Karen A. Boegler

Flea Diversity as an Element for Persistence of Plague Bacteria in an East African Plague Focus

Plague is a flea-borne rodent-associated zoonotic disease that is caused by Yersinia pestis and characterized by long quiescent periods punctuated by rapidly spreading epidemics and epizootics. How plague bacteria persist during inter-epizootic periods is poorly understood, yet is important for predicting when and where epizootics are likely to occur and for designing interventions aimed at local elimination of the pathogen. Existing hypotheses of how Y. pestis is maintained within plague foci typically center on host abundance or diversity, but little attention has been paid to the importance of flea diversity in enzootic maintenance. Our study compares host and flea abundance and diversity along an elevation gradient that spans from low elevation sites outside of a plague focus in the West Nile region of Uganda (∼725–1160 m) to higher elevation sites within the focus (∼1380–1630 m). Based on a year of sampling, we showed that host abundance and diversity, as well as total flea abundance on hosts was similar between sites inside compared with outside the plague focus. By contrast, flea diversity was significantly higher inside the focus than outside. Our study highlights the importance of considering flea diversity in models of Y. pestis persistence.

Transmission of Borrelia miyamotoi sensu lato relapsing fever group spirochetes in relation to duration of attachment by Ixodes scapularis nymphs

Borrelia miyamotoi sensu lato relapsing fever group spirochetes are emerging as causative agents of human illness (Borrelia miyamotoi disease) in the United States. Host-seeking Ixodes scapularis ticks are naturally infected with these spirochetes in the eastern United States and experimentally capable of transmitting B. miyamotoi. However, the duration of time required from tick attachment to spirochete transmission has yet to be determined. We therefore conducted a study to assess spirochete transmission by single transovarially infected I. scapularis nymphs to outbred white mice at three time points post-attachment (24, 48, and 72h) and for a complete feed (>72-96h). Based on detection of B. miyamotoi DNA from the blood of mice fed on by an infected nymph, the probability of spirochete transmission increased from 10% by 24h of attachment (evidence of infection in 3/30 mice) to 31% by 48h (11/35 mice), 63% by 72h (22/35 mice), and 73% for a complete feed (22/30 mice). We conclude that (i) single I. scapularis nymphs effectively transmit B. miyamotoi relapsing fever group spirochetes while feeding, (ii) transmission can occur within the first 24h of nymphal attachment, and (iii) the probability of transmission increases with the duration of nymphal attachment.

Identification of Risk Factors for Plague in the West Nile Region of Uganda

Plague is an often fatal, primarily flea-borne rodent-associated zoonosis caused by Yersinia pestis. We sought to identify risk factors for plague by comparing villages with and without a history of human plague cases within a model-defined plague focus in the West Nile Region of Uganda. Although rat (Rattus rattus) abundance was similar inside huts within case and control villages, contact rates between rats and humans (as measured by reported rat bites) and host-seeking flea loads were higher in case villages. In addition, compared with persons in control villages, persons in case villages more often reported sleeping on reed or straw mats, storing food in huts where persons sleep, owning dogs and allowing them into huts where persons sleep, storing garbage inside or near huts, and cooking in huts where persons sleep. Compared with persons in case villages, persons in control villages more commonly reported replacing thatch roofing, and growing coffee, tomatoes, onions, and melons in agricultural plots adjacent to their homesteads. Rodent and flea control practices, knowledge of plague, distance to clinics, and most care-seeking practices were similar between persons in case villages and persons in control villages. Our findings reinforce existing plague prevention recommendations and point to potentially advantageous local interventions.

Isolation of the Lyme Disease Spirochete Borrelia mayonii From Naturally Infected Rodents in Minnesota

Abstract Borrelia mayonii is a newly described member of the Borrelia burgdorferi sensu lato complex that is vectored by the black-legged tick (Ixodes scapularis Say) and a cause of Lyme disease in Minnesota and Wisconsin. Vertebrate reservoir hosts involved in the enzootic maintenance of B. mayonii have not yet been identified. Here, we describe the first isolation of B. mayonii from naturally infected white-footed mice (Peromyscus leucopus Rafinesque) and an American red squirrel (Tamiasciurus hudsonicus Erxleben) from Minnesota, thus implicating these species as potential reservoir hosts for this newly described spirochete.

