Methicillin Resistant Staphylococcus aureus ST398 in Veal Calf Farming: Human MRSA Carriage Related with Animal Antimicrobial Usage and Farm Hygiene

Introduction Recently a specific MRSA sequence type, ST398, emerged in food production animals and farmers. Risk factors for carrying MRSA ST398 in both animals and humans have not been fully evaluated. In this cross-sectional study, we investigated factors associated with MRSA colonization in veal calves and humans working and living on these farms. Methods A sample of 102 veal calf farms were randomly selected and visited from March 2007–February 2008. Participating farmers were asked to fill in a questionnaire (n = 390) to identify potential risk factors. A nasal swab was taken from each participant. Furthermore, nasal swabs were taken from calves (n = 2151). Swabs were analysed for MRSA by selective enrichment and suspected colonies were confirmed as MRSA by using slide coagulase test and PCR for presence of the mecA-gene. Spa types were identified and a random selection of each spa type was tested with ST398 specific PCR. The Sequence Type of non ST398 strains was determined. Data were analyzed using logistic regression analysis. Results Human MRSA carriage was strongly associated with intensity of animal contact and with the number of MRSA positive animals on the farm. Calves were more often carrier when treated with antibiotics, while farm hygiene was associated with a lower prevalence of MRSA. Conclusion This is the first study showing direct associations between animal and human carriage of ST398. The direct associations between animal and human MRSA carriage and the association between MRSA and antimicrobial use in calves implicate prudent use of antibiotics in farm animals

Rhesus Macaques (Macaca mulatta) Are Natural Hosts of Specific Staphylococcus aureus Lineages

Currently, there is no animal model known that mimics natural nasal colonization by Staphylococcus aureus in humans. We investigated whether rhesus macaques are natural nasal carriers of S. aureus. Nasal swabs were taken from 731 macaques. S. aureus isolates were typed by pulsed-field gel electrophoresis (PFGE), spa repeat sequencing and multi-locus sequence typing (MLST), and compared with human strains. Furthermore, the isolates were characterized by several PCRs. Thirty-nine percent of 731 macaques were positive for S. aureus. In general, the macaque S. aureus isolates differed from human strains as they formed separate PFGE clusters, 50% of the isolates were untypeable by agr genotyping, 17 new spa types were identified, which all belonged to new sequence types (STs). Furthermore, 66% of macaque isolates were negative for all superantigen genes. To determine S. aureus nasal colonization, three nasal swabs from 48 duo-housed macaques were taken during a 5 month period. In addition, sera were analyzed for immunoglobulin G and A levels directed against 40 staphylococcal proteins using a bead-based flow cytometry technique. Nineteen percent of the animals were negative for S. aureus, and 17% were three times positive. S. aureus strains were easily exchanged between macaques. The antibody response was less pronounced in macaques compared to humans, and nasal carrier status was not associated with differences in serum anti-staphylococcal antibody levels. In conclusion, rhesus macaques are natural hosts of S. aureus, carrying host-specific lineages. Our data indicate that rhesus macaques are useful as an autologous model for studying S. aureus nasal colonization and infection prevention

Rare Occurrence of Methicillin-Resistant Staphylococcus aureus CC130 with a Novel mecA Homologue in Humans in Germany

MRSA CC130 containing the mecA homologue mecALGA251 were reported from the UK and from Denmark so far from cattle and humans. Here we report on 11 MRSA CC130 among a sample of 12691 isolates of human origin collected from January 2006 until June 2011. MRSA CC130 grew insufficiently on chromogernic agar plates for detection of MRSA; the agglutination test for presence of PBP2a was negative. We designed primers for specific detection of mecALGA251 as well as for concomitant detection of both, mecLGA251 and mecA. As already described, the isolates exhibited spa-types t843, t1736, and t1773. The ccrA homologue indicated the presence SCCmecXI. When subjected to further characterization by means of a commercially available microarray the isolates were negative for sak chp, and scn, and as expected positive for hla, untruncated hlb, and hld. They furthermore contained edinB, aur, slpA, slpB, slpE. From genes coding for surface and cell wall associated products the ica-operon, cap8, clfA, clfF, ebpS, fnbA, fnbB, sdrC were detected but not cna. The isolates were negative for enterotoxin genes and tst, as well as for eta, and etb; agr-type was III

Capturing the cloud of diversity reveals complexity and heterogeneity of MRSA carriage, infection and transmission.

