Distribution of Staphylococcal Cassette Chromosome mec Types and Correlation with Comorbidity and Infection Type in Patients with MRSA Bacteremia

Background

Molecular epidemiological definitions that are based on staphylococcal cassette chromosome mec (SCCmec) typing and phylogenetic analysis of methicillin-resistant Staphylococcus aureus (MRSA) isolates are considered a reliable way to distinguish between healthcare-associated MRSA (HA-MRSA) and community-associated MRSA (CA-MRSA). However, there is little information regarding the clinical features and outcomes of bacteremia patients with MRSA carrying different SCCmec types.

Methods

From January 1 through December 31, 2006, we recorded the demographic data and outcomes of 159 consecutive adult MRSA bacteremia patients from whom isolates for SCCmec analysis were collected. All participants were patients at a tertiary care center in Taiwan.

Principal Findings

The following SCCmec types were identified in MRSA isolates: 30 SCCmec II (18.9%), 87 SCCmec III (54.7%), 22 SCCmec IV (13.8%), and 20 SCCmec V (12.6%). The time from admission to the first MRSA-positive blood culture for patients infected with isolates with the SCCmec III element (mean/median, 50.7/26 days) was significantly longer than for patients infected with isolates carrying SCCmec IV or V (mean/median, 6.7/3 days for SCCmec IV; 11.1/10.5 days for SCCmec V) (P<0.05). In univariate analysis, community onset, soft tissue infection, and deep-seated infection were predictors for SCCmec IV/V. In multivariate analysis, length of stay before index culture, diabetes mellitus, and being bedridden were independent risk factors associated with SCCmec II/III.

Conclusions

These findings are in agreement with previous studies of the genetic characteristics of CA-MRSA. MRSA bacteremia with SCCmec II/III isolates occurred more among patients with serious comorbidities and prolonged hospitalization. Community onset, skin and soft tissue infection, and deep-seated infection best predicted SCCmec IV/V MRSA bacteremia.     

Genetic Diversity of Arginine Catabolic Mobile Element in Staphylococcus epidermidis

Background

The methicillin-resistant Staphylococcus aureus clone USA300 contains a novel mobile genetic element, arginine catabolic mobile element (ACME), that contributes to its enhanced capacity to grow and survive within the host. Although ACME appears to have been transferred into USA300 from S. epidermidis, the genetic diversity of ACME in the latter species remains poorly characterized.

Methodology/Principal Findings

To assess the prevalence and genetic diversity of ACME, 127 geographically diverse S. epidermidis isolates representing 86 different multilocus sequence types (STs) were characterized. ACME was found in 51% (65/127) of S. epidermidis isolates. The vast majority (57/65) of ACME-containing isolates belonged to the predominant S. epidermidis clonal complex CC2. ACME was often found in association with different allotypes of staphylococcal chromosome cassette mec (SCCmec) which also encodes the recombinase function that facilities mobilization ACME from the S. epidermidis chromosome. Restriction fragment length polymorphism, PCR scanning and DNA sequencing allowed for identification of 39 distinct ACME genetic variants that differ from one another in gene content, thereby revealing a hitherto uncharacterized genetic diversity within ACME. All but one ACME variants were represented by a single S. epidermidis isolate; the singular variant, termed ACME-I.02, was found in 27 isolates, all of which belonged to the CC2 lineage. An evolutionary model constructed based on the eBURST algorithm revealed that ACME-I.02 was acquired at least on 15 different occasions by strains belonging to the CC2 lineage.

Conclusions/Significance

ACME-I.02 in diverse S. epidermidis isolates were nearly identical in sequence to the prototypical ACME found in USA300 MRSA clone, providing further evidence for the interspecies transfer of ACME from S. epidermidis into USA300.     

High genetic diversity among community-associated Staphylococcus aureus in Europe: results from a multicenter study

Background

Several studies have addressed the epidemiology of community-associated Staphylococcus aureus (CA-SA) in Europe; nonetheless, a comprehensive perspective remains unclear. In this study, we aimed to describe the population structure of CA-SA and to shed light on the origin of methicillin-resistant S. aureus (MRSA) in this continent.

