and oak titmice (Baelophus inornatus) in woodland-grass. Birds hosted 35 Ixodes pacificus (15 larvae, 20 nymphs) and 9 Haemaphysalis leporispalustris (3 larvae, 5 nymphs, 1 adult) ticks, of which 32 were removed from chaparral birds and 12 from woodland birds. The prevalence of tick infestation was 13% (21/167) in chaparral and 5% (3/67) in woodland-grass, but the relative and mean tick intensities of 0.19 and 1.5 for chaparral birds, and 0.18 and 4.0 for woodland birds, respectively, did not differ significantly by habitat. Spirochetes were not detected in either bird-blood or tick-tissue samples when tested by culture, immunofluorescence, or Giemsa-staining. In contrast, over 90% (86/94) of western fence lizards (Sceloporus occidentalis) collected in June or July were infested with an average of 6.9 and 8.9 immature I. pacificus in chaparral and woodland-grass, respectively. We conclude that birds contribute little to the enzootiology of B. burgdorferi in chaparral and woodland-grass habitats in northwestern California because of their limited parasitism by tick vectors and lack of detectable spirochetemias.     

Description and epidemiology of Theileria youngi n. sp. from a northern Californian dusky-footed woodrat (Neotoma Fuscipes) population

An epidemiologic study designed to identify the small mammal reservoir for the zoonotic WA1-type babesial parasite resulted in the discovery of a small, intraerythrocytic piroplasm in smeared blood from dusky-footed woodrats (Neotoma fuscipes) in northern California. The woodrat parasites were isolated and compared to other piroplasm parasites based on their morphology, antigenicity, and genetic characteristics. These studies indicated that the woodrat parasites were not the WA1-type babesial agent but were of the genus Theileria. We accordingly named it Theileria youngi. The prevalence in the woodrat population was high (61%). Infection was unrelated to gender or age of the woodrats. Potential vectors for this tick-transmitted parasite included 3 species of ticks recovered from the woodrats. Dermacentor occidentalis, Ixodes woodi, and Ixodes pacificus. Mostly larval or nymphal stages were recovered, suggesting transstadial transmission is possible. This is the first piroplasm fully characterized from a dusky-footed woodrat.     

Wild turkey (Meleagris gallopavo) as a host of ixodid ticks, lice, and Lyme disease spirochetes (Borrelia burgdorferi sensu lato) in California state parks

Rio Grande wild turkeys (Meleagris gallopavo intermedia) were evaluated as potential hosts of ixodid ticks, lice, and Lyme disease spirochetes (Borrelia burgdorferi sensu lato [s.l.]) in three state parks in Sonoma County, California, USA, during 2003 and 2004. In total, 113 birds were collected, 50 (44.2%) of which were found to be infested by 361 ixodid ticks representing three species: the western black-legged tick (Ixodes pacificus, n=248), the rabbit tick (Haemaphysalis leporispalustris, n=112), and one American dog tick (Dermacentor variabilis). Year-round the prevalence of all ticks combined was unrelated to the age or sex of turkeys, and the prevalence of infestation by I. pacificus (35.4%) was significantly higher than it was for either H. leporispalustris (14.2%) or D. variabilis (0.9%). The proportion of the two prevalent tick species differed significantly by life stage with 86.3% of the I. pacificus and 82.1% of the H. leporispalustris enumerated being nymphs and larvae, respectively. Three species of lice were collected, including the chicken body louse Menacanthus stramineus (12.5% of total), Chelopistes meleagridis (37.5% of total), and Oxylipeurus polytrapezius (50% of total). The records for all three ticks are the first ever from wild turkeys, and those for the lice are the first from this host in the far-western United States. Wild turkeys potentially were exposed to the feeding activities of I. pacificus nymphs infected with B. burgdorferi s.l. as 15% of host-seeking nymphs (n=200) collected in woodlands used by turkeys as roosting or foraging areas were infected mainly with B. burgdorferi sensu stricto (s.s.). However, only one (1%) of 90 turkey blood specimens tested by PCR contained B. burgdorferi s.s., and four in vitro, complement-protein assays demonstrated that domestic turkey serum is moderately bacteriolytic for this spirochete. Taken together, these findings indicate that wild turkeys are important avian hosts of I. pacificus nymphs, but they appear to be inconsequential hosts of B. burgdorferi s.l.     

