Epidemic Methicillin-resistant Staphylococcus Aureus Research Articles

Increase of genetic diversity and clonal replacement of epidemic methicillin-resistant Staphylococcus aureus strains in South-East Austria.

Spa-typing and microarray techniques were used to study epidemiological changes in methicillin-resistant Staphylococcus aureus (MRSA) in South-East Austria. The population structure of 327 MRSA isolated between 2002 and 2012 was investigated. MRSA was assigned to 58 different spa types and 14 different MLST CC (multilocus sequence type clonal complexes); in particular, between 2007 and 2012, an increasing diversity in MRSA clones could be observed. The most abundant clonal complex was CC5. On the respective SCCmec cassettes, the CC5 isolates differed clearly within this decade and CC5/SCCmecI, the South German MRSA, predominant in 2002, was replaced by CC5/SCCmecII, the Rhine-Hesse MRSA in 2012. Whereas in many European countries MLST CC22-MRSA (EMRSA 15, the Barnim epidemic MRSA) is predominant, this clone occurred in Austria nearly 10 years later than in neighbouring countries. CC45, the Berlin EMRSA, epidemic in Germany, was only sporadically found in South-East Austria. The Irish ST8-MRSA-II represented by spa-type t190 was frequently found in 2002 and 2007, but disappeared in 2012. Our results demonstrate clonal replacement of MRSA clones within the last years in Austria. Ongoing surveillance is warranted for detection of changes within the MRSA population.

Caged xanthones: Potent inhibitors of global predominant MRSA USA300

Total of 22 caged xanthones were subjected to susceptibility testing of global epidemic MRSA USA300. Natural morellic acid showed the strongest potency (MIC of 12.5μM). However, its potent toxicity diminishes MRSA therapeutic potential. We synthetically modified natural morellic acid to yield 13 derivatives (3a–3m). Synthetically modified 3b retained strong potency in MRSA growth inhibition, yet the toxicity was 20-fold less than natural morellic acid, permitting the possibility of using caged xanthones for MRSA therapeutic.

Optimizing the detection of methicillin-resistant Staphylococcus aureus with elevated vancomycin minimum inhibitory concentrations within the susceptible range.

BackgroundDetermination of vancomycin minimum inhibitory concentration (MIC) can influence the agent used to treat methicillin-resistant Staphylococcus aureus (MRSA) infection. We studied diagnostic accuracy using E-test and VITEK® 2 against a gold standard broth microdilution (BMD) methodology, the correlation between methods, and associations between vancomycin MIC and MRSA phenotype from clinical isolates.MethodsMRSA isolates were obtained from April 2012 to December 2013. Vancomycin MIC values were determined prospectively on all isolates by gradient diffusion E-test and automated VITEK® 2. The Clinical and Laboratory Standards Institute reference BMD method was performed retrospectively on thawed frozen isolates. Diagnostic accuracy for detecting less susceptible strains was calculated at each MIC cutoff point for E-Test and VITEK® 2 using BMD ≥1 µg/mL as a standard. The correlation between methods was assessed using Spearman’s rho (ρ). The association between MRSA phenotype and MIC for the three methods was assessed using Fisher’s exact test.ResultsOf 148 MRSA isolates, all except one (E-test =3 µg/mL) were susceptible to vancomycin (MIC of ≤2 µg/mL) irrespective of methodology. MICs were ≥1.0 µg/mL for 9.5% of BMD, 50.0% for VITEK® 2, and 27.7% for E-test. Spearman’s ρ showed weak correlations between methods: 0.29 E-test vs VITEK® 2 (P=0.003), 0.27 E-test vs BMD (P=0.001), and 0.31 VITEK® 2 vs BMD (P=0.002). The optimal cutoff points for detecting BMD-defined less susceptible strains were ≥1.0 µg/mL for E-test and VITEK® 2. E-test sensitivity at this cutoff point was 0.85 and specificity 0.29, while VITEK® 2 sensitivity and specificity were 0.62 and 0.51, respectively. Multiresistant MRSA strains tended to have higher MIC values compared to nonmultiresistant MRSA or epidemic MRSA 15 phenotypes by E-test (Fisher’s exact P<0.001) and VITEK® 2 (Fisher’s exact P<0.001).ConclusionOverall diagnostic accuracy and correlations between MIC methods used in routine diagnostic laboratories and the gold standard BMD showed limited overall agreement. This study helps optimize guidance on the effective use of vancomycin.

