Quantification of β-lactamase producing bacteria in German surface waters with subsequent MALDI-TOF MS-based identification and β-lactamase activity assay

• Published in: Heliyon Vol. 10; no. 5; p. e27384
• Main Authors: Stelmaszyk, Lara, Stange, Claudia, Hügler, Michael, Sidhu, Jatinder P.S., Horn, Harald, Tiehm, Andreas
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 15-03-2024; Elsevier
• Subjects: Antibiotic resistance; Culture-based methods; Environmental bacteria; MALDI-TOF MS; Oligotrophic bacteria; Pathogens; qPCR; β-lactamases; β-lactamases; Environmental bacteria; MALDI-TOF MS; Pathogens; Culture-based methods; Antibiotic resistance; Oligotrophic bacteria; qPCR
• ISSN: 2405-8440; 2405-8440
• DOI: 10.1016/j.heliyon.2024.e27384

Environmental oligotrophic bacteria are suspected to be highly relevant carriers of antimicrobial resistance (AMR). However, there is a lack of validated methods for monitoring in the aquatic environment. Since extended-spectrum β-lactamases (ESBLs) play a particularly important role in the clinical sector, a culturing method based on R2A-medium spiked with different combinations of β-lactams was applied to quantify β-lactamase-producing environmental bacteria from surface waters. In German surface water samples (n = 28), oligotrophic bacteria ranging from 4.0 × 103 to 1.7 × 104 CFU per 100 mL were detected on the nutrient-poor medium spiked with 3rd generation cephalosporins and carbapenems. These numbers were 3 log10 higher compared to ESBL-producing Enterobacteriales of clinical relevance from the same water samples. A MALDI-TOF MS identification of the isolates demonstrated, that the method leads to the isolation of environmentally relevant strains with Pseudomonas, Flavobacterium, and Janthinobacterium being predominant β-lactam resistant genera. Subsequent micro-dilution antibiotic susceptibility tests (Micronaut-S test) confirmed the expression of β-lactamases. The qPCR analysis of surface waters DNA extracts showed the presence of β-lactamase genes (blaTEM, blaCMY-2, blaOXA-48, blaVIM-2, blaSHV, and blaNDM-1) at concentrations of 3.7 (±1.2) to 1.0 (±1.9) log10 gene copies per 100 mL. Overall, the results demonstrate a widespread distribution of cephalosporinase and carbapenemase enzymes in oligotrophic environmental bacteria that have to be considered as a reservoir of ARGs and contribute to the spread of antibiotic resistance.
[Display omitted]
•β-lactam resistant oligotrophic bacteria are widespread in German surface waters.•ESBL-Enterobacteriales were detected in 3 log-level lower concentration.•Pseudomonas, Flavobacterium and Janthinobacterium were identified with MALDI-TOF MS.•Expression of β-lactamases was proofed in resistant environmental isolates.•Cephalosporine and carbapenem resistance genes were detected in water samples.