Peromyscus maniculatus, a possible reservoir host of Borrelia garinii from the Gannet Islands off Newfoundland and Labrador

Thirty-five deer mice, Peromyscus maniculatus, were trapped on Gannet Cluster 2 (GC-2), one of a group of islands numbered by convention in the Gannet Island Archipelago, and examined for ectoparasites. One species each of Acari (Ixodes uriae) and Siphonaptera (Orchopeas leucopus) were recovered. Samples of mice favored males to females (3.4∶1). Twenty-nine percent (10) of the mice were free of ectoparasites. Males were more heavily parasitized than females when both parasites were considered. No ticks were recovered from the female mice, while the males that were parasitized carried adult Ixodes uriae. These 2 ectoparasites parasitizing P. maniculatus, which is a known reservoir host for Lyme disease (Borrelia burgdorferi), may carry B. garinii and their presence would have serious implications for the spread of this human pathogen northward in continental North America.     

Borrelia Burgdorferi Sensu Lato in Ixodes Ricinus Ticks and Rodents in a Recreational Park in South-Western Ireland

Ixodes ricinus ticks infected with Borrelia burgdorferi sensu lato were numerous on the edges of paths and roads in a recreational park in south-western Ireland. The abundance of ticks at different sites was related to the presence of deer, but a negative relationship was shown between tick abundance and tick infection rates. This is thought to be due to the deposition of large numbers of uninfected ticks by deer, which are apparently not good reservoir hosts of B. burgdorferi s.l. Blood meal analysis only detected deer DNA in uninfected nymphs. Reservoir competent rodents, Apodemus sylvaticus and Clethrionomys glareolus, were abundant at all sites and a high proportion of captured specimens were infested with larval ticks. However, very few rodents were infected with B. burgdorferi s.l. and none of the unfed infected nymphs analysed for the identity of their larval blood meal had fed on rodents. The spirochaetes detected in I. ricinus in the study area may be poorly adapted to rodents or are not transmitted readily because of the absence of nymphal infestation. The majority of spirochaetes in these ticks were apparently acquired from non-rodent hosts, such as birds.     

Complement-mediated killing of Borrelia burgdorferi by nonimmune sera from sika deer

Various species of cervid deer are the preferred hosts for adult, black-legged ticks (Ixodes scapularis and Ixodes pacificus) in the United States. Although frequently exposed to the agent of Lyme disease (Borrelia burgdorferi), these animals, for the most part, are incompetent as transmission reservoirs. We examined the borreliacidal activity of normal and B. burgdorferi-immune sera from sika deer (Cervus nippon) maintained in a laboratory setting and compared it to that of similar sera from reservoir-competent mice and rabbits. All normal deer sera (NDS) tested killed > 90% of B. burgdorferi cells. In contrast, normal mouse and rabbit sera killed < or = 22% of the Borrelia. Anti-B. burgdorferi antibodies could not be detected in any normal sera by indirect fluorescent antibody assay (IFA). Sera collected from deer 6 wk after exposure to B. burgdorferi by tick feeding exhibited IFA titers of 1:256, whereas sera from mice and rabbits similarly exposed had titers of > 1:1,024. Heat treatment (56 C, 30 min) of NDS reduced borreliacidal activity, with < 20% of the B. burgdorferi cells killed, suggesting complement-mediated killing. The chelators EGTA and EDTA were used to block the classical or both the classical and alternative complement pathways, respectively. Addition of 10 mM EGTA to NDS had a negligible effect on borreliacidal activity, with > 90% of the cells killed. Addition of 10 mM EDTA reduced the killing to approximately 30%, whereas the addition of Mg2+ (10 mM) restored borreliacidal activity to NDS. The addition of zymosan A, an activator of the alternative pathway, increased the survival of B. burgdorferi cells to approximately 80% in NDS. These data suggest that the alternative complement activation pathway plays a major role in the borreliacidal activity of NDS. Additionally, 10 mM EGTA had almost no effect on the killing activity of B. burgdorferi-exposed deer sera, suggesting that the classical pathway is not involved in Borrelia killing, even in sera from B. burgdorferi-exposed deer.     

