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Intestinal carriage of extended-spectrum beta-lactamase (ESBL) -producing bacteria in food-producing animals and contamination of retail meat may contribute to increased incidences of infections with ESBL-producing bacteria in humans. Therefore, distribution of ESBL genes, plasmids and strain genotypes in Escherichia coli obtained from poultry and retail chicken meat in the Netherlands was determined and defined as ‘poultry-associated’ (PA). Subsequently, the proportion of E. coli isolates with PA ESBL genes, plasmids and strains was quantified in a representative sample of clinical isolates. The E. coli were derived from 98 retail chicken meat samples, a prevalence survey among poultry, and 516 human clinical samples from 31 laboratories collected during a 3-month period in 2009. Isolates were analysed using an ESBL-specific microarray, sequencing of ESBL genes, PCR-based replicon typing of plasmids, plasmid multi-locus sequence typing (pMLST) and strain genotyping (MLST). Six ESBL genes were defined as PA (blaCTX-M-1, blaCTX-M-2, blaSHV-2, blaSHV-12, blaTEM-20, blaTEM-52): 35% of the human isolates contained PA ESBL genes and 19% contained PA ESBL genes located on IncI1 plasmids that were genetically indistinguishable from those obtained from poultry (meat). Of these ESBL genes, 86% were blaCTX-M-1 and blaTEM-52 genes, which were also the predominant genes in poultry (78%) and retail chicken meat (75%). Of the retail meat samples, 94% contained ESBL-producing isolates of which 39% belonged to E. coli genotypes also present in human samples. These findings are suggestive for transmission of ESBL genes, plasmids and E. coli isolates from poultry to humans, most likely through the food chain. [ABSTRACT FROM AUTHOR]

Objectives: MRSA carrying the mecC gene (mecC-MRSA) have been found in humans and animals worldwide. A high carriage rate of mecC-MRSA has been described among hedgehogs in different countries. We performed genomic comparison of mecC-MRSA from hedgehogs and humans using next-generation sequencing (NGS) to investigate possible zoonotic transmission in the Netherlands., Methods: Nasal swabs from hedgehogs (n = 105) were cultured using pre-enrichment and selective plates. Isolates were sequenced using Illumina NGS platforms. These data were compared with sequence data of mecC-MRSA (n = 62) from the Dutch national MRSA surveillance in humans., Results: Fifty hedgehogs were found to be MRSA positive, of which 48 carried mecC. A total of 60 mecC-MRSA isolates derived from 50 hedgehogs were compared with the human isolates. Fifty-nine mecC-MRSA from hedgehogs and all but one isolate from humans belonged to clonal complexes CC130 and CC1943. The mecC gene was located within the SCCmec XI element. Most mecC-MRSA did not carry other resistance genes besides mecC and blaZ. Two human isolates carried erm(C). Isolates differed in the presence of various virulence genes, which were linked to distinct STs and clonal complexes. Some isolates had up to 17 virulence genes, which underlines their pathogenic potential. No genetic clusters of hedgehog and human isolates were found., Conclusions: mecC-MRSA from hedgehogs and humans mainly belonged to the same two clonal complexes, indicating a common source. No firm evidence for recent zoonotic transmission was found. Further studies are needed to investigate the role of hedgehogs in the occurrence of mecC-MRSA in humans., (© The Author(s) 2023. Published by Oxford University Press on behalf of British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Salmonella Infantis is a poultry-adapted Salmonella enterica serovar that is increasingly reported in broilers and is also regularly identified among human salmonellosis cases. An emerging S. Infantis mega-plasmid (pESI), carrying fitness, virulence and antimicrobial resistance genes, is also increasingly found. We investigated the prevalence, genetic characteristics and risk factors for (pESI-carrying) S. Infantis in broilers. Faecal samples from 379 broiler flocks (in 198 farms with ≥3000 birds) in the Netherlands were tested. A questionnaire about farm characteristics was also administered. Sampling was performed in July 2018-May 2019, three weeks before slaughter. Fourteen flocks (in 10 farms) were S. Infantis-positive, resulting in a 3.7 % flock-level and 5.1 % farm-level prevalence. Based on multi-locus sequence typing (MLST), all isolates belonged to sequence type 32. All but one isolate carried a pESI-like mega-plasmid. Core-genome MLST showed considerable heterogeneity among the isolates, even within the same farm, with a few small clusters detected. The typical pESI-borne multi-resistance pattern to aminoglycosides, sulphonamide and tetracycline (93 %), as well as trimethoprim (71 %), was found. Additionally, resistance to (fluoro)quinolones based on gyrA gene mutations was detected. S. Infantis was found more often in flocks using salinomycin as coccidiostat, where flock thinning was applied or litter quality was poor, whereas employing external cleaning companies, wheat in feed, and vaccination against infectious bronchitis, were protective. Suggestive evidence for vertical transmission from hatcheries was found. A heterogeneous (pESI-carrying) S. Infantis population has established itself in Dutch broiler flocks, calling for further monitoring of its spread and a comprehensive appraisal of control options., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

