An “unoccupied niche” is susceptible to colonization, with or without human assistance, and identifying that niche leads to predictions about which organisms will invade and how they will spread. One prediction, for example, is that a species having “a broader niche” will be more invasive than another with “a narrower niche” (Vazquez, 2006). The hypothesis that a “broader” niche allows the organism to display a more extensive set of adaptations to varying ecological conditions, so that a larger geographic area will be potentially available to it (Slatyer et al., 2013). The idea that invasiveness is correlated with overall “niche breadth” can be tested at a simple level; the hypothesis is not universally validated by evidence but has received strong support in some cases (e.g., Sol, 2016; Granot et al., 2017). Unfortunately, however, it is not self-evident what is meant by overall “niche breadth” nor how it can be measured or compared. The concept of the Hutchinsonian niche with its multiple dimensions helps to visualize the niche's complexity, but since each dimension varies in breadth, possibly but not necessarily independently of all other dimensions, it is often not clear how the various dimensions can be summed to give a single measure of niche breadth. For example, what units should be used to describe a niche's dimensions? It is thus difficult to characterize the overall “breadth” of a niche in a simple way.

A further problem is that niche breadth may be maintained through evolutionary adaptation to more than one niche dimension, in which case, local populations of a species may each be separately adapted to local conditions, so that “niche breadth” is not actually a characteristic of the genome of the species in general at all, but a reflection of its genetic diversity. As a consequence, an invasion into a new geographic region would most likely be achieved by migration from only a suitably pre-adapted subset of the original species and this might be expected to leave the signature of a selective sweep in the genome of the invading population. Alternatively, the original niche breadth of a species may be the result of phenotypic plasticity, a situation that would be likely to favor invasiveness without genetic change. Such phenotypic plasticity is likely to be present in species that originally inhabit rapidly fluctuating environments (Lee and Gelembiuk, 2008).

Finally, it is simplistic to suppose that the existence of a correlation between niche breadth and geographic range size necessarily implies a causal relationship in the direction of the former to the latter. It may also be the case that larger geographic range actually drives adaptation to greater variety of niches (Jahner et al., 2011).

The above discussion indicates that there are problems in using niche theory to understand invasiveness (i.e., the tendency to invasion-like geographic range expansion when the opportunity for this arises). Parasites, however, offer a way out of these difficulties. Obligate vector-borne parasites live their entire lives within their hosts and vectors and do not occur free in the environment. Niches occupied by such parasites are therefore almost all entirely associated with selective forces exerted by (or at least mediated through) their vectors and hosts. Because related parasites tend to have vectors and hosts that share many characteristics, and can thus be measured in the same way, parasites offer an exceptionally favorable opportunity to test niche-related ideas about invasiveness; we have a much better chance of realistically comparing “niche breadth” between related parasites. We might say that a broader parasitic niche would be one that has a larger number of dimensions (i.e., corresponding to a larger number of hosts/vectors) or dimensions that are spatially more extensive (i.e., the geographic ranges of their hosts/vectors are larger). The precision with which the parasite's niche is described might be increased by considering each of the different physiological or biochemical interactions between the parasite and its host (for example the various immune defenses deployed by the host, and associated countermeasures used by the parasite) to be a separate dimension of the niche. As far as we are aware, however, the hypothesis that the geographical invasiveness of obligate parasites is related to the breadth of host- and/or vector-associated niche space has not been tested (e.g., Vazquez, 2006; Slatyer et al., 2013).

On the other hand, parasites co-evolve with hosts and vectors (Woolhouse et al., 2002; Wade, 2007; Toft and Andersson, 2010), because parasites exert strong selective pressures on host(s) (and vectors) which in turn evolve to reject the parasite, effectively narrowing the niche available to the parasite. Much of the program of parasitological research is concerned with identifying such co-evolutionary adaptations (Penczykowski et al., 2016). Co-evolution is notoriously tricky to study, because it is a system that is subject to positive feedback. It is often found that co-evolution is local, rapid and episodic, affecting several different gene pairs that coevolve episodically and occur unpredictably in time and space (this is Thompson's “geographic mosaic” model; Thompson, 1999).