Spirochetes in ticks and antibodies to Borrelia burgdorferi in white-tailed deer from Connecticut, New York State, and North Carolina

Ticks were screened for spirochetes and serum samples from white-tailed deer (Odocoileus virginianus) were assayed for antibodies to Borrelia burgdorferi during 1983-1984. Using fluorescein isothiocyanate-labeled rabbit antibodies produced to B. burgdorferi, the etiologic agent of Lyme disease, spirochetes were detected in Ixodes dammini (10.5% of 1,193) and Dermacentor albipictus (0.6% of 157) adults from Connecticut, I. dammini nymphs (49.1% of 108) and adults (64.7% of 99) from Armonk, New York, and in I. scapularis (0.4% of 531) and Amblyomma americanum (3.5% of 173) adults from North Carolina. Infected ticks were either seeking hosts or feeding on deer during the summer and fall. Direct fluorescent antibody staining also revealed spirochetes in two larvae of I. scapularis that emerged from eggs deposited by separate females in the laboratory. Using indirect immunofluorescence tests, antibodies to B. burgdorferi were identified in white-tailed deer living in tick-infested areas of all three states. Aside from minor cross-reactivity, there was no serologic evidence of Treponema or Leptospira infections. Ixodes dammini is a primary vector of B. burgdorferi in northeastern United States, but in North Carolina, other ixodid ticks may transmit this spirochete to humans and wildlife.     

A molecular marker for the identification of the zoonotic reservoirs of Lyme borreliosis by analysis of the blood meal in its European vector Ixodes ricinus.

The efficacy of the mitochondrially encoded cytochrome b gene as a molecular marker for the discrimination of the reservoir host species of the Lyme borreliosis spirochete, Borrelia burgdorferi sensu lato (s.l.), in its European vector Ixodes ricinus (Acari: Ixodidae) was determined. Degenerate PCR primers were designed which amplified orthologous regions of the cytochrome b gene in several animal species which act as B. burgdorferi s.l. reservoirs and hosts for I. ricinus. PCR products were amplified and characterized by hybridization and restriction fragment length polymorphism analysis. Restriction fragment length polymorphism analysis of a 638-bp PCR product with HaeIII and DdeI revealed unique restriction fragment profiles, which allowed the taxonomic identification of animals to the genus level. A system was devised for the detection of the larval host blood meal from the remnants in unfed nymphal I. ricinus ticks by nested PCR amplification. An inverse correlation was demonstrated between amplicon size and successful PCR amplification of host DNA from the nymphal stage of the tick. The stability of the cytochrome b product as a marker for the identification of the larval host species in the nymphal instar was demonstrated up to 200 days after larval ingestion (approximately 165 days after molting) by reverse line blotting with a host-specific probe. This assay has the potential for the determination of the reservoir hosts of B. burgdorferi s.l. by using extracts from the same individual ticks for both the identification of the host species and the detection of the Lyme borreliosis spirochete.     

