Emergence of virulent methicillin-resistant Staphylococcus aureus strains carrying Panton-Valentine leucocidin genes in The Netherlands

Methicillin-resistant Staphylococcus aureus (MRSA) strains carrying the Panton-Valentine leucocidin (PVL) genes have been reported worldwide and are a serious threat to public health. The PVL genes encode a highly potent toxin which is involved in severe skin infections and necrotizing pneumonia, even in previously healthy individuals. We assessed the prevalence of PVL-positive MRSA in The Netherlands for two periods of time: (i) 1987 through 1995 and (ii) 2000 and 2002, and determined their characteristics by using multilocus sequence typing and staphylococcal chromosome cassette (SCCmec) typing. It was found that up to 15% of all MRSA isolates detected in The Netherlands harbored the PVL genes. Most PVL-positive MRSA isolates were obtained from severe soft tissue infections in relatively young individuals. The first PVL-positive MRSA described in The Netherlands, isolated in 1988, was a single-locus variant of the "Berlin" epidemic MRSA clone. The 20 PVL-positive MRSA isolates studied in 2000 and 2002 consisted of five different sequence types (STs) that belonged to four clonal complexes. One of the STs, ST80, is considered to be a widespread European clone and was the most predominant ST (60%) in this study, while ST37 had never been found to be associated with PVL-positive MRSA. Most isolates harbored SCCmec type IV, a supposed marker for community-acquired MRSA. The number and type of virulence-associated genes varied among the different STs.     

High percentage of methicillin-resistant Staphylococcus aureus isolates with reduced susceptibility to glycopeptides in The Netherlands

While testing the in vitro activities of 14 antimicrobial agents against 107 methicillin-susceptible Staphylococcus aureus (MSSA) and 250 methicillin-resistant S. aureus (MRSA) isolates collected in The Netherlands, we found to our surprise that 19 (7.6%) MRSA isolates were suspected of having reduced susceptibilities to the glycopeptides when the Etest system (AB Biodisk, Solna, Sweden) was used with a large inoculum (no. 2 McFarland standard) and an extended incubation time (48 h) on brain heart infusion agar for MIC testing. Eventually, 15 of these isolates were classified as heterogeneously resistant to glycopeptides (heterogeneously glycopeptide-intermediate S. aureus [hGISA] isolates) according to the population analysis profile-area under the curve analysis. The MICs at which 50 and 90% of isolates are inhibited obtained with the Etest system with the large inoculum were as follows: for MSSA isolates, 3.0 and 4.0 micro g/ml, respectively, for both teicoplanin and vancomycin; for MRSA isolates, 3.0 and 8.0 micro g/ml, respectively, for teicoplanin, and 3.0 and 4.0 micro g/ml, respectively, for vancomycin. This is the first report of hGISA isolates in The Netherlands.     

The molecular evolution of methicillin-resistant Staphylococcus aureus

Staphylococcus aureus is a potentially pathogenic bacterium that causes a broad spectrum of diseases. S. aureus can adapt rapidly to the selective pressure of antibiotics, and this has resulted in the emergence and spread of methicillin-resistant S. aureus (MRSA). Resistance to methicillin and other β-lactam antibiotics is caused by the mecA gene, which is situated on a mobile genetic element, the Staphylococcal Cassette Chromosome mec (SCC mec ). To date, five SCC mec types (I–V) have been distinguished, and several variants of these SCC mec types have been described. All SCC mec elements carry genes for resistance to β-lactam antibiotics, as well as genes for the regulation of expression of mecA . Additionally, SCC mec types II and III carry non-β-lactam antibiotic resistance genes on integrated plasmids and a transposon. The epidemiology of MRSA has been investigated by pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), spa typing and SCC mec typing. Numerous MRSA clones have emerged and disseminated worldwide. SCC mec has been acquired on at least 20 occasions by different lineages of methicillin-sensitive S. aureus . Although most MRSA strains are hospital-acquired (HA-MRSA), community-acquired MRSA (CA-MRSA) strains have now been recognised. CA-MRSA is both phenotypically and genotypically different from HA-MRSA. CA-MRSA harbours SCC mec types IV or V, and is associated with the genes encoding Panton–Valentine leukocidin. The prevalence of MRSA ranges from 0.6% in The Netherlands to 66.8% in Japan. This review describes the latest developments in knowledge concerning the structure of SCC mec , the molecular evolution of MRSA, the methods used to investigate the epidemiology of MRSA, and the risk-factors associated with CA-MRSA and HA-MRSA.     

