Extensive Phylogenomic Discordance and the Complex Evolutionary History of the Neotropical Cat Genus Leopardus.

Even in the genomics era, the phylogeny of Neotropical small felids comprised in the genus Leopardus remains contentious. We used whole-genome resequencing data to construct a time-calibrated consensus phylogeny of this group, quantify phylogenomic discordance, test for interspecies introgression, and assess patterns of genetic diversity and demographic history. We infer that the Leopardus radiation started in the Early Pliocene as an initial speciation burst, followed by another in its subgenus Oncifelis during the Early Pleistocene. Our findings challenge the long-held notion that ocelot (Leopardus pardalis) and margay (L. wiedii) are sister species and instead indicate that margay is most closely related to the enigmatic Andean cat (L. jacobita), whose whole-genome data are reported here for the first time. In addition, we found that the newly sampled Andean tiger cat (L. tigrinus pardinoides) population from Colombia associates closely with Central American tiger cats (L. tigrinus oncilla). Genealogical discordance was largely attributable to incomplete lineage sorting, yet was augmented by strong gene flow between ocelot and the ancestral branch of Oncifelis, as well as between Geoffroy's cat (L. geoffroyi) and southern tiger cat (L. guttulus). Contrasting demographic trajectories have led to disparate levels of current genomic diversity, with a nearly tenfold difference in heterozygosity between Andean cat and ocelot, spanning the entire range of variability found in extant felids. Our analyses improved our understanding of the speciation history and diversity patterns in this felid radiation, and highlight the benefits to phylogenomic inference of embracing the many heterogeneous signals scattered across the genome.

Drivers of habitat availability for terrestrial mammals: Unravelling the role of livestock, land conversion and intrinsic traits in the past 50 years.

The global decline of terrestrial species is largely due to the degradation, loss and fragmentation of their habitats. The conversion of natural ecosystems for cropland, rangeland, forest products and human infrastructure are the primary causes of habitat deterioration. Due to the paucity of data on the past distribution of species and the scarcity of fine-scale habitat conversion maps, however, accurate assessment of the recent effects of habitat degradation, loss and fragmentation on the range of mammals has been near impossible. We aim to assess the proportions of available habitat within the lost and retained parts of mammals' distribution ranges, and to identify the drivers of habitat availability. We produced distribution maps for 475 terrestrial mammals for the range they occupied 50 years ago and compared them to current range maps. We then calculated the differences in the percentage of 'area of habitat' (habitat available to a species within its range) between the lost and retained range areas. Finally, we ran generalized linear mixed models to identify which variables were more influential in determining habitat availability in the lost and retained parts of the distribution ranges. We found that 59% of species had a lower proportion of available habitat in the lost range compared to the retained range, thus hypothesizing that habitat loss could have contributed to range declines. The most important factors negatively affecting habitat availability were the conversion of land to rangeland and high density of livestock. Significant intrinsic traits were those related to reproductive timing and output, habitat breadth and medium body size. Our findings emphasize the importance of implementing conservation strategies to mitigate the impacts caused by human activities on the habitats of mammals, and offer evidence indicating which species have the potential to reoccupy portions of their former range if other threats cease to occur.

Assessing micro-macroparasite selective pressures and anthropogenic disturbance as drivers of immune gene diversity in a Neotropical wild cat.

Anthropogenic environmental change is reducing available habitat for wild species, providing novel selection pressures such as infectious diseases and causing species to interact in new ways. The potential for emerging infectious diseases and zoonoses at the interface between humans, domestic animals, and wild species is a key global concern. In vertebrates, diversity at the major histocompatibility complex MHC is critical to disease resilience, and its study in wild populations provides insights into eco-evolutionary dynamics that human activities alter. In natural populations, variation at MHC loci is partly maintained by balancing selection, driven by pathogenic selective pressures. We hypothesize that MHC genetic diversity differs between guigna populations inhabiting human-dominated landscapes (higher pathogen pressures) versus more natural habitats (lower pathogen pressures). We predict that MHC diversity in guignas would be highest in human-dominated landscapes compared with continuous forest habitats. We also expected to find higher MHC diversity in guignas infected with micro and macro parasites (higher parasite load) versus non infected guignas. We characterized for the first time the genetic diversity at three MHC class I and II exons in 128 wild guignas (Leopardus guigna) across their distribution range in Chile (32-46° S) and Argentina, representing landscapes with varying levels of human disturbance. We integrated MHC sequence diversity with multiple measures of anthropogenic disturbance and both micro and macro parasite infection data. We also assessed signatures of positive selection acting on MHC genes. We found significantly higher MHC class I diversity in guignas inhabiting landscapes where houses were present, and with lower percentage of vegetation cover, and also in animals with more severe cardiorespiratory helminth infection (richness and intensity) and micro-macroparasite co-infection. This comprehensive, landscape-level assessment further enhances our knowledge on the evolutionary dynamics and adaptive potential of vertebrates in the face of emerging infectious disease threats and increasing anthropogenic impacts.

