This study presents the first comprehensive molecular characterization of Escherichia coli producing extended-spectrum beta-lactamases (ESBL-Ec) in surface waters in Albania, focusing on the Shkumbini river. Antimicrobial resistance (AMR) in aquatic ecosystems poses a significant threat to public health, yet data from Albania remain scarce. Thirty water samples were collected from six locations near Elbasan between September 2022 and February 2024. Following the WHO Tricycle protocol, 52 ESBL-Ec isolates were recovered and characterized for antimicrobial susceptibility, biofilm formation, resistance genotypes and clonal relatedness via pulsed-field gel electrophoresis (PFGE). ESBL-Ec was detected in 80% of the samples analyzed, with 94.2% of the isolates classified as multidrug-resistant (MDR). High resistance frequencies were observed for ampicillin (98.1%) and cefotaxime (86.5%), while 7.7% of the isolates displayed colistin resistance associated with the mcr-3 gene. The bla_CTX-M-1 genotype was the most prevalent (57.7%), and almost half of the isolates harbored multiple ESBL genes. Phylogroup A (46.2%) predominated, followed by the high-risk extraintestinal lineages B2 (23.1%) and D (11.5%). PFGE revealed high genetic heterogeneity, with 51 distinct pulsotypes indicating multiple sources of contamination, such as untreated municipal, agricultural and industrial waste. Additionally, 55.8% of the isolates were capable of forming biofilms. These results highlight the critical role of the Shkumbini river as a reservoir for highly resistant pathogens and emphasize the urgent need for integrated environmental surveillance and improved wastewater management in Albania.

This study presents the first comprehensive molecular characterization of Escherichia coli producing extended-spectrum beta-lactamases (ESBL-Ec) in surface waters in Albania, focusing on the Shkumbini river. Antimicrobial resistance (AMR) in aquatic ecosystems poses a significant threat to public health, yet data from Albania remain scarce. Thirty water samples were collected from six locations near Elbasan between September 2022 and February 2024. Following the WHO Tricycle protocol, 52 ESBL-Ec isolates were recovered and characterized for antimicrobial susceptibility, biofilm formation, resistance genotypes and clonal relatedness via pulsed-field gel electrophoresis (PFGE). ESBL-Ec was detected in 80% of the samples analyzed, with 94.2% of the isolates classified as multidrug-resistant (MDR). High resistance frequencies were observed for ampicillin (98.1%) and cefotaxime (86.5%), while 7.7% of the isolates displayed colistin resistance associated with the mcr-3 gene. The bla_CTX-M-1 genotype was the most prevalent (57.7%), and almost half of the isolates harbored multiple ESBL genes. Phylogroup A (46.2%) predominated, followed by the high-risk extraintestinal lineages B2 (23.1%) and D (11.5%). PFGE revealed high genetic heterogeneity, with 51 distinct pulsotypes indicating multiple sources of contamination, such as untreated municipal, agricultural and industrial waste. Additionally, 55.8% of the isolates were capable of forming biofilms. These results highlight the critical role of the Shkumbini river as a reservoir for highly resistant pathogens and emphasize the urgent need for integrated environmental surveillance and improved wastewater management in Albania.