No abstract

Dissemination of extended-spectrum-cephalosporin (ESC)-resistant Salmonella, especially extended-spectrum-β-lactamase (ESBL)-producing Salmonella, is a concern worldwide. Here, we assessed Salmonella carriage by food workers in Japan to clarify the prevalence of ESC-resistant Salmonella harboring blaCTX-M. We then characterized the genetic features, such as transposable elements, of blaCTX-M-harboring plasmids using whole-genome sequencing. A total of 145,220 stool samples were collected from food workers, including cooks and servers from several restaurants, as well as food factory workers, from January to October 2017. Isolated salmonellae were subjected to antimicrobial susceptibility testing (disk diffusion method), and whole-genome sequencing was performed for Salmonella strains harboring blaCTX-M. Overall, 164 Salmonella isolates (0.113%) were recovered from 164 samples, from which we estimated that at least 0.113% (95% confidence interval [CI]: 0.096 to 0.132%) of food workers may carry Salmonella. Based on this estimation, 3,473 (95% CI = 2,962 to 4,047) individuals among the 3,075,330 Japanese food workers are likely to carry Salmonella. Of the 158 culturable isolates, seven showed resistance to ESCs: three isolates harbored blaCMY-2 and produced AmpC β-lactamase, while four ESBL-producing isolates harbored blaCTX-M-14 (n = 1, Salmonella enterica serovar Senftenberg) or blaCTX-M-15 (n = 3, S. enterica serovar Haardt). blaCTX-M-15 was chromosomally located in the S. Haardt isolates, which also contained ISEcp1, while the S. Senftenberg isolate contained an IncFIA(HI1)/IncHI1A/IncHI1B(R27) hybrid plasmid carrying blaCTX-M-14 along with ISEcp1. This study indicates that food workers may be a reservoir of ESBL-producing Salmonella and associated genes. Thus, these workers may contribute to the spread of blaCTX-M via plasmids or mobile genetic elements such as ISEcp1. IMPORTANCE Antimicrobial-resistant Salmonella bacteria arise in farm environments through imprudent use of antimicrobials. Subsequently, these antimicrobial-resistant strains, such as extended-spectrum-β-lactamase (ESBL)-producing Salmonella, may be transmitted to humans via food animal-derived products. Here, we examined Salmonella carriage among food handlers in Japan. Overall, 164 of 145,220 fecal samples (0.113%) were positive for Salmonella. Among the 158 tested isolates, four were identified as ESBL-producing isolates carrying ESBL determinants blaCTX-M-15 or blaCTX-M-14. In all cases, the genes coexisted with ISEcp1, regardless of whether they were located on the chromosome or on a plasmid. Our findings suggest that food workers may be a reservoir of ESBL-producing strains and could contribute to the spread of resistance genes from farm-derived Salmonella to other bacterial species present in the human gut.

Salmonella enterica serovar Agona strains isolated from human cases were compared to strains that were derived from a clone caused a serovar shift in broilers. Pulsed field gel electrophoresis (PFGE) analysis with XbaI or BlnI digestion showed that three of seven strains from human case strains and most of the 81 strains from broilers were clustered in single complex in a minimum spanning tree (MST) reconstructed from the PFGE data. All the strains from human cases and 22 randomly selected strains from broilers were also analyzed by whole genome sequencing (WGS). Analysis of single nucleotide polymorphism (SNP) in the S. Agona core genes showed that four strains from human cases and all the strains from broilers were clustered in a maximum likelihood phylogenetic tree (ML tree) and an MST. These results indicated that the strains derived from the clone caused the serovar shift had already spread to humans. PFGE analysis with XbaI showed that four strains from broilers did not cluster with the other strains in an MST, though all those strains clustered in an ML tree and an MST reconstructed from SNP data. Moreover, three strains from broilers did not cluster in an MST reconstructed from PFGE with BlnI digestion, though those strains clustered in an ML tree and an MST reconstructed from SNP data. Therefore, it was suggested that S. Agona strains derived from a particular clone could not be traced by PFGE analysis but can be investigated by WGS analysis.

Our previous studies found that a dominant serovar of Salmonella enterica isolates from three farms raising broilers in 2014 and 2015 was serovar Agona and the number of Infantis isolates decreased (the serovar shift). In this study, 52 S. Agona strains which isolated between 1993 and 2008, were compared to the serovar shift clone by molecular epidemiology and phylogenetic analyses, using pulsed field gel electrophoresis and whole genome sequence analyses. Of the 52 strains, one strain isolated from a human case in 1995 was genetically identical to the serovar shift clone, even though it was isolated prior to the serovar shift. These results suggested that the S . Agona serovar shift clone had existed in a source other than chicken penetrated chicken population.

Hospital-acquired infections caused by extended-spectrum β-lactamase (ESBL)-producing Escherichia coli are a global problem. Healthy people can carry ESBL-producing E. coli in their intestines; thus, E. coli from healthy people can potentially cause hospital-acquired infections. Therefore, the transmission routes of ESBL-producing E. coli from healthy persons should be determined.A foodborne outbreak of human norovirus GII (HuNoV) occurred at a restaurant in Shizuoka, Japan, in 2018. E. coli O25:H4 was isolated from some of the HuNoV-infected customers. Pulsed-field gel electrophoresis showed that these E. coli O25:H4 strains originated from one clone. Because the only epidemiological link among the customers was eating food from this restaurant, the customers were concurrently infected with E. coli O25:H4 and HuNoV GII via the restaurant food. Whole-genome analysis revealed that the E. coli O25:H4 strains possessed genes for regulating intracellular iron and expressing the flagellum and flagella. Extraintestinal pathogenic E. coli often express these genes on the chromosome. Additionally, the E. coli O25:H4 strains had plasmids harboring nine antimicrobial-resistance genes. These strains harbored ESBL-encoding blaCTX-M-14 genes on two loci of the chromosome and had higher ESBL activity. Multilocus sequence typing and fimH subtyping revealed that the E. coli O25:H4 strains from the outbreak belonged to the subclonal group, ST131-fimH30R, which has been driving ESBL epidemics in Japan.Because the E. coli O25:H4 strains isolated in the outbreak belonged to a subclonal group spreading in Japan, foods contaminated with ESBL-producing E. coli might contribute to spreading these strains among healthy persons. The isolated E. coli O25:H4 strains produced ESBL and contained plasmids with multiple antimicrobial-resistance genes, which may make it difficult to select antimicrobials for treating extraintestinal infections caused by these strains.