Hydrothermal carbonization (HTC), utilizing high temperature and pressure, has the potential to treat agricultural waste via inactivating pathogens, antibiotic resistance genes (ARG), and contaminants of emerging concern (CEC) in a environmental and economical manner. Livestock mortality is one facet of agricultural waste that can pose a threat to the surrounding environment. While several methods are utilized to treat livestock mortality, there remains a paucity of data on the elimination of microbially-derived DNA in these treatment practices. This DNA, most notably ARGs, if it survives treatment can be reintroduced in agricultural environments where it could potentially be passed to pathogens, posing a risk to animal and human populations. HTC treatments have been successfully utilized for the treatment of CECs, however very little is understood on how ARGs survive HTC treatment. This study aims to fill this knowledge gap by examining the survivability of microbially-derived DNA in the HTC treatment of livestock mortality. We examined three treatment temperatures (100°C, 150°C, and 200°C) at autogenic pressures at three treatment times (30, 60, and 240 min). We examined the amplification of a plasmid-borne reporter gene carried by DH10B introduced to both beef bone and tissue. Results indicate that while all three temperatures, at all treatment times, were suitable for complete pathogen kill, only temperatures of 150°C and 200°C were sufficient for eliminating microbial DNA. These results serve as the basis for future potential HTC treatment recommendations for livestock mortality when considering the elimination of pathogens and ARGs.