Many vector-borne pathogens consist of multiple strains that circulate in both the vertebrate host and the arthropod vector. Characterization of the community of pathogen strains in the arthropod vector is therefore important for understanding the epidemiology of mixed vector-borne infections. Borrelia afzelii and B. garinii are two species of tick-borne bacteria that cause Lyme disease in humans. These two sympatric pathogens use the same tick, Ixodes ricinus , but are adapted to different classes of vertebrate hosts. Both Borrelia species consist of multiple strains that are classified using the highly polymorphic ospC gene. Vertebrate cross-immunity against the OspC antigen is predicted to structure the community of multiple-strain Borrelia pathogens. Borrelia isolates were cultured from field-collected I. ricinus ticks over a period spanning 11 years. The Borrelia species of each isolate was identified using a reverse line blot (RLB) assay. Deep sequencing was used to characterize the ospC communities of 190 B. afzelii isolates and 193 B. garinii isolates. Infections with multiple ospC strains were common in ticks, but vertebrate cross-immunity did not influence the strain structure in the tick vector. The pattern of genetic variation at the ospC locus suggested that vertebrate cross-immunity exerts strong selection against intermediately divergent ospC alleles. Deep sequencing found that more than 50% of our isolates contained exotic ospC alleles derived from other Borrelia species. Two alternative explanations for these exotic ospC alleles are cryptic coinfections that were not detected by the RLB assay or horizontal transfer of the ospC gene between Borrelia species.

Mixed or multiple-strain infections are common in vector-borne diseases and have important implications for the epidemiology of these pathogens. Previous studies have mainly focused on interactions between pathogen strains in the vertebrate host, but little is known about what happens in the arthropod vector. Borrelia afzelii and Borrelia garinii are two species of spirochete bacteria that cause Lyme borreliosis in Europe and that share a tick vector, Ixodes ricinus . Each of these two tick-borne pathogens consists of multiple strains that are often differentiated using the highly polymorphic ospC gene. For each Borrelia species, we studied the frequencies and abundances of the ospC strains in a wild population of I. ricinus ticks that had been sampled from the same field site over a period of 3 years. We used quantitative PCR (qPCR) and 454 sequencing to estimate the spirochete load and the strain diversity within each tick. For B. afzelii , there was a negative relationship between the two most common ospC strains, suggesting the presence of competitive interactions in the vertebrate host and possibly the tick vector. The flat relationship between total spirochete abundance and strain richness in the nymphal tick indicates that the mean abundance per strain decreases as the number of strains in the tick increases. Strains with the highest spirochete load in the nymphal tick were the most common strains in the tick population. The spirochete abundance in the nymphal tick appears to be an important life history trait that explains why some strains are more common than others in nature. IMPORTANCE Lyme borreliosis is the most common vector-borne disease in the Northern Hemisphere and is caused by spirochete bacteria that belong to the Borrelia burgdorferi sensu lato species complex. These tick-borne pathogens are transmitted among vertebrate hosts by hard ticks of the genus Ixodes . Each Borrelia species can be further subdivided into genetically distinct strains. Multiple-strain infections are common in both the vertebrate host and the tick vector and can result in competitive interactions. To date, few studies on multiple-strain vector-borne pathogens have investigated patterns of cooccurrence and abundance in the arthropod vector. We demonstrate that the abundance of a given strain in the tick vector is negatively affected by the presence of coinfecting strains. In addition, our study suggests that the spirochete abundance in the tick is an important life history trait that can explain why some strains are more common in nature than others.

Background: Lyme borreliosis (LB) is the most widespread hard tick-borne zoonosis in the northern hemisphere. Existing studies in Europe have focused mainly on acarological risk assessment, with few investigations exploring human LB occurrence. Aim: We explored the determinants of spatial and seasonal LB variations in France from 2016 to 2021 by integrating environmental, animal, meteorological and anthropogenic factors, and then mapped seasonal LB risk predictions. Methods: We fitted 2016–19 LB national surveillance data to a two-part spatio-temporal statistical model. Spatial and temporal random effects were specified using a Besag-York-Mollie model and a seasonal model, respectively. Coefficients were estimated in a Bayesian framework using integrated nested Laplace approximation. Data from 2020–21 were used for model validation. Results: A high vegetation index (≥ 0.6) was positively associated with seasonal LB presence, while the index of deer presence (> 60%), mild soil temperature (15–22 °C), moderate air saturation deficit (1.5–5 mmHg) and higher tick bite frequency were associated with increased incidence. Prediction maps show a higher risk of LB in spring and summer (April–September), with higher incidence in parts of eastern, midwestern and south-western France. Conclusion: We present a national level spatial assessment of seasonal LB occurrence in Europe, disentangling factors associated with the presence and increased incidence of LB. Our findings yield quantitative evidence for national public health agencies to plan targeted prevention campaigns to reduce LB burden, enhance surveillance and identify further data needs. This approach can be tested in other LB endemic areas.

Multiple-strain pathogens often establish mixed infections inside the host that result in competition between strains. In vector-borne pathogens, the competitive ability of strains must be measured in both the vertebrate host and the arthropod vector to understand the outcome of competition. Such studies could reveal the existence of trade-offs in competitive ability between different host types. We used the tick-borne bacteriumBorrelia afzeliito test for competition between strains in the rodent host and the tick vector, and to test for a trade-off in competitive ability between these two host types. Mice were infected via tick bite with either one or two strains, and these mice were subsequently used to create ticks with single or mixed infections. Competition in the rodent host reduced strain-specific host-to-tick transmission and competition in the tick vector reduced the abundance of both strains. The strain that was competitively superior in host-to-tick transmission was competitively inferior with respect to bacterial abundance in the tick. This study suggests that in multiple-strain vector-borne pathogens there are trade-offs in competitive ability between the vertebrate host and the arthropod vector. Such trade-offs could play an important role in the coexistence of pathogen strains.

Alteration in anti-predatory behaviour following geographic isolation has been observed in a number of taxa. Such alteration was attributed to the effect of relaxed selection in the novel environment, reinforced by the cost of anti-predatory behaviours. We studied aspects of anti-snake behaviour in 987 adult and juvenile wall lizards Podarcis muralis from two mainland areas (heavy snake predatory pressure) and two islands (low snake predatory pressure), isolated from the mainland 5000 and 7000 years ago. We conducted a scented retreat site choice experiment using the odours of five different snake species (saurophagous, piscivorous or generalist feeder). Mainland lizards avoided shelters scented by saurophagous snakes, but not those scented by non saurophagous snake species. Long isolated lizards (7000 years ago) showed no anti-predator response to any snake, suggesting a total loss of anti-predatory behaviour towards saurophagous snakes. More recently isolated lizards (5000 years ago) however showed anti-snake behaviour towards a former sympatric adder species, and a tendency to avoid the scent of a sympatric generalist feeder snake. There was no difference in the anti-snake responses between adult and juvenile wall lizards from all four sites, suggesting a limited role for experience (behavioural plasticity) in the expression of anti-snake behaviour in wall lizards.