Another aspect of niche theory that is relevant here is that the fundamental niche of an organism, which extends to the furthest limits of all dimensions that can be tolerated by that species, is almost always greater in extent than the realized niche, which is that part of the niche that is actually occupied. The greater the mismatch between the realized and fundamental niche, the greater is the potential for invasion (Soberon and Arroyo-Pena, 2017). It is obvious that the extent to which the size of the realized niche approaches that of the fundamental niche is subject to limitations due to local physical environmental conditions; for example, if it is too hot, too cold, too wet, or too dry. In the case of a parasite, this kind of environmental limitation may be mediated through biotic interactions (because of their own physical limitations, hosts, and vectors are not present everywhere). It is less intuitively obvious that in the case of parasites, biotic limitations may not be due to unsuitable environmental conditions, but due to phylogeographic contingency: any one of a set of potentially interacting parasites, hosts and vectors may be able to persist in a particular geographic zone, but is nevertheless presently absent simply because the organism in question has not yet expanded its range to occupy all of its fundamental niche. This absence of either the host or the vector may then limit the occurrence of the parasite.

Such a mismatch between the fundamental and realized niche can be corrected by geographic range expansion. Considering this from the point of view of the parasite, such realization of potential niche space can happen in two ways: first the parasite may invade a new environment when an existing host or vector expands its own range, taking the parasite with it. In this case, successful range expansion of the parasite depends completely on the invasiveness of the existing host or vector. Alternatively, a parasite may start utilizing a new species as host. This parasite-host combination might not have been utilized previously simply because that host species did not co-occur with the parasite in its existing range. Removal of a former landscape or climatic barrier to geographic range expansion of that host or vector may allow the parasite to expand its geographic range within what was actually always the fundamental niche simply because it can now exploit a new host or vector in this previously unexploited environment. For obligate parasites, the opportunity to enter the new geographic zone will likely depend on being transported there within an existing host or vector, but once it is there its success may depend on a switch in its host(s) or vector(s). An example of this is the invasion of European fresh-water habitats by the fungal parasite Aphanomyces astaci, a parasite of the introduced North American signal crayfish, Pacifastacus leniusculus. As a result, the parasite has increased the extent of its realized niche to include the European crayfish host Austropotamobius pallipes, which is now revealed as a part of its fundamental niche that was previously only latent (Edgerton et al., 2004; Vrålstad et al., 2011).

The increase in size of the realized niche by geographic range expansion to occupy a greater fraction of the fundamental niche necessarily involves an increase in population size. This is an ecological process that need not involve evolutionary change at all. The implication is that once an existing barrier to exploitation within one or more existing niche dimensions has been removed, population dynamic feedbacks will allow population expansion to occur; density-dependent limitations will be temporarily reduced and allow an increase in the number of surviving progeny. As the fraction of the fundamental niche that is actually realized increases, these feedbacks will act to slow and eventually stop further population growth. By contrast, an invading species may expand its geographic range (i.e., invade a new adaptive zone) because of some genetically based adaptive change (in the case of a parasite this might be the ability to exploit a new host or vector). In this case we see a change in the extent of the fundamental niche, which may or may not be accompanied by a change in the proportion of the fundamental niche that is actually realized.

In the latter case, it is appropriate to ask what sets limits to the size (geographic extent) of the fundamental niche? Why can parasites not simply adapt to colonize more and more kinds of host or vector? Of course, the answer is that there are trade-offs to the acquisition of new adaptations. In the case of a parasite, the fitness advantage that accrues as a result of the ability to utilize a new host or vector species may be offset by a loss of fitness that results from a decreased ability to exploit existing hosts or vectors.

The dependence of Borrelia on both vector and reservoir hosts determines the geographic distribution of the Lyme borreliosis group of spirochetes. With few exceptions such as B. chilensis (Ivanova et al., 2014), most B. burgdorferi s.l. species are found between 40 and 60° latitude where Lyme borreliosis is reported to occur in humans (Figure 2).