MRSA in equine hospitals and its significance for infections in humans

MRSA infections in equine clinics were reported from Northern America, Europe, Australia, and Japan. The majority of nosocomial infections in horses is obviously associated with particular MRSA clonal lineages. As already observed for epidemic MRSA in human hospitals more than 10 years ago, a dynamics of MRSA clonal lineages is also observed in European equine clinics: clonal lineages belonging to clonal complex (CC) 8 are on the retreat whereas MRSA attributed to CC398 become increasingly prevalent. The majority of CC398 isolates belong to a subpopulation which is particularly associated with equine hospitals as indicated by molecular typing. When emerging in equine clinics, MRSA from horses were also found as nasal colonizers of veterinary personnel. MRSA exhibiting the typing characteristics of MRSA known from equine clinics are obviously rare among MRSA from infections in humans. Although rare so far epidemic MRSA from human hospitals (HA-MRSA, e.g., ST22, ST225) have been isolated from nosocomial infections in horses and need particular attention in further surveillance.

Emergence of cfr-Mediated Linezolid Resistance in a Methicillin-Resistant Staphylococcus aureus Epidemic Clone Isolated from Patients with Cystic Fibrosis.

Resistance to linezolid (LZD) in methicillin-resistant Staphylococcus aureus (MRSA) isolates from patients with cystic fibrosis (CF) is due mainly to ribosomal mutations. We report on four CF patients with LZD-resistant MRSA bronchopulmonary infections by strains carrying the cfr gene. Strains from one patient also harbored the G2576U mutation (23S rRNA) and the G139R substitution (L3 protein). All strains belonged to the epidemic clone ST125 MRSA IVc. Our results support the monitoring of LZD resistance emergence in CF and non-CF MRSA isolates.

Systematic Review and Meta-Analysis of the Epidemiology of Vancomycin-Intermediate and Heterogeneous Vancomycin-Intermediate Staphylococcus aureus Isolates.

BackgroundVancomycin-intermediate Staphylococcus aureus (VISA) and heterogeneous VISA (hVISA) are associated with vancomycin treatment failure, and are becoming an increasing public health problem. Therefore, we undertook this study of 91 published studies and made subgroup comparisons of hVISA/VISA incidence in different study years, locations, and types of clinical samples. We also analyzed the genetic backgrounds of these strains.MethodsA systematic literature review of relevant articles published in PubMed and EMBASE from January 1997 to August 2014 was conducted. We selected and assessed journal articles reporting the prevalence rates of hVISA/VISA.ResultsThe pooled prevalence of hVISA was 6.05% in 99,042 methicillin-resistant S. aureus (MRSA) strains and that of VISA was 3.01% in 68,792 MRSA strains. The prevalence of hVISA was 4.68% before 2006, 5.38% in 2006–2009, and 7.01% in 2010–2014. VISA prevalence was 2.05%, 2.63%, and 7.93%, respectively. In a subgroup analysis of different isolation locations, the prevalence of hVISA strains was 6.81% in Asia and 5.60% in Europe/America, and that of VISA was 3.42% and 2.75%, respectively. The frequencies of hVISA isolated from blood culture samples and from all clinical samples were 9.81% and 4.68%, respectively, and those of VISA were 2.00% and 3.07%, respectively. The most prevalent genotype was staphylococcal cassette chromosome mec (SCCmec) II, which accounted for 48.16% and 37.74% of hVISA and VISA, respectively. Sequence Type (ST) 239 was most prevalent.ConclusionThe prevalence of hVISA/VISA has been increasing in recent years, but has been grossly underestimated. Its incidence is higher in Asia than in Europe/America. hVISA is isolated from blood culture samples more often than from other samples. These strains are highly prevalent in epidemic MRSA strains. This study clarifies the epidemiology of hVISA/VISA and indicates that the detection of these strains and the control of nosocomial infections must be strengthened.