Objectives: To investigate the occurrence and characteristics of ESBL/AmpC-producing Escherichia coli in faecal samples from horses at one equine clinic in the Netherlands., Methods: A total of 91 horses, including residents and patients, were sampled. ESBL/AmpC-producing E. coli were identified by a combination disc diffusion test. Phylogenetic groups and MLST were determined. ESBL/AmpC genes were analysed using PCR and sequencing. Plasmids were characterized by transformation and PCR-based replicon typing. Subtyping of plasmids was done by plasmid MLST., Results: At least one E. coli isolate with a confirmed ESBL/AmpC gene was found in samples from 76 horses (84%). Although phylogenetic group B1 E. coli bla CTX-M-1 predominated, a diverse E. coli population was found, indicating that clonal nosocomial spread was not the only reason for the high occurrence found. MLST analysis revealed the presence of 47 E. coli STs, organized in four clusters of genetically related strains. ST10, ST641, ST1079 and ST1250 were most commonly found. With regard to the genes, bla CTX-M-1 was most prevalent ( n  =   91), followed by bla CTX-M-2 ( n  =   26). The most frequently found plasmid type was IncHI1, but plasmids belonging to the IncF, IncI1 and IncN groups were also identified., Conclusions: A high occurrence of ESBL-producing E. coli in faecal samples was found among horses in an equine clinic and the variety of STs, ESBL genes and plasmid types suggests nosocomial transmission. ESBL E. coli can cause difficult-to-treat infections in horses and prudent use of antimicrobials is warranted. A further assessment of the risks of transmission to persons in close contact with horses, such as caretakers or veterinarians, is crucial., (© The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Objectives: This study aimed to compare ESBL-producing Escherichia coli causing infections in humans with infecting or commensal isolates from animals and isolates from food of animal origin in terms of the strain types, the ESBL gene present and the plasmids that carry the respective ESBL genes., Methods: A collection of 353 ESBL-positive E. coli isolates from the UK, the Netherlands and Germany were studied by MLST and ESBL genes were identified. Characterization of ESBL gene-carrying plasmids was performed using PCR-based replicon typing. Moreover, IncI1-Iγ and IncN plasmids were characterized by plasmid MLST., Results: The ESBL-producing E. coli represented 158 different STs with ST131, ST10 and ST88 being the most common. Overall, blaCTX-M-1 was the most frequently detected ESBL gene, followed by blaCTX-M-15, which was the most common ESBL gene in the human isolates. The most common plasmid replicon type overall was IncI1-Iγ followed by multiple IncF replicons., Conclusions: ESBL genes were present in a wide variety of E. coli STs. IncI1-Iγ plasmids that carried the blaCTX-M-1 gene were widely disseminated amongst STs in isolates from animals and humans, whereas other plasmids and STs appeared to be more restricted to isolates from specific hosts., (© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

