Salmonellosis is an important zoonotic infection, and exposure to pet reptiles has been implicated in several human outbreaks. Although several studies report a low prevalence of salmonellae in free-ranging chelonians, they may serve as a reservoir. In spring and summer of 2013 and 2019, free-ranging eastern box turtles () from populations in Illinois (rural) and Tennessee (urban) were collected through canine and visual search. Cloacal swab samples were collected from each turtle, selectively enriched with tetrathionate broth, then plated on selective and differential media to isolate spp. Genus was confirmed via MALDI-TOF MS and antibiotic sensitivities were performed. Isolates were serotyped by the National Veterinary Services Laboratory. Of the 341 turtles sampled, spp. were detected in nine individuals (2.64%; 95% CI: 1.2-5.0%). The isolates included five different serovars: Anatum ( = 2), Newport ( = 2), Thompson ( = 1), Bareilly ( = 2), and Hartford ( = 2). spp. were detected from six animals in 2013 (3.19%, 95% CI: 1.2-6.8%) and three in 2019 (1.96%, 95% CI: 0.4-5.6%). There was no significant difference in prevalence between state, ( = 0.115), Illinois locations ( = 0.224), season ( = 0.525), year ( = 0.297), sex ( = 0.435), or age class ( = 0.549). The health of -positive and -negative turtles was not significantly different, as assessed through hematology and plasma biochemistry ( > 0.05), indicating asymptomatic carrier status. The low prevalence detected in this study likely concludes that free-ranging eastern box turtles play a minimal role in the spread of salmonellae. However, the identified serotypes are potentially human- and animal-pathogenic. Documenting the prevalence of serotypes in animal indicators furthers our understanding of their spread between humans, animal agriculture, and the environment.

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