Methods and Findings

A total of 568 colonization and infection isolates, comprising both MRSA and methicillin-susceptible S. aureus (MSSA), were recovered in 16 European countries, from community and community-onset infections. The genetic background of isolates was characterized by molecular typing techniques (spa typing, pulsed-field gel electrophoresis and multilocus sequence typing) and the presence of PVL and ACME was tested by PCR. MRSA were further characterized by SCCmec typing. We found that 59% of all isolates were associated with community-associated clones. Most MRSA were related with USA300 (ST8-IVa and variants) (40%), followed by the European clone (ST80-IVc and derivatives) (28%) and the Taiwan clone (ST59-IVa and related clonal types) (15%). A total of 83% of MRSA carried Panton-Valentine leukocidin (PVL) and 14% carried the arginine catabolic mobile element (ACME). Surprisingly, we found a high genetic diversity among MRSA clonal types (ST-SCCmec), Simpson’s index of diversity = 0.852 (0.788–0.916). Specifically, about half of the isolates carried novel associations between genetic background and SCCmec. Analysis by BURP showed that some CA-MSSA and CA-MRSA isolates were highly related, suggesting a probable local acquisition/loss of SCCmec.

Conclusions

Our results imply that CA-MRSA origin, epidemiology and population structure in Europe is very dissimilar from that of USA.     

Potential associations between severity of infection and the presence of virulence-associated genes in clinical strains of Staphylococcus aureus

Background

The clinical spectrum of Staphylococcus aureus infection ranges from asymptomatic nasal carriage to osteomyelitis, infective endocarditis (IE) and death. In this study, we evaluate potential association between the presence of specific genes in a collection of prospectively characterized S. aureus clinical isolates and clinical outcome.

Methodology/Principal Findings

Two hundred thirty-nine S. aureus isolates (121 methicillin-resistant S. aureus [MRSA] and 118 methicillin-susceptible S. aureus [MSSA]) were screened by array comparative genomic hybridization (aCGH) to identify genes implicated in complicated infections. After adjustment for multiple tests, 226 genes were significantly associated with severity of infection. Of these 226 genes, 185 were not in the SCCmec element. Within the 185 non-SCCmec genes, 171 were less common and 14 more common in the complicated infection group. Among the 41 genes in the SCCmec element, 37 were more common and 4 were less common in the complicated group. A total of 51 of the 2014 sequences evaluated, 14 non-SCCmec and 37 SCCmec, were identified as genes of interest.

Conclusions/Significance

Of the 171 genes less common in complicated infections, 152 are of unknown function and may contribute to attenuation of virulence. The 14 non-SCCmec genes more common in complicated infections include bacteriophage-encoded genes such as regulatory factors and autolysins with potential roles in tissue adhesion or biofilm formation.     

Molecular Characterization of Staphylococcus aureus from Patients with Surgical Site Infections at Mulago Hospital in Kampala,Uganda

Background

The prevalence of Methicillin resistant Staphylococcus aureus (MRSA) is progressively increasing globally with significant regional variation. Understanding the Staphylococcus aureus lineages is crucial in controlling nosocomial infections. Recent studies on S. aureus in Uganda have revealed an escalating burden of MRSA. However, the S. aureus genotypes circulating among patients are not known. Here, we report S. aureus lineages circulating in patients with surgical site infections (SSI) at Mulago National hospital, Kampala, Uganda.

Methods

A cross-sectional study involving 314 patients with SSI at Mulago National Hospital was conducted from September 2011 to April 2012. Pus swabs from the patients’ SSI were processed using standard microbiological procedures. Methicillin sensitive Staphylococcus aureus (MSSA) and MRSA were identified using phenotypic tests and confirmed by PCR-detection of the nuc and mecA genes, respectively. SCCmec genotypes were determined among MRSA isolates using multiplex PCR. Furthermore, to determine lineages, spa sequence based-genotyping was performed on all S. aureus isolates.

