EPIDEMIOLOGY

GEOGRAPHICAL DISTRIBUTION AND HOSTS

Rickettsia species are widely distributed across Europe. Their distribution is determined by that of the arthropods to which they are adapted.

Anaplasma phagocytophilum is widely distributed across Europe as it exploits, primarily, several Ixodes species as vectors, and has been encountered in most countries within the continent. Anaplasma platys is associated with the brown dog tick (Rhipicephalus sanguineus), which is also widely distributed across Europe, particularly in the warmer countries bordering the Mediterranean.

Of the five Anaplasma species encountered in European wildlife, A. phagocytophilum has been the most frequently reported. Evidence of infection has been reported in small mammals, including the bank vole (Myodes glareolus), rodent species in the genera Mus, Apodemus and Microtus, and shrews of the genera Crocidura and Sorex. Larger mammals, in particular deer, are also recognized as hosts for A. phagocytophilum. Infections have been reported in roe deer (Capreolous capreolus), red deer (Cervus elaphus), fallow deer (Dama dama) and sika deer (Cervus nippon), and in other ungulates including wild boar (Sus scrofa), bison (Bison bonasus) and mouflon ( Ovis orientalis). Recently, infection has also been reported in a brown bear (Ursus arctos) in Slovakia. Anaplasma phagocytophilum infections have been reported in raccoons (Procyon lotor) in their native North America, so it is likely that animals introduced into Europe are also susceptible. Elsewhere in the world, infections have been found in other groups of mammals, including leporids and members of the Sciuri- dae family, but surveys of these animals in Europe have yet to be reported.

The other A naplasma species appear to have a more limited host range; A. platys is primarily associated with dogs, although infections in wild- living canids have not been described. Anaplasma centrale and A. marginale are most frequently associated with domestic cattle; however, wild ruminants are thought to also play a role in the epizootiology of bovine anaplasmosis in some regions. Although the role of specific wild ruminant species has been studied in greatest depth in North America, some work has also been carried out in Europe. Anaplasma marginale DNA was detected in the blood of red deer in a region of Spain where infection is common in cattle. Anaplasma ovis infections are primarily reported in domes­ticated sheep and goats; however, the bacterium has also been detected in the blood of roe deer. Anaplasma ovis has been detected in reindeer (Rangifer tarandus) in East Asia, but infections in European populations have yet to appear.

Ehrlichia canis is a pathogen of domestic dogs, and is transmitted, like R. conorii and A. platys, by Rh. sanguineus. As yet, there is no evidence for infections in wild- living European canids; a survey of over 1500 red foxes (Vulpes vulpes) in Switzerland did not detect antibodies to E. canis in any animals, however, E. canis is not enzootic in the Swiss domestic canine population, and Rh. sanguineus infestation is rare. Conversely, a survey in Israel, where E. canis is enzootic in dogs, revealed 36% seropositivity among red foxes, suggesting that these hosts may serve as reservoirs for E.

Ehrlichia muris was discovered to be infecting red- backed voles (Eothenomys kageus) in Japan, but isolates were also subsequently obtained from several other small rodent species in the same country. Evidence for the pres­ence of E. muris in Europe was first obtained from a survey of yellow-necked mice (Apodemus flavicollii) inhabiting central Slovakia in 2007. DNA from E. muris, or at least E. muris-like organisms, has been detected in questing I. ricinus ticks in Slovakia and France.

ENVIRONMENTAL FACTORS

The distribution of Rickettsia, Anaplasma and Ehrlichia species is clearly restricted to that of their reservoir hosts and their competent vectors. The range of both is deter­mined by environmental factors such as habitat, altitude, temperature and rainfall. Considerable effort has gone into understanding the environmental determinants of the dis­tribution of ticks, and the temperature and humidity sen­sitivity of these arthropods has been quantified across Europe. As ticks serve as a reservoir as well as a vector for many Rickettsia, Anaplasma and Ehrlichia species, their ability to survive during periods when questing is not pos­sible (primarily through winter) is of the utmost impor­tance for the natural maintenance of the bacteria. Recently, work has shown that in infected ticks A. phagocytophilum induces the expression of a gene encoding a tick antifreeze glycoprotein, which is important for tick survival in a cold environment; hence it appears that A. phagocytophilum is enhancing its own persistence over winter by increasing the cold tolerance and survival of its vector.

EPIDEMIOLOGICAL ROLE OF WILD ANIMALS

Mammalian reservoir hosts have been identified for Ana- plasma and Ehrlichia species, and for TG Rickettsia species, but the role of vertebrates in the natural maintenance of SFG Rickettsia species is less well known. The role of Rickettsia, Anaplasma and Ehrlichia species in wildlife disease is also uncertain.

As discussed above, the role of wildlife in the natural maintenance of SFG Rickettsia species remains unclear. Despite the prevalence of SFG Rickettsia species in various European tick populations being relatively high (often over 10%), the prevalence of infections in the wildlife species that serve as major hosts for these ticks appears to be low. For example, in a recent survey in Poland, whereas 12.5% of I. ricinus ticks collected from 44 deer were found to contain R. helvetica, no evidence of infection was found in any of the deer themselves. Similarly, no evidence of infection was found in rodents or birds collected in the same region. Evidence for R. helvetica infection has been found in a roe deer in Slovakia, although again, this was only one of over 100 animals surveyed. However, very recently, a survey of woodland mammals in southern Germany revealed serological evidence of exposure to SFG rickettsiae in almost 30% of individuals tested and polymerase chain reaction (PCR)-based evidence of infec­tion in 8% of animals.

