This study investigated the prevalence, serovar distribution, antimicrobial resistance, and pulsed field gel electrophoresis (PFGE) typing of Salmonella enterica isolated from Lake Zapotlán, Jalisco, Mexico. Additionally, the association of the presence of Salmonella with physicochemical and environmental parameters was analyzed using Pearson correlation analysis and principal component analysis (PCA). Salmonella spp. were identified in 19 of 63 (30.15%) samples. The prevalence of Salmonella was positively correlated with air temperature, electrical conductivity, pH, and dissolved oxygen and negatively correlated with relative humidity, water temperature, turbidity, and precipitation. The predominant serotype identified was Agona (68.48%), followed by Weltevreden (5.26%), Typhimurium (5.26%), and serogroup B (21.05%). Overall, the highest detected antimicrobial resistance was toward colistin (73.68%), followed by sulfamethoxazole (63.15%), tetracycline (57.89%), nalidixic acid (52.63%), and trimethoprim (52.63%). All Salmonella strains were genetically diverse, with a total of 11 XbaI and four BlnI profiles on PFGE. The use of these two enzymes allowed differentiate strains of Salmonella of the same serotype. The results obtained in this study contribute to a better understanding of the Salmonella spp. ecology in an endorheic subtropical lake and provide information for decision makers to propose and implement effective strategies to control point and non-point sources of pathogen contamination.