The geographic ranges of the various B. burgdorferi s.l. species (Margos et al., 2011) are in each case limited to those locations in which both reservoir hosts and vector ticks are able to maintain natural transmission cycles (Kurtenbach et al., 2006, 2010) and thus we should be able to define the fundamental niche of each Borrelia species simply by taking account of where its vectors and hosts occur. So long as at least one associated vector and one associated host are present in a particular geographic zone, a Borrelia spirochete utilizing them could theoretically be maintained there. But because most B. burgdorferi s.l. species can utilize multiple vertebrate host species and a number can utilize more than one vector, and because the ecological associations between borreliae, ticks, and reservoir hosts are not all equivalent in strength, the realized niche actually achieved by each B. burgdorferi s.l. species will almost always in practice be considerably less than the simple overlap of the geographic measures (Tsao, 2009). The actual spatial limitation for each spirochete species (i.e., its realized niche) will be roughly equivalent to the sum of all those areas in which both at least one vector species and one host species occur at sufficiently high density to maintain its transmission cycle. In more formal terms, the value of the basic reproduction number R₀ for every local population of the parasite, summed over all its hosts and vectors, must exceed unity (Randolph, 1998; Hartemink et al., 2008; Tsao, 2009). The relative contributions to the overall R₀ of particular host and vector associations may of course vary between particular locations within its distribution.

Importantly, however, we cannot simply add up values of R₀ that have been determined for each vector and each host under laboratory conditions, because the presence of less efficient vectors and hosts will impact negatively on the value of R₀ achieved by the “best” vectors and hosts (Randolph and Craine, 1995; Ostfeld et al., 2006). These negative effects may be very important. Some potential mammalian hosts (mostly large animals such as ruminants) can be colonized by B. burgdorferi s.l. spirochetes when they are bitten by an infected tick vector, but are completely non-permissive when it comes to transmission of the bacteria to a new tick. In fact, feeding on a deer may actually clear a Borrelia infection in a tick (Telford et al., 1988; Kurtenbach et al., 1998). The presence of such “dead end hosts” can influence the parasite's success in diametrically opposite ways. On one hand, the presence of deer in a particular geographic zone may permit the population density of the vector ticks that transmit Borrelia to increase; this increases the likelihood of successful transmission of spirochetes from infected reservoir hosts to ticks and thus increases R₀. On the other hand, the presence of large numbers of deer may drive ticks to feed with greater probability on deer than on small mammals and this may actually suppress the spirochete infection rate of true reservoir hosts in that location (reviewed by Ogden and Tsao, 2009; Kilpatrick et al., 2017).

Furthermore, the application of multilocus sequence typing to B. burgdorferi sensu stricto in combination with landscape ecology has shown that variation in host availability or landscape connectivity can strongly impact the spatial pattern of population distribution (Hoen et al., 2009; Margos et al., 2012b; Mechai et al., 2018). In the late nineteenth century, human-induced changes to habitats and severe hunting in the northeastern USA almost led to regional extinction of deer, which are important hosts for tick survival (Barbour and Fish, 1993). As a consequence of a likely decline of vector populations, Borrelia populations were reduced to spatially divided refuge populations in Northeast and Midwest. Independent expansion of these populations correlated with reintroduction of hosts for vector and Borrelia as well as environmental ecological changes such as reforestation (Hoen et al., 2009). Other population boundaries correlated with landscape barriers such as the Great Plains and/or the Rocky Mountains (Margos et al., 2012b). In a more recent study attempting to understand spatial spread pattern of B. burgdorferi s.s. into a zone of emerging Lyme borreliosis risk, the importance of landscape connectivity for the spatial distribution of B. burgdorferi s.s. of certain sequence types was underlined (Mechai et al., 2018).

