3.1 bTB history in the park
From the investigation of official records, we delineated a timeline of all TB cases that culminated in animal deaths from 2003 to 2018 in the safari park (Figure 2). In brief, from 2003 to 2015, at least 16 animals, from 8 different species, died or were humanely euthanized due to TB. Except for two llamas in 2012, TB was only diagnosed based on necropsy findings and Ziehl-Neelsen staining of tissue samples (Figure 2). It is possible to observe an increase in the number of cases starting around 2005, which coincides with management changes in the park (personal communication of a previous employee). These changes resulted in overpopulation (particularly of deer) and poor feeding practices. In 2013, due to widespread TB, other sanitary problems and selling of deer to properties in different Brazilian states, the official veterinary service, through a court order, demanded the closure of the park to visitations and halting of animal trade. From 2013 onwards, many tuberculin skin tests were performed in deer, cattle and buffalo. However, as the PNCEBT solely covers the latter two species, only these were mandatorily culled. The deer tested positive by TST in 2013, 2014 and 2015, and recommendations to contain the outbreak were made to the park. In 2018, after the official veterinary service noted the park did not follow through with the recommendations, they ordered depopulation of the park deer based on interstate trade of potentially infected animals. Tissue from necropsy of the two llamas and the culled deer were the only animal samples ever subjected to M. bovisculture. A more complete history of all TB cases of the park is described in Supplementary material.