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The Neotropical otter (Lontra longicaudis) is a medium-sized mustelid with semiaquatic habits, inhabiting rivers, lagoons, lakes and small water bodies (de Almeida Rodrigues et al., 2013) across all Brazilian biomes (Rheingantz et al., 2014; Rosas-Ribeiro et al., 2017). It is considered a solitary species, despite females can be found with their offspring for several months (de Almeida Rodrigues et al., 2013). It is listed as a “Near Threatened” species at Red List from International Union for Conservation of Nature (IUCN), historically due to the fur trade and currently due to retaliation by fishers and fish farmers, pollution, cattle ranching, urban expansion, hydroelectric network, illegal trade as pets, and diseases (Rheingantz et al., 2022).

Nearly half of the mustelid species are experiencing population declines, with higher risks for some subfamilies, such as Lutrinae (Wright et al., 2022). The high rate of habitat degradation and the increasing proximity between wildlife and human-modified areas pose significant threats to the species conservation (Scanes, 2018). In this context, the exposure of wild animals to domestic dogs and cats have been shown significant impact on the decline of wild carnivore populations (Roelke-Parker et al., 1996). Beyond predation and direct competition, domestic animals may spread a wide range of pathogens to wildlife (Beineke et al., 2015). The detection of viral and protozoan pathogens originating from domestic animals in wild species has increased, and their effects have already been reported to cause disease and mortality in mustelids in various regions of the world (Kimber et al., 2000; De Bosschere et al., 2005; Echenique et al., 2018; Calvo-Mac et al., 2020; Thomas et al., 2020).

The aim of this study was to assess the exposure of Lontra longicaudis to domestic carnivores and pathogens associated with these species within a protected area in southern Brazil. To this end, we investigated the overlapping occurrence of otters, domestic dogs, and cats in the studied area, as well as the exposure to major pathogens commonly associated with dogs (Canine Distemper Virus - CDV, Canine Parvovirus type 2 - CPV, and Neospora caninum) and cats (Toxoplasma gondii and Feline Leukemia Virus - FeLV) in Neotropical otters.

During biological sample collection, one adult male otter was live-captured during 110 trap-nights and five freshly road-killed carcasses (2 adult males and 3 adult females) were recovered over a six-month period.

Serological tests detected antibodies against T. gondii in four of the six otters sampled (66%). The antibody titer for T. gondii in otters ranged from 64 to 2048 (geometric mean 181). Using the competitive ELISA, all otters were seronegative for anti-Neospora caninum antibodies. Antibodies against Canine Distemper Virus (CDV) were detected in five of the six otters sampled (83%). Four of the six otters (66%) were considered seropositive for the presence of antibodies for Canine Parvovirus type 2 (CPV).

All domestic dogs and otters tested negative for the presence of CDV RNA and CPV DNA in molecular analyses using real-time PCR. Whereas all otters were negative for FeLV antigen and pro-viral DNA, two domestic cats sampled tested positive for FeLV in both real-time PCR and antigen tests. Table 1 summarizes data regarding pathogen investigation in the six sampled otters.

A total of 640 km was surveyed using the linear transect method, which allowed the identification of nine distinct otter individuals based on characteristics such as size, body shape and visible scars, resulting in a total of 22 direct observations. Eleven dogs and five cats were observed inside the reserve along the road that crosses the area. Other domestic species such as cows, horses and pigs were also commonly seen inside the protected area.

A total of 1,019 night-trap sessions were analyzed. Camera trap monitoring detected otters at three of the nineteen successfully monitored sites, with a total of nine recorded events ( Table 2 ). Domestic cats were detected at three sites, totaling eleven events, with four distinct individuals identified. Domestic dogs were recorded at four sites, with a total of 27 detections events and 16 individuals identified. In some cases, dogs were recorded in the company of humans engaged in cattle and horses-related activities. Domestic cats were recorded only at night, while domestic dogs were registered in the day time, mainly at the early morning. Some camera traps recorded the presence of otters and domestic carnivores at the same site within the reserve ( Figure 1 ). Table 2 outlines the data on all records of otters and domestic carnivorans in the studied area.

The spatial distribution maps of otter, domestic dog, and domestic cat records, according to the methodologies employed, show a concentration pattern at the extremes of the reserve ( Figure 2 ). Overlapping detection of otters and domestic carnivorans was observed at the same site in three of the camera traps ( Figure 2A ). Furthermore, two otter hotspots were evident through direct observation, reinforcing the overlap of domestic dogs and cats over the hotspots of otters ( Figure 2B ) Finally, Figure 2C showed the location of the collection sites.

We investigated the exposure of L. longicaudis to domestic carnivorans and their pathogens inside a protected area using different methodologies. This issue is an emerging topic for otter conservation worldwide (Thomas et al., 2020; Calvo-Mac et al., 2020).

Although the capture rate of otters may be considered lower to similar methodologies or studies using different trap types, the captured individual showed no injuries related to trapping, different to what is frequently reported for other types of traps (Rutter et al., 2020). Carcass collection method proved to be an effective approach for obtaining biological samples from L. longicaudis in this reserve, which might be a potential way to obtain samples in other regions where wetlands are intersected by roads. This finding aligns with previous studies reporting that several species, particularly semi-aquatic ones, are susceptible of roadkill in this area (Bager and Rosa, 2010).

