Abstract
The aim of this article was to define the sampling level and method combination that captures antibiotic resistance at pig herd level utilizing qPCR antibiotic resistance gene quantification and culture-based quantification of antibiotic resistant coliform indicator bacteria. Fourteen qPCR assays for commonly detected antibiotic resistance genes were developed, and used to quantify antibiotic resistance genes in total DNA from swine fecal samples that were obtained using different sampling and pooling methods. In parallel, the number of antibiotic resistant coliform indicator bacteria was determined in the same swine fecal samples. The results showed that the qPCR assays were capable of detecting differences in antibiotic resistance levels in individual animals that the coliform bacteria colony forming units (CFU) could not. Also, the qPCR assays more accurately quantified antibiotic resistance genes when comparing individual sampling and pooling methods. qPCR on pooled samples was found to be a good representative for the general resistance level in a pig herd compared to the coliform CFU counts. It had significantly reduced relative standard deviations compared to coliform CFU counts in the same samples, and therefore differences in antibiotic resistance levels between samples were more readily detected. To our knowledge, this is the first study to describe sampling and pooling methods for qPCR quantification of antibiotic resistance genes in total DNA extracted from swine feces.
Highlights
Antibiotic resistance in pathogenic bacteria is an increasing problem challenging disease treatment in humans and animals globally [1,2,3]
The sampling methods were included for the following reasons: Shoe cover sampling is used in the Danish Salmonella control program in broiler flocks [12] and it is of interest to investigate whether shoe cover sampling can be applied to pig herds; pen floor sampling is currently used in the Danish Salmonella control program in pig herds [13,14]; slurry tank sampling is of interest as the antibiotic resistance levels within the slurry tank may reflect the herd’s levels rendering it a convenient method for herd sampling potentially representing resistance levels for the past 6 months
Quantification of antibiotic resistance levels facilitates antibiotic resistance surveillance, helping to contain and prevent infections caused by antibiotic resistant bacteria
Summary
Antibiotic resistance in pathogenic bacteria is an increasing problem challenging disease treatment in humans and animals globally [1,2,3]. It is important to minimize antibiotic use in intensive agricultural practices where widespread antibiotic use is common e.g. in the pig production, as PLOS ONE | DOI:10.1371/journal.pone.0131672 June 26, 2015
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