Genome sequencing is revolutionizing clinical microbiology and our understanding of infectious diseases. Previous studies have largely relied on the sequencing of a single isolate from each individual. However, it is not clear what degree of bacterial diversity exists within, and is transmitted between individuals. Understanding this 'cloud of diversity' is key to accurate identification of transmission pathways. Here, we report the deep sequencing of methicillin-resistant Staphylococcus aureus among staff and animal patients involved in a transmission network at a veterinary hospital. We demonstrate considerable within-host diversity and that within-host diversity may rise and fall over time. Isolates from invasive disease contained multiple mutations in the same genes, including inactivation of a global regulator of virulence and changes in phage copy number. This study highlights the need for sequencing of multiple isolates from individuals to gain an accurate picture of transmission networks and to further understand the basis of pathogenesis.Thanks to Dr Alex O’Neill, University of Leeds and Dr Matthew Ellington, Public Health England for provision of RN4220 and RN4200mutS. We thank the core sequencing and informatics team at the Wellcome Trust Sanger Institute for sequencing of the isolates described in this study. This work was supported by a Medical Research Council Partnership grant (G1001787/1) held between the Department of Veterinary Medicine, University of Cambridge (M.A.H.), the School of Clinical Medicine, University of Cambridge (S.J.P.), the Moredun Research Institute, and the Wellcome Trust Sanger Institute (J.P. and S.J.P). S.J.P. receives support from the NIHR Cambridge Biomedical Research Centre. M.T.G.H., S.R.H. and J.P. were funded by Wellcome Trust grant no. 098051. G.G.R.M. was funded by an MRC studentship.This is the final version of the article. It first appeared from Nature Publishing Group via http://dx.doi.org/10.1038/ncomms756

Intestinal carriage of Staphylococcus aureus: How does its frequency compare with that of nasal carriage and what is its clinical impact?

The bacterial species Staphylococcus aureus, including its methicillin-resistant variant (MRSA), finds its primary ecological niche in the human nose, but is also able to colonize the intestines and the perineal region. Intestinal carriage has not been widely investigated despite its potential clinical impact. This review summarizes literature on the topic and sketches the current state of affairs from a microbiological and infectious diseases' perspective. Major findings are that the average reported detection rate of intestinal carriage in healthy individuals and patients is 20% for S. aureus and 9% for MRSA, which is approximately half of that for nasal carriage. Nasal carriage seems to predispose to intestinal carriage, but sole intestinal carriage occurs relatively frequently and is observed in 1 out of 3 intestinal carriers, which provides a rationale to include intestinal screening for surveillance or in outbreak settings. Colonization of the intestinal tract with S. aureus at a young age occurs at a high frequency and may affect the host's immune system. The frequency of intestinal carriage is generally underestimated and may significantly contribute to bacterial dissemination and subsequent risk of infections. Whether intestinal rather than nasal S. aureus carriage is a primary predictor for infections is still ill-defined

Livestock-associated methicillin-resistant Staphylococcus aureus responsible for human colonization and infection in an area of Italy with high density of pig farming

BACKGROUND: Livestock-Associated MRSA (LA-MRSA) belonging to ST398 lineage, common among pigs and other animals, emerged in Central and Northern Europe, becoming a new risk factor for MRSA among farm workers. Strains belonging to ST398 can be responsible for human colonization and infection, mainly in areas with high livestock-farming. The aim of this study was to investigate the occurrence of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) human colonization and infections in an area of the Lombardy Region (Italy), the Italian region with the highest density of pig farming. METHODS: In the period March-April 2010, 879 nasal swabs were taken from subjects at admission to a local hospital serving an area of the Lombardy Region devoted to agriculture and farming. In the period March 2010-February 2011, all MRSA strains from community-acquired infection (CAI) observed in the same hospital, were collected. Molecular characterization of the isolates included SCCmec typing, spa typing and multilocus sequence typing (MLST). RESULTS: Out of 879 nasal swabs examined, 9 (1%) yielded MRSA. Five strains were assigned to sequence type (ST)398 (spa t899, 3 isolates; t108 and t2922, 1 isolate each) and were therefore categorized as LA-MRSA. The other 4 isolates were likely of hospital origin. No strains were positive for Panton-Valentine Leukocidin genes. Twenty MRSA isolates were detected from CAI, 17 were from skin and soft-tissue infections and 3 from other infections. An MRSA isolate from otitis externa was t899/ST398 and PVL-negative, hence categorized as LA-MRSA. Four isolates were assigned to t127/ST1. Eight strains were PVL-positive community acquired (CA)-MRSA and belonged to different clones, the most frequent being ST8. CONCLUSIONS: In an area of Italy with high density of pig farming, LA-MRSA is able to colonize the population and rarely to produce infections. Typical CA-MRSA is more common than LA-MRSA among CAI

