Examining Prevalence and Diversity of Tick-Borne Pathogens in Questing Ixodes pacificus Ticks in California.

Tick-borne diseases in California include Lyme disease (caused by Borrelia burgdorferi), infections with Borrelia miyamotoi, and human granulocytic anaplasmosis (caused by Anaplasma phagocytophilum). We surveyed multiple sites and habitats (woodland, grassland, and coastal chaparral) in California to describe spatial patterns of tick-borne pathogen prevalence in western black-legged ticks (Ixodes pacificus). We found that several species of Borrelia-B. burgdorferi, Borrelia americana, and Borrelia bissettiae-were observed in habitats, such as coastal chaparral, that do not harbor obvious reservoir host candidates. Describing tick-borne pathogen prevalence is strongly influenced by the scale of surveillance: aggregating data from individual sites to match jurisdictional boundaries (e.g., county or state) can lower the reported infection prevalence. Considering multiple pathogen species in the same habitat allows a more cohesive interpretation of local pathogen occurrence. IMPORTANCE Understanding the local host ecology and prevalence of zoonotic diseases is vital for public health. Using tick-borne diseases in California, we show that there is often a bias to our understanding and that studies tend to focus on particular habitats, e.g., Lyme disease in oak woodlands. Other habitats may harbor a surprising diversity of tick-borne pathogens but have been neglected, e.g., coastal chaparral. Explaining pathogen prevalence requires descriptions of data on a local scale; otherwise, aggregating the data can misrepresent the local dynamics of tick-borne diseases.

Leptospira in river and soil in a highly endemic area of Ecuador.

BACKGROUND: Leptospira are shed into the environment via urine of infected animals. Rivers are thought to be an important risk factor for transmission to humans, though much is unknown about the types of environment or characteristics that favor survival. To address this, we screened for Leptospira DNA in two rivers in rural Ecuador where Leptospirosis is endemic. RESULTS: We collected 112 longitudinal samples and recorded pH, temperature, river depth, precipitation, and dissolved oxygen. We also performed a series of three experiments designed to provide insight into Leptospira presence in the soil. In the first soil experiment, we characterized prevalence and co-occurrence of Leptospira with other bacterial taxa in the soil at dispersed sites along the rivers (n = 64). In the second soil experiment, we collected 24 river samples and 48 soil samples at three points along eight transects to compare the likelihood of finding Leptospira in the river and on the shore at different distances from the river. In a third experiment, we tested whether Leptospira presence is associated with soil moisture by collecting 25 soil samples from two different sites. In our river experiment, we found pathogenic Leptospira in only 4 (3.7%) of samples. In contrast, pathogenic Leptospira species were found in 22% of shore soil at dispersed sites, 16.7% of soil samples (compared to 4.2% of river samples) in the transects, and 40% of soil samples to test for associations with soil moisture. CONCLUSIONS: Our data are limited to two sites in a highly endemic area, but the scarcity of Leptospira DNA in the river is not consistent with the widespread contention of the importance of river water for leptospirosis transmission. While Leptospira may be shed directly into the river, onto the shores, or washed into the river from more remote sites, massive dilution and limited persistence in rivers may reduce the environmental load and therefore, the epidemiological significance of such sources. It is also possible that transmission may occur more frequently on shores where people are liable to be barefoot. Molecular studies that further explore the role of rivers and water bodies in the epidemiology of leptospirosis are needed.

Isolation and Molecular Characterization of Tick-Borne Relapsing Fever Borrelia Infecting Ornithodoros (Pavlovskyella) verrucosus Ticks Collected in Ukraine.

BACKGROUND: Tick-borne relapsing fever (TBRF) is a neglected zoonotic bacterial disease known to occur on 5 continents. We report a laboratory-acquired case of TBRF caused by Borrelia caucasica, which is endemic in Ukraine and transmitted by Ornithodoros verrucosus ticks. METHODS: We isolated spirochetes and characterized them by partially sequencing the 16s ribosomal ribonucleic acid (rrs), flagellin (flaB), and deoxyribonucleic acid gyrase (gyrB) genes and conducting a phylogenetic analysis. RESULTS: These analyses revealed a close relationship of Ukrainian spirochetes with the Asian TBRF species, Borrelia persica. The taxonomic and nomenclature problems related to insufficient knowledge on the spirochetes and their vectors in the region are discussed. CONCLUSIONS: Although these findings enhance our understanding of species identities for TBRF Borrelia in Eurasia, further work is required to address the neglected status of TBRF in this part of the world. Public health practitioners should consider TBRF and include the disease into differential diagnosis of febrile illnesses with unknown etiology.

