This study investigated 624 cats in Germany for the presence of Hepatozoon spp. and Dirofilaria spp. via direct detection methods as well as for the presence of antibodies against Ehrlichia spp., Rickettsia spp. and Leishmania spp. via indirect detection methods. A background history was available for 371 cats, the majority of which had either been imported or had spent time outside of Germany (363/371 cats, 98%). These numbers can be attributed to the fact that the testing panel used as the basis for this study to detect different vector-borne pathogens is offered as a “Feline Travel Profile” to veterinarians. The majority of the 363 cats with a known background history of time spent abroad had done so in other European countries (88%), but several non-European countries were also implicated (22%, table 2). Spain (n=158) and Greece (n=52) were most commonly involved, and many of the cats with a background history implicating either one of these countries had positive test results (Spain: 32%, Greece: 33%). Imports by animal welfare organizations may play a significant role for both these countries (Spain: 20 animal welfare imports, Greece 6 animal welfare imports). Similarly, 6 of 25 cats that had spent time in Bulgaria were imported to Germany by animal welfare organizations (table 2). The number of imported cats greatly outweighs that of cats accompanying their owners‘ travels, which contrasts with the findings of previous studies in dogs [14, 15]. The rising numbers of cats tested between 2012 and 2020 (table 1) may indicate that the import of cats is gaining importance in Germany. Together with the change in climate in many parts of Europe, this could contribute to an increase in the spread of pathogens and their potential vectors into previously non-endemic areas such as Germany, where they may spread further and form reservoirs for infection. Under suitable conditions, pathogens transmitted by imported vectors may cause infection in competent hosts endemic to Germany, of which cats are only one example. Moreover, endemic vectors which are potentially competent may be infected with previously non-endemic pathogens during a blood meal on infected cats and could proceed to contribute to the spread of these pathogens. One example of this phenomenon are presently isolated cases of autochthonous infections with D. repens [16-18] and L. infantum [19] in dogs in Germany.
To the knowledge of the authors, the prevalence of many vector-borne infectious pathogens in cats in Germany is still unknown, as for example for Hepatozoon spp. In this study, 9% of the cats tested for this pathogen had positive PCR results. Direct detection methods demonstrate the presence of deoxyribonucleic acid or the antigen of a pathogen. Apart from infections with H. canis and H. silvestris, H. felis seems to be the primary infecting pathogen in cats [20-25]. Species differentiation showed the presence of H. felis in 7/53 cats infected with Hepatozoon spp. in this study. They had been imported from Spain (n=5), Greece, and Malta (n=1 respectively), which is consistent with the above cited literature. There is little knowledge about the pathogenesis, replication cycle, host range, and modes of transmission of Hepatozoon spp in cats. In addition to vector transmission, there are reports of transplacental transmission of H. canis and H. felis in cats [6]. Therefore, any female cat which was tested positive in this study and was not spayed (n=7) could transmit the pathogen in Germany to its kittens, regardless of any contact with a vector. Autochthonous infection with H. felis has been reported in a cat in Austria [9]. This may indicate the spread of the pathogen and/or vectors from historically endemic countries in the Mediterranean to more northern regions of Central Europe. In this study, 39/53 cats with positive test results had a history of travel/import to a known endemic area, and time spent abroad could not be excluded for any of the animals with positive test results. Consequently, this study provides no evidence of autochthonous infections in cats within Germany.