A single natural nucleotide mutation alters bacterial pathogen host tropism

The capacity of microbial pathogens to alter their host tropism leading to epidemics in distinct host species populations is a global public and veterinary health concern. To investigate the molecular basis of a bacterial host-switching event in a tractable host species, we traced the evolutionary trajectory of the common rabbit clone of Staphylococcus aureus. We report that it evolved through a likely human-to-rabbit host jump over 40 years ago and that only a single naturally occurring nucleotide mutation was required and sufficient to convert a human-specific S. aureus strain into one that could infect rabbits. Related mutations were identified at the same locus in other rabbit strains of distinct clonal origin, consistent with convergent evolution. This first report of a single mutation that was sufficient to alter the host tropism of a microorganism during its evolution highlights the capacity of some pathogens to readily expand into new host species populations

Phylogenetic Analysis of Staphylococcus aureus CC398 Reveals a Sub-Lineage Epidemiologically Associated with Infections in Horses

In the early 2000s, a particular MRSA clonal complex (CC398) was found mainly in pigs and pig farmers in Europe. Since then, CC398 has been detected among a wide variety of animal species worldwide. We investigated the population structure of CC398 through mutation discovery at 97 genetic housekeeping loci, which are distributed along the CC398 chromosome within 195 CC398 isolates, collected from various countries and host species, including humans. Most of the isolates in this collection were received from collaborating microbiologists, who had preserved them over years. We discovered 96 bi-allelic polymorphisms, and phylogenetic analyses revealed that an epidemic sub-clone within CC398 (dubbed 'clade (C)') has spread within and between equine hospitals, where it causes nosocomial infections in horses and colonises the personnel. While clade (C) was strongly associated with S. aureus from horses in veterinary-care settings (p = 2 × 10(-7)), it remained extremely rare among S. aureus isolates from human infections

Levels of alpha-toxin correlate with distinct phenotypic response profiles of blood mononuclear cells and with agr background of community-associated Staphylococcus aureus isolates

Epidemiological studies of Staphylococcus aureus have shown a relation between certain clones and the presence of specific virulence genes, but how this translates into virulence-associated functional responses is not fully elucidated. Here we addressed this issue by analyses of community-acquired S. aureus strains characterized with respect to antibiotic resistance, ST types, agr types, and virulence gene profiles. Supernatants containing exotoxins were prepared from overnight bacterial cultures, and tested in proliferation assays using human peripheral blood mononuclear cells (PBMC). The strains displayed stable phenotypic response profiles, defined by either a proliferative or cytotoxic response. Although, virtually all strains elicited superantigen-mediated proliferative responses, the strains with a cytotoxic profile induced proliferation only in cultures with the most diluted supernatants. This indicated that the superantigen-response was masked by a cytotoxic effect which was also confirmed by flow cytometry analysis. The cytotoxic supernatants contained significantly higher levels of α-toxin than did the proliferative supernatants. Addition of α-toxin to supernatants characterized as proliferative switched the response into cytotoxic profiles. In contrast, no effect of Panton Valentine Leukocidin, δ-toxin or phenol soluble modulin α-3 was noted in the proliferative assay. Furthermore, a significant association between agr type and phenotypic profile was found, where agrII and agrIII strains had predominantly a proliferative profile whereas agrI and IV strains had a predominantly cytotoxic profile. The differential response profiles associated with specific S. aureus strains with varying toxin production could possibly have an impact on disease manifestations, and as such may reflect specific pathotypes