The putative virulence and antimicrobial resistance gene contents of extended spectrum β-lactamase (ESBL)-positive E. coli (n=629) isolated between 2005 and 2009 from humans, animals and animal food products in Germany, The Netherlands and the UK were compared using a microarray approach to test the suitability of this approach with regard to determining their similarities. A selection of isolates (n=313) were also analysed by multilocus sequence typing (MLST). Isolates harbouring bla(CTX-M-group-1) dominated (66%, n=418) and originated from both animals and cases of human infections in all three countries; 23% (n=144) of all isolates contained both bla(CTX-M-group-1) and bla(OXA-1-like) genes, predominantly from humans (n=127) and UK cattle (n=15). The antimicrobial resistance and virulence gene profiles of this collection of isolates were highly diverse. A substantial number of human isolates (32%, n=87) did not share more than 40% similarity (based on the Jaccard coefficient) with animal isolates. A further 43% of human isolates from the three countries (n=117) were at least 40% similar to each other and to five isolates from UK cattle and one each from Dutch chicken meat and a German dog; the members of this group usually harboured genes such as mph(A), mrx, aac(6')-Ib, catB3, bla(OXA-1-like) and bla(CTX-M-group-1). forty-four per cent of the MLST-typed isolates in this group belonged to ST131 (n=18) and 22% to ST405 (n=9), all from humans. Among animal isolates subjected to MLST (n=258), only 1.2% (n=3) were more than 70% similar to human isolates in gene profiles and shared the same MLST clonal complex with the corresponding human isolates. The results suggest that minimising human-to-human transmission is essential to control the spread of ESBL-positive E. coli in humans.

Objectives: The presence of ESBL/AmpC-producing E. coli in cattle has been reported previously, however information on veal calves is limited. This study describes the prevalence and molecular characteristics of E. coli with non-wild type susceptibility to cefotaxime in veal calves at slaughter., Methods: Faecal samples from 100 herds, 10 individual animals per herd, were screened for E. coli with non-wild type susceptibility for cefotaxime. Molecular characterization of ESBL/AmpC genes and plasmids was performed on one isolate per herd by microarray, PCR and sequence analysis., Results: 66% of the herds were positive for E. coli with non-wild type susceptibility for cefotaxime. Within-herd prevalence varied from zero to 90%. 83% of E. coli producing ESBL/AmpC carried bla(CTX-M) genes, of which bla(CTX-M-1), bla(CTX-M-14) and bla(CTX-M-15) were most prevalent. The dominant plasmids were IncI1 and IncF-type plasmids., Conclusions: A relatively high prevalence of various bla(CTX-M) producing E. coli was found in veal calves at slaughter. The genes were mainly located on IncI1 and IncF plasmids.

Quinolone resistance is studied and reported increasingly in isolates from humans, food-producing animals and companion animals. Resistance can be caused by chromosomal mutations in topoisomerase genes, plasmid-mediated resistance genes, and active transport through efflux pumps. Cross sectional data on quinolone resistance mechanisms in non-pathogenic bacteria from healthy veal calves is limited. The purpose of this study was to determine the prevalence and characteristics of quinolone resistance mechanisms in Escherichia coli isolates from veal calves, after more than 20 years of quinolone usage in veal calves. MIC values were determined for all isolates collected as part of a national surveillance program on antimicrobial resistance in commensal bacteria in food-producing animals in The Netherlands. From the strains collected from veal calves in 2007 (n=175) all isolates with ciprofloxacin MIC ≥ 0.125 mg/L (n=25) were selected for this study, and screened for the presence of known quinolone resistance determinants. In this selection only chromosomal mutations in the topoisomerase type II and IV genes were detected. The number of mutations found per isolate correlated with an increasing ciprofloxacin MIC. No plasmid-mediated quinolone resistance genes were found. The contribution of efflux pumps varied from no contribution to a 16-fold increase in susceptibility. No correlation was found with the presence of resistance genes of other antimicrobial classes, even though all quinolone non-wild type isolates were resistant to 3 or more classes of antibiotics other than quinolones. Over twenty years of quinolone usage in veal calves in The Netherlands did not result in a widespread occurrence of plasmid-mediated quinolone resistance, limiting the transmission of quinolone resistance to clonal distribution., (Copyright © 2011 Elsevier B.V. All rights reserved.)