MRSAspat1081, a Highly Transmissible Strain Endemic to Hong Kong, China, in the Netherlands

MRSA<i>spa</i>t1081, a Highly Transmissible Strain Endemic to Hong Kong, China, in the Netherlands

Monitoring meticillin resistant Staphylococcus aureus and its spread in Copenhagen, Denmark, 2013, through routine whole genome sequencing.

Typing of meticillin resistant Staphylococcus aureus (MRSA) by whole genome sequencing (WGS) is performed routinely in Copenhagen since January 2013. We describe the relatedness, based on WGS data and epidemiological data, of 341 MRSA isolates. These comprised all MRSA (n = 300) identified in Copenhagen in the first five months of 2013. Moreover, because MRSA of staphylococcal protein A (spa)-type 304 (t304), sequence type (ST) 6 had been associated with a continuous neonatal ward outbreak in Copenhagen starting in 2011, 41 t304 isolates collected in the city between 2010 and 2012 were also included. Isolates from 2013 found to be of t304, ST6 (n=14) were compared to the 41 earlier isolates. In the study, isolates of clonal complex (CC) 22 were examined in detail, as this CC has been shown to include the hospital-acquired epidemic MRSA (EMRSA-15) clone. Finally, all MRSA ST80 were also further analysed, as representatives of an important community-acquired MRSA in Europe. Overall the analysis identified 85 spa-types and 35 STs from 17 CCs. WGS confirmed the relatedness of epidemiologically linked t304 neonatal outbreak isolates. Several non-outbreak related patients had isolates closely related to the neonatal isolates suggesting unrecognised community chains of transmission and insufficient epidemiological data. Only four CC22 isolates were related to EMRSA-15. No community spread was observed among the 13 ST80 isolates. WGS successfully replaced conventional typing and added information to epidemiological surveillance. Creation of a MRSA database allows clustering of isolates based on single nucleotide polymorphism (SNP) calling and has improved our understanding of MRSA transmission.

Molecular epidemiology of methicillin resistant staphylococcus aureus colonizing the anterior Nares of school children of Udupi Taluk

Context: Community associated methicillin resistant Staphylococcus aureus (CA-MRSA) cause serious skin and soft tissue infections including necrotizing fasciitis and necrotizing pneumonia. Production of Panton Valentine Leucocidine (PVL) toxin is implicated in its enhanced virulence. A variant of epidemic MRSA-15 (EMRSA-15) which produces PVL toxin has been isolated and characterized by pulsed-field gel electrophoresis (PFGE) method from the Indian population both in hospital and community settings. Aims: Identify the epidemiological type of MRSA colonizing the anterior nares of school children in Udupi taluk. Settings and Design: The study population included children of the age group of 5-16 years belonging to the Udupi taluk of Karnataka, India. A total of 1503 children were screened for MRSA colonization during July 2009 to December 2010. Materials and Methods: PVL assay, Staphylococcal Cassette Chromosome (SCC) mec typing and PFGE typing were carried out with all the MRSA isolates. Statistical Analysis Used: Frequency distribution of different variables was assessed by SPSS. Results: Among the 1.1% of MRSA, 58.8% (10/17) of isolates were positive for pvl and 41.7% (7/17) were identified as SCC mec type IV. PFGE patterns of all the strains were identical with Indian variant EMRSA-15; however they were different from classical EMRSA-15 in 3-4 bands. Conclusions: The Indian variant EMRSA-15 gains much epidemiological relevance owing to the acquisition of pvl gene. In spite of low prevalence of nasal colonization of MRSA, emergence of the virulent Indian variant EMRSA-15 in our community is a worrisome fact to be reckoned with.

Methicillin-resistant Staphylococcus aureus strains from Ghana include USA300.