Blood Meal Identification in Off-Host Cat Fleas (Ctenocephalides felis) from a Plague-Endemic Region of Uganda

The cat flea, Ctenocephalides felis, is an inefficient vector of the plague bacterium (Yersinia pestis) and is the predominant off-host flea species in human habitations in the West Nile region, an established plague focus in northwest Uganda. To determine if C. felis might serve as a Y. pestis bridging vector in the West Nile region, we collected on- and off-host fleas from human habitations and used a real-time polymerase chain reaction-based assay to estimate the proportion of off-host C. felis that had fed on humans and the proportion that had fed on potentially infectious rodents or shrews. Our findings indicate that cat fleas in human habitations in the West Nile region feed primarily on domesticated species. We conclude that C. felis is unlikely to serve as a Y. pestis bridging vector in this region.

Infection Prevalence, Bacterial Loads, and Transmission Efficiency in Oropsylla montana (Siphonaptera: Ceratophyllidae) One Day After Exposure to Varying Concentrations of Yersinia pestis in Blood

Abstract Unblocked fleas can transmit Yersinia pestis, the bacterium that causes plague, shortly (≤4 d) after taking an infectious bloodmeal. Investigators have measured so-called early-phase transmission (EPT) efficiency in various fleas following infection with highly bacteremic blood (≥108 cfu/ml). To date, no one has determined the lower limit of bacteremia required for fleas to acquire and transmit infection by EPT, though knowing this threshold is central to determining the length of time a host may be infectious to feeding fleas. Here, we evaluate the ability of Oropsylla montana (Baker) to acquire and transmit Y. pestis after feeding on blood containing 103 to 109 cfu/ ml. We evaluated the resulting infection prevalence, bacterial loads, and transmission efficiency within the early-phase time period at 1 d postinfection. Fleas acquired infection from bacteremic blood across a wide range of concentrations, but transmission was observed only when fleas ingested highly bacteremic blood.

Flea-Associated Bacterial Communities across an Environmental Transect in a Plague-Endemic Region of Uganda

The vast majority of human plague cases currently occur in sub-Saharan Africa. The primary route of transmission of Yersinia pestis, the causative agent of plague, is via flea bites. Non-pathogenic flea-associated bacteria may interact with Y. pestis within fleas and it is important to understand what factors govern flea-associated bacterial assemblages. Six species of fleas were collected from nine rodent species from ten Ugandan villages between October 2010 and March 2011. A total of 660,345 16S rRNA gene DNA sequences were used to characterize bacterial communities of 332 individual fleas. The DNA sequences were binned into 421 Operational Taxonomic Units (OTUs) based on 97% sequence similarity. We used beta diversity metrics to assess the effects of flea species, flea sex, rodent host species, site (i.e. village), collection date, elevation, mean annual precipitation, average monthly precipitation, and average monthly temperature on bacterial community structure. Flea species had the greatest effect on bacterial community structure with each flea species harboring unique bacterial lineages. The site (i.e. village), rodent host, flea sex, elevation, precipitation, and temperature also significantly affected bacterial community composition. Some bacterial lineages were widespread among flea species (e.g. Bartonella spp. and Wolbachia spp.), but each flea species also harbored unique bacterial lineages. Some of these lineages are not closely related to known bacterial diversity and likely represent newly discovered lineages of insect symbionts. Our finding that flea species has the greatest effect on bacterial community composition may help future investigations between Yersinia pestis and non-pathogenic flea-associated bacteria. Characterizing bacterial communities of fleas during a plague epizootic event in the future would be helpful.

Prevalence and Diversity of Tick-Borne Pathogens in Nymphal Ixodes scapularis (Acari: Ixodidae) in Eastern National Parks