Citizen science informs human-tick exposure in the Northeastern United States.

BACKGROUND: Tick-borne disease is the result of spillover of pathogens into the human population. Traditionally, literature has focused on characterization of tick-borne disease pathogens and ticks in their sylvatic cycles. A limited amount of research has focused on human-tick exposure in this system, especially in the Northeastern United States. Human-tick interactions are crucial to consider when assessing the risk of tick-borne disease since a tick bite is required for spillover to occur. METHODS: Citizen scientists collected ticks from the Northeastern US through a free nationwide program. Submitted ticks were identified to species, stage, and sex. Blacklegged ticks, Ixodes scapularis, were tested for the presence of Borrelia burgdorferi sensu lato (s.l.) and hard-tick relapsing fever Borrelia. Seasonality of exposure and the citizen science activity during tick exposure was recorded by the citizen scientist. A negative binomial model was fit to predict county level CDC Lyme disease cases in 2016 using citizen science Ixodes scapularis submissions, state, and county population as predictor variables. RESULTS: A total of 3740 submissions, comprising 4261 ticks, were submitted from the Northeastern US and were reported to be parasitizing humans. Of the three species submitted, blacklegged ticks were the most prevalent followed by American dog ticks and lone star ticks. Submissions peaked in May with the majority of exposure occurring during every-day activities. The most common pathogen in blacklegged ticks was B. burgdorferi s.l. followed by hard-tick relapsing fever Borrelia. Negative binomial model performance was best in New England states followed by Middle Atlantic states. CONCLUSIONS: Citizen science provides a low-cost and effective methodology for describing the seasonality and characteristics of human-tick exposure. In the Northeastern US, everyday activities were identified as a major mechanism for tick exposure, supporting the role of peri-domestic exposure in tick-borne disease. Citizen science provides a method for broad pathogen and tick surveillance, which is highly related to human disease, allowing for inferences to be made about the epidemiology of tick-borne disease.

Borrelia miyamotoi Infections in Small Mammals, California, USA.

Surveillance to investigate the wildlife-vector transmission cycle of the human pathogen Borrelia miyamotoi in California, USA, revealed infections in dusky-footed woodrats, brush mice, and California mice. Phylogenetic analyses suggest a single, well-supported clade of B. miyamotoi is circulating in California.

Where Are the Ticks? Solving the Mystery of a Tickborne Relapsing Fever Outbreak at a Youth Camp.

During the summer of 2014 an outbreak of tickborne relapsing fever (TBRF) occurred in a group of high school students and staff at a youth camp, which was reported to Coconino County Public Health Services District. Six confirmed and five probable cases of TBRF occurred. During the environmental investigation two rodents tested positive for TBRF, but the vector, soft ticks, could not be found in their "normal" habitat. Ticks were finally located in areas not typical for soft ticks.

Notes from the Field: Tickborne Relapsing Fever Outbreak at an Outdoor Education Camp - Arizona, 2014.

Tickborne relapsing fever (TBRF) is a bacterial infection characterized by recurring episodes of fever, headache, muscle and joint aches, and nausea. In North America, TBRF primarily is caused by Borrelia hermsii spirochetes transmitted by Ornithodoros hermsii ticks. Once infected, these soft ticks are infectious for life and transmit the spirochete to sleeping humans quickly (possibly within 30 seconds) during short feeds (15-90 minutes). On August 10, 2014, the Coconino County Public Health Services District in Arizona was notified by a local hospital that five high school students who attended the same outdoor education camp had been hospitalized with fever, headache, and myalgias. Hantavirus infection initially was suspected because of reported exposure to rodent droppings, but after detecting spirochetes on peripheral blood smears from all five hospitalized students, TBRF was diagnosed. The camp was instructed to close immediately, and the health department, in collaboration with local university experts, investigated to identify additional cases, determine the cause, and prevent further infections. A total of 11 cases (six confirmed and five probable) were identified.

Relapsing fever group Borrelia in Southern California rodents.

