Seasonal dynamics of tick burden and associated Borrelia burgdorferi s.l. and Borrelia miyamotoi infections in rodents in a Dutch forest ecosystem.

Ixodes ricinus ticks transmit Borrelia burgdorferi sensu lato (s.l.) as well as Borrelia miyamotoi. Larvae become infected when feeding on infected rodents, with horizontal transmission of B. burgdorferi and horizontal and vertical transmission of B. miyamotoi. We studied seasonal dynamics of infection rates of I. ricinus and their rodent hosts, and hence transmission risk of these two distinctly different Borrelia species. Rodents were live-trapped and inspected for ticks from May to November in 2013 and 2014 in a forest in The Netherlands. Trapped rodents were temporarily housed in the laboratory and detached ticks were collected. Borrelia infections were determined from the trapped rodents and collected ticks. Borrelia burgdorferi s.l. and B. miyamotoi were found in ticks as well as in rodents. Rodent density was higher in 2014, whereas tick burden as well as the Borrelia infection rates in rodents were higher in 2013. The density of B. miyamotoi-infected nymphs did not differ between the years. Tick burdens were higher on Apodemus sylvaticus than on Myodes glareolus, and higher on males than on females. Borrelia-infection rate of rodents varied strongly seasonally, peaking in summer. As the larval tick burden also peaked in summer, the generation of infected nymphs was highest in summer. We conclude that the heterogeneity of environmental and host-specific factors affects the seasonal transmission of Borrelia spp., and that these effects act more strongly on horizontally transmitted B. burgdorferi spp. than on the vertically transmitted B. miyamotoi.

Effect of rodent density on tick and tick-borne pathogen populations: consequences for infectious disease risk.

BACKGROUND: Rodents are considered to contribute strongly to the risk of tick-borne diseases by feeding Ixodes ricinus larvae and by acting as amplifying hosts for pathogens. Here, we tested to what extent these two processes depend on rodent density, and for which pathogen species rodents synergistically contribute to the local disease risk, i.e. the density of infected nymphs (DIN). METHODS: In a natural woodland, we manipulated rodent densities in plots of 2500 m2 by either supplementing a critical food source (acorns) or by removing rodents during two years. Untreated plots were used as controls. Collected nymphs and rodent ear biopsies were tested for the presence of seven tick-borne microorganisms. Linear models were used to capture associations between rodents, nymphs, and pathogens. RESULTS: Investigation of data from all plots, irrespective of the treatment, revealed a strong positive association between rodent density and nymphal density, nymphal infection prevalence (NIP) with Borrelia afzelii and Neoehrlichia mikurensis, and hence DIN's of these pathogens in the following year. The NIP, but not the DIN, of the bird-associated Borrelia garinii, decreased with increasing rodent density. The NIPs of Borrelia miyamotoi and Rickettsia helvetica were independent of rodent density, and increasing rodent density moderately increased the DINs. In addition, NIPs of Babesia microti and Spiroplasma ixodetis decreased with increasing rodent density, which had a non-linear association with DINs of these microorganisms. CONCLUSIONS: A positive density dependence for all rodent- and tick-associated tick-borne pathogens was found, despite the observation that some of them decreased in prevalence. The effects on the DINs were variable among microorganisms, more than likely due to contrasts in their biology (including transmission modes, host specificity and transmission efficiency). The strongest associations were found in rodent-associated pathogens that most heavily rely on horizontal transmission. Our results draw attention to the importance of considering transmission mode of a pathogen while developing preventative measures to successfully reduce the burden of disease.

The genetic diversity of Borrelia afzelii is not maintained by the diversity of the rodent hosts.

BACKGROUND: Small mammals are essential in the enzootic cycle of many tick-borne pathogens (TBP). To understand their contribution to the genetic diversity of Borrelia afzelii, the most prevalent TBP in questing Ixodes ricinus, we compared the genetic variants of B. afzelii at three distinct genetic loci. We chose two plasmid loci, dbpA and ospC, and a chromosomal one, IGS. RESULTS: While the larvae that fed on shrews (Sorex sp.) tested negative for B. afzelii, those fed on bank voles (Myodes glareolus) and wood mice (Apodemus sylvaticus) showed high infection prevalences of 0.13 and 0.27, respectively. Despite the high genetic diversity within B. afzelii, there was no difference between wood mice and bank voles in the number and types of B. afzelii haplotypes they transmit. CONCLUSIONS: The genetic diversity in B. afzelii cannot be explained by separate enzootic cycles in wood mice and bank voles.

A Borrelia afzelii Infection Increases Larval Tick Burden on Myodes glareolus (Rodentia: Cricetidae) and Nymphal Body Weight of Ixodes ricinus (Acari: Ixodidae).

Several microorganisms have been shown to manipulate their host or vector to enhance their own transmission. Here we examined whether an infection with Borrelia afzelii affects its transmission between its bank vole (Myodes glareolus, Schreber, 1780) host and tick vector. Captive-bred bank voles were inoculated with B. afzelii or control medium, after which host preference of Ixodes ricinus L. nymphs was determined in a Y-tube olfactometer. Thereafter, infected and uninfected bank voles were placed in a semifield arena containing questing larvae to measure larval tick attachment. Engorged larvae were collected from these bank voles, molted into nymphs, weighed, and analyzed for infection by PCR.Nymphs were attracted to the odors of a bank vole compared to ambient air and preferred the odors of an infected bank vole over that of an uninfected bank vole. In the semifield arena, infected male bank voles had greater larval tick burdens then uninfected males, while similar larval tick burdens were observed on females regardless of infection status. Nymphal ticks that acquired a B. afzelii infection had higher body weight than nymphs that did not acquire an infection regardless of the infection status of the vole. These results show that a B. afzelii infection in bank voles increases larval tick burden and that a B. afzelii infection in larvae increases nymphal body weight. This finding provides novel ecological insights into the enzootic cycle of B. afzelii.