Results

Of the 314 patients with SSI, S. aureus accounted for 20.4% (64/314), of which 37.5% (24/64) were MRSA. The predominant SCCmec types were type V (33.3%, 8/24) and type I (16.7%, 4/24). The predominant spa lineages were t645 (17.2%, 11/64) and t4353 (15.6%, 10/64), and these were found to be clonally circulating in all the surgical wards. On the other hand, lineages t064, t355, and t4609 were confined to the obstetrics and gynecology wards. A new spa type (t10277) was identified from MSSA isolate. On multivariate logistic regression analysis, cancer and inducible clindamycin resistance remained as independent predictors of MRSA-SSI.

Conclusion

SCCmec types I and V are the most prevalent MRSA mecA types from the patients’ SSI. The predominant spa lineages (t645 and t4353) are clonally circulating in all the surgical wards, calling for strengthening of infection control practices at Mulago National Hospital.     

High prevalence of EMRSA-15 in Portuguese public buses: a worrisome finding

Background

The nosocomial prevalence of methicillin resistant Staphylococcus aureus (MRSA) in Portugal remains one of the highest in Europe and is currently around 50%. Transmission of S. aureus, including MRSA, occurs principally by direct human-to-human skin contact. However, S. aureus can survive for long periods on inanimate objects, which may represent an important reservoir for dissemination as well.

Methodology/Principal Findings

Between May 2009 and February 2010, handrails of 85 public urban buses circulating in Oporto, Portugal, were screened for the occurrence of MRSA. Twenty-two (26%) buses showed MRSA contamination. The molecular characterization of a total of 55 MRSA, by pulsed-field gel electrophoresis (PFGE), staphylococcal cassette chromosome (SCC) mec typing, spa typing, and multilocus sequence typing (MLST), clustered the isolates into three clonal types. However, the overwhelming majority (n = 50; 91%) of the isolates belonged to a single clone (PFGE A, spa types t747, t032, t025 or t020, ST22, SCCmec type IVh) that exhibits the characteristics of the pandemic EMRSA-15, currently the major lineage circulating in Portuguese hospitals, namely in the Oporto region. Two additional clones were found but in much lower numbers: (i) PFGE B, ST5, spa type t002, SCCmec IVa (n = 3), and (ii) PFGE C, spa type t008, ST8, SCCmec IVa (n = 2). None of the 55 isolates was PVL positive.

Conclusions/Significance

Public buses in Oporto seem to be an important reservoir of MRSA of nosocomial origin, providing evidence that the major hospital-associated MRSA clone in Portugal is escaping from the primary ecological niche of hospitals to the community environment. Infection control measures are urgently warranted to limit the spread of EMRSA-15 to the general population and future studies are required to assess the eventual increase of MRSA in the Portuguese community, which so far remains low.     

SCC<Emphasis Type="Italic">mec</Emphasis> Type IV,PVL-Negative,Methicillin-Resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> in Cystic Fibrosis Patients from Brazil

Twenty seven S. aureus isolates were obtained from cystic fibrosis (CF) patients at a tertiary care hospital in Brazil. Nineteen (70.4%) were methicillin-susceptible S. aureus (MSSA) and eight (29.6%) methicillin-resistant S. aureus (MRSA). Of the MRSA isolates, four had SCCmec type III and four had SCCmec type IV. PVL genes were not detected in any of the MSSA or MRSA isolates. New studies are necessary to evaluate the exact impact of these different MRSA clones in CF patients.     

Comparison of Biofilm Formation between Major Clonal Lineages of Methicillin Resistant Staphylococcus aureus

Objectives

Epidemic methicillin-resistant S. aureus (MRSA) clones cause infections in both hospital and community settings. As a biofilm phenotype further facilitates evasion of the host immune system and antibiotics, we compared the biofilm-forming capacities of various MRSA clones.

Methods

Seventy-six MRSA classified into 13 clones (USA300, EMRSA-15, Hungarian/Brazilian etc.), and isolated from infections or from carriers were studied for biofilm formation under static and dynamic conditions. Static biofilms in microtitre plates were quantified colorimetrically. Dynamic biofilms (Bioflux 200, Fluxion, USA) were studied by confocal laser-scanning and time-lapse microscopy, and the total volume occupied by live/dead bacteria quantified by Volocity 5.4.1 (Improvision, UK).