Wild rodent reservoir host species were surveyed prospectively for infection with Borrelia hermsii, the causative agent of tick-borne relapsing fever in the western United States. Trapping occurred during the summer of 2009-2012 at field sites surrounding Big Bear Lake, CA, a region where human infection has been reported for many years. Using quantitative polymerase chain reaction (qPCR), we tested 207 rodents from 11 species and found chipmunks (Tamias spp.) and a woodrat (Neotoma macrotis) infected. Chipmunks represented the majority of captures at these sites. Sixteen of the 207 (7.7%; CI = 4.6-12.4) animals were qPCR-positive for Borrelia spp. associated with relapsing fever, and of those, we obtained bacterial DNA sequences from eight. The phylogram made from these sequences depict a clear association with B. hermsii genomic group I. In addition, we identified an infection with Borrelia coriaceae in a Tamias merriami, a potentially nonpathogenic member of the tick-borne relapsing fever group. Our findings support the hypothesis that chipmunk species play an important role in the maintenance of Borrelia species that cause tick-borne relapsing fever in the western United States, and therefore the risk of infection to people.

Citizen Science Provides an Efficient Method for Broad-Scale Tick-Borne Pathogen Surveillance of Ixodes pacificus and Ixodes scapularis across the United States.

Tick-borne diseases have expanded over the last 2 decades as a result of shifts in tick and pathogen distributions. These shifts have significantly increased the need for accurate portrayal of real-time pathogen distributions and prevalence in hopes of stemming increases in human morbidity. Traditionally, pathogen distribution and prevalence have been monitored through case reports or scientific collections of ticks or reservoir hosts, both of which have challenges that impact the extent, availability, and accuracy of these data. Citizen science tick collections and testing campaigns supplement these data and provide timely estimates of pathogen prevalence and distributions to help characterize and understand tick-borne disease threats to communities. We utilized our national citizen science tick collection and testing program to describe the distribution and prevalence of four Ixodes-borne pathogens, Borrelia burgdorferi sensu lato, Borrelia miyamotoi, Anaplasma phagocytophilum, and Babesia microti, across the continental United States. IMPORTANCE In the 21st century, zoonotic pathogens continue to emerge, while previously discovered pathogens continue to have changes within their distribution and prevalence. Monitoring these pathogens is resource intensive, requiring both field and laboratory support; thus, data sets are often limited within their spatial and temporal extents. Citizen science collections provide a method to harness the general public to collect samples, enabling real-time monitoring of pathogen distribution and prevalence.

Predicting the current and future distribution of the western black-legged tick, Ixodes pacificus, across the Western US using citizen science collections.

In the twenty-first century, ticks and tick-borne diseases have expanded their ranges and impact across the US. With this spread, it has become vital to monitor vector and disease distributions, as these shifts have public health implications. Typically, tick-borne disease surveillance (e.g., Lyme disease) is passive and relies on case reports, while disease risk is calculated using active surveillance, where researchers collect ticks from the environment. Case reports provide the basis for estimating the number of cases; however, they provide minimal information on vector population or pathogen dynamics. Active surveillance monitors ticks and sylvatic pathogens at local scales, but it is resource-intensive. As a result, data are often sparse and aggregated across time and space to increase statistical power to model or identify range changes. Engaging public participation in surveillance efforts allows spatially and temporally diverse samples to be collected with minimal effort. These citizen-driven tick collections have the potential to provide a powerful tool for tracking vector and pathogen changes. We used MaxEnt species distribution models to predict the current and future distribution of Ixodes pacificus across the Western US through the use of a nationwide citizen science tick collection program. Here, we present niche models produced through citizen science tick collections over two years. Despite obvious limitations with citizen science collections, the models are consistent with previously-predicted species ranges in California that utilized more than thirty years of traditional surveillance data. Additionally, citizen science allows for an expanded understanding of I. pacificus distribution in Oregon and Washington. With the potential for rapid environmental changes instigated by a burgeoning human population and rapid climate change, the development of tools, concepts, and methodologies that provide rapid, current, and accurate assessment of important ecological qualities will be invaluable for monitoring and predicting disease across time and space.

Equids engineer desert water availability.

Megafauna play important roles in the biosphere, yet little is known about how they shape dryland ecosystems. We report on an overlooked form of ecosystem engineering by donkeys and horses. In the deserts of North America, digging of ≤2-meter wells to groundwater by feral equids increased the density of water features, reduced distances between waters, and, at times, provided the only water present. Vertebrate richness and activity were higher at equid wells than at adjacent dry sites, and, by mimicking flood disturbance, equid wells became nurseries for riparian trees. Our results suggest that equids, even those that are introduced or feral, are able to buffer water availability, which may increase resilience to ongoing human-caused aridification.

Borreliosis Transmission from Ticks Associated with Desert Tortoise Burrows: Examples of Tick-Borne Relapsing Fever in the Mojave Desert.

