Abstract
A nationwide study on the occurrence of ESBL/AmpC in non-hospitalized horses in the Netherlands was performed. Molecular characterization was done and questionnaires were analyzed to identify factors associated with carriage. In total, 796 horse owners were approached, of which 281 submitted a fecal sample of their horse(s), resulting in 362
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samples. All samples were cultured qualitatively in LB broth and subsequently on MacConkey agar, both supplemented with 1 mg/L cefotaxime (LB+ and MC+). Positive samples were subsequently cultured quantitatively on MC+. Initial ESBL/AmpC screening was performed by PCR, followed by whole genome sequencing on selected strains. Association between ESBL/AmpC carriage and questionnaire items were analyzed using a univariate generalized estimating equation (GEE) regression analysis, followed by a multiple GEE model for relevant factors. In total, 39 out of 362 samples (11%) were positive for ESBL/AmpC. blaCTX-M-1-carrying isolates were obtained from 77% of positive samples (n=30). Other ESBL/AmpC genes observed were blaCTX-M-2, blaCTX-M-14, blaCTX-M-15, blaCTX-M-32, blaSHV-12, blaCMY-2, blaACT-10. A high association between the presence of blaCTX-M-1 andIncHI1 plasmids was observed (46% of samples; n=18). Based on core genome analysis (n=48 isolates), six E. coli clusters were identified, of which three represented 80% of the isolates. A negative association between ESBL/AmpC carriage and horses being in contact with other horses at a different site was observed. The presence of a dog on the premises and housing in a more densely human-populated region were positively associated. IMPORTANCE Extended spectrum ß-lactamases (ESBL) are widespread in the human and animal population and the environment. Many different ESBL-variants exist. The dissemination of ESBLs within and between populations and the environment is also largely influenced by genetic mobile elements (e.g. plasmids) that facilitate spread of these ESBLs. In order to identify potential attributable ESBL-sources for e.g. the human population, it is important to identify the different ESBL-variants, the bacteria carrying them and the potential risk factors for ESBL carriage from other potential sources. This nationwide study focuses on ESBL-carriage in the open horse population and investigated the molecular characteristics, geographical distribution throughout the Netherlands and potential risk factors for fecal ESBL-carriage in horses. This data can be used for future attribution studies in order to reduce potential transmission of ESBL-producing bacteria between sources.
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