Results

MRSA harbouring SCCmec IV produced significantly more biomass under static conditions than SCCmec I–III (P = 0.003), and those harbouring SCCmec II significantly less than those harbouring SCCmec I or III (P<0.001). In the dynamic model, SCCmec I–III harbouring MRSA were significantly better biofilm formers than SCCmec IV (P = 0.036). Only 16 strains successfully formed biofilms under both conditions, of which 13 harboured SCCmec IV and included all tested USA300 strains (n = 3). However, USA300 demonstrated remarkably lower percentages of cell-occupied space (6.6%) compared to the other clones (EMRSA-15 = 19.0%) under dynamic conditions. Time-lapse microscopy of dynamic biofilms demonstrated that USA300 formed long viscoelastic tethers that stretched far from the point of attachment, while EMRSA-15 consisted of micro-colonies attached densely to the surface.

Conclusions

MRSA harbouring SCCmec types IV and I–III demonstrate distinct biofilm forming capacities, possibly owing to their adaptation to the community and hospital settings, respectively. USA300 demonstrated abundant biofilm formation under both conditions, which probably confers a competitive advantage, contributing to its remarkable success as a pathogen.     

Difference in agr Dysfunction and Reduced Vancomycin Susceptibility between MRSA Bacteremia Involving SCCmec Types IV/IVa and I–III

Background

Dysfunction of agr, with reduced susceptibility or hetero-resistance to vancomycin, is thought to be associated with a worse outcome of methicillin-resistant Staphylococcus aureus (MRSA) bacteremia (MRSAB). However, the difference in agr dysfunction according to the SCCmec type in MRSA infection is undetermined. We compared the prevalence of agr dysfunction, reduced vancomycin susceptibility and the outcomes of SCCmec IV/IVa and I–III MRSAB.

Methods

The study included 307 cases of MRSAB. SCCmec types were determined by multiplex PCR. The clinical and microbiological features and outcomes of 58 SCCmec IV/IVa MRSAB were compared with those of 249 SCCmec I–III MRSAB.

Results

Compared with SCCmec I–III MRSAB, SCCmec IV/IVa MRSAB was associated with lower rates of agr dysfunction (3% vs. 43%), vancomycin minimum inhibitory concentration (MIC) = 2 µg/mL (3% vs. 15%), and hetero-resistance to vancomycin (0% vs. 8%) (all P<0.05). However, the 30-day and S. aureus-related mortality in patients with SCCmec IV/IVa MRSAB were not different from those in patients with SCCmec I–III MRSAB in multivariate analyses (HR 1.168, 95% CI 0.705–1.938; HR 1.025, 95% CI 0.556–1.889).

Conclusions

SCCmec IV/IVa MRSAB was associated with lower rates of agr dysfunction and hetero-resistance to vancomycin and a lower vancomycin MIC, compared with SCCmec I–III MRSAB. However, the outcomes of SCCmec IV/IVa MRSAB did not differ from those of SCCmec I–III MRSAB.     

Molecular typing and phenotype characterization of methicillin-resistant Staphylococcus aureus isolates from blood in Taiwan

Background

Staphylococcus aureus causes a variety of severe infections such as bacteremia and sepsis. At present, 60–80% of S. aureus isolates from Taiwan are methicillin resistant (MRSA). It has been shown that certain MRSA clones circulate worldwide. The goals of this study were to identify MRSA clones in Taiwan and to correlate the molecular types of isolates with their phenotypes.

Methods

A total of 157 MRSA isolates from bacteremic patients were collected from nine medical centers. They were typed based on polymorphisms in agr, SCCmec, MLST, spa, and dru. Phenotypes characterized included Panton-Valentine leucocidin (pvl), inducible macrolide-lincosamide-streptogramin B resistance (MLSBi), vancomycin (VA) and daptomycin (DAP) minimal inhibitory concentrations (MIC), and superantigenic toxin gene profiles. Difference between two consecutive samples was determined by Mann-Whitney-U test, and difference between two categorical variables was determined by Fisher''s exact test.