Serological results showed that four out of six otters tested positive for antibodies against T. gondii, indicating past exposure to the pathogen. The high seroprevalence, combined with frequent cat detections (recorded through multiple methodologies) suggests widespread fecal contamination of aquatic ecosystems with oocysts. Toxoplasma gondii is a globally distributed zoonotic pathogen known to cause significant morbidity and mortality in both humans and animals (Hill et al., 2005). To the best of our knowledge, this is the first report of free-ranging L. longicaudis with positive serology for T. gondii; since the occurrence of this pathogen in Neotropical otters was only recorded previously in captive animals (Alvarado-Esquivel et al., 2013; Silva et al., 2018). However, T. gondii has been frequently reported in several otter species, such as marine otter (Lontra felina), Eurasian otter (Lutra lutra), and particularly in sea otters (Enhydra lutris) with lethal outbreaks (Miller et al., 2004, 2008; Conrad et al., 2005; Chadwick et al., 2013; Calvo-Mac et al., 2020; Dubey et al., 2020).

Although all otter tested negative in serological tests for N. caninum, the parasite is known to be widespread in areas where domestic dogs and cattle are present (Wouda et al., 1999), as supported by camera trap records in the reserve. Previous studies have also reported N. caninum in wild carnivores, including mustelids (Almería, 2013). Although N. caninum has already been detected in the brain of a free-ranging L. lutra (Stuart et al., 2013), its pathogenic potential and conservation risk for otters have not yet been elucidated. Therefore, continued investigation into the presence of this parasite in otter populations is needed.

The presence of antibodies against CDV and CPV, alongside with the absence of viral RNA/DNA in molecular analyses, suggests that the sampled otters were exposed to these viruses in the past but were not actively shedding them at the time of sampling. The high seroprevalence of these viruses, typically associated with domestic dogs, supports the hypothesis of widespread exposure among otters in this region. This is further corroborated by frequent dog sightings recorded during the study. It also suggests that non-lethal exposure to these viruses occurs in Neotropical otters. Both CDV and CPV have caused significant past outbreaks and mortality in wild animals, including reported cases in L. longicaudis and other otter species (Roelke-Parker et al., 1996; Kimber et al., 2000; Tamukai et al,. 2021; De Bosschere et al., 2005; Echenique et al., 2018; Reck et al., 2025b). Indeed, Gras et al. (2018) assessing the role of landscape structures in disease spreading among wildlife found that areas in direct proximity to water bodies were identified as high-risk areas for CDV transmission. It may have a significant impact for semi-aquatic wild animals, such as otters.

Negative qPCR results in domestic dogs do not necessarily indicate an absence of viral infection in dogs roaming in the reserve but suggest that, at the time of sampling, it seems that these animals were not shedding the virus. This is an expected result, as the animals appeared healthy. Notably, during visits to surrounding farms, most of residents stated that they did not regularly vaccinate their dogs and cats (data not shown). Thus, considering that there are several vaccines against CDV and CPV available, the vaccination of dogs and cats from households near to protected areas may be considered also as a critical component of conservation strategies for wild carnivores.

Reports of FeLV infection in free-ranging and captive wild felids have been documented worldwide; however, the virus was only classically associated with felid species (Sacristán et al., 2021; Guimaraes et al., 2009; Reck et al., 2025a). For Lontra canadensis (North American river otter), there was one documented case showing that an individual with clinical signs tested positive for FeLV antibodies, raising the possibility of broader host susceptibility among other carnivoran species (Dalton et al., 1989). Although all L. longicaudis individuals in the present study tested negative for the FeLV, two domestic cats were positive, by both molecular and antigen tests. It may be hypothesized that the potential involvement of another retrovirus leading to false-positive FeLV antibody results in the L. canadensis cannot be ruled out. This finding underscores the need for further studies on retroviral infections in otters.

Camera trap footage revealed a high frequency of domestic dogs and cats moving within the reserve, a protected area where the presence of such animals is prohibited by law (Brasil, 1998). Some of the dogs recorded by the cameras were also observed on nearby farms where domestic animal samples were collected, indicating that these were free-ranging owned animals capable of moving long distances, such as between farms and inside the reserve. The occurrence of dogs and cats in protected areas has been increasingly reported across Brazil, raising concerns due to their negative impacts on wildlife, including predation, competition, disease transmission, and behavioral disturbance (Lessa et al., 2016). Our findings support this concern, given the serological evidence of high exposure to canine-associated pathogens in otters.

Spatial analyses highlighted the overlap of otter habitat with areas frequented by domestic dogs and cats. Kernel data showed that areas with the highest frequency of otter sightings had a higher frequency of domestic animals. This overlap suggests that domestic dogs and cats are moving inside key areas used by otters. The presence of domestic animals at the edges of the reserve is likely associated with their closer proximity to human settlements. In the case of otters, their higher occurrence in these areas warrants further investigation. When considered together with the camera trap data and the serological findings showing a high number of otters with antibodies against pathogens commonly associated with domestic animals, it reinforces the potential risk of inter-species pathogen transmission. The abundance of domestic carnivores on nearby farms pointed out the potential risk of their incursion into protected areas, as demonstrated by the camera trap results.

Our results support the hypothesis that otter’s exposure to CDV, CPV and T. gondii may result from spillover events originating from dogs and cats that roam near or inside the reserve. Although at this time we cannot definitely prove this hypothesis, further studies may help to solve this issue by conducting long-term monitoring and pathogen genetic characterization in otters. Nevertheless, the results made clear that Neotropical otters, even in protected areas, are certainly exposed to domestic carnivorans and their pathogens. It also reinforces the need for more effective strategies to prevent the presence of domestic animals in protected areas. As a conservation strategy, collaborative efforts with local communities may begin by sharing the results of studies like this, followed by the implementation of free or low-cost vaccination and sterilization campaigns for domestic animals. Moreover, it highlights the potential of the Neotropical otter as a sentinel species for health risk assessment, contributing to the monitoring of pathogen spillover and overall environmental quality in conservation areas.