The objective of this study was to provide baseline information on circulating methicillin-resistant Staphylococcus aureus (MRSA) clones in Ghana. Thirty MRSA isolates collected between 2010 and 2013 from patients and healthy carriers were characterised by DNA microarray analysis, staphylococcal protein A (spa) typing, multilocus sequence typing (MLST) and minimum inhibitory concentration (MIC) determination to 21 antimicrobial agents. Phenotypic resistance was detected to tetracycline (67%), norfloxacin (40%), moxifloxacin (37%), erythromycin (37%), clindamycin (33%), gentamicin (30%), kanamycin (30%) and ceftaroline (20%), whereas no resistance was observed for glycopeptides, linezolid, daptomycin and tigecycline. DNA microarray analysis showed that tet(M) (43%), tet(K) (33%), aphA3 (23%), aacA-aphD (17%) and erm(C) (13%) were the most prevalent resistance genes. ST88-IV (WA MRSA-2) (n=8), ST8-IV (USA300) (n=5) containing arginine catabolic mobile element (ACME) and Panton-Valentine leukocidin (PVL), and ST247-I (North German/Iberian EMRSA) (n=4) were the most frequent clones detected. All MRSA contained sak and scn genes, one isolate (ST36-II) harboured the gene encoding the toxic shock syndrome toxin (TSST) and none contained exfoliative toxin genes. In conclusion, the relatively high levels of resistance to easily accessible non-β-lactam agents further complicate the treatment of MRSA infections in Ghana. The occurrence of USA300 and other epidemic multidrug-resistant MRSA clones in this African country is a matter of public health concern due to the lack of adequate infrastructures for MRSA surveillance and control in this geographical setting.

Population structure analyses of Staphylococcus aureus at Tygerberg Hospital, South Africa, reveals a diverse population, a high prevalence of Panton–Valentine leukocidin genes, and unique local methicillin-resistant S. aureus clones

Studies reporting on the population structure of Staphylococcus aureus in South Africa have focused only on methicillin-resistant S. aureus (MRSA). This study describes the population structure of S. aureus, including methicillin-susceptible S. aureus (MSSA) isolated from patients at Tygerberg Academic Hospital, Western Cape province. Pulsed-field gel electrophoresis (PFGE), detection of Panton–Valentine leukocidin (PVL), spa typing, multilocus sequence typing (MLST), agr typing and SCCmec typing were used to characterize strains. Of 367 non-repetitive S. aureus isolates collected over a period of 1 year, 56 (15.3%) were MRSA. Skin and soft tissue infections were the most frequent source (54.8%), followed by bone and joint (15.3%) and respiratory tract infections (7.7%). For strain typing, PFGE was the most discriminative method, and resulted in 31 pulsotypes (n = 345, 94.0%), as compared with 16 spa clonal complexes (CCs) (n = 344, 93.4%). Four MLST CCs were identified after eBURST of sequence types (STs) of selected isolates. One hundred and sixty isolates (MSSA, n = 155, 42.2%) were PVL-positive, and agr types I–IV and SCCmec types I–V were identified. Our S. aureus population consisted of genotypically diverse strains, with PVL being a common characteristic of MSSA. MSSA and MRSA isolates clustered in different clones. However, the dominant MRSA clone (ST612) also contained an MSSA isolate, and had a unique genotype. Common global epidemic MRSA clones, such as ST239-MRSA-III and ST36-MRSA-II, were identified. A local clone, ST612-MRSA-IV, was found to be the dominant MRSA clone.