One cat in this study had positive PCR results for Dirofilaria spp., but further species differentiation was not done and a travel history/information on any time spent abroad was not available. This cat also had a positive IFAT for Leishmania spp., so contact with the pathogens in an endemic country in the Mediterranean is likely. Infections with Dirofilaria spp. in cats and dogs historically occurs in Mediterranean countries but have recently spread within these countries as for example Italy, Spain, France, Greece and Turkey [10]. Dirofilaria repens [26-28] has been the primary pathogen reported in Central and Eastern Europe, and it is currently considered an emerging zoonotic agent in all of Europe [29]. The prevalence of Dirofilaria spp. in cats varies from 0% to 33% across Europe [11, 30-40]. According to predictive models developed for dirofilariasis, temperatures during the summer may be suitable for the life cycle of larvae in mosquitoes even in colder regions like the United Kingdom, provided that reservoirs are present [10, 27, 41, 42]. It is to be noted that due to some diagnostic peculiarities, the true prevalence of D. immitis may be higher than indicated by the relatively low number of cats with positive test results in this study. Many of the immature pathogens are destroyed shortly after reaching the pulmonary arteries in cats, and the lifespan of the surviving pathogens is shorter in cats (2-4 years) than in most other species, such as dogs (5-7 years) [43]. Cats are rarely infected with more than five roundworms, which can be missed even in a post-mortem examination [44]. Microfilaremia is rare in cats, as less male worms are present [44]. Data on the prevalence of Dirofilaria spp. in cats in Germany is not yet available. A single case report from Central Europe describes a cat in Poland which was infected with D. repens and Wolbachia spp. [24].
Indirect detection methods were used to detect Ehrlichia spp., Rickettsia spp. and Leishmania spp. They demonstrate the presence of antibodies produced in response to the pathogen contact, but this does not necessarily correlate with the presence of disease. Seroconversion may not occur until two to three weeks after exposure and antibodies may be detectable for up to several years after disease resolution, depending on the pathogen. It is generally possible to distinguish more recent infections from those in the past by means of simultaneous Immunoglobulin M levels, or paired serum samples taken at intervals of two to four weeks. However, the former is unusual in routine diagnostics for the pathogens discussed and the latter is often not feasible in practice. The indirect IFAT utilised in this study detected Immunoglobulin G antibodies for all pathogens. Furthermore, the interpretation of IFAT can be subjective and so sensitivity can be low, especially where titres are low or borderline. Limitations may also include the possibility of cross reactivity with other pathogens, as well as false negative results in very young or immunosuppressed animals, or where investigations were done early in the natural history of the disease and therefore prior to seroconversion [45].
The IFAT used in this study detected antibodies to Leishmania spp. in 22/624 cats (4%). Cats in Mediterranean countries are generally infected by L. infantum. There is much variation in the reported prevalence of Leishmania spp. in cats tested by indirect assays not only between different European countries but also across different regions within one country, ranging from 0.1% to 60% [1, 30, 40, 46-68]. Dogs are currently the only known primary reservoir of infection [69]. It has been speculated that cats may be an additional reservoir, but this has not been confirmed [70]. Sandflies can be infected with L. infantum during a blood meal on an infected cat, and so cats could be instrumental in the spread of the pathogen in areas with a high prevalence [71]. Consequently, cats with antibodies to the pathogen could be a reservoir for infection within Germany, provided they were also still infected. The presence of competent vectors like P. perniciosus has been reported in the South of Germany [72], as that of the potentially competent vector P. mascitti [72, 73]. There is little evidence on the susceptibility or resistance of cats to natural infection. Cats have a more efficient T-helper 1 cell immune response compared to dogs, which may be the cause of the lower prevalence of the pathogen in cats [46]. Twelve of the 22 cats with positive IFAT results (55%) in this study were imported to Germany from Mediterranean countries and Southeast Europe, where L. infantum is endemic. One of the 22 cats (5%) was imported from Brazil, where cats may be infected by not only L. infantum but also L. amazonensis or L. braziliensis [74-77].