A comparative study of mammalian and reptilian alternative pathway of complement-mediated killing of the Lyme disease spirochete (Borrelia burgdorferi)

The potential bactericidal activity of the alternative complement pathway of mammalian and reptilian sera to Borrelia burgdorferi sensu stricto (s.s.) was evaluated in vitro. Complement-mediated killing was observed when cultured spirochetes were inoculated into sera from the western fence lizard (Sceloporus occidentalis) and from the southern alligator lizard (Elgaria multicarinata), but not when they were inoculated into serum from either the deer mouse (Peromyscus maniculatus) or from humans. Spirochetes were still alive after 4 hr in lizard serum that had been preheated at 56 C for 30 min to inactivate complement. Furthermore, when lizard serum was chelated with 10 mM ethylenediaminetetraacetic acid to block all complement activation, borreliacidal activity was arrested. When lizard serum was chelated with 10 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid plus 4 mM MgCl2 to block only classical complement pathway activation, >85% of spirochetes were immobilized within 1 hr. Differences in B. burgdorferi s.s. mortality were not observed when chelators with or without MgCl2 were added to serum from either deer mice or humans. Proteins comprising the alternative complement pathway are responsible for the borreliacidal activity observed in the blood of S. occidentalis and E. multicarinata.     

The role of lizards in the ecology of Lyme disease in two endemic zones of the northeastern United States

We examined the role of lizards in the ecology of Lyme disease in New York and Maryland. We collected data on vector tick infestations, measured lizard "realized" reservoir competence for the Lyme disease spirochete Borrelia burgdorferi, and estimated lizard population density. These data were incorporated into a model that predicts a host's ability to influence the prevalence of B. burgdorferi in the tick population, a primary risk factor in the epidemiology of Lyme disease. Published data on other northeastern hosts were included in the model to provide a reference for interpreting the importance of lizards as hosts. The model results indicate that 5-lined skinks (Eumeces fasciatus) are dilution hosts, capable of reducing infection prevalence in the tick population by 10.7-51.5 percentage points, whereas eastern fence lizards (Sceloporus undulatus) are not dilution hosts in the areas studied. Owing to moderate burdens of larval ticks, relatively high population densities, and reservoir incompetence, E. fasciatus may play an important role in the ecology of Lyme disease by reducing vector infection prevalence and associated human risk of infection.     

Role of small mammals in the ecology of Borrelia burgdorferi in a peri-urban park in north coastal California

The role of small mammals other than woodrats in the enzootiology of the Lyme disease spirochete, Borrelia burgorferi, was assessed in a peri-urban park. Mammals were collected monthly from September through to April. Following tick removal, the animals were tested for B. burgdorferi by culture of ear-punch biopsies. Larvae and nymphs that were intermediate in morphology between Ixodes spinipalpis and Ixodes neotomae occurred on several species of rodents (Peromyscus truei, Peromyscus californicus, Microtus californicus, Rattus rattus and Reithrodontomys megalotis) and the brush rabbit (Sylvilagus bachmani). Morphometric analyses of these I. spinipalpis-like ticks and the offspring from two I. neotomae females from this site suggest that I. neotomae may be conspecific with I. spinipalpis. Borrelia burgdorferi was isolated from eight out of 109 (7.3%), three out of 16 (18.8%), two out of 38 (5.3%) and two out of six (33.3%) P. truei, P. maniculatus, M. californicus and R. rattus, respectively. One brush rabbit yielded the first isolate of B. burgdorferi from a lagomorph in western North America. This isolate and three others derived from unfed I. spinipalpis-like nymphs failed to produce infection when inoculated intradermally into 11–12 P. maniculatus each. Likewise, no spirochetes were detected in 420 Ixodes pacificus nymphs derived from larvae fed on animals inoculated with these isolates. An additional isolate, derived from an I. spinipalpis-like nymph, was recovered by ear-punch biopsies from five out of 12 (42%) needle-inoculated P. maniculatus. However, spirochetes were not detected in 20 I. pacificus nymphs fed as larvae on each of five mice (two infected and three uninfected) inoculated with this isolate. We conclude that brush rabbits and several species of rodents besides woodrats may contribute to the maintenance of B. burgdorferi because they harbour the spirochete and are fed upon by competent enzootic and bridge vectors.