Molecular detection of extended-spectrum β-Lactamase genes in multi-drug-resistant bacteria isolated from drinking water supply and surface water systems in Addis Ababa, Ethiopia

BACKGROUND: The emergence of β-lactamase-producing bacteria is a critical global public health threat that undermines the efficacy of last-resort antibiotics. In Ethiopia, studies exploring resistance genes in drinking water systems are scarce. This study investigated bacterial profiles, antimicrobial susceptibility, and associated Extended-Spectrum β-Lactamase (ESBL) genes, including blaTEM, blaCTXM and blaSHV. METHODS: A cross-sectional study was conducted between March 2023 and April 2024 by sampling raw, finished, reservoir, tap, and storage tank water in Addis Ababa. The samples were processed aseptically with routine microbiological culture and characterization methods. Antibiotic susceptibility testing was conducted using standardized Kirby-Bauer disc diffusion method as described by Clinical Laboratory Standards Institute (CLSI) version 33 guidelines. All data recorded were validated and then analyzed by Statistical Package for the Social Sciences version 25.0 Armonk. The associated parameters were compared using Fishers' exact test to determine their correlation. In all cases, a p-value less than or equal to 0.05 was considered statistically significant. RESULTS: From 45 samples, 94 bacterial species were characterized. Dominant isolates included E. coli (23.4%), Enterobacter spp. (16%), Citrobacter spp. (10.6%), K. pneumoniae (10.6%), Proteus spp. (8.5%), and Coagulase negative Staphylococcus species. Multi-drug resistant (MDR) is a non-susceptibility of an isolate to at least one antibiotic agent in three or more antimicrobial classes. Disc diffusion tests revealed that 63.8% (60/94) of isolates were MDR. The highest resistance levels were observed for ampicillin (86%), tetracycline (72.7%), cefuroxime (66.2%), and cefoxitin (53.5%). The principal MDR isolates were E. coli, Enterobacter, and Klebsiella species. Multiplex PCR on genomic DNA of MDR isolates showed a 26.6% (16/60) occurrence of ESBL genes; the combination of blaCTXM + blaTEM 11.7% (7/60) being the most frequent, followed by blaTEM (10%), blaCTXM (3.3%), and blaSHV (1.7%). In plasmid DNA of Gram-negative MDR isolates, ESBL genes occurred in 10.9% (6/54), with blaTEM (7.3%) being dominant, followed by blaCTXM (0.18%) and co-occurring blaSHV + blaTEM (0.18%). CONCLUSION: This study confirms the presence of ESBL genes in both the genomic and plasmid DNA of MDR bacteria in the water supply, particularly in storage tanks, tap and raw water. These findings indicate the transmission of β-lactamase-producing bacteria into drinking water supply systems. Regular monitoring, surveillance, and rigorous studies to identify contamination sources and mitigation strategies are urgently recommended.