Antibodies against Ehrlichia spp. were detected via IFAT in 12% of the tested cats. Previous studies involving indirect detection methods (IFAT) report a 1-18% prevalence of Ehrlichia spp. in cats in the Mediterranean [30, 50, 52, 54, 57, 78-82]. Data on the prevalence of antibodies against Ehrlichia spp. in cats in Germany is not currently available. IFAT may show some cross-reactivity with E. chaffensis (found in cats in the United States and Brazil) and E. ewingii (found in cats in the United States), as well as with A. phagocytophilum and A. platys at lower titres. Cross-reactivity due to contact with A. phagocytophilum in Germany cannot be excluded, especially in the 44 cats with a low titre of 1:40 in this study. Seropositivity in the remaining 29 animals with higher titres is most likely due to infection with Ehrlichia. A study in 479 cats in Southern Germany did not demonstrate any Ehrlichia spp. DNA [83]. Therefore, autochthonous natural infections in cats in Germany are unlikely and the infections most likely occurred abroad.
Eleven percent of 467 cats had positive IFAT results for Rickettsia spp. Seroprevalence in cats has been researched in Italy, Spain, and Portugal (IFAT/ELISA: 0-48.7%) [30, 46, 50, 52, 78, 84, 85]. Cats may be instrumental in the transmission cycle of some rickettsiae of the spotted fever group (SFG), especially R. conorii and R. felis [86, 87]. Antibodies for R. conorii have been detected in cats after infections with Rhipicephalus sanguineus [78, 84, 87]. Rickettsia felis is a well-established cause of the emerging flea-borne spotted fever, of which there have been several cases in humans worldwide [88, 89]. Cats will have antibodies for Rickettsia spp. after being infected (either natural or experimental) with fleas of the species Ctenocephalides (C.) felis [90]. The pathogen has also been detected via PCR in previously non-infected fleas after a blood meal on infected cats [91]. Consequently, C. felis can be considered a competent vector and autochthonous infections within Germany are possible. This study detected antibodies by means of IFAT, which is regarded as the gold standard for serological confirmation of pathogen contact in dogs and cats. There are, however, cross-reactions between any of the more than 20 species in the spotted fever group [87]. We detected antibodies to Rickettsia spp. in 52/467 cats (11%). There were 29 cats which were seropositive and had been imported from abroad, and it is unclear whether they were infected in Germany or in their country of origin. The four seropositive cats which had never left Germany were most likely infected with R. felis. The clinical importance of Rickettsia spp. infections in cats is still unknown. A study in clinically symptomatic cats found no association between positive antibody titres and fever, and no febrile cats in this study had positive PCR results for R. felis or R. rickettsi [92].
In this study, 22 of 624 cats (4%) had positive test results for more than one pathogen. It is known that co-infections may complicate diagnosis and treatment in dogs and may worsen their prognosis [2]. Coinfections with multiple vector-borne pathogens may occur in cats as well as dogs and humans, but their clinical consequences are still unknown and should be evaluated in further studies, especially in cats [93]. In this study, nine cats infected with Hepatozoon spp. also had antibodies against Leishmania spp. (n=4), Rickettsia spp. (n=3), and Ehrlichia spp. (n=2). Antibodies to Leishmania and Ehrlichia spp. were present in 12 cats infected with Hepatozoon spp., respectively. This indicates a pathogen contact with concurrent immunosuppression to be discussed in case of persistent infection, as it may result in increased susceptibility of infected animals to other pathogens [2].
Limitations of this study
Limitations of this study are mainly its retrospective design (e.g. no consistent histories) and the limited number of pathogens included. Certain vector-borne infectious pathogens such as Cytauxzoon spp. could not be included. Furthermore, species differentiation for specific pathogens included in the study was not performed, except in the case of seven cats with positive test results for H. felis. There was also no information on the extent of ectoparasite prophylaxis in the cats, which may impact the prevalence of certain vector-borne pathogens. In the cats which had travelled with their owners to endemic countries, it was not possible to reliably document the duration or the time of the year of these travels. As many of the relevant vectors show pronounced seasonality, the time of year may significantly influence both incidence and prevalence of the pathogens they may transmit. The histories taken from the veterinarians only included the countries of stays abroad.