Results

Four major MRSA clone complexes CC1, CC5, CC8, and CC59 were found, including 4 CC1, 9 CC5, 111 CC8, and 28 CC59 isolates. These clones had the following molecular types: CC1: SCCmecIV and ST573; CC5: SCCmecII and ST5; CC8: SCCmecIII, ST239, and ST241, and CC59: SCCmecIV, SCCmecV_T, ST59, and ST338. The toxin gene profiles of these clones were CC1: sec-seg-(sei)-sell-selm-(seln)-selo; CC5: sec-seg-sei-sell-selm-(seln)-selp-tst1; CC8: sea-selk-selq, and CC59: seb-selk-selq. Most isolates with SCCmecV_T, ST59, spat437, and dru11 types were pvl ⁺ (13 isolates), while multidrug resistance (≥4 antimicrobials) were associated with SCCmecIII, ST239, spa t037, agrI, and dru14 (119 isolates) (p<0.001). One hundred and twenty four isolates with the following molecular types had higher VA MIC: SCCmecII and SCCmecIII; ST5, ST239, and ST241; spa t002, t037, and t421; dru4, dru10, dru12, dru13, and dru14 (p<0.05). No particular molecular types were found to be associated with MLSBi phenotype.

Conclusions

Four major MRSA clone complexes were found in Taiwan. Further studies are needed to delineate the evolution of MRSA isolates.     

Emergence of Community-Associated Methicillin-Resistant Staphylococcus aureus Associated with Pediatric Infection in Cambodia

Background

The incidence of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infection is rising in the developed world but appears to be rare in developing countries. One explanation for this difference is that resource poor countries lack the diagnostic microbiology facilities necessary to detect the presence of CA-MRSA carriage and infection.

Methodology and Principal Findings

We developed diagnostic microbiology capabilities at the Angkor Hospital for Children, Siem Reap, western Cambodia in January 2006 and in the same month identified a child with severe community-acquired impetigo caused by CA-MRSA. A study was undertaken to identify and describe additional cases presenting between January 2006 and December 2007. Bacterial isolates underwent molecular characterization using multilocus sequence typing, staphylococcal cassette chromosome mec (SCCmec) typing, and PCR for the presence of the genes encoding Panton-Valentine Leukocidin (PVL). Seventeen children were identified with CA-MRSA infection, of which 11 had skin and soft tissue infection and 6 had invasive disease. The majority of cases were unrelated in time or place. Molecular characterization identified two independent MRSA clones; fifteen isolates were sequence type (ST) 834, SCCmec type IV, PVL gene-negative, and two isolates were ST 121, SCCmec type V, PVL gene-positive.

Conclusions

This represents the first ever report of MRSA in Cambodia, spread of which would pose a significant threat to public health. The finding that cases were mostly unrelated in time or place suggests that these were sporadic infections in persons who were CA-MRSA carriers or contacts of carriers, rather than arising in the context of an outbreak.     

A Novel Gene,fudoh, in the SCCmec Region Suppresses the Colony Spreading Ability and Virulence of Staphylococcus aureus

Staphylococcus aureus colonies can spread on soft agar plates. We compared colony spreading of clinically isolated methicillin-sensitive S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA). All MSSA strains showed colony spreading, but most MRSA strains (73%) carrying SCCmec type-II showed little colony spreading. Deletion of the entire SCCmec type-II region from these MRSA strains restored colony spreading. Introduction of a novel gene, fudoh, carried by SCCmec type-II into Newman strain suppressed colony spreading. MRSA strains with high spreading ability (27%) had no fudoh or a point-mutated fudoh that did not suppress colony spreading. The fudoh-transformed Newman strain had decreased exotoxin production and attenuated virulence in mice. Most community-acquired MRSA strains carried SCCmec type-IV, which does not include fudoh, and showed high colony spreading ability. These findings suggest that fudoh in the SCCmec type-II region suppresses colony spreading and exotoxin production, and is involved in S. aureus pathogenesis.     

Methicillin-Susceptible Staphylococcus aureus as a Predominantly Healthcare-Associated Pathogen: A Possible Reversal of Roles?

Background

Methicillin-resistant Staphylococcus aureus (MRSA) strains have become common causes of skin and soft tissue infections (SSTI) among previously healthy people, a role of methicillin-susceptible (MSSA) isolates before the mid-1990s. We hypothesized that, as MRSA infections became more common among S. aureus infections in the community, perhaps MSSA infections had become more important as a cause of healthcare-associated infection.