The Subantarctic Rayadito (Aphrastura subantarctica), a new bird species on the southernmost islands of the Americas.

We describe a new taxon of terrestrial bird of the genus Aphrastura (rayaditos) inhabiting the Diego Ramírez Archipelago, the southernmost point of the American continent. This archipelago is geographically isolated and lacks terrestrial mammalian predators as well as woody plants, providing a contrasted habitat to the forests inhabited by the other two Aphrastura spp. Individuals of Diego Ramírez differ morphologically from Aphrastura spinicauda, the taxonomic group they were originally attributed to, by their larger beaks, longer tarsi, shorter tails, and larger body mass. These birds move at shorter distances from ground level, and instead of nesting in cavities in trees, they breed in cavities in the ground, reflecting different life-histories. Both taxa are genetically differentiated based on mitochondrial and autosomal markers, with no evidence of current gene flow. Although further research is required to define how far divergence has proceeded along the speciation continuum, we propose A. subantarctica as a new taxonomic unit, given its unique morphological, genetic, and behavioral attributes in a non-forested habitat. The discovery of this endemic passerine highlights the need to monitor and conserve this still-pristine archipelago devoid of exotic species, which is now protected by the recently created Diego Ramírez Islands-Drake Passage Marine Park.

New Records of the Andean Cat (Leopardus jacobita) in the Central Andes of Chile: Filling Gaps in the Distribution Range through Private-Social Partnerships.

The Andean cat (Leopardus jacobita) is one of the most endangered and least known wild cat species in the Americas and the world in general. We describe new records of the Andean cat in the central Andes of Chile, in Parque Andino Juncal, obtained as part of a monitoring program conducted from December 2020 to May 2021 using eight trail cameras. The cameras were active for 135 days (sampling effort 1080 camera-trap days). We recorded Andean cats in two different cameras, corresponding to two independent events in January and March 2021, respectively (0.19% capture success). Our new records are relevant since they fill a gap on the species distribution map, specifically in the area located between the two previously identified evolutionarily significant units (ESU) (26-35° S) which has been prioritized by the Andean Cat Alliance (AGA). We highlight the relevance of private protected areas in ecosystems of high biodiversity and fragility such as Parque Andino Juncal and also of strategic private-social partnerships for successful collaborative efforts to monitor the presence of rare, elusive, and endangered species. Our next steps will be to collect scats from this newly identified site and conduct genetic analyses to determine whether these animals are part of previously identified ESUs or a yet unidentified potentially distinct ESU or MU requiring special conservation measures.

SEROSURVEY FOR SELECTED PARASITIC AND BACTERIAL PATHOGENS IN DARWIN'S FOX (LYCALOPEX FULVIPES): NOT ONLY DOG DISEASES ARE A THREAT.

The Darwin's fox (Lycalopex fulvipes) is one of the most endangered carnivores worldwide, with the risk of disease spillover from domestic dogs being a major conservation threat. However, lack of epidemiologic information about generalist, non-dog-transmission-dependent protozoal and bacterial pathogens may be a barrier for disease prevention and management. To determine the exposure of some of these agents in Darwin's fox populations, 54 serum samples were collected from 47 Darwin's foxes in Southern Chile during 2013-18 and assessed for the presence of antibodies against Brucella abortus, Brucella canis, Coxiella burnetii, pathogenic Leptospira (serovars Grippotyphosa, Pomona, Canicola, Hardjo, and Copehageni), Toxoplasma gondii, and Neospora caninum. The highest seroprevalence was detected for T. gondii (78%), followed by pathogenic Leptospira (14%). All the studied Leptospira serovars were confirmed in at least one animal. Two foxes seroconverted to Leptospira and one to T. gondii during the study period. No seroconversions were observed for the other pathogens. No risk factors, either intrinsic (sex, age) or extrinsic (season, year, and degree of landscape anthropization), were associated with the probability of being exposed to T. gondii. Our results indicate that T. gondii exposure is widespread in the Darwin's fox population, including in areas with minimal anthropization, and that T. gondii and pathogenic Leptospira might be neglected threats to the species. Further studies identifying the causes of morbidity and mortality in Darwin's fox are needed to determine if these or other pathogens are having individual or population-wide effects in this species.

