The prevalence of bacteria with multidrug-resistance (MDR) is a significant threat to public health globally. Listeria spp. are naturally ubiquitous, with L. monocytogenes particularly being ranked as important foodborne disease-causing microorganisms. This study aimed to evaluate the incidence and determine the antimicrobial resistance (AMR) profiles of multidrug-resistant Listeria spp. (MDRL) isolated from different environmental samples (river and irrigation water) in the Sarah Baartman District Municipality (SBDM), Eastern Cape Province (ECP), South Africa. Molecular identification and characterization were carried out using polymerase chain reaction (PCR) and isolates that exhibited phenotypic resistance were further screened for relevant antimicrobial-resistant genes (ARGs). Findings revealed a total of 124 presumptive Listeria isolates; 69 were molecularly confirmed Listeria species. Out of the confirmed species, 41 isolates (59%) were classified as L. monocytogenes while 9 (13%) were classified as L. welshimeri. All Listeria spp. exhibited phenotypic resistance against ampicillin, penicillin, and trimethoprim-sulphamethoxazole and further screening revealed ARGs in the following proportions: sulI (71%), blaTEM (66%), tetA (63%), and blaCIT (33%). Results confirmed the occurrence of ARGs among Listeria inhabiting surface waters of ECP. The present study indicates that the river water samples collected from SBDM are highly contaminated with MDRL, hence, constituting a potential health risk.
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