Apart from host associations that impact the distribution of Borrelia species and populations, the non-uniform distribution pattern of Borrelia genospecies observed in field studies suggests that vector associations (based on differential vector competence) do indeed play an important role in limiting their geographic distribution ranges (Figure 3; Margos et al., 2011). Evidence for this is provided by the ability of some Borrelia species to utilize a wide range of vectors (Masuzawa, 2004; Margos et al., 2012a), for example B. garinii can be vectored by I. persulcatus, I.pavlovskyi, I. ricinus, and I. uriae. Accordingly, B. garinii's geographic distribution ranges from Japan to France and to sea bird colonies in both the Northern and Southern Hemispheres (Figure 2). B. garinii has been found in sea bird colonies in Newfoundland (Munro et al., 2017) but has not been discovered in I. scapularis dominated regions or in I. pacificus (Hamer et al., 2007; Hoen et al., 2009; Ogden et al., 2010; Mechai et al., 2013; Fedorova et al., 2014). Similarly, B. burgdorferi s.s. is able to utilize I. scapularis, I. pacificus, I. spinipalpis, and I. affinis as vectors in North America (Burgdorfer et al., 1982; Maupin et al., 1994; Clover and Lane, 1995; Maggi et al., 2010), as well as I. ricinus in Europe (Rauter and Hartung, 2005; Sertour et al., 2018), but has not been found in I. persulcatus (Korenberg et al., 2002; Mukhacheva and Kovalev, 2013). B. valaisiana, a bird adapted Borrelia species, is frequently found in Europe associated with I. ricinus but only a single occurrence in Russia has been recorded (Kurilshikov et al., 2014) suggesting that I. persulcatus is not a competent vector. Consistent with this, in the overlapping zone of I. ricinus and I. persulcatus, the prevalence of B. valaisiana is higher in I. ricinus and drops to much lower values in I. persulcatus (Bormane et al., 2004).

Previous investigations using RFLP analysis had suggested that strains of ospA type 4 (i.e., Western European B. bavariensis) were genetically highly homogenous (Marconi et al., 1999), a finding that was confirmed using MLSA (Margos et al., 2009) and NGS (Gatzmann et al., 2015). The presence of extensive genetic variation within Asian strains of B. bavariensis (comparable in extent to that within B. garinii itself; Figure 4) implies that the split between B. garinii and B. bavariensis took place relatively long ago, although clearly more recently than the original radiation of the B. burgdorferi s.l. complex as a whole (Becker et al., 2016). The invasion-like range expansion of B. bavariensis into Europe, on the other hand, appears to have happened much more recently, and to have involved a selective sweep that imposed a severe bottleneck on genetic variation in the genome of the surviving European invaders (Figure 4). Since Asian B. bavariensis are associated only with I. persulcatus, a tick species which does not occur in Western Europe, while European B. bavariensis are associated solely with the European species, I. ricinus (Postic et al., 1997; Korenberg et al., 2002; Masuzawa et al., 2005; Fingerle et al., 2008; Takano et al., 2011; Geller et al., 2013; Mukhacheva and Kovalev, 2013, 2014; Scholz et al., 2013), it seems likely that colonization of Europe by the spirochete must have been coincident with, or driven by, a switch in its vector association.

Importantly, the genetic homogeneity of European B. bavariensis, the close genetic relationship of European and Asian B. bavariensis and the recent divergence from B. garinii have all now been confirmed by data derived from next generation sequencing and analyses of the core genome, the main chromosome and the two plasmids lp54 and cp26 (Gatzmann et al., 2015; Becker et al., 2016). In European B. bavariensis no more than 400 single nucleotide polymorphisms (SNPs) were found on the main chromosome between the most divergent isolates while approximately 10 times more chromosomal SNPs were noticed in Asian B. bavariensis, giving rise to the deep branching pattern seen in the phylogeny of the latter population (Figure 4; Gatzmann et al., 2015).

