Salmonella spp. are gram-negative bacteria capable of causing diseases in a wide range of aquatic and terrestrial animals, including humans. Sea and terrestrial turtles have been recognized as carriers of this zoonotic pathogen. In this project, conventional and molecular diagnostic methods were combined to investigate the prevalence of Salmonella enterica in leatherback sea turtles (Dermochelys coriacea) that used the island of St. Kitts, West Indies as a nesting ground during 2011 (n = 21). Isolates obtained from selective media were screened and colonies suspected of being Salmonella spp. were confirmed by fluorescence resonance energy transfer polymerase chain reaction. The prevalence of S. enterica within this sample population during this period was found to be 14.2%. Moreover, due to the increasing risk of antibiotic resistance in enteric bacteria, antimicrobial susceptibility was investigated in all recovered Salmonella spp. isolates utilizing the broth microdilution method. All isolates were susceptible to the lowest concentration of kanamycin, gentamicin, ciprofloxacin, enrofloxacin, nalidixic acid, and trimethoprim/sulfamethoxazole tested. Further research should be pursued to understand the interaction of this bacterial pathogen with the environment, host, and other microbial communities, and to further develop faster, more sensitive, and more specific diagnostic methods.