Ticks transmit pathogens and parasitize wildlife in turn causing zoonotic diseases in many ecosystems. Argasid ticks, such as Ornithodoros spp., harbor and transmit Borrelia spp., resulting in tick-borne relapsing fever (TBRF) in people. In the western United States, TBRF is typically associated with the bite of an infected Ornithodoros hermsi tick found in habitats at high elevations (>1500 ft). This report describes the first TBRF cases in people in the Mojave Desert (Clark County, NV). Individuals documented in these case studies were exposed to Ornithodoros ticks during excavation of soil burrows associated with Mojave Desert tortoises (Gopherus agassizii), with bacteria from one of the human case's blood sample genetically matching to Borrelia turicatae as determined by quantitative PCR and sequencing. Our findings should serve as a precaution to individuals working with tortoises or animal burrows, or those in contact with Ornithodoros ticks in this region.

Integration of choline geranate into electrospun protein scaffolds affords antimicrobial activity to biomaterials used for cutaneous wound healing.

Wound healing attempts to maintain homeostasis in the wound while minimizing the risk of infection to the tissue by foreign agents, such as opportunistic bacterial pathogens. Biofilms established by these pathogens are a common cause of chronic infections that slow the healing process. Preparation of skin wound healing devices comprised of electrospun proteins associated with skin have been shown to accelerate the healing process relative to conventional wound dressings. In this work, we have developed electrospinning methods to incorporate the antimicrobial ionic liquid/deep eutectic solvent choline geranate (CAGE) into these devices. Integration of CAGE into the dressing material was verified via 1 H nuclear magnetic resonance spectrometry, and the effect on the material property of the resultant devices were assessed using scanning electron microscopy. CAGE-containing devices demonstrate a concentration-dependent inactivation of exogenously applied solutions of both gram-positive and gram-negative pathogens (Enterococcus sp and Pseudomonas aeruginosa, respectively), but maintain their ability to serve as a compatible platform for proliferation of human dermal neonatal fibroblasts.

Pilot immunization of mice infected with an equine strain of Corynebacterium pseudotuberculosis.

This pilot study evaluated protection of an equine autogenous bacterin-toxoid vaccine against Corynebacterium pseudotuberculosis infection. Twenty-four BALB/c mice were inoculated with two doses of bacterin-toxoid vaccine or two injections of a placebo. Clinical, microbiologic, and pathologic outcomes were assessed after intradermal infection with one of two equine-origin C. pseudotuberculosis strains. Mice receiving bacterin-toxoid from fast-growing C. pseudotuberculosis showed significant protection from challenge infection, as evidenced by a higher survival rate, fewer gross and histopathologic lesions, and lower bacterial levels on culture. Successful protection via a vaccine against equine internal abscesses might provide supplementary management options against an important, potentially fatal disease.

COYOTES (CANIS LATRANS) IN ARIZONA, USA, EXHIBIT IMMUNE AND GENETIC EVIDENCE OF RICKETTSIAL INFECTIONS.

Rocky Mountain spotted fever (RMSF), caused by the bacterium Rickettsia rickettsii, was recognized as endemic in Arizona, US after a 2002 outbreak and has since been a public health concern. The brown dog tick (Rhipicephalus sanguineus sensu lato) is the principal vector of this pathogen in Arizona. Domesticated dogs (Canis lupus familiaris) are the tick's main host, so free-roaming dogs in peridomestic areas have been named the primary risk factor for human cases of RMSF. However, the sudden emergence and long-distance dispersal of the pathogen have not been adequately explained, and one possible mechanism could include wildlife. Coyotes (Canis latrans) are wide ranging in Arizona and closely related to dogs, so it is possible that brown dog ticks parasitize coyotes and infect them. Although R. rickettsii is the most severe spotted fever group (SFG) rickettsial pathogen in humans, others occur in Arizona, and antibodies raised against them are cross-reactive, so we more-broadly hypothesized that coyotes in Arizona are exposed to SFG rickettsiae. We collected coyote tissues in spring 2016 and 2017. We tested sera for antibodies to R. rickettsii and found 9% (8/94) of samples were antibody-positive with titers of ≥256. Subsequent quantitative PCR analyses of skin showed evidence for Rickettsia spp. in 2.9% (4/138) of samples. These data suggest that coyotes have a role in the maintenance of SFG rickettsiae in Arizona. Further investigation is warranted to reveal which specific pathogen-vector complexes act on coyotes in the region and whether they represent a risk to human health.

Evaluation of hepatic disease in mice following intradermal inoculation with Corynebacterium pseudotuberculosis.