Gentamicin resistance in methicillin-resistant Staphylococcus aureus

Gentamicin resistance has been studied in methicillin-resistant Staphylococcus aureus (MRSA) strains, from Royal Melbourne Hospital (RMH) and Sydney. Gentamicin resistance was transferred in mixed cultures to a plasmid free strain, and the determinants were examined. The Sydney strain had high level resistance to gentamicin, tobramycin, kanamycin and neomycin which was carried on a c.34 megadalton plasmid. The gentamicin resistant RMH isolates all had a determinant which conferred low level resistance to gentamicin, tobramycin and kanamycin and appeared to be chromosomal in one isolate, on a plasmid of c.28.5 megadaltons in another and on a plasmid of c.18 megadaltons in the other isolates. It is suggested that a gentamicin resistance transposon is being transferred in the MRSA at RMH.     

Strain differentiation in methicillin-resistant Staphylococcus aureus

Three different systems were used to test 236 isolates of methicillin-resistant Staphylococcus aureus in an attempt to ascertain if more than one strain is responsible for the current problem of cross-infection by this organism in N.S.W. hospitals. The biochemical tests used were of little assistance. Phage typing, using the Basic International Set of typing phages at 100 x routine test dilution (RTD), provided evidence of the presence of several different strains. Phage type 83A/85/95/90/88 was the typing pattern of the predominant strain and the nest most frequent group was not typable. These results were often difficult to read. Five new phages were therefore isolated and found to be valuable as they produced easily identifiable patterns at RTD.     

The changing face of community-acquired methicillin-resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (MRSA) is an important cause of infection, both in hospitalised patients with significant healthcare exposure and in patients without healthcare risk factors. Community-acquired methicillin-resistant S. aureus (CA-MRSA) are known for their rapid community transmission and propensity to cause aggressive skin and soft tissue infections and community-acquired pneumonia. The distinction between the healthcare-associated (HA)-MRSA and CA-MRSA is gradually fading owing to the acquisition of multiple virulence factors and genetic elements. The movement of CA-MRSA strains into the nosocomial setting limits the utility of using clinical risk factors alone to designate community or HA status. Identification of unique genetic characteristics and genotyping are valuable tools for MRSA epidemiological studies. Although the optimum pharmacotherapy for CA-MRSA infections has not been determined, many CA-MRSA strains remain broadly susceptible to several non-β-lactam antibacterial agents. This review aimed at illuminating the characteristic features of CA-MRSA, virulence factors, changing clinical settings and molecular epidemiology, insurgence into the hospital settings and therapy with drug resistance.     

婴幼儿耐甲氧西林金黄色葡萄球菌感染状况和耐药基因的研究

目的了解感染婴幼儿的耐甲氧西林金黄色葡萄球菌（MRSA）耐药性和耐药基因,明确检测青霉素结合蛋白2a（PBP2a）和甲氧西林耐药基因（mecA）的临床价值。方法用金黄色葡萄球菌乳胶凝集试验和梅里埃鉴定金黄色葡萄球菌,同时用纸片扩散法完成12种常用抗生素的药敏试验。对头孢西丁的结果,按当年CLSI标准执行,用于判断菌株甲氧西林的耐药性。采用PBP2检测试剂盒检测PBP2a蛋白,PCR检测mecA基因。结果 2007-2009年婴幼儿共检出245株金黄色葡萄球菌,其中MRSA检出率为17.6%（43/245）。MRSA对庆大霉素、复方新诺明、克林霉素、红霉素、氯霉素的耐药性均显著高于甲氧西林敏感金黄色葡萄球菌（P〈0.05）,其余抗生素的差异无统计学意义（P〉0.05）,43株MRSA的PBP2a蛋白和mecA基因的检测结果全为阳性,阳性率为100%。结论用头孢西丁（30μg）能有效地筛查MRSA,检测PBP2a蛋白和mecA基因均能正确地确认MRSA。且检测PBP2a蛋白方便快捷,特异性好,值得临床推广。     

Phenotypic prediction rule for community-associated methicillin-resistant Staphylococcus aureus

Recent studies have suggested that community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections are encroaching upon nosocomial settings. We assessed the performance characteristics of a rule using the antimicrobial phenotype to predict genotype. This rule could be applied for epidemiologic purposes to describe the trend in CA-MRSA infections over time.     

