Detection and genetic characterization of blaESBL-carrying plasmids of cloacal Escherichia coli isolates from white stork nestlings (Ciconia ciconia) in Spain
Sandra Martínez-Álvarez, Pierre Châtre, Teresa Cardona-Cabrera, Pauline François, Alberto Sánchez‐Cano, Úrsula Höfle, Myriam Zarazaga, Jean-Yves Madec, Marisa Haenni, Cármen Torres
Abstract
This study aims to characterize Escherichia coli isolates from cloacal samples of white stork nestlings, with a special focus on ESBL-producing E. coli isolates and their plasmid content. Cloacal samples of 88 animals were seeded on MacConkey-agar and chromogenic-ESBL plates to recover Escherichia coli and ESBL-producing E. coli. Antimicrobial susceptibility was screened by disc-diffusion-method and the genotypic characterization were assessed by PCR-sequencing. S1-nuclease-PFGE, Southern blot and conjugation essays were performed on ESBL-E. coli, as well as whole-genome-sequencing by short- and long-reads. The four blaESBL-carrying plasmids were completely sequenced. A total of 113 non-ESBL-producing E. coli isolates were collected on antibiotic-free MacConkey-agar, of which 27 (23.9%) showed a multidrug-resistance phenotype, mainly associated with beta-lactam-phenicol-sulfonamide resistance (blaTEM/cmlA/floR/sul1/sul2/sul3). Moreover, four white stork nestlings carried ESBL-producing E. coli (4.5%) with the following characteristics: blaSHV-12/ST38-D, blaSHV-12/ST58-B1, blaCTX-M-1/ST162-B1 and blaCTX-M-32/ST155-B1. Whole-genome sequencing followed by Southern blot hybridization on S1-PFGE gels on ESBL-positive isolates proved that the blaCTX-M-1 and one blaSHV-12 gene were carried by IncI1/pST3 plasmids, while the second blaSHV-12 and the blaCTX-M-32 gene located on IncF plasmids. The two blaSHV-12 and the two blaCTX-M genes had similar but non-identical close genetic environments, all four genes being flanked by insertion sequences. The role played by several genetic platforms in the mobility of ESBL genes, allows for interchangeability on a remarkably small scale (gene-plasmid-clones), which may support the spread of ESBL genes.