Analyses of the core genome (chromosome, lp54, and cp26) of European and Japanese B. bavariensis revealed genes that were under positive selection in the European population (Gatzmann et al., 2015). Among those were two genes flhB (BG0275) and antigen S1 (BGA04) (a homolog of BBA05) which had previously been shown to be up-regulated under conditions that simulate vector conditions (Ojaimi et al., 2003) or in feeding nymphs (Xu et al., 2010). The results presented by Gatzmann et al. (2015) were based on short-read NGS data which did not allow complete assembly of all plasmids in B. bavariensis and thus our knowledge of the comparative genomics of plasmid encoded genes in this species remains incomplete. This is an avenue for future consideration.

In this context it is important to emphasize that there is so far no direct experimental evidence on the vector competence of different tick species for the different B. bavariensis populations. However, strong support for differential vector adaptation of European vs. Asian B. bavariensis populations comes from field studies. The occurrence of B. burgdorferi s.l. spirochetes in ticks from the European and Asian parts of Russia has been studied since the early 1980s, soon after the description of B. burgdorferi s.s. (Korenberg et al., 2002). Ixodes ricinus, I. persulcatus, and I. trianguliceps ticks were used to culture Borrelia. Intriguingly, B. bavariensis (then designated as B. garinii) NT29-type strains were only ever isolated from I. persulcatus ticks and never from I. ricinus ticks, while B. afzelii and bird associated B. garinii were found in both tick species. Postic et al. (1997) concluded from this work that NT29-type strains (now correctly designated as Asian B. bavariensis) are associated only with I. persulcatus. Remarkably, in terms of vector adaptation, B. burgdorferi s.s., B. lusitaniae, and B. valaisiana were also found only in I. ricinus, and not in I. persulcatus (Korenberg et al., 2002). Subsequent molecular studies conducted in Asia, Mongolia, Russia and Siberia all found only Asian type B. bavariensis in I. persulcatus (Masuzawa et al., 2005; Takano et al., 2011; Mukhacheva and Kovalev, 2013, 2014; Scholz et al., 2013). The most compelling evidence yet for the contrasting vector association/adaptations of the European and Asian populations of B. bavariensis has now been provided by a study conducted in the overlapping zone of I. ricinus and I. persulcatus in Estonia (Geller et al., 2013). These authors reported that Asian type B. bavariensis was associated only with I. persulcatus, and not with I. ricinus. Moreover, a European type of B. bavariensis has recently been found in I. ricinus in Latvia (Mercy Okeyo, NRZ Borrelia, personal communication).

Although several Borrelia genes have been identified that are important for persistence of Borrelia in the tick or for receptor interaction, how these genes differ between species and which genes allow different Borrelia species to utilize certain tick vectors is unknown. To further characterize such genes and to pinpoint their involvement in differential vector compatibility will require comparative genomics studies of Borrelia species that utilize different tick species as vector. Gene upregulation in Borrelia during or following transmission to the vector has been demonstrated and several genes have been shown to be crucial for transmission to, or survival of, Borrelia in the vector (e.g., Pal and Fikrig, 2010; Radolf et al., 2012). A comprehensive study showed that 246 gene transcripts were upregulated and transcribed in Borrelia during or after a blood meal was taken by ticks; 88 of these were present in Borrelia taken up by both, larvae and nymphs, while 17 Borrelia transcripts were only found in Borrelia taken up by larvae during blood feeding (Iyer et al., 2015). Although transcription does not equal presence of proteins, this study does suggest that some of these gene products are likely to be important for survival of Borrelia in the vector. In addition, studies on the gene expression regulation system RpoS identified 41 genes that are repressed in B. burgdorferi in vitro and under conditions representing the vertebrate host. Although a good number encoded hypothetical proteins, many of the identified genes were known to be important for survival in, and interaction with, the vector (Caimano et al., 2019). Whether variations in these molecules contribute to differential vector competence needs to be further investigated.

