Traditional zoonotic disease research focuses on detection of recognized pathogens and may miss opportunities to understand broader microbial transmission dynamics between humans, animals, and the environment. We studied human-macaque microbiome overlap in Kosum Phisai District, Maha Sarakham Province, Thailand, where a growing population of long-tailed macaques (Macaca fascicularis) in Kosumpee Forest Park interact with humans from an adjacent village. We surveyed workers in or near the park with elevated exposure to macaques to characterize tasks resulting in exposure to macaque feces in addition to dietary and lifestyle factors that influence gut microbiome composition. Fecal samples were collected from 12 exposed workers and 6 controls without macaque exposure, as well as 8 macaques from Kosumpee Forest Park and 4 from an isolated forest patch with minimal human contact. The V4 region of the 16S rRNA gene from fecal sample extracted DNA was amplified and sequenced using Illumina MiSeq to characterize the microbial community. A permuted betadisper test on the weighted UniFrac distances revealed significant differences in the dispersion patterns of gut microbiota from exposed and control macaques (p = 0.03). The high variance in gut microbiota composition of macaques in contact with humans has potential implications for gut microbiome stability and susceptibility to disease, described by the Anna Karenina principle (AKP). Human samples had homogenous variance in beta diversity but different spatial medians between groups (p = 0.02), indicating a shift in microbial composition that may be explained by fundamental lifestyle differences between the groups unrelated to exposure status. SourceTracker was used to estimate the percent of gut taxa in exposed humans that was contributed by macaques. While one worker showed evidence of elevated contribution, the overall trend was not significant. Task observations among workers revealed opportunities to employ protective measures or training to reduce exposure to occupational hazards. These results suggest the potential for hygiene measures to mitigate negative aspects of contact between humans and macaques in order to optimize the health of both populations.