Puma (Puma concolor) in the Neighborhood? Records Near Human Settlements and Insights into Human-Carnivore Coexistence in Central Chile.

The wildland-urban interface lies at the confluence of human-dominated and wild landscapes-creating a number of management and conservation challenges. Wildlife sightings near human settlements have appeared to increase in the last years. This article reports 51 records of presences, sightings, and livestock attacks of Puma concolor, a large-sized felid, collected from 2012 to 2020 across the O'Higgins region in central Chile. Puma records were concentrated in the east of the region in the Andes Range and foothills (90%). The number of puma records is higher in the last four to six years than in previously studied years. Of the 51 records, 23.5% are between 0 and 999 m from the nearest human settlement (classified as very close), 25.5% are between 1000 and 4999 m (moderately close), and 51% are over 5000 m (distant). Most of the sightings are recorded in the summer (35%) and spring (29%). We identify an area of approximately 9000 km2 of suitable habitat as the most probable corridor effectively connecting pumas moving between eastern and western areas, encompassing the Angostura de Paine mountain range. Our results contribute to the understanding of the presence and movements of P. concolor near urban areas and human settlements, confirming their persistence in and adaptation to human-dominated landscapes. We also provide insights into human-carnivore coexistence in the current global context in the densely populated central Chile.

Cross-species transmission of retroviruses among domestic and wild felids in human-occupied landscapes in Chile.

Human transformation of natural habitats facilitates pathogen transmission between domestic and wild species. The guigna (Leopardus guigna), a small felid found in Chile, has experienced habitat loss and an increased probability of contact with domestic cats. Here, we describe the interspecific transmission of feline leukemia virus (FeLV) and feline immunodeficiency virus (FIV) between domestic cats and guignas and assess its correlation with human landscape perturbation. Blood and tissue samples from 102 free-ranging guignas and 262 domestic cats were collected and analyzed by PCR and sequencing. Guigna and domestic cat FeLV and FIV prevalence were very similar. Phylogenetic analysis showed guigna FeLV and FIV sequences are positioned within worldwide domestic cat virus clades with high nucleotide similarity. Guigna FeLV infection was significantly associated with fragmented landscapes with resident domestic cats. There was little evidence of clinical signs of disease in guignas. Our results contribute to the understanding of the implications of landscape perturbation and emerging diseases.

Widespread Infection with Hemotropic Mycoplasmas in Free-Ranging Dogs and Wild Foxes Across Six Bioclimatic Regions of Chile.

Blood samples of 626 rural dogs, 140 Andean foxes (Lycalopex culpaeus), and 83 South American grey foxes (L. griseus) from six bioregions of Chile spanning 3000 km were screened for Mycoplasma DNA by conventional PCR and sequencing. Risk factors of infection were inferred using Generalized Linear Mixed Models and genetic structure by network analyses. Overall, Mycoplasma haemocanis/Mycoplasma haemofelis (Mhc/Mhf) and Candidatus Mycoplasma haematoparvum (CMhp) observed prevalence was 23.8% and 12.8% in dogs, 20.1% and 7.2% in Andean foxes, and 26.5% and 8.4% in grey foxes, respectively. Both hemoplasmas were confirmed in all the bioregions, with higher prevalence in those where ticks from the Rhipicephalus sanguineus species group were absent. Candidatus M. haematominutum and a Mycoplasma sp. previously found in South American carnivores were detected in one fox each. Although the most prevalent Mhc/Mhf and CMhp sequence types were shared between dogs and foxes, network analysis revealed genetic structure of Mhc/Mhf between hosts in some regions. Male sex was associated with a higher risk of Mhc/Mhf and CMhp infection in dogs, and adult age with CMhp infection, suggesting that direct transmission is relevant. No risk factor was identified in foxes. Our study provides novel information about canine hemoplasmas with relevance in distribution, transmission routes, and cross-species transmission.

Molecular Survey of Parvoviruses and Mycoplasma spp. in Invasive American Mink (Neovison vison) from Southern Chile.