Some molecules that have been shown to have a function within the tick vector include the Outer surface protein A (OspA) which binds to the vector midgut molecule TROSPA (Pal et al., 2004); BBE31 which binds to the vector molecule Tre31, a protein secreted by tick midgut epithelial cells (Zhang et al., 2011); PdeB, a phosphodiesterase required for survival in, and transmission by, ticks; Lpm1, a surface molecule, depletion of which inhibits Borrelia in the vector (Koci et al., 2018); BB0365, BB0690, BptA, lp6.6, BBk32 are all required for persistence in the tick, while antigen P35 (BBA64) may play a role in tick-to-host transmission (Fikrig et al., 2000; Radolf et al., 2012; Brandt et al., 2014). In a recent study using transposon-based gene interruption approach, >100 genes were shown to be involved in Borrelia survival in larval ticks (Phelan et al., 2019). These included previously reported genes and novel genes important for tick survival of Borrelia. For 46 of these genes complete absence of transmission was observed after knockout, while for 56 other genes a drastic reduction (90%) of transmission occurred. Of the genes inducing complete interruption of transmission 22% were predicted lipoproteins, but for 41% was no predicted function.

From the vector's point of view, it is likely that physiological differences between tick species have an impact on vector competence. Generally, Ixodes species are three-host ticks and as such, both biotic and abiotic factors influence where they can survive. On-host feeding periods are short compared to off-host periods, which include periods of diapause (genetically programmed) or quiescence (due to unfavorable conditions). Abiotic factors that influence tick physiology include temperature, humidity, and day-length, all of which have an impact on host seeking phenology, success of egg hatching, overwintering of immature and adult stages. During off-host periods, hot conditions may result in overheating, freezing in sub-zero temperatures, or dehydration in dry conditions. Such conditions require ticks to seek refuges for cooling down or re-hydration and appropriate (micro-)habitats with suitable vegetation or leaf litter layer therefore play an important role in the tick's life cycle (Medlock et al., 2013; Eisen et al., 2016). Ixodes ricinus and I. persulcatus overlap in a region ranging from Estonia to Russia and, although some fluctuation of populations may occur (Filipova, 2017), the general geographic range distribution of the two tick species persists. Differences in host seeking phenology due to differences in tick physiology may affect feeding success in such overlapping zones and it is conceivable that this has an influence on vector competence (Korenberg et al., 1991, 2002; Korenberg, 1995; Alekseev et al., 1998). An impressive example of changed host-seeking phenology was reported for I. scapularis by Gatewood et al. (2009). Ixodes scapularis populations in Midwestern regions of the USA showed more seasonal synchrony of nymphal and larval feeding pattern. Investigations of B. burgdorferi s.s. infection pattern revealed that lineages which carried the 16S-23S rRNA intergenic spacer restriction fragment length polymorphism sequence type 1 were more frequently found in northeastern Ixodes populations (Gatewood et al., 2009). There are several excellent reviews on these topics (Lindgren and Jaenson, 2006; Ogden et al., 2006, 2013b; Medlock et al., 2013; Eisen et al., 2016) and it would go beyond the scope of this review to expand further on this.

An attractive hypothesis is that tick immunity plays a role in differential vector competence. In recent years progress has been made in recognizing the complexity of the tick's immune system (reviewed in Hajdusek et al., 2013; Kitsou and Pal, 2018) and it has been shown that Ixodes spp. possess a number of immune effectors and modulators such as recognition molecules, among them thioester-containing proteins (T-TEPS, that serve as lectins labeling cells for immune attack), phagocytotic hemocytes, defensins, lysozymes, antimicrobial peptides, a dityrosine network (DTN), and (atypical) signaling pathways. The signaling pathways regulating the immune system include Toll, IMD (Immunodeficiency) and JAK-STAT (Janus Kinase/Signal Transducers and Activators of Transcription) as well as an indirect, cross-species signaling pathway that recognizes the cytokine interferon gamma in the blood of the vertebrate host (Johns et al., 2001; Hajdusek et al., 2013; Yang et al., 2014; Sonenshine and Macaluso, 2017; Urbanova et al., 2017). RNA interference studies on genes involved in the tick's immune response have shown that depletion of expression may lead to suppression of Borrelia colonization in ticks (Narasimhan et al., 2017) suggesting that Borrelia can exploit tick molecules to its own advantage. In addition, the induction by Borrelia in I. scapularis of a protein with a Reeler domain (PIXR) limits bacterial biofilm formation in the tick's gut, thereby preventing alterations in the microbiome and promoting colonization by Borrelia (Narasimhan et al., 2017). Thus, it is highly likely that immune effectors play an important role in determining the competence of Ixodes species for Borrelia species and vice versa.