To gain more information on the genetic basis of the rapid increase in the number of isolates exhibiting non-wild type Minimum Inhibitory Concentrations (MICs) for cefotaxime observed since 2003, beta-lactamase genes of 22 Salmonella enterica and 22 Escherichia coli isolates from broilers in 2006 showing this phenotype were characterized by miniaturized micro-array, PCR and DNA-sequencing. Presence and size of plasmids were determined by S1-digest pulsed-field gel electrophoresis and further characterized by PCR-based replicon typing. Transfer of resistance plasmids was tested by conjugation and transformation experiments. To link resistance genes and plasmid type, Southern blot hybridization experiments were conducted. In 42 isolates, five (bla(CTX-M-1), bla(CTX-M-2), bla(TEM-20), bla(TEM-52), bla(SHV-2)) different extended spectrum beta-lactamase (ESBL)-genes and two (bla(ACC-1), bla(CMY-2)) AmpC-genes were present. Three of the detected ESBL-genes (bla(CTX-M-1), bla(TEM-52) and bla(CTX-M-2)) were located on similar types of plasmids (IncI1 and IncHI2/P) in both E. coli and Salmonella. Two other detected ESBL- and AmpC-genes bla(SHV-2) and bla(CMY-2) respectively (on IncK plasmids), were only found in E. coli, whereas the AmpC-gene bla(ACC-1) (on non-typable plasmids), and the ESBL-gene bla(TEM-20) (on IncI1 plasmids), were only detected in Salmonella. In two isolates, no ESBL- or AmpC-gene could be detected through these methods. The increase in the number of E. coli and S. enterica isolates from the gastro-intestinal tract of broilers exhibiting non-wild type MICs for cefotaxime is mainly due to an increase in IncI1 plasmids containing bla(CTX-M-1). The reason for the successful spread of this plasmid type in these species is not yet understood., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Objectives: To characterize plasmids and resistance genes of multidrug-resistant (MDR) Salmonella Senftenberg and Salmonella Concord isolated from patients in the Netherlands., Methods: The resistance genes of four MDR Salmonella isolates (three Salmonella Concord and one Salmonella Senftenberg) were identified by miniaturized microarray, PCR and sequencing. Plasmids were characterized by S1 nuclease-PFGE and PCR-based replicon typing (PBRT). Linkage between plasmids and genes was determined by conjugation experiments and microarray analysis. The genetic relationship between the three Salmonella Concord isolates was determined by XbaI-PFGE., Results: A large variety of resistance genes was detected, including qnrB2 and the beta-lactamase genes bla(TEM-1) and bla(SHV-12) in all isolates; moreover all Salmonella Concord isolates also harboured bla(CTX-M-15). Salmonella Senftenberg harboured a large IncHI2 plasmid. The three Salmonella Concord isolates harboured two large plasmids typed as IncHI2 and IncA/C., Conclusions: We detected the first plasmid-mediated MDR Salmonella isolates in the Netherlands harbouring both qnr and extended-spectrum beta-lactamase (ESBL) genes. In Salmonella Senftenberg one large plasmid (IncHI2) and in Salmonella Concord two large plasmids (IncHI2 and IncA/C) were responsible for the multidrug resistance.

This case study describes the isolation ofa multiresistant strain ofBrachyspira hyodysenteriae in April 2007 in a Dutch sow herd with recurrent diarrhoea. Examination of faecal samples taken from 7-month-old breeding gilts with diarrhoea revealed the presence of resistance against tiamulin, lincomycin, tylosin, doxycycline, and tylvalosin (the active substance in Aivlosin) in four of five samples. Tiamulin resistance has not been reported in The Netherlands before. The repeated use of tiamulin on the affected farm was assumed to be the main cause of the development of resistance to the drug. The farmer was advised to adopt a medication strategy and to implement management practices that would prevent an ongoing cycle of infection on the farm. It is important that the Dutch swine industry appreciates that tiamulin-resistant strains of B. hyodysenteriae may be found on other farms as well. The appropriate and prudent use of antibiotics is essential in order to prevent the development of resistance against the last option left to cure B. hyodysenteriae infections: valnemulin.