Using PCR, we evaluated the presence of parvoviruses and Mycoplasma spp. in 123 American mink (Neovison vison), an introduced invasive carnivore in Chile. Our results showed all analyzed animals were negative for both pathogen groups. We cannot completely dismiss their presence, but if present, their prevalence should be lower than 2%.

Epidemiology and molecular characterization of Carnivore protoparvovirus-1 infection in the wild felid Leopardus guigna in Chile.

Landscape anthropization has been identified as one of the main drivers of pathogen emergence worldwide, facilitating pathogen spillover between domestic species and wildlife. The present study investigated Carnivore protoparvovirus-1 infection using molecular methods in 98 free-ranging wild guignas (Leopardus guigna) and 262 co-occurring owned, free-roaming rural domestic cats. We also assessed landscape anthropization variables as potential drivers of infection. Protoparvovirus DNA was detected in guignas across their entire distribution range, with observed prevalence of 13.3% (real-time PCR) and 9% (conventional PCR) in guignas, and 6.1% (conventional PCR) in cats. Prevalence in guigna did not vary depending on age, sex, study area or landscape variables. Prevalence was higher in juvenile cats (16.7%) than in adults (4.4%). Molecular characterization of the virus by amplification and sequencing of almost the entire vp2 gene (1,746 bp) from one guigna and five domestic cats was achieved, showing genetic similarities to canine parvovirus 2c (CPV-2c) (one guigna and one cat), feline panleukopenia virus (FPV) (one cat), CPV-2 (no subtype identified) (two cats), CPV-2a (one cat). The CVP-2c-like sequence found in a guigna clustered together with domestic cat and dog CPV-2c sequences from South America, suggesting possible spillover from a domestic to a wild species as the origin of infection in guigna. No clinical signs of disease were found in PCR-positive animals except for a CPV-2c-infected guigna, which had haemorrhagic diarrhoea and died a few days after arrival at a wildlife rescue centre. Our findings reveal widespread presence of Carnivore protoparvovirus-1 across the guigna distribution in Chile and suggest that virus transmission potentially occurs from domestic to wild carnivores, causing severe disease and death in susceptible wild guignas.

High seroprevalence of feline morbilliviruses in free-roaming domestic cats in Chile.

Feline morbillivirus infections have gained increased attention due to repeated reports of their association with urinary tract disease in cats. In the present study, 112 serum samples from free-roaming domestic cats in Chile were tested for antibodies against feline morbillivirus genotypes 1 and 2 (FeMV-1 and FeMV-2) using an indirect immunofluorescence assay. In total, 63% of the animals showed antibodies against one or both FeMV genotypes. Antibodies directed exclusively against FeMV-2 were significantly more prevalent in male cats. The correlation of sex and FeMV-2 infection might give insight into potential routes of transmission. We provide, for the first time, serological data on FeMV in Chile.

Domestic dog origin of Carnivore Protoparvovirus 1 infection in a rescued free-ranging guiña (Leopardus guigna) in Chile.

Carnivore protoparvovirus 1 is one of the most important pathogens affecting both wild and domestic carnivores. Here, we reported the genetic characterization of canine parvovirus (CPV-2) strains from a rescued guiña (Leopardus guigna) and domestic dogs from Chile. Guiña strain was classified as CPV-2c, and phylogenetic analysis of the complete coding genome showed that the guiña CPV-2c strain shares a recent common ancestor with Chilean domestic dogs' strains. These viruses showed >99% identity and exhibited three changes in the NS1 protein (V596A, E661K and L582F). This is the first detection and genetic characterization of CPV-2c infection in guiña worldwide, and one of the few comparative studies that show the source of infection was domestic dogs. The current findings highlight the fact that guiña is a susceptible species to protoparvovirus infection and that domestic dogs represent an important threat to its conservation. The CPV-2 cross-species transmission between domestic dogs and guiña should be taken into account for protection programmes of this endangered species.

Identification of Novel Feline Paramyxoviruses in Guignas (Leopardus guigna) from Chile.