In the past decade much attention has been invested to study the tick's microbiome in more detail. These studies have shown that the microbiome of ticks consists of microorganisms associated with the outer surface of ticks, the gut microbiome, and endosymbiontic bacteria (reviewed by Narasimhan and Fikrig, 2015). Initial studies on different Ixodes species (e.g., I. scapurlaris, I. ricinus, I. pacificus, and I. persulcatus) using high throughput sequencing methods have discovered a whole range of bacterial taxa that are associated with ticks. Some studies found differences in bacterial genera contributing to the tick microbiome between geographical locations, tick species and stages (nymphs, female, and males) (Van Treuren et al., 2015), but others did not (Clow et al., 2018). In most studies bacterial genera were found that constitute known tick symbionts like Coxiella, Midichloria, Francisella, Wolbachia, Cardinium, Arsenophonus, Spiroplasma, Rickettsia, Rickettsiella, and Lariskella (Benson et al., 2004; Carpi et al., 2011; Abraham et al., 2017; Bonnet et al., 2017; Mukhacheva and Kovalev, 2017; Swei and Kwan, 2017). Recently in a more refined approach, dissected tick tissues of questing I. scapularis were used to determine what constitutes the “internal” microbiome as opposed to the “surface” microbiome (Ross et al., 2018). This showed that the gut microbiome of I. scapularis was of limited diversity in the majority of adults being dominated by Rickettsia and B. burgdorferi s.s. Only a minority of tick samples contained high microbiome diversity, harboring bacteria of the genera Bacillus and Pseudomonas, and the family Enterobacteriaceae in their midguts.

The distribution of microbial symbionts, in hybrid ticks in the overlapping zone of I. ricinus and I. persulcatus was very interesting. The endosymbiont Midichloria mitochondrii (Sassera et al., 2006) was found to 100% in female I. ricinus ticks. A different endosymbiont, Lariskella arthropodarum, was identified in I. persulcatus (Mediannikov et al., 2004; Mukhacheva and Kovalev, 2017). Hybrid progeny of these ticks harbored only the respective symbionts of their mothers underpinning the specificity of association (Mukhacheva and Kovalev, 2017).

As pointed out below, hybrid offspring of different tick vector species such as I. ricinus and I. persulcatus in regions where these species occur in sympatry may offer an opportunity for vector switching. Such hybrid ticks will have genetic inheritance from both parents and receptor molecules in the midgut or molecules of the immune system may phenotypically represent both parents, which we speculate might allow the survival of an invading Borrelia species and facilitate its adaptation to a new vector.

Our first suggestion is that the vector-switch required a mixed infection within a tick between B. bavariensis and some other Borrelia species that was already adapted to use I. ricinus as a vector.

As noted above, different Borrelia species can be found in the same Ixodes tick in mixed infections; the different spirochetes may have been acquired by the tick together at the same time when feeding from a multiply-parasitized host. Whilst standard niche theory would predict that competition between the parasite species would occur in such a scenario, it might also be envisaged that both species may benefit from the secretion by just one of the two species of adaptive “public goods” goods (i.e., secreted proteins that are beneficial to both parasites under these conditions). We note that the evolutionary outcomes of such interactions between parasites in multiple infections may be complex (Alizon and Lion, 2011).