Mechanism of beta-lactam-resistance in methicillin-resistant Staphylococcus aureus

The role of penicillin-binding protein (PBP) 2' in the expression of beta-lactam-resistance was investigated using methicillin-resistant Staphylococcus aureus (MRSA) strains with different level of resistance. Both high- and moderate-level MRSA produced very similar PBP 2' with low affinities for beta-lactam antibiotics. Affinities of antibiotics for PBP 2' (I50, concentration which inhibits [14C] benzylpenicillin-binding by 50%) correlated well with their antibacterial activities (MIC) in a high-level MRSA, but did not in a moderate-level MRSA. High-level MRSA contained a larger amount of PBP 2' than moderate-level MRSA, and the amount of PBP 2' decreased by increasing the temperature of the culture; the extent of decrease was larger in a strain which was sensitive at 37 degrees C than a strain which exerted relatively high level resistance even at 40 degrees C. A cephamycin-resistant, methicillin-sensitive strain began to synthesize PBP 2' by adding cephamycin-type antibiotics to the medium and consequently acquired resistance to methicillin. Latent MRSA producing no PBP 2' generated clones which produced PBP 2' constitutively and were highly resistant to all beta-lactams. These results suggest that the presence of PBP 2' is critical for the expression of beta-lactam-resistance in MRSA and the degree of the resistance depends mainly on the amount of PBP 2' which differs from strain to strain and is influenced by environments such as temperature and the presence of inducer.     

Nosocomial infection with methicillin-resistant Staphylococcus aureus

Methicillin-resistant Staphylococcus aureus (MRSA) isolated in our hospital between 1986 and 1989 were mainly examined for their susceptibility to various antibiotics and coagulase types. The isolation frequency of MRSA among S. aureus isolated from clinical specimens has been steadily increasing; 37.8% in 1986, 49.8% in 1987, 60.6% in 1988 and 63.2% in 1989. This trend was particularly noticeable in the specimens associated with the respiratory tracts. The isolation rate of MRSA in the surgery and pediatrics wards was higher than that in the internal medicine ward. More than 80% of MRSA were coagulase type II, which were multi-resistant to penicillins, cephems, aminoglycosides and macrolides, and sensitive to MINO, new quinolones and VCM. These epidemic strains were also isolated from the nose of medical staff and from air samples in the wards. These findings suggest that the hospital environment including the patients and hospital personnel is extensively contaminated with multi-resistant MRSA of coagulase type II. Measures should be taken for prevention and control of nosocomial infection with MRSA in the whole hospital.     

Molecular genetics of methicillin-resistant Staphylococcus aureus

A large and growing proportion of Staphylococcus aureus clinical isolates are methicillin resistant, and are resistant to practically all beta-lactam antibiotics. Methicillin-resistant S. aureus (MRSA) strains harbor mecA, which is carried by a unique mobile genetic element, staphylococcal cassette chromosome mec (SCCmec) integrated into the S. aureus chromosome. The mecA gene encodes a methicillin-insensitive transpeptidase, the production of which confers resistance to otherwise inhibitory concentrations of beta-lactam antibiotics. Several distinct clones have been identified among MRSA that apparently have been generated by integration of distinct types of SCCmec. While MRSA are primarily nosocomial pathogens, recent observations indicate that other MRSA clones are colonizing a significant proportion of healthy individuals in the community as well. Community-acquired MRSA (C-MRSA), may become a new threat to humans, and international cooperation of researchers and clinicians will be of cardinal importance in addressing this problem.     

Community-acquired methicillin-resistant Staphylococcus aureus in Taiwan.

Staphylococcus aureus is a major cause of infections in both hospitals and communities, and is exhibiting increasing resistance to methicillin (methicillin-resistant S. aureus, MRSA) and related beta-lactams. MRSA is usually considered a nosocomial pathogen, but increasingly it is acquired in the community. In Taiwan, MRSA was colonized in a substantial proportion of healthy children and accounted for 25% to 75% of childhood community-acquired (CA) S. aureus infections. From the preliminary data, the isolates of sequence type (ST) 59 by multilocus sequence typing method appeared to be the major clone of CA-MRSA in northern Taiwan. Compared with those reported from the US and other countries, CA-MRSA isolates in Taiwan did not always harbor type IV staphylococcal cassette chromosome (SCCmec) and were resistant to multiple non-beta-lactam antibiotics, including clindamycin and macrolides. Molecular evidence suggested transmission of the community strain of MRSA into the hospital setting, and that the community strain had became a health care-associated pathogen. The treatment of putative CA S. aureus infection should be stratified according to the severity and the disease entity.     

Lectin typing of methicillin-resistant Staphylococcus aureus

This study reports the patterns of agglutination of 77 clinical isolates of methicillin-resistant Staphylococcus aureus by 32 commercially available lectins. Cell suspensions were not pre-treated. Each isolate was cultured on three media: Columbia blood agar, trypticase-soy agar and Chapman Stone