The family of paramyxoviruses has received growing attention as several new species have been identified recently, notably two different clusters in domestic cats, designated as feline morbillivirus (FeMV) and feline paramyxovirus (FPaV). Their phylogenetic origin and whether wild felids also harbor these viruses are currently unknown. Kidney samples from 35 guignas (Leopardus guigna), a wild felid from Chile, were investigated for paramyxoviruses using consensus-RT-PCR. In addition, thirteen serum samples of guignas were screened for the presence of FeMV-specific antibodies by an immunofluorescence assay (IFA). Viral RNA was detected in 31% of the kidney samples. Phylogenetic analyses revealed two well-supported clusters, related to isolates from domestic cats, rodents and bats. No significant histopathology changes were recorded in infected guignas. Serology identified two samples which were positive for FeMV-specific antibodies. Our study highlights the diversity of paramyxovirus infections in felids with special emphasis on guignas from Chile.

Gastrointestinal and cardiorespiratory endoparasites in the wild felid guigna (Leopardus guigna) in Chile: Richness increases with latitude and first records for the host species.

Guignas (Leopardus guigna) are small felids closely associated with native forest habitats. In fragmented landscapes, they use vegetation corridors and forest remnants to move across the landscape. In these contexts, guignas may increase contact probabilities with domestic animals, being therefore relevant to assess their pathogens and parasites. The aim of this study was to characterize the helminth fauna in the gastrointestinal tract and cardiorespiratory system of guignas from central and southern Chile. Between 2015 and 2018, 33 dead free-ranging guignas were found road-killed or were collected from wildlife rescue centers. Thirty-two gastrointestinal tracts and 32 cardiorespiratory organs were analyzed through direct analysis and artificial digestion. We found 81.8% (27/33) guignas were positive for helminth endoparasites (84.4% (27/32) positive for gastrointestinal parasites, 37.5% (12/32) positive for cardiorespiratory parasites). Fourteen parasites were identified (7 at genus level and 7 at species level), with Angiostrongylus sp., Molineus sp., Oslerus sp. and Troglostrongylus sp. as first records in guignas. The most prevalent parasites were the species Toxascaris leonina, Toxocara cati and Uncinaria stenocephala. Uncinaria stenocephala showed the highest intensity of infection. Multiparasitism was observed in 76% of the animals. Significant differences in richness of endoparasites and prevalence of cardiorespiratory parasites were found between geographic zones; higher values in the southern zone are possibly due to favorable environmental characteristics for endoparasite development. There were no statistically significant differences between sexes. All the parasites found in this study have been previously reported in domestic cats. These results are valuable to understand parasite transmission at the domestic-wildlife interface; the possibility of endoparasite transmission between domestic cats and guignas should be clarified with molecular analysis.

Hemoplasmas Are Endemic and Cause Asymptomatic Infection in the Endangered Darwin's Fox (Lycalopex fulvipes).

Mycoplasma haemocanis is prevalent in the endangered Darwin's fox (Lycalopex fulvipes) in its main stronghold, Chiloé Island (Chile). The origin of the infection, its dynamics, its presence in other fox populations and the potential consequences for fox health remain unexplored. For 8 years, hemoplasmal DNA was screened and characterized in blood from 82 foxes in Chiloé and two other fox populations and in 250 free-ranging dogs from Chiloé. The prevalence of M. haemocanis in foxes was constant during the study years, and coinfection with "Candidatus Mycoplasma haematoparvum" was confirmed in 30% of the foxes. Both hemoplasma species were detected in the two mainland fox populations and in Chiloé dogs. M. haemocanis was significantly more prevalent and more genetically diverse in foxes than in dogs. Two of the seven M. haemocanis haplotypes identified were shared between these species. Network analyses did not show genetic structure by species (foxes versus dogs), geographic (island versus mainland populations), or temporal (years of study) factors. The probability of infection with M. haemocanis increased with fox age but was not associated with sex, season, or degree of anthropization of individual fox habitats. Some foxes recaptured years apart were infected with the same haplotype in both events, and no hematological alterations were associated with hemoplasma infection, suggesting tolerance to the infection. Altogether, our results indicate that M. haemocanis is enzootic in the Darwin's fox and that intraspecific transmission is predominant. Nevertheless, such a prevalent pathogen in a threatened species represents a concern that must be considered in conservation actions.IMPORTANCEMycoplasma haemocanis is enzootic in Darwin's foxes. There is a higher M. haemocanis genetic diversity and prevalence in foxes than in sympatric dogs, although haplotypes are shared between the two carnivore species. There is an apparent tolerance of Darwin's foxes to Mycoplasma haemocanis.

A forest-specialist carnivore in the middle of the desert?Comments on https://onlinelibrary.wiley.com/doi/full/10.1002/ece3.5230.