Abstract Tick-borne pathogens transmitted by Ixodes scapularis Say (Acari: Ixodidae), also known as the deer tick or blacklegged tick, are increasing in incidence and geographic distribution in the United States. We examined the risk of tick-borne disease exposure in 9 national parks across six Northeastern and Mid-Atlantic States and the District of Columbia in 2014 and 2015. To assess the recreational risk to park visitors, we sampled for ticks along frequently used trails and calculated the density of I. scapularis nymphs (DON) and the density of infected nymphs (DIN). We determined the nymphal infection prevalence of I. scapularis with a suite of tick-borne pathogens including Borrelia burgdorferi, Borrelia miyamotoi, Anaplasma phagocytophilum, and Babesia microti. Ixodes scapularis nymphs were found in all national park units; DON ranged from 0.40 to 13.73 nymphs per 100 m2. Borrelia burgdorferi, the causative agent of Lyme disease, was found at all sites where I. scapularis was documented; DIN with B. burgdorferi ranged from 0.06 to 5.71 nymphs per 100 m2. Borrelia miyamotoi and A. phagocytophilum were documented at 60% and 70% of the parks, respectively, while Ba. microti occurred at just 20% of the parks. Ixodes scapularis is well established across much of the Northeastern and Mid-Atlantic States, and our results are generally consistent with previous studies conducted near the areas we sampled. Newly established I. scapularis populations were documented in two locations: Washington, D.C. (Rock Creek Park) and Greene County, Virginia (Shenandoah National Park). This research demonstrates the potential risk of tick-borne pathogen exposure in national parks and can be used to educate park visitors about the importance of preventative actions to minimize tick exposure.

Transmission of the Lyme Disease Spirochete Borrelia mayonii in Relation to Duration of Attachment by Nymphal Ixodes scapularis (Acari: Ixodidae)

Abstract The recently recognized Lyme disease spirochete, Borrelia mayonii, has been detected in host-seeking Ixodes scapularis Say ticks and is associated with human disease in the Upper Midwest. Although experimentally shown to be vector competent, studies have been lacking to determine the duration of time from attachment of a single B. mayonii-infected I. scapularis nymph to transmission of spirochetes to a host. If B. mayonii spirochetes were found to be transmitted within the first 24 h after tick attachment, in contrast to Borrelia burgdorferi spirochetes (>24 h), then current recommendations for tick checks and prompt tick removal as a way to prevent transmission of Lyme disease spirochetes would need to be amended. We therefore conducted a study to determine the probability of transmission of B. mayonii spirochetes from single infected nymphal I. scapularis ticks to susceptible experimental mouse hosts at three time points postattachment (24, 48, and 72 h) and for a complete feed (>72–96 h). No evidence of infection with or exposure to B. mayonii occurred in mice that were fed upon by a single infected nymph for 24 or 48 h. The probability of transmission by a single infected nymphal tick was 31% after 72 h of attachment and 57% for a complete feed. In addition, due to unintended simultaneous feeding upon some mice by two B. mayonii-infected nymphs, we recorded a single occasion in which feeding for 48 h by two infected nymphs resulted in transmission and viable infection in the mouse. We conclude that the duration of attachment of a single infected nymphal I. scapularis tick required for transmission of B. mayonii appears to be similar to that for B. burgdorferi: transmission is minimal for the first 24 h of attachment, rare up to 48 h, but then increases distinctly by 72 h postattachment.

An Acarological Risk Model Predicting the Density and Distribution of Host-Seeking Ixodes scapularis Nymphs in Minnesota

Abstract. Ixodes scapularis is the vector of at least seven human pathogens in Minnesota, two of which are known to cause Lyme disease (Borrelia burgdorferi sensu stricto and Borrelia mayonii). In Minnesota, the statewide incidence of Lyme disease and other I. scapularis–borne diseases and the geographic extent over which cases have been reported have both increased substantially over the last two decades. These changes correspond with an expanding distribution of I. scapularis over a similar time frame. Because the risk of exposure to I. scapularis–borne pathogens is likely related to the number of ticks encountered, we developed an acarological risk model predicting the density of host-seeking I. scapularis nymphs (DON) in Minnesota. The model was informed by sampling 81 sites located in 42 counties in Minnesota. Two main foci were predicted by the model to support elevated densities of host-seeking I. scapularis nymphs, which included the seven-county Minneapolis-St. Paul metropolitan area and counties in northern Minnesota, including Lake of the Woods and Koochiching counties. There was substantial heterogeneity observed in predicted DON across the state at the county scale; however, counties classified as high risk for I. scapularis–borne diseases and counties with known established populations of I. scapularis had the highest proportion of the county predicted as suitable for host-seeking nymphs (≥ 0.13 nymphs/100 m2). The model provides insight into areas of potential I. scapularis population expansion and identifies focal areas of predicted suitable habitat within counties where the incidence of I. scapularis–borne diseases has been historically low.