Methods

We compared patients, including children and adults, with MRSA and MSSA infections at the University of Chicago Medical Center (UCMC) from all clinical units from July 1, 2004-June 30, 2005; we also compared the genotypes of the MRSA and MSSA infecting bacterial strains.

Results

Compared with MRSA patients, MSSA patients were more likely on bivariate analysis to have bacteremia, endocarditis, or sepsis (p = 0.03), to be an adult (p = 0.005), to be in the intensive care unit (21.9% vs. 15.6%) or another inpatient unit (45.6% vs. 40.7%) at the time of culture. MRSA (346/545) and MSSA (76/114) patients did not differ significantly in the proportion classified as HA-S. aureus by the CDC CA-MRSA definition (p = 0.5). The genetic backgrounds of MRSA and MSSA multilocus sequence type (ST) 1, ST5, ST8, ST30, and ST59 comprised in combination 94.5% of MRSA isolates and 50.9% of MSSA isolates. By logistic regression, being cared for in the Emergency Department (OR 4.6, CI 1.5-14.0, p = 0.008) was associated with MRSA infection.

Conclusion

Patients with MSSA at UCMC have characteristics consistent with a health-care-associated infection more often than do patients with MRSA; a possible role reversal has occurred for MSSA and MRSA strains. Clinical MSSA and MRSA strains shared genotype backgrounds.     

Microbiological and Molecular Characterization of Staphylococcus hominis Isolates from Blood

Background

Among Coagulase-Negative Staphylococci (CoNS), Staphylococcus hominis represents the third most common organism recoverable from the blood of immunocompromised patients. The aim of this study was to characterize biofilm formation, antibiotic resistance, define the SCCmec (Staphylococcal Chromosomal Cassette mec) type, and genetic relatedness of clinical S. hominis isolates.

Methodology

S. hominis blood isolates (n = 21) were screened for biofilm formation using crystal violet staining. Methicillin resistance was evaluated using the cefoxitin disk test and the mecA gene was detected by PCR. Antibiotic resistance was determined by the broth microdilution method. Genetic relatedness was determined by pulsed-field gel electrophoresis (PFGE) and SCCmec typed by multiplex PCR using two different methodologies described for Staphylococcus aureus.

Results

Of the S. hominis isolates screened, 47.6% (10/21) were categorized as strong biofilm producers and 23.8% (5/21) as weak producers. Furthermore, 81% (17/21) of the isolates were methicillin resistant and mecA gene carriers. Resistance to ampicillin, erythromycin, and trimethoprim was observed in >70% of isolates screened. Each isolate showed a different PFGE macrorestriction pattern with similarity ranging between 0–95%. Among mecA-positive isolates, 14 (82%) harbored a non-typeable SCCmec type: eight isolates were not positive for any ccr complex; four contained the mec complex A ccrAB1 and ccrC, one isolate contained mec complex A, ccrAB4 and ccrC, and one isolate contained the mec complex A, ccrAB1, ccrAB4, and ccrC. Two isolates harbored the association: mec complex A and ccrAB1. Only one strain was typeable as SCCmec III.

Conclusions

The S. hominis isolates analyzed were variable biofilm producers had a high prevalence of methicillin resistance and resistance to other antibiotics, and high genetic diversity. The results of this study strongly suggested that S. hominis isolates harbor new SCCmec structural elements and might be reservoirs of ccrC1 in addition to ccrAB1 and mec complex A.     

Staphylococcus aureus Ocular Infection: Methicillin-Resistance, Clinical Features, and Antibiotic Susceptibilities

Background

Methicillin-resistant Staphylococcus aureus (MRSA) infection is an important public health issue. The study aimed to determine the prevalence of ocular infections caused by MRSA and to identify the clinical characteristics and antibiotic susceptibility of ocular MRSA infections by comparing those of ocular methicillin-sensitive S. aureus (MSSA) infections.