Serological evidence of exposure to rickettsiae in Euro­pean wildlife populations is also lacking, although strong support for host exposure to SFG group Rickettsia species can be drawn from studies on dogs, which are the primary host for Rh. sanguineus ticks, the vectors of R conorii. In regions where R conorii is endemic, the seroprevalence of R. conorii antibodies in dogs has been found to be high, and seasonal variation in seroprevalence correlates well with the active period of Rh. sanguineus. Early reports of isolation of R. akari (associated with peridomestic rodent­infesting Lyponyssoides sanguineus mites) from house mice in the USA have not been followed up in Europe, although it is widely considered that these rodents do play a direct role in the transmission of the bacterium. Rickettsia felii, most commonly associated with cat fleas, has been detected in numerous other blood- feeding arthropods, suggesting that they are being exposed to the bacterium even if they do not have a major role in its transmission. The role of mammals in the maintenance of TG rickettsiae appears to be clearer, as it is generally accepted that TG members exploit mammals as reservoir hosts. Rickettsia typhi is associated with wild rodents, in particular Rattui species, and anti-R. typhi antibodies have been observed in rats and other rodents in several European surveys. Thus, in summary, mammals are undoubtedly exposed to Rickettsia species through contact with blood- feeding arthropods. However, the frequency with which mammals become infected with SFG members, the kinetics of these infec­tions, and their impact on host wellbeing, remains uncertain.

It is not entirely clear which of the mammalian species discussed above as being susceptible to A naplasma or E hr- lichia infections are important as reservoir hosts in the natural maintenance of the bacteria, and which are merely accidental hosts. Small mammals, deer and livestock are thought to act as reservoir hosts for A. phagocytophilum, on the basis of either longitudinal studies of natural popu­lations or experimental infections. Interestingly, in parts of Europe at least, A. phagocytophilum strains present in small mammals are genotypically distinct from those present in deer, suggesting that these two hosts form part of distinct, enzootic cycles. It appears likely that transmission of A. phagocytophilum in the enzootic cycle involving deer relies on I. ricinus, whereas transmission in the cycle involving small mammals relies on Ixodes triangulicepi, suggesting that subpopulations of A. phagocytophilum have evolved to exploit, exclusively, different vectors. Livestock are consid­ered reservoir hosts for A. centrale, A. marginale and A. ovis, but the importance of wildlife in this role remains untested. Likewise, dogs are reservoir hosts for A. platys and E. canis, but wildlife species that are also parasitized by Rh. sanguineus may be able to fulfil this role too. It can only be assumed that small mammals act as reservoirs for E. muris, but, given that this species remains virtually unstudied, clarification of this role remains some way off.

TRANSMISSION

Rickettsia, Anaplasma and E hrlichia species can be main­tained in populations of arthropods by trans-stadial trans­mission, but only Rickettsia species can be transmitted transovarially. A naplasma and E hrlichia species are passed from infected vectors to non-infected vectors via the blood of mammalian hosts. Although this transmission route may also be employed by Rickettsia species, its relative importance in the natural maintenance of these bacteria is not known. Transmission between vectors may also occur via co-feeding, a process that involves an infected arthro­pod transmitting microorganisms in saliva or by regurgita­tion to an uninfected arthropod feeding nearby. Several Rickettsia species use co-feeding as a transmission strategy. For those Rickettsia species associated with fleas, R typhi and R. felis, it has also been proposed that bacteria may be naturally shed in faecal material and that this material may be infectious either to susceptible hosts or flea larvae. Indeed, for R. typhi, this has long been considered the primary mode of transmission. However, recent experi­mental work has demonstrated that transmission via flea bite, at least between rats, also occurs. Interestingly, experi­ments have also revealed that some Rickettsia species can be transmitted by aerosol. Although the importance of this route in nature is unknown, its existence has led to con­cerns being raised that Rickettsia species could effectively

be used in bioterrorism or biological warfare. There is also increasing evidence that mechanical transmission, via arthropod mouthparts or other vehicles, have a role in the transmission of Anaplasma species. This mode of transmis­sion may explain why A. marginale infections are encoun­tered in places where tick vectors are apparently absent.

The mechanisms by which Rickettsia, Anaplasma and Ehrlichia species exploit their arthropod reservoirs/vectors have been explored to some degree. Rickettsia species can be acquired by arthropods by one of two routes — either via the midgut from infected blood meal or transovarially. Infection via the first of these routes occurs by rickettsial invasion of the tick midgut epithelial cells, followed by dissemination of the bacteria throughout the rest of the tick’s body. Replication of rickettsiae occurs in salivary glands and ovaries, thereby facilitating transmission by feeding or to eggs. For Anaplasma and Ehrlichia species, the midgut is the first site of infection, as bacteria are acquired with ingestion of the blood meal into the midgut lumen. From here, bacteria enter the midgut epithelial cells and begin to replicate within membrane-bound vacu­oles. Subsequently, bacteria migrate to and invade the sali­vary glands, and following this invasion, bacteria again begin to replicate in salivary gland acinar cells. This second round of replication is probably dependent on resumption of tick feeding on a mammalian host, and is followed by transmission via the saliva.