We present comments on an article recently published in Ecology and Evolution ("High-resolution melting of the cytochrome B gene in fecal DNA: A powerful approach for fox species identification of the Lycalopex genus in Chile") by Anabalon et al. that reported the presence of Darwin's fox (Lycalopex fulvipes), a temperate forest specialist, in the hyperarid Atacama Desert of northern Chile. We argue that this putative record lacks ecological support in light of ongoing research on this endangered species, and contains numerous methodological flaws and omissions related to the molecular identification of the species. Based on these issues, we suggest the scientific community and conservation decision-makers disregard the alleged presence of the Darwin's fox in the Atacama Desert.

Antibiotic resistance genes as landscape anthropization indicators: Using a wild felid as sentinel in Chile.

Antimicrobial resistance is a global emerging public health issue whose presence and impact in wildlife are widely unknown. Antimicrobial resistance genes (ARGs) are considered environmental contaminants, suitable to evaluate the degree of anthropic impact on wildlife and the environment. We used a wild felid, the guigna (Leopardus guigna), as a sentinel for the presence of ARGs in anthropized and pristine areas across their entire distribution range in Chile. We evaluated fecal samples from 51 wild guignas, collected between 2009 and 2018. Real-time PCR essays were employed to detect and quantify 22 selected ARGs in their fecal microbiome. All animals (100%) were positive for at least one ARG. The most prevalent ARG families were those that confer resistance to tetracycline (88.2%) and beta-lactamase (68.9%), with tet(Q) (60.8%), tet(W) (60.8%), and blaTEM (66.7%) as the most prevalent ARGs. Multi-resistance profiles were observed in 43% of the guignas. Statistically significant differences were found between anthropized and pristine areas for tet(Q) (p = 0.014), tet(W) (p = 0.0037), tetracycline family (p = 0.027), multi-resistance profile prevalence (p = 0.043) and tet(W) quantification (p = 0.004). Two animals from anthropized landscapes were positive for mecA, a gene associated with Staphylococcus aureus and other staphylococci resistant to methicillin, while three animals from anthropized areas were positive for blaCTX-M, that encodes class A extended-spectrum beta-lactamase. Both genes have been identified in bacteria causing relevant nosocomial infections worldwide. This is the first study on ARGs in wild felids from Chile and the first detection of mecA in South American wild felids. We observed an association between the degree of landscape anthropization and ARG prevalence, confirming that ARGs are important indicators of wildlife exposure to human activity/presence, with a widespread distribution.

An Eight-Year Survey for Canine Distemper Virus Indicates Lack of Exposure in the Endangered Darwin's Fox (Lycalopex fulvipes).

No evidence of exposure to canine distemper virus (CDV) was detected in 70 samples corresponding to 58 wild-trapped Darwin's foxes (Lycalopex fulvipes) in Chile. Given its current endangered status and it being immunologically naïve, in the event of a CDV spillover from dogs to foxes, high population mortality is expected.

Molecular and serological survey of carnivore pathogens in free-roaming domestic cats of rural communities in southern Chile.

Owned, free-roaming domestic cats are abundant in the Chilean countryside, having high probability of contact with wildlife and potentially participating as reservoirs of zoonotic pathogens. In the present study, 131 cats from two remote study areas (Valdivia and Chiloe Island) in southern Chile were analyzed for infection/exposure to eight pathogens. Serum samples from 112 cats were tested for antigens against feline leukemia virus (FeLV antigen-ELISA) and antibodies against feline immunodeficiency virus (FIV-ELISA) and canine distemper virus (CDV-serum neutralization), yielded occurrence of 8.9, 1.7 and 0.8% respectively. The presence of DNA of five vector-borne pathogens, piroplasmids, Ehrlichia spp., Anaplasma spp., Rickettsia spp. and Bartonella spp. was investigated in thirty cats. Overall observed occurrence was 6.6% (2/30) for both Anaplasma platys, and B. henselae, and 3.3% (1/30) for both Bartonella sp. and Theileria equi. Observed occurrence for all vector-borne pathogens in Valdivia area was significantly higher than in Chiloe Island (5/15 vs 0/15; P=0.04). Our results represent the first description of exposure to CDV and DNA detection of T. equi and A. platys in domestic cats in Chile. The results highlight the importance of performing pathogen screening in owned, free-roaming rural cats to evaluate their potential role as reservoirs of infection and vectors for disease transmission to wildlife.