Methodology/Principal Findings

The medical records of the patients (n = 519) with culture-proven S. aureus ocular infections seen between January 1, 1999 and December 31, 2008 in Chang Gung Memorial Hospital were retrospectively reviewed. Two hundred and seventy-four patients with MRSA and 245 with MSSA ocular infections were identified. The average rate of MRSA in S. aureus infections was 52.8% and the trend was stable over the ten years (P value for trend  = 0.228). MRSA ocular infections were significantly more common among the patients with healthcare exposure (P = 0.024), but 66.1% (181/274) patients with MRSA ocular infections had no healthcare exposure. The most common clinical presentation for both MRSA and MSSA ocular infections was keratitis; MRSA and MSSA caused a similar disease spectrum except for lid infections. MRSA was significantly more resistant than MSSA to clindamycin, erythromycin and sulfamethoxazole/trimethoprim (all P<0.001).

Conclusions/significance

We demonstrated a paralleled trend of ocular MRSA infection in a highly prevalent MRSA country by hospital-based survey. Except for lid disorder, MRSA shared similar spectrum of ocular pathology with MSSA. Since S. aureus is a common ocular pathogen, our results raise clinician’s attention to the existence of highly prevalent MRSA.     

Diversity of SCCmec elements in methicillin-resistant coagulase-negative staphylococci clinical isolates

Background

Methicillin-resistant coagulase-negative staphylococci (MR-CoNS) are opportunistic pathogens and serve as a large reservoir of staphylococcal cassette chromosome mec (SCCmec). Characterization of SCCmec in MR-CoNS can generate useful information on the mobilization and evolution of this element.

Methodology/Principal Findings

Non-repetitive MR-CoNS clinical isolates (n = 84; 39 S. epidermidis, 19 S. haemolyticus, 9 S. hominis, 6 S. capitis, 4 S. warneri, 2 S. cohnii, 2 S. saprophyticus, 1 S. kloosii, 1 S. simulans and 1 S. massiliensis) were collected. All isolates could grow on plates with 4 mg/L cefoxitin and all had mecA as detected by PCR. Strain typing using RAPD and ERIC-PCR revealed that almost all isolates were of different strains. SCCmec typing was performed using multiplex PCR published previously. For isolates in which SCCmec could not be typed, the mec complex classes were determined by additional PCR and the ccr genes were amplified with published or newly-designed primers and then sequenced. SCCmec types were assigned for 63 isolates by multiplex PCR and were assigned for 14 other isolates by PCR targeting mec and ccr. Among 77 isolates with determined SCCmec types, 54 had a single type, including type III (n = 19), IV (n = 14), V (n = 10), II (n = 2), I (n = 1), VIII (n = 1) and five unnamed types (n = 7), while 23 isolates had two types, III+V (n = 12), II+V (n = 8), II+IV (n = 2) or IV+V (n = 1). The five unnamed types were assigned UT1 (class A mec, ccrA1/ccrB4), UT2 (class C1 mec, ccrA4/ccrB4), UT3 (class A mec, ccrA5/ccrB3), UT4 (class C2 mec, ccrA2/ccrB2 plus ccrC1) and UT5 (class A mec, ccrA1/ccrB1 plus ccrC1).

Conclusions/Significance

SCCmec types III, IV and V were prevalent in MR-CoNS and many isolates could harbor more than one type. Several new types of SCCmec were identified, highlighting the great genetic diversity and the need of developing classification schemes for SCCmec in MR-CoNS.     

Molecular epidemiology of methicillin-resistant Staphylococcus hominis (MRSHo): low clonality and reservoirs of SCCmec structural elements

Background

Methicillin resistant Staphylococcus hominis (MRSHo) are important human pathogens in immunocompromised patients. However, little is known regarding its population structure and staphylococcal chromosomal cassette mec (SCCmec) content.

