title: Short communication: Heat-resistant Escherichia coli as potential persistent reservoir of extended-spectrum β-lactamases and Shiga toxin-encoding phages in dairy.
abstract: Here we report the isolation of heat-resistant Escherichia coli from raw milk cheeses. Detection of the heat-resistance markers clpK and orfI by PCR was followed by phenotypical confirmation of increased heat-resistance. These strains were Shiga toxin-negative and, although several were found to be multidrug resistant, no plasmids encoding extended-spectrum β-lactamases (ESBL) were found in any of the isolates. The aim of this study was to assess the potential of these strains to acquire ESBL plasmids and a modified Shiga toxin-encoding phage. Only 4 ESBL-encoding, heat-sensitive E. coli strains were isolated from 1,251 dairy samples (2/455 raw milk and 2/796 raw milk cheese samples). One incompatibility group FII plasmid (CTX-M-14, 79.0 kb) and 3 incompatibility group I1 plasmids (CTX-M-15, 95.2, 96.1, and 97.8 kb) were fully sequenced and de novo assembled. All 4 plasmids are readily transferred to heat-resistant E. coli isolates in plate matings (9.7×10-5to 3.7×10-1exconjugants per recipient) and, to a lesser extent, in milk (up to 7.4×10-5exconjugants per recipient). Importantly, the plasmids are stably maintained during passaging in liquid media without antimicrobial pressure. The heat-resistant isolate FAM21805 was also shown to be capable of acting as donor of all 4 ESBL plasmids. In addition, 3 of 11 tested ESBL exconjugants of heat-resistant strains were lysogenized by the modified Shiga toxin-encoding phage 933W ∆stx::gfp::cat. The higher fraction of heat-resistant E. coli (93 of 256 isolates) compared with the estimated 2% previously predicted based on genomic prevalence of heat resistance genes seems to indicate a selection advantage in the raw milk cheese production environment. The combination of 2 factors may lead to said advantage: increased survival during thermization of raw milk (heating to subpasteurization temperatures) and increased survival rates during cheese ripening. Should these strains acquire ESBL-encoding plasmids, Shiga toxin-encoding phages, or both, these genetic elements would profit from the selection advantage of their host and become more abundant in this particular environment, which in turn could lead to an increased threat to consumers of raw milk products.
authors: Agroscope, Institute for Food Sciences IFS, Schwarzenburgstrasse 161, CH-3003 Bern, Switzerland.
authors: Roger
authors: R
authors: Marti
authors: Department of Microbiology, University of Barcelona, 08007 Barcelona, Spain.
authors: Maite
authors: M
authors: Muniesa
authors: Agroscope, Institute for Plant Production Sciences IPS and Swiss Institute of Bioinformatics, Schloss 1, 8820 Wädenswil, Switzerland.
authors: Michael
authors: M
authors: Schmid
authors: Agroscope, Institute for Plant Production Sciences IPS and Swiss Institute of Bioinformatics, Schloss 1, 8820 Wädenswil, Switzerland.
authors: Christian H
authors: CH
authors: Ahrens
authors: Agroscope, Institute for Food Sciences IFS, Schwarzenburgstrasse 161, CH-3003 Bern, Switzerland.
authors: Javorka
authors: J
authors: Naskova
authors: Agroscope, Institute for Food Sciences IFS, Schwarzenburgstrasse 161, CH-3003 Bern, Switzerland. Electronic address: joerg.hummerjohann@agroscope.admin.ch.
authors: Jörg
authors: J
authors: Hummerjohann
mesh_terms: D000818:Animals; D002611:Cheese / Q000382:microbiology*; D024901:Drug Resistance, Multiple, Bacterial / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D005798:Genes, Bacterial; D006358:Hot Temperature; D008892:Milk / Q000382:microbiology; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D022621:Shiga Toxin / Q000235:genetics* / Q000378:metabolism; D054323:Shiga-Toxigenic Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*; D001618:beta-Lactamases / Q000235:genetics* / Q000378:metabolism
chemical_list: D029968:Escherichia coli Proteins; D022621:Shiga Toxin; D001618:beta-Lactamases
keywords: Escherichia coli; dairy; extended spectrum β-lactamase; heat resistance; horizontal gene transfer