First description of PVL-positive methicillin-resistant Staphylococcus aureus (MRSA) in wild boar meat

Staphylococcus aureus is an important food-borne pathogen due to the ability of enterotoxigenic strains to produce staphylococcal enterotoxins (SEs) in food. Methicillin-resistant S. aureus (MRSA) is also an important pathogen for humans, causing severe and hard to treat diseases in hospitals and in the community due to its multiresistance against antimicrobials. In particular, strains harbouring genes encoding for the Panton–Valentine leukocidin (PVL) toxin are of concern from a public health perspective as they are usually capable of causing severe skin and soft tissue infections (sSSTIs) and occasionally necrotizing pneumonia which is associated with high mortality. This is the first report on the detection of MRSA with genes encoding for PVL in wild boar meat. Among the 28 MRSA isolated from wild boar meat in the course of a national monitoring programme in Germany, seven harboured PVL-encoding genes. Six of the isolates were identical according to the results of spa-, MLST-, microarray- and PFGE-typing. They could be assigned to the epidemic MRSA clone USA300. Epidemiological investigations revealed that people handling the food were the most likely common source of contamination with these MRSA. These findings call again for suitable hygienic measures at all processing steps of the food production chain. The results of the study underline that monitoring along the food chain is essential to closely characterise the total burden of MRSA for public health.

A shared population of epidemic methicillin-resistant Staphylococcus aureus 15 circulates in humans and companion animals.

ABSTRACTMethicillin-resistant Staphylococcus aureus (MRSA) is a global human health problem causing infections in both hospitals and the community. Companion animals, such as cats, dogs, and horses, are also frequently colonized by MRSA and can become infected. We sequenced the genomes of 46 multilocus sequence type (ST) 22 MRSA isolates from cats and dogs in the United Kingdom and compared these to an extensive population framework of human isolates from the same lineage. Phylogenomic analyses showed that all companion animal isolates were interspersed throughout the epidemic MRSA-15 (EMRSA-15) pandemic clade and clustered with human isolates from the United Kingdom, with human isolates basal to those from companion animals, suggesting a human source for isolates infecting companion animals. A number of isolates from the same veterinary hospital clustered together, suggesting that as in human hospitals, EMRSA-15 isolates are readily transmitted in the veterinary hospital setting. Genome-wide association analysis did not identify any host-specific single nucleotide polymorphisms (SNPs) or virulence factors. However, isolates from companion animals were significantly less likely to harbor a plasmid encoding erythromycin resistance. When this plasmid was present in animal-associated isolates, it was more likely to contain mutations mediating resistance to clindamycin. This finding is consistent with the low levels of erythromycin and high levels of clindamycin used in veterinary medicine in the United Kingdom. This study furthers the “one health” view of infectious diseases that the pathogen pool of human and animal populations are intrinsically linked and provides evidence that antibiotic usage in animal medicine is shaping the population of a major human pathogen.

Characterization of Staphylococcus aureus from distinct geographic locations in China: an increasing prevalence of spa-t030 and SCCmec type III.

Staphylococcus aureus belongs to one of the most common bacteria causing healthcare and community associated infections in China, but their molecular characterization has not been well studied. From May 2011 to June 2012, a total of 322 non-duplicate S. aureus isolates were consecutively collected from seven tertiary care hospitals in seven cities with distinct geographical locations in China, including 171 methicillin sensitive S. aureus (MSSA) and 151 MRSA isolates. All isolates were characterized by spa typing. The presence of virulence genes was tested by PCR. MRSA were further characterized by SCCmec typing. Seventy four and 16 spa types were identified among 168 MSSA and 150 MRSA, respectively. One spa type t030 accounted for 80.1% of all MRSA isolates, which was higher than previously reported, while spa-t037 accounted for only 4.0% of all MRSA isolates. The first six spa types (t309, t189, t034, t377, t078 and t091) accounted for about one third of all MSSA isolates. 121 of 151 MRSA isolates (80.1%) were identified as SCCmec type III. pvl gene was found in 32 MSSA (18.7%) and 5 MRSA (3.3%) isolates, with ST22-MSSA-t309 as the most commonly identified strain. Compared with non-epidemic MRSA clones, epidemic MRSA clones (corresponding to ST239) exhibited a lower susceptibility to rifampin, ciprofloxacin, gentamicin and trimethoprim-sulfamethoxazole, a higher prevalence of sea gene and a lower prevalence of seb, sec, seg, sei and tst genes. The increasing prevalence of multidrug resistant spa-t030 MRSA represents a major public health problem in China.