Methodology/Principal Findings

To assess the population structure and the SCCmec content of S. hominis, 34 MRSHo and 11 methicillin-susceptible S. hominis (MSSHo) from neutropenic patients collected over a 3-year period were studied. The genetic backgrounds of S. hominis isolates were analyzed by pulsed-field gel electrophoresis (PFGE) and SCCmec types were determined by PCR. Cassette chromosome recombinases (ccr) were characterized by PCR and ccrB sequencing. The 34 S. hominis isolates were classified into as many as 28 types and 32 subtypes (SID = 99.82%); clonal dissemination was occasionally observed. The main SCCmec structures identified were SCCmec type VI (4B) (20%), SCCmec VIII (4A) (15%), and a new SCCmec composed of mec complex A in association with ccrAB1 (38%); 27% of the isolates harbored non-typeable SCCmec. Overall, a high prevalence of mec complex A (73.5%), ccrAB1 (50%) and ccrAB4 (44%) were found. Importantly, ccrB1 and ccrB4 from both MRSHo and MSSHo showed a high nucleotide sequence homology with those found in S. aureus SCCmec I, VI and VIII respectively (>95%).

Conclusions/Significance

The S. hominis population showed a limited clonality and a low genetic diversity in the allotypes of ccr and classes of mec complex. Moreover, our data suggest that S. hominis might have been a privileged source of mec complex A, ccrB1 and ccrB4, for the assembly of primordial SCCmec types.     

Gene acquisition at the insertion site for SCCmec, the genomic island conferring methicillin resistance in Staphylococcus aureus

Staphylococcus aureus becomes resistant to methicillin by acquiring a genomic island, known as staphylococcal chromosome cassette mec (SCCmec), which contains the methicillin resistance determinant, mecA. SCCmec is site-specifically integrated into the staphylococcal chromosome at a locus known as the SCCmec attachment site (attB). In an effort to gain a better understanding of the potential that methicillin-sensitive S. aureus (MSSA) isolates have for acquiring SCCmec, the nucleotide sequences of attB and surrounding DNA regions were examined in a diverse collection of 42 MSSA isolates. The chromosomal region surrounding attB varied among the isolates studied and appears to be a common insertion point for acquired foreign DNA. Insertions of up to 15.1 kb were found containing open reading frames with homology to enterotoxin genes, restriction-modification systems, transposases, and several sequences that have not been previously described in staphylococci. Two groups, containing eight and four isolates, had sequences found in known SCCmec elements, suggesting SCCmec elements may have evolved through repeated DNA insertions at this locus. In addition, the attB sequences of the majority of MSSA isolates in this collection differ from the attB sequences of strains for which integrase-mediated SCCmec insertion or excision has been demonstrated, suggesting that some S. aureus isolates may lack the ability to site-specifically integrate SCCmec into their chromosomes.     

Nasal Carriage of Methicillin-Resistant Staphylococcus aureus among Pediatricians in Taiwan

Background

Health care workers (HCWs) are at the interface between hospitals and communities. The survey for methicillin-resistant Staphylococcus aureus (MRSA) carriage among HCWs has mostly been conducted to investigate outbreaks or endemics. Community-associated MRSA are prevalent among children in Taiwan. We conducted this study to better understand the carriage rate of MRSA among pediatricians in non-outbreak situations in Taiwan,.

Methods

A total of 220 pediatricians from Taiwan who attended the annual meeting of Taiwan Pediatric Association in April, 2010 were recruited to participate in this study and were sampled from the nares for the detection of MRSA by polymerase chain reaction (PCR) and further by culture. The following molecular analyses were performed, including pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), typing of staphylococcal cassette chromosome mec (SCCmec) and the presence of Panton-Valentine leukocidin (PVL) genes.

Results

MRSA was detected from 15 attendees (6.8%) by PCR. MRSA-colonized attendees had a significantly lower rate (0.041) of working in the medical center, while borderline significantly higher rate of working in the Regional Hospital (p=0.056), than those without MRSA colonization. From those 15 samples, 12 MRSA isolates were identified by culture and molecularly characterized. Three PFGE patterns, two sequence types (ST 59, ST 508), and two SCCmec types (IV and V_T) were identified, respectively. Five isolates, including three carrying SCCmec types V_T, were PVL-positive. All 12 isolates were susceptible to vancomycin, teicoplanin, linezolid, fusidic acid, trimethoprim/sulfamethoxazole, and doxycyclin, and resistant to penicillin.

Conclusion/significance

Around seven percent of pediatricians in Taiwan harbored CA-MRSA in their nares.     

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.