Exploring the Pathogen Profiles of Ancient Feces.

Analysis of ancient desiccated feces - termed paleofeces or coprolites - can unlock insights into the lives of ancient people. We collected desiccated feces from caves in the Rio Zape Valley in Mexico (725-920 CE). First, we extracted DNA with methods previously optimized for paleofeces. Then, we applied highly sensitive modern molecular tools (i.e., PCR pre-amplification followed by multi-parallel qPCR) to assess the presence of 30 enteric pathogens. We detected ≥1 pathogen associated gene in each of the ten samples and a mean of 3.9 pathogens per sample. The targets detected included spp. (n=7), atypical enteropathogenic (n=7), (n=6), spp. (n=5), enterotoxigenic (n=5), spp./enteroinvasive (n=3), spp. (n=2), and O157:H7 (n=1). The protozoan pathogens we detected (i.e., spp. and spp.) have been previously detected in paleofeces via enzyme-linked immunoassay (ELISA), but have not via PCR. This work represents the first detection of spp. atypical enteropathogenic , enterotoxigenic spp./enteroinvasive , and O157:H7 in paleofeces. These results suggest that sensitive modern molecular tools, such as PCR, can be used to evaluate ancient materials for genes of interest.