Thus, a simple model for Borrelia adaptation to a new vector would be that co-infection of a single tick with a Borrelia species already adapted to transmission by I. ricinus (e.g., B. garinii, B. afzelii) would enable not only that species, but also B. bavariensis to attach to the midgut of the new tick vector, and/or to be protected against immune attack of the new vector, and/or to be successfully transmitted by the new vector to the reservoir host (Figure 6). This would have enabled B. bavariensis to colonize the new vector, despite lacking the necessary adaptive genes within its own genome. This would not in itself have been enough to make a full transition to the new vector. Importantly, however, the opportunity for B. bavariensis to exploit adaptive genes resident in another Borrelia species during mixed infections in this way would also provide an opportunity whereby those same genes could eventually be acquired by the invading Borrelia strain. This could occur via horizontal gene transfer (HGT) from the other co-infecting species of Borrelia. Phylogenomic studies of Borrelia plasmids have revealed that HGT is common for genes on some plasmids (Qiu et al., 2004; Casjens et al., 2012, 2017, 2018).

Although it is likely that several genes and gene products are involved in the stepwise adaptation of Borrelia spirochetes to their journey through the vector tick, it should be remembered that every Borrelia species is already well-adapted to utilize at least one species of tick. It is thus conceivable that only one of these steps would actually be deficient in the interaction between B. bavariensis and I. ricinus, much improving the probability that co-infection with another already-adapted Borrelia species would enable B. bavariensis to colonize and exploit the new vector species. Since the acquisition of a single new gene by HGT could theoretically confer the ability to colonize a new vector species with the help of a co-infecting agent, the probability of the evolution of a complete vector switch would also be enhanced. If the necessary HGT event was rare, then this would explain why all B. bavariensis strains that utilize I. ricinus as a vector also show evidence of a historic selective sweep.

Our second, speculative suggestion is that adaptation to a new vector required the presence of hybrid I. persulcatus-I. ricinus ticks that allowed the parental strain of B. bavariensis to complete its life cycle. Although the coexistence of the two tick vector species in the Baltic/Western Russian zone would have led to the continually repeated introduction of Asian-type B. bavariensis into I. ricinus ticks, these spirochetes may have been unable to survive in the tick, or (just as likely) were unable successfully to enter the tick's salivary glands and thus to be transferred into the small mammal reservoir hosts utilized by I. ricinus.

The two tick species I. ricinus and I. persulcatus have an overlapping zone in Estonia, Latvia and Western Russia (Figure 5; Swanson et al., 2006). In such overlapping zones cross-species mating occurs, leading to hybrid offspring that likely possess phenotypic traits of both parents. Cross-mating has not only observed between I. ricinus and I. persulcatus but also between I. persulcatus and I. pavlovskyi (Balashov Iu et al., 1998; and references therein Kovalev et al., 2015, 2016) and between I. scapularis and I. cookei in North America (Patterson et al., 2017) suggesting that cross-mating between tick species may be more frequent than once thought.

The presence of hybrid ticks might be a predisposing factor for Borrelia vector -switching. In the Eurasian I. ricinus-I. persulcatus hybrid zone, up to 13% of ticks have been found to be hybrids (Kovalev et al., 2016). Such hybrids presumably arise afresh in each generation through cross-species mating, but it is also possible that hybrids might themselves cross with either parental species for a limited number of generations (Kovalev et al., 2016). Although it has been considered that hybrid offspring produced under laboratory conditions are sterile (Bugmyrin et al., 2015, 2016) the introgression of genes in natural populations suggests that hybrid offspring of these two tick species are at least sometimes fertile (Kovalev et al., 2016). Whether or not hybrid ticks have appreciable fertility might not matter much as far as vector-switching goes, since even sterile hybrid ticks can probably transmit Borrelia.

The significance of tick hybridization for vector switching is that it would allow “mixed” infections to occur in which Borrelia specifically adapted to one tick species can come into intimate contact with another Borrelia species specifically adapted to a different tick vector. This would permit the exchange of bacterial genes conferring colonizing ability between the two Borrelia species, which are not normally able to utilize the “wrong” host (as is the case with B. afzelii adapted to I. ricinus, and Asian populations of B. bavariensis adapted to I. persulcatus) This is illustrated in Figure 7.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The reviewer, SM, declared a past collaboration with one of the authors, GM, to the handling editor.