Transferable Extended-Spectrum β-Lactamase (ESBL) Plasmids in Enterobacteriaceae from Irrigation Water

Extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae are considered serious threats to human health. Water used for irrigation of fresh produce can transmit such antibiotic resistant bacteria (ARB) directly to edible plant parts, which in turn can transmit ARB to the consumer, where the ingested ARB can interact with the gut microbiome. The aim of our study therefore was to assess the potential of ESBL-producing Enterobacteriaceae from irrigation water to transfer resistance to susceptible Escherichia coli.

We screened 19 ESBL-producing E. coli, Enterobacter cloacae, and Citrobacter freundii isolated from irrigation water for their ability to transmit resistance to third-generation cephalosporins on solid agar as well as in liquid culture. Additionally, donor strains and resulting transconjugants were genome-sequenced (PacBio and Illumina) to identify the underlying antibiotic resistance genes (ARGs) and plasmids. Additionally, phenotypic resistance patterns of donors and corresponding transconjugant strains were compared.

Of the 19 ESBL-producing Enterobacteriaceae, five E. coli were able to transfer resistance to susceptible recipient E. coli, with transmission efficiency ranging from 2.3 × 10⁻⁹ to 4.1 × 10⁻⁵ transconjugants per recipient cell. The obtained transconjugants all displayed ESBL-producing phenotype, as determined by broth microdilution. Genome-sequencing revealed that each of the five donors which yielded transconjugants carried a plasmid containing a β-lactamase gene of the CTX-M family, which encode class A ESBLs conferring resistance to third-generation cephalosporins. The most frequent ESBL gene was bla_CTX-M-1 (3 of 5), and the corresponding plasmids all belonged to incompatibility group I1 (IncI1), while the other two plasmids carried IncFIA and/or IncFIB. Importantly, all but one of the five plasmids encoded additional ARGs (up to nine in total). Finally, all ARGs encoded by the bla_CTX-M-carrying plasmid could be detected in the genome sequences of the respective transconjugants. Overall, our findings highlight the importance of monitoring water quality to avoid contamination of irrigated produce with conjugative ESBL-encoding plasmids.