Description
Introduction: The rise of antibiotic resistance poses a global public health challenge. Wastewater environments, often overlooked, serve as reservoirs for antibiotic-resistant bacteria, fostering the dissemination of resistance genes and posing health risks through various exposure routes. Our study, conducted in Morocco, focuses on the phenotypic and genotypic characterization of antibiotic resistance and biofilm formation in Escherichia coli (E. coli) strains isolated from wastewater samples. This research underscores the pivotal role of wastewater in the perpetuation and dissemination of antibiotic resistance, emphasizing the necessity for nuanced wastewater management strategies to curb resistance propagation and safeguard public health.
Methodology: Over a period of two months, from the 1st February to 31st March 2023, wastewater samples were systematically collected from two distinct sites, Anza and Oued Lahouar, situated in the Agadir city. These sites were purposefully chosen for their representativeness in terms of effluents originating from both human and animal sources. The geographical coordinates of the sampling sites are as follows: Site A: Anza (Latitude: 30.45431; Longitude: -9.65362) and Site B: Oued Lahouar (Latitude: 30.40016; Longitude: -9.60008). Isolation of E. coli strains involved the use of a chromogenic medium Brilliance UTI Clarity Agar (Oxoid). Identification was achieved through biochemical and 16S rRNA methods. The antimicrobial susceptibility by disc diffusion method on Mueller-Hinton (MH) agar (Bio-Rad), and the VITEK® 2 system (bioMérieux), was conducted according to EUCAST 2022 guidelines. The extraction of total DNA was carried out using magnetic bead technology (MagSi-NA Pathogens 96 preps, magtivio), PCR assays were conducted to identify genes encoding resistance to β-lactams (blactx-M, blaTEM, blaSHV), elucidating the genetic basis of resistance patterns. Furthermore, the kinetics of biofilm formation were studied on an abiotic surface (polystyrene microplate). E. coli isolates were incubated on the microplate at various time points ranging from 2 to 24 hours. To assess the strength and kinetics of biofilm formation, absorbance at 620 nm was measured using a microplate reader (800 TS Microplate Reader) and corrected by the absorbance of control wells.
Results: In the E. coli strains isolated, consisting of a total of 14 isolates, varied resistance rates were observed towards different β-lactams (AMP, AMC, CTX, FEP, ETP), with the lowest resistance recorded for Ertapenem (7.14%). Reduced resistance was noted for aminoglycosides (14.29%). Fluoroquinolones maintained activity at 28.57% for Ciprofloxacin and Norfloxacin. However, a relatively higher resistance rate of 50.00% was observed for Trimethoprim-Sulfamethoxazole. Genetic analysis revealed the presence of blactx-M1 in one strain. Biofilm formation analysis indicated moderate formation (20%) after 4h and 22h of incubation. Most environmental strain biofilms initiated between 4h to 18h, resuming normal cycles. Weak biofilm formation peaked at 6h, 14h, and 16h without reaching maximum adherence.
Conclusion: This study underscores the critical role of wastewater as a reservoir for antibiotic-resistant E. coli, emphasizing the need for tailored wastewater management strategies to mitigate resistance dissemination and protect public health.
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