Healthcare-associated outbreak of meticillin-resistant Staphylococcus aureus bacteraemia: role of a cryptic variant of an epidemic clone

SummaryBackgroundNew strains of meticillin-resistant Staphylococcus aureus (MRSA) may be associated with changes in rates of disease or clinical presentation. Conventional typing techniques may not detect new clonal variants that underlie changes in epidemiology or clinical phenotype.AimTo investigate the role of clonal variants of MRSA in an outbreak of MRSA bacteraemia at a hospital in England.MethodsBacteraemia isolates of the major UK lineages (EMRSA-15 and -16) from before and after the outbreak were analysed by whole-genome sequencing in the context of epidemiological and clinical data. For comparison, EMRSA-15 and -16 isolates from another hospital in England were sequenced. A clonal variant of EMRSA-16 was identified at the outbreak hospital and a molecular signature test designed to distinguish variant isolates among further EMRSA-16 strains.FindingsBy whole-genome sequencing, EMRSA-16 isolates during the outbreak showed strikingly low genetic diversity (P < 1 × 10−6, Monte Carlo test), compared with EMRSA-15 and EMRSA-16 isolates from before the outbreak or the comparator hospital, demonstrating the emergence of a clonal variant. The variant was indistinguishable from the ancestral strain by conventional typing. This clonal variant accounted for 64/72 (89%) of EMRSA-16 bacteraemia isolates at the outbreak hospital from 2006.ConclusionsEvolutionary changes in epidemic MRSA strains not detected by conventional typing may be associated with changes in disease epidemiology. Rapid and affordable technologies for whole-genome sequencing are becoming available with the potential to identify and track the emergence of variants of highly clonal organisms.

Methicillin-ResistantStaphylococcus aureus

Despite the availability of effective preventative and therapeutic strategies, methicillin-resistant Staphylococcus aureus (MRSA) remains amajor cause of health care–associated and, more recently, community-acquired infections. Since the introduction of methicillin in 1959, epidemics due to different clones ofMRSAhave occurred in diverse geographic regions.1 The result has been the persistence of MRSA as an important pathogen worldwide. In the United States, themost recentMRSA epidemic has been due to the clone USA300. USA300 has been responsible for themajorityof communityandhealth care–associatedoutbreaks since the early 1990s.1 Until recently, thenumber of infections has continued to rise, especially in urban centers. While themajority of these have been skin and soft-tissue infections (SSTIs), a notable minority have been invasive and life-threatening.1 Two articles in this issueof JAMAInternalMedicineprovide additional insight into both the incidence and the risks associatedwithdevelopmentofMRSA infections.Danteset al2provide evidence that the overall incidence of invasiveMRSA infections nationally is declining. Casey et al3 perform an ecological investigation; they find that individuals livingnear high-density farming operations (ie, farms having an animal density of 2 animals or 1000-lbs live weight per acre) or near crop fieldsusingmanure exported from thehigh-density livestock farms are at increased risk of developingMRSA or other bacterial SSTIs. Both studies raise questions thatwarrant further investigation. Dantes et al2 useddata from theActiveBacterial Core Surveillance Program (ABCSP) to compare the national incidence rate of invasive MRSA infections for the years 2005 through 2011. The findings are of special importance because of the surveymethods that areusedand the size of the sample population (approximately 15 million). The ABCSP is an active surveillance program that is population based and includes a reviewofmedical records. It serves as amodel for this type of surveillance. The results show that the incidence of health care–associated community-onset and hospital-onset invasive MRSA infections have declined by 27.7% and 54.2%, respectively. The findings are consistent with several earlier reports including one from the same group. This impressive declinemaywellbeduetothe implementationof infectioncontrol efforts at thehospital level; however, it is alsopossible that the epidemic MRSA strains circulating in the health care setting have evolved, becoming less virulent. In contrast to the decline inhealth care–associatedMRSA, the estimated reduction in incidence of community-associated infections was a modest 5%. In addition to the evidence that the incidenceofMRSA infections is declining, the study highlights 2 important points. The first is the observation that there is an increased risk of recurrent infection among recentlyhospitalizedpatientswith healthcare–associatedcommunity-onset infections.Thesepatients had a 64% risk of infection at 3 months or less followingdischarge.Others have also reported ahigh risk of reinfection in discharged patients either infected or colonized with MRSA.4 Possible explanations for the patients’ increased risk include underlyingmedical comorbidities, persistent colonizationor recolonizationwithMRSA,orexposure tosimilar risks of MRSA infection following discharge. This subset of patients would appear to be candidates for studies directed at reducing their high risk of recurrent infection. The second finding is the limited change in the incidence of communityassociated infections. This is in marked contrast with the reduction in the incidence of health care–associated infections. As noted by the authors, the study highlights the need for a greater understanding of how these epidemic MRSA strains spread and initiate infection within the community. Related articles pages 1970 and 1980 Research Original Investigation National Burden of Invasive MRSA Infections

