Differential Analysis of the Nasal Microbiome of Pig Carriers or Non-Carriers of Staphylococcus aureus

Staphylococcus aureus is presently regarded as an emerging zoonotic agent due to the spread of specific methicillin-resistant S. aureus (MRSA) clones in pig farms. Studying the microbiota can be useful for the identification of bacteria that antagonize such opportunistic veterinary and zoonotic pathogen in animal carriers. The aim of this study was to determine whether the nasal microbiome of pig S. aureus carriers differs from that of non-carriers. The V3-V5 region of the 16S rRNA gene was sequenced from nasal swabs of 44 S. aureus carriers and 56 non-carriers using the 454 GS FLX titanium system. Carriers and non-carriers were selected on the basis of quantitative longitudinal data on S. aureus carriage in 600 pigs sampled at 20 Danish herds included in two previous studies in Denmark. Raw sequences were analysed with the BION meta package and the resulting abundance matrix was analysed using the DESeq2 package in R to identify operational taxonomic units (OTUs) with differential abundance between S. aureus carriers and non-carriers. Twenty OTUs were significantly associated to non-carriers, including species with known probiotic potential and antimicrobial effect such as lactic acid-producing isolates described among Leuconostoc spp. and some members of the Lachnospiraceae family, which is known for butyrate production. Further 5 OTUs were significantly associated to carriage, including known pathogenic bacteria such as Pasteurella multocida and Klebsiella spp. Our results show that the nasal microbiome of pigs that are not colonized with S. aureus harbours several species/taxa that are significantly less abundant in pig carriers, suggesting that the nasal microbiota may play a role in the individual predisposition to S. aureus nasal carriage in pigs. Further research is warranted to isolate these bacteria and assess their possible antagonistic effect on S. aureus for the pursuit of new strategies to control MRSA in pig farming

Genome-wide association study reveals a locus for nasal carriage of <i>Staphylococcus aureus</i> in Danish crossbred pigs

BACKGROUND: Staphylococcus aureus is an important human opportunistic pathogen residing on skin and mucosae of healthy people. Pigs have been identified as a source of human colonization and infection with methicillin-resistant Staphylococcus aureus (MRSA) and novel measures are needed to control zoonotic transmission. A recent longitudinal study indicated that a minority of pigs characterized by high nasal load and stable carriage may be responsible for the maintenance of S. aureus within farms. The primary objective of the present study was to detect genetic loci associated with nasal carriage of S. aureus in Danish crossbred pigs (Danish Landrace/Yorkshire/Duroc). RESULTS: Fifty-six persistent carriers and 65 non-carriers selected from 15 farms surveyed in the previous longitudinal study were genotyped using Illumina’s Porcine SNP60 beadchip. In addition, spa typing was performed on 126 S. aureus isolates from 37 pigs to investigate possible relationships between host and S. aureus genotypes. A single SNP (MARC0099960) on chromosome 12 was found to be associated with nasal carriage of S. aureus at a genome-wide level after permutation testing (p = 0.0497) whereas the association of a neighboring SNP was found to be borderline (p = 0.114). Typing of S. aureus isolates led to detection of 11 spa types belonging to the three main S. aureus clonal complexes (CC) previously described in pigs (CC9, CC30 and CC398). Individual carriers often harbored multiple S. aureus genotypes and the host-pathogen interaction seems to be independent of S. aureus genotype. CONCLUSION: Our results suggest it may be possible to select pigs genetically resistant to S. aureus nasal colonization as a tool to control transmission of livestock-associated MRSA to humans. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12917-015-0599-y) contains supplementary material, which is available to authorized users

MRSA carrying <i>mecC</i> in captive mara

This is the final published version. It first appeared at http://jac.oxfordjournals.org/content/early/2015/02/24/jac.dkv024.full.Objectives: To characterize the staphylococcal cassette chromosome mec (SCCmec), virulence and antimicrobial\ud susceptibility of Staphylococcus aureus ST130 isolated from mara (Dolichotis patagonum), a large rodent species\ud native to South America and kept in captivity at Copenhagen Zoo.\ud Methods: The presence of mecCwas confirmed by PCR in 15 S. aureus ST130 isolated from mara during a previous\ud study. WGS was performed on two randomly selected isolates to characterize their genomes with respect to\ud SCCmec, virulence and resistance gene content. Antimicrobial susceptibility was tested using commercial\ud broth microdilution tests.\ud Results: All the isolates belonged to spa type t528 ST130 and carried mecC. Based on WGS, mecC was 100%\ud identical to the prototype sequence of S. aureus strain LGA251. The sequence of SCCmec type XI in the mara\ud isolates had 23 SNPs compared with the one described in LGA251. The two sequenced strains harboured a set\ud of virulence factors and other genomic features previously observed in ST130. Both strains carried norA as the\ud only putative antimicrobial resistance gene in addition to mecC.\ud Conclusions: Our findings support the notion that a genetically conserved mecC-carryingMRSA ST130 clone iswidespread\ud in a variety of unrelated hosts in Denmark. Since the mara at Copenhagen Zoo have limited contact with\ud humans and other animal species, it remains unclear whether mara are natural hosts of ST130 or acquired this lineage\ud from unknown sources. The broad host range of MRSA ST130 supports its designation as a generalist lineage.This study was supported by internal funding (C. E.-G., A. M. and L. G.) and by the Medical Research Council (E. M. H. and M. A. H.; G1001787/1)

Horses in Denmark Are a Reservoir of Diverse Clones of Methicillin-Resistant and -Susceptible Staphylococcus aureus

Denmark is a country with high prevalence of livestock-associated methicillin-resistant Staphylococcus aureus (MRSA) clonal complex (CC) 398 in pigs. Even though pig farming is regarded as the main source of human infection or colonization with MRSA CC398, 10–15% of the human cases appear not to be linked to pigs. Following the recent reports of MRSA CC398 in horses in other European countries and the lack of knowledge on S. aureus carriage in this animal species, we carried out a study to investigate whether horses constitute a reservoir of MRSA CC398 in Denmark, and to gain knowledge on the frequency and genetic diversity of S. aureus in horses, including both methicillin-resistant and -susceptible S. aureus (MSSA). Nasal swabs were collected from 401 horses originating from 74 farms, either at their farms or prior to admission to veterinary clinics. Following culture on selective media, species identification by MALDI-TOF MS and MRSA confirmation by standard PCR-based methods, S. aureus and MRSA were detected in 54 (13%) and 17 (4%) horses originating from 30 (40%) and 7 (9%) farms, respectively. Based on spa typing, MSSA differed genetically from MRSA isolates. The spa type prevalent among MSSA isolates was t127 (CC1), which was detected in 12 horses from 11 farms and represents the most common S. aureus clone isolated from human bacteremia cases in Denmark. Among the 17 MRSA carriers, 10 horses from three farms carried CC398 t011 harboring the immune evasion cluster (IEC), four horses from two farms carried IEC-negative CC398 t034, and three horses from two farms carried a mecC-positive MRSA lineage previously associated with wildlife and domestic ruminants (CC130 t528). Based on whole-genome phylogenetic analysis of the 14 MRSA CC398, t011 isolates belonged to the recently identified horse-adapted clone in Europe and were closely related to human t011 isolates from three Danish equine veterinarians, whereas t034 isolates belonged to pig-adapted clones. Our study confirms that horses carry an equine-specific clone of MRSA CC398 that can be transmitted to veterinary personnel, and reveals that these animals are exposed to MRSA and MSSA clones that are likely to originate from livestock and humans, respectively