Horizontal and Vertical Transmission Excluded as Primary Causes of Trypanosoma cruzi in Rhesus Macaques (Macaca mulatta) Housed Within a Texas Breeding Colony

Trypanosoma cruzi (T. cruzi) is a kinetoplasid protozoan parasite that infects both invertebrates and vertebrate hosts during its life cycle and is the causative agent for Chagas disease. Triatomine insects (kissing bugs) serve as vectors of T. cruzi and are distributed across the Americas including the southern United States. Kissing bugs feed on diverse wild animals, domestic animals, and humans, and hundreds of wild mammals serve as reservoirs of T. cruzi in nature. Across the southern US in regions where kissing bugs occur, biomedical research facilities housing non-human primates (NHPs) have increasingly been faced with T. cruzi infections among the NHPs, presenting major concerns for both the integrity of the animal models and for animal welfare. For this project, we tracked a population of T. cruzi seropositive rhesus macaques (Macaca mulatta) housed in an outdoor breeding colony in Texas to determine the potential for horizontal and vertical transmission. We determined the physical location of each seropositive animal within the colony at the time of their infection and also identified all animals housed with seropositive animals, including any offspring of seropositive dams. The animals housed with seropositive animals were deemed to be T. cruzi-associated monkeys (TAMs). The serostatus of all TAMs was determined through medical records or from the testing of archived banked serum samples. These serological data, along with spatial analyses of the seropositive and seronegative animals over time, were used to look for associations between animals that could support horizontal transfer, vertical transfer, or implicate environmental factors in the transmission of T. cruzi with the colony. A total of 80 breeding colony animals housed between 1999 and 2018 were identified to be seropositive for T. cruzi antibodies. From these 80 animals, 688 individual TAMs were identified to have interacted in a total of 925 animal-to-animal association events during the period covered in this study. These events were explored through the use of husbandry records, PCR, and identification of different discrete typing units (DTUs) to look for the potential of vertical and horizontal transmission within the colony. In review of the data related to the possibility of vertical transmission, there were 54 breeding-age, seropositive females identified in the colony that collectively produced 196 offspring. None of the offspring from these dams were found to be seropositive suggesting that vertical transmission either does not occur or is a rare event within this colony. In review of the data related to the possibility of horizontal transmission, it was found that horizontal transmission of T. cruzi may have occurred in as many as 25 of the 925 (2.70%) associations between naïve and seropositive animals, and that horizontal transmission may be the route of infection for as many as 25 of the 80 (31.25%) seropositive animals within the colony. These last two values are interpreted to represent the maximum number of horizontal transmission events that could have possibly occurred within the colony between 1999 and 2018, as these data are the result of a study-design that favored sensitivity over specificity, with regard to its inclusion of animals that were suspected to have become infected through horizontal transmission. Despite the fact that these values are likely overestimates of horizontal transmission within the colony, it is notable that even these data suggest horizontal transmission accounts for less than one-third of all T. cruzi infections and that horizontal transmission appears to be a very inefficient means of T. cruzi infection. Spatial analysis using the location of the T. cruzi seropositive animals at the time of their seroconversion was undertaken to identify potential spatial clusters (hot spots) of infections within the colony and to determine if there were aspects of the environment that influenced the incidence of infection or if new infections occurred at equal rates throughout the colony as a whole. Data from these analyses identified a few specific animal rooms within the colony that had increased incidences of infection. Notably, these hot spot rooms were adjacent to a wooded area and elevated storage sheds, both of which contained abundant leaf-litter and low growing vegetation which is presumed to have served as habitats for kissing bug vectors of the parasite. This presumption, that the colony infection rates are affected by increased exposure of animals to kissing bugs, was supported by the finding that there has been a marked decrease in the incidence of T. cruzi infections in the hot spot rooms adjacent to the wooded area since 2012 when the leaf-litter and low growing vegetation was cleared from this area as part of fire-mitigation efforts by the Center. In conclusion, this study found that hot spots of T. cruzi infection within the breeding colony from 1999-2018 correlate with areas of the local environment that are appreciated to be prime habitats for the insect vectors that transmit the parasite. The potential contribution of the horizontal route of transmission to the overall number of T. cruzi infections within the colony was determined to be 31.25% of infections, at most, and there was no evidence for the vertical route of T. cruzi transmission identified within the colony. Collectively, these data suggest that exposure to the triatomine insect vectors is the primary route of T. cruzi infection in rhesus macaques and that direct animal-to-animal transmission of T. cruzi is likely to be minimal concern for captive colonies in south Texas. The findings from this study are instrumental towards advancing the management practices of NHP biomedical research facilities and breeding programs in the southern United States by providing novel data as to the risks associated with maintaining seropositive animals in a research setting and by contributing to the literature in support of environmental modifications to limit the exposure of NHPs to infected triatomines