Larvae of Ixodes ricinus transmit Borrelia afzelii and B. miyamotoi to vertebrate hosts.

BACKGROUND: Lyme borreliosis is the most common tick-borne human disease and is caused by Borrelia burgdorferi sensu lato (s.l.). Borrelia miyamotoi, a relapsing fever spirochaete, is transmitted transovarially, whereas this has not been shown for B. burgdorferi (s.l). Therefore, B. burgdorferi (s.l) is considered to cycle from nymphs to larvae through vertebrates. Larvae of Ixodes ricinus are occasionally B. burgdorferi (s.l) infected, but their vector competence has never been studied. METHODS: We challenged 20 laboratory mice with field-collected larvae of I. ricinus. A subset of these larvae was analysed for infections with B. burgdorferi (s.l) and B. miyamotoi. After three to four challenges, mice were sacrificed and skin and spleen samples were analysed for infection by PCR and culture. RESULTS: Field-collected larvae were naturally infected with B. burgdorferi (s.l) (0.62%) and B. miyamotoi (2.0%). Two mice acquired a B. afzelii infection and four mice acquired a B. miyamotoi infection during the larval challenges. CONCLUSION: We showed that larvae of I. ricinus transmit B. afzelii and B. miyamotoi to rodents and calculated that rodents have a considerable chance of acquiring infections from larvae compared to nymphs. As a result, B. afzelii can cycle between larvae through rodents. Our findings further imply that larval bites on humans, which easily go unnoticed, can cause Lyme borreliosis and Borrelia miyamotoi disease.

Multi-trophic interactions driving the transmission cycle of Borrelia afzelii between Ixodes ricinus and rodents: a review.

The tick Ixodes ricinus is the main vector of the spirochaete Borrelia burgdorferi sensu lato, the causal agent of Lyme borreliosis, in the western Palearctic. Rodents are the reservoir host of B. afzelii, which can be transmitted to I. ricinus larvae during a blood meal. The infected engorged larvae moult into infected nymphs, which can transmit the spirochaetes to rodents and humans. Interestingly, even though only about 1% of the larvae develop into a borreliae-infected nymph, the enzootic borreliae lifecycle can persist. The development from larva to infected nymph is a key aspect in this lifecycle, influencing the density of infected nymphs and thereby Lyme borreliosis risk. The density of infected nymphs varies temporally and geographically and is influenced by multi-trophic (tick-host-borreliae) interactions. For example, blood feeding success of ticks and spirochaete transmission success differ between rodent species and host-finding success appears to be affected by a B. afzelii infection in both the rodent and the tick. In this paper, we review the major interactions between I. ricinus, rodents and B. afzelii that influence this development, with the aim to elucidate the critical factors that determine the epidemiological risk of Lyme borreliosis. The effects of the tick, rodent and B. afzelii on larval host finding, larval blood feeding, spirochaete transmission from rodent to larva and development from larva to nymph are discussed. Nymphal host finding, nymphal blood feeding and spirochaete transmission from nymph to rodent are the final steps to complete the enzootic B. afzelii lifecycle and are included in the review. It is concluded that rodent density, rodent infection prevalence, and tick burden are the major factors affecting the development from larva to infected nymph and that these interact with each other. We suggest that the B. afzelii lifecycle is dependent on the aggregation of ticks among rodents, which is manipulated by the pathogen itself. Better understanding of the processes involved in the development and aggregation of ticks results in more precise estimates of the density of infected nymphs, and hence predictions of Lyme borreliosis risk.

Circulation of four Anaplasma phagocytophilum ecotypes in Europe.

BACKGROUND: Anaplasma phagocytophilum is the etiological agent of granulocytic anaplasmosis in humans and animals. Wild animals and ticks play key roles in the enzootic cycles of the pathogen. Potential ecotypes of A. phagocytophilum have been characterized genetically, but their host range, zoonotic potential and transmission dynamics has only incompletely been resolved. METHODS: The presence of A. phagocytophilum DNA was determined in more than 6000 ixodid ticks collected from the vegetation and wildlife, in 289 tissue samples from wild and domestic animals, and 69 keds collected from deer, originating from various geographic locations in The Netherlands and Belgium. From the qPCR-positive lysates, a fragment of the groEL-gene was amplified and sequenced. Additional groEL sequences from ticks and animals from Europe were obtained from GenBank, and sequences from human cases were obtained through literature searches. Statistical analyses were performed to identify A. phagocytophilum ecotypes, to assess their host range and their zoonotic potential. The population dynamics of A. phagocytophilum ecotypes was investigated using population genetic analyses. RESULTS: DNA of A. phagocytophilum was present in all stages of questing and feeding Ixodes ricinus, feeding I. hexagonus, I. frontalis, I. trianguliceps, and deer keds, but was absent in questing I. arboricola and Dermacentor reticulatus. DNA of A. phagocytophilum was present in feeding ticks and tissues from many vertebrates, including roe deer, mouflon, red foxes, wild boar, sheep and hedgehogs but was rarely found in rodents and birds and was absent in badgers and lizards. Four geographically dispersed A. phagocytophilum ecotypes were identified, that had significantly different host ranges. All sequences from human cases belonged to only one of these ecotypes. Based on population genetic parameters, the potentially zoonotic ecotype showed significant expansion. CONCLUSION: Four ecotypes of A. phagocytophilum with differential enzootic cycles were identified. So far, all human cases clustered in only one of these ecotypes. The zoonotic ecotype has the broadest range of wildlife hosts. The expansion of the zoonotic A. phagocytophilum ecotype indicates a recent increase of the acarological risk of exposure of humans and animals.