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This pathogen is considered endemic in the Kruger National Park (KNP), South Africa, where it threatens the health and conservation of African wildlife, including endangered species. This mycobacterial pathogen is spread through close contact with an infected host by aerosols or ingestion but may also be transmitted indirectly through the environment. Detecting environmental\n                      <jats:italic toggle=\"yes\">M. bovis<\/jats:italic>\n                      is challenging due to the complexity of the sample matrix and may require culture-independent techniques due to biosecurity restrictions associated with sample movement. In this study, DNA was extracted from soil near water sources in the KNP (\n                      <jats:italic toggle=\"yes\">n<\/jats:italic>\n                      = 180) and screened for\n                      <jats:italic toggle=\"yes\">Mycobacterium tuberculosis<\/jats:italic>\n                      complex (MTBC) DNA using the\n                      <jats:italic toggle=\"yes\">hsp65<\/jats:italic>\n                      polymerase chain reaction (PCR) followed by Sanger amplicon sequencing and the GeneXpert MTB\/RIF Ultra (GXU) qPCR assay. The region of difference (RD)-PCR was used to confirm whether\n                      <jats:italic toggle=\"yes\">M. bovis<\/jats:italic>\n                      DNA was present. Sanger\n                      <jats:italic toggle=\"yes\">hsp65<\/jats:italic>\n                      amplicon sequencing and GXU detected MTBC DNA in three and zero samples, respectively. Moreover, Sanger sequencing detected organisms belonging to the\n                      <jats:italic toggle=\"yes\">Mycobacterium<\/jats:italic>\n                      genus in 44 samples and non-tuberculous\n                      <jats:italic toggle=\"yes\">Mycobacterium<\/jats:italic>\n                      species in 21 samples. The RD-PCR confirmed\n                      <jats:italic toggle=\"yes\">M. bovis<\/jats:italic>\n                      DNA presence in 2\/3 MTBC-positive samples. The presence of\n                      <jats:italic toggle=\"yes\">M. bovis<\/jats:italic>\n                      DNA in KNP soil suggests potential environmental contamination by shedding from infected animals. Further research is required to confirm the viability of MTBC and the role of environmental contamination in the TB epidemiology of this multi-host system.\n                    <\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>IMPORTANCE<\/jats:title>\n                    <jats:p>\n                      This article describes the first evidence that DNA from the bacterial pathogen,\n                      <jats:italic toggle=\"yes\">Mycobacterium bovis<\/jats:italic>\n                      , which causes animal tuberculosis (TB) in wildlife, can be detected in soil from the Kruger National Park (KNP). Animal TB threatens wildlife conservation, including threatened and endangered species, in areas such as the KNP. Pathogenic\n                      <jats:italic toggle=\"yes\">Mycobacterium<\/jats:italic>\n                      are spread primarily through direct contact with infected hosts. However, the presence of\n                      <jats:italic toggle=\"yes\">M. bovis<\/jats:italic>\n                      DNA in KNP soil could indicate a role for the environment in disease transmission. This complements the growing evidence from European regions that\n                      <jats:italic toggle=\"yes\">M. bovis<\/jats:italic>\n                      can be shed by infected animals into water, soil, or plant material and potentially infect animals in the surrounding environment. This indirect route of spread has implications for disease management strategies and warrants further scientific investigation. Moreover, the direct DNA-based detection techniques described in this study may provide a tool for detecting\n                      <jats:italic toggle=\"yes\">Mycobacterium<\/jats:italic>\n                      pathogens using non-invasive sampling (sampling the environment rather than animals directly) when culturing is not possible.\n                    <\/jats:p>\n                  <\/jats:sec>","DOI":"10.1128\/spectrum.01658-25","type":"journal-article","created":{"date-parts":[[2025,12,3]],"date-time":"2025-12-03T14:00:31Z","timestamp":1764770431000},"update-policy":"https:\/\/doi.org\/10.1128\/asmj-crossmark-policy-page","source":"Crossref","is-referenced-by-count":2,"title":["A microbial safari: finding evidence of\n                    <i>Mycobacterium bovis<\/i>\n                    DNA in soil from the Kruger National Park, South Africa"],"prefix":"10.1128","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-8994-2293","authenticated-orcid":true,"given":"M. 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