OBJECTIVE: To evaluate clinical, microbiologic, and pathologic outcomes in mice after inoculation with 4 equine-origin Corynebacterium pseudotuberculosis strains. ANIMALS: 15 C3H/HeJ mice. PROCEDURES: In a preliminary study, the optimum route of inoculation was determined. In the main study, mice were allocated to 4 treatment groups (3 mice/group). One slow- or rapid-growing equine-origin C pseudotuberculosis strain was inoculated ID into the mice of each treatment group. RESULTS: All 4 strains had distinct tropism for the liver. Histologic lesions associated with rapid-growing strains included focally extensive unencapsulated areas of acute, massive coagulative necrosis of hepatocytes with intralesional colonies of bacteria and variable portal hepatitis characterized by accumulations of mononuclear and polymorphonuclear inflammatory cells. In contrast, the livers of mice inoculated with slow-growing strains had multiple discrete, randomly distributed foci of hepatocellular necrosis and neutrophilic hepatitis that were considerably less severe than the lesions in the mice inoculated with the rapid-growing strains. Significantly more bacterial colonies were recovered from the organs of mice inoculated with rapid-growing than with slow-growing strains of bacteria. Bacteria were isolated from the liver, spleen, lungs, and mesenteric lymph nodes of mice inoculated with rapid-growing strains and from the liver and lymph nodes of mice inoculated with slow-growing strains. CONCLUSIONS AND CLINICAL RELEVANCE: Study of host-bacteria interactions in hosts that are naturally infected with C pseudotuberculosis is difficult because of underlying genetic variability among animals, expense, and requirements for multiple replicates and control animals. The C3H/HeJ mice may provide a useful means for studying virulence mechanisms of C pseudotuberculosis.

Time of year and outdoor recreation affect human exposure to ticks in California, United States.

Interactions between humans and ticks are often measured indirectly, using surveillance of tick population abundance and pathogen prevalence, or reported human disease data. We used data garnered as part of a free national citizen science research effort to describe actual human exposures to ticks in California. Human-biting ticks (n = 1,905) submitted for identification were predominantly western black-legged ticks (Ixodes pacificus) (68%), American dog ticks (Dermacentor variabilis) (24%), and Pacific Coast ticks (Dermacentor occidentalis) (7%). Tick exposure occurred predominantly during recreational use of the outdoors, rather than exposure near the home environment. Tick submissions peaked in May, but human exposure to ticks occurred throughout the year. Adult I. pacificus were most frequently found on humans during March-May, though previous research demonstrates that questing adults on vegetation are more abundant earlier in the winter.

Anaplasma phagocytophilum infection in small mammal hosts of Ixodes ticks, western United States.

A total of 2,121 small mammals in California were assessed for Anaplasma phagocytophilum from 2006 through 2008. Odds ratios were >1 for 4 sciurids species and dusky-footed woodrats. High seroprevalence was observed in northern sites. Ten tick species were identified. Heavily infested rodent species included meadow voles, woodrats, deer mice, and redwood chipmunks.

Evaluation of squirrels (Rodentia: Sciuridae) as ecologically significant hosts for Anaplasma phagocytophilum in California.

Granulocytic anaplasmosis (GA), caused by Anaplasma phagocytophilum, is a potentially fatal, emerging rickettsial disease of humans, domestic animals, and wildlife. The purpose of this study was to determine whether sciurids from multiple areas of northern California were infested with ticks or exposed to or infected with A. phagocytophilum using polymerase chain reaction (PCR) and indirect-fluorescent antibody (IFA) serology. Sciurids of nine different tree- and ground-dwelling species were assessed: arboreal squirrels (western and eastern gray squirrels, Sciurus griseus and S. carolinensis, and Douglas squirrels, Tamiasciurus douglasii) but not northern flying squirrels (Glaucomys sabrinus) had greater evidence of exposure and current infection than did semiarboreal or ground dwelling sciurids (California ground squirrels, Spermophilus beecheyi, and chipmunks, Tamias spp.). Western gray squirrels had the most extensive exposure (70.7% seroprevalence and 12.1% PCR prevalence). Positive squirrels were identified in all regions where squirrels were collected. A logistic regression identified being a western gray squirrel (OR = 20.5, P = 2.95 X 10(-8)) and from the north coastal region of California (OR = 9.052, P = 1.41 X 10(-6)) as having the highest risk of exposure to A. phagocytophilum. Five of nine sciurid species had evidence of infestation with Ixodes pacificus or I. spinipalpis that could vector A. phagocytophilum. Extensive exposure from multiple areas suggests sciurids may be important in the maintenance of GA in California and indicates that studies of reservoir competence of these species are warranted.