Genotype diversity of methicillin-resistant Staphylococcus aureus in Shenyang, China

The aim of this study was to better understand methicillin-resistant Staphylococcus aureus (MRSA) at the molecular level by investigating the genotypic characteristics and evolutionary patterns of MRSA clones in Shenyang, China. We analyzed the molecular epidemiology of 60 MRSA isolates in Shenyang, China, between 2002 and 2008, using pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), staphylococcal cassette chromosome mec (SCCmec) typing, and S. aureus protein A (spa) typing. They were examined for their antimicrobial susceptibilities. Among the 60 isolates, ST239 was identified most frequently (34 isolates; 58%), followed by ST5 (20 isolates; 34%). Nine spa types were obtained and 4 PFGE strain families (A, B, C, and D) were resolved. Spa type t030, which corresponded to PFGE genotypes A1, A3, and A4, constituted 45% (27/60) of all isolates; spa type t037, which corresponded to PFGE type A2, accounted for 13% (8/60) of all isolates. These 2 spa genotypes belonged to ST239 and carried SCCmec type III. Isolates genotyped as spa type t002 comprised 27% (16/60) of the study set and included isolates typed as PFGE B1 and B2, ST5, and SCCmec II. Most of MRSA isolates belonging to ST239 were susceptible to trimethoprim-sulfamethoxazole. The minimum inhibitory concentration (MIC50) of vancomycin among MRSA isolates belonging to ST5 (2 mg/l) was higher than that for other isolates (1 mg/l). These data document 2 major epidemic MRSA clones in Shenyang, China: ST239-MRSA-SCCmec type III-t037/t030 and ST5-MRSA-SCCmec type II-t002.

Evaluation of the affinity of various species and strains of Staphylococcus to adhere to equine corneocytes

Meticillin-resistant Staphylococcus aureus (MRSA) strain USA 500 predominately colonizes horses and people working with them. Previous studies demonstrate that some Staphylococcus species exhibit higher affinity for corneocytes of specific mammalian species. The objective was to determine the relative affinities of various MRSA strains, meticillin-susceptible S.aureus (MSSA) strains and a meticillin-susceptible Staphylococcuspseudintermedius (MSSP) for equine corneocytes. We hypothesized that MRSA strain USA 500 would exhibit greater adhesion than other staphylococcal strains tested. Epidemic MRSA strains (USA 100, USA 300, USA 500 and USA 800), two MSSA control strains and an MSSP field strain were tested on corneocytes from 15 client-owned horses. Isolates were incubated with corneocytes in conditions (bacterial concentration of 10(8) colony-forming units/mL for 45min) recently shown to maximize adherence of S.aureus without competitive interference. A validated image-analysis system was used to quantify the cell surface density of bacterial adhesion. The MSSP strain adhered with significantly higher affinity (P<0.0015) to corneocytes than did MSSA strains. All MRSA strains other than USA 500 had significantly higher affinity than MSSA strains (P range <0.03 to <0.0015). There were no statistical differences in adhesion between strain USA 500 and the other MRSA strains tested. Meticillin-resistant S.aureus strain USA 500 did not adhere more robustly than other strains of Staphylococcus; therefore, its affinity to colonize horses may not be solely attributed to corneocyte adhesion. Additional studies are required to explain the epidemiological role of this strain as the predominant cause of colonization and infections of horses in North America.

Molecular epidemiology of hospital-onset methicillin-resistant Staphylococcus aureus infections in Southern Chile

Methicillin-resistant Staphylococcus aureus (MRSA) is a pathogen of public health importance. In Chile, the Cordobes/Chilean clone was the predominant healthcare-associated MRSA (HA-MRSA) clone in 1998. Since then, the molecular epidemiological surveillance of MRSA has not been performed in Southern Chile. We aimed to investigate the molecular epidemiology of HA-MRSA infections in Southern Chile to identify the MRSA clones involved, and their evolutionary relationships with epidemic international MRSA lineages. A total of 303 single inpatient isolates of S. aureus were collected in the Valdivia County Hospital (2007-2008), revealing 33% (100 MRSA/303) prevalence for HA-MRSA infections. The SCCmec types I and IV were identified in 97% and 3% of HA-MRSA, respectively. All isolates lacked the pvl genes. A random sample (n = 29) of all MRSA was studied by pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), SCCmec subtyping, agr and spa typing, and virulence genes profiling. PFGE analysis revealed the predominance (89%, 26/29) of pulsotype A and three additional pulsotypes, designated H1, I33, and G1. Pulsotype A (ST5-SCCmecI-spa-t149) is clonally related to the Cordobes/Chilean clone. Pulsotype H1 (ST5-SCCmecIVNT-spa-t002) is genetically related to the Pediatric clone (ST5-SCCmecIV). Pulsotype I33 (ST5-SCCmecIVc-spa-t002) is clonally related by PFGE to the community-associated MRSA (CA-MRSA) clone spread in Argentina, I-ST5-IVa-PVL(+). The G1 pulsotype (ST8-SCCmecIVc-spa-t024) is clonally related to the epidemic USA300 CA-MRSA. Here, we demonstrate the stability of the Cordobes/Chilean clone over time as the major HA-MRSA clone in Southern Chile. The identification of two CA-MRSA clones might suggest that these clones have entered into the healthcare setting from the community. These results emphasize the importance of the local surveillance of MRSA infections in the community and hospital settings.

Analysis of the Small RNA Transcriptional Response in Multidrug-Resistant Staphylococcus aureus after Antimicrobial Exposure

The critical role of noncoding small RNAs (sRNAs) in the bacterial response to changing conditions is increasingly recognized. However, a specific role for sRNAs during antibiotic exposure has not been investigated in Staphylococcus aureus. Here, we used Illumina RNA-Seq to examine the sRNA response of multiresistant sequence type 239 (ST239) S. aureus after exposure to four antibiotics (vancomycin, linezolid, ceftobiprole, and tigecycline) representing the major classes of antimicrobials used to treat methicillin-resistant S. aureus (MRSA) infections. We identified 409 potential sRNAs and then compared global sRNA and mRNA expression profiles at 2 and 6 h, without antibiotic exposure and after exposure to each antibiotic, for a vancomycin-susceptible strain (JKD6009) and a vancomycin-intermediate strain (JKD6008). Exploration of this data set by multivariate analysis using a novel implementation of nonnegative matrix factorization (NMF) revealed very different responses for mRNA and sRNA. Where mRNA responses clustered with strain or growth phase conditions, the sRNA responses were predominantly linked to antibiotic exposure, including sRNA responses that were specific for particular antibiotics. A remarkable feature of the antimicrobial response was the prominence of antisense sRNAs to genes encoding proteins involved in protein synthesis and ribosomal function. This study has defined a large sRNA repertoire in epidemic ST239 MRSA and shown for the first time that a subset of sRNAs are part of a coordinated transcriptional response to specific antimicrobial exposures in S. aureus. These data provide a framework for interrogating the role of staphylococcal sRNAs in antimicrobial resistance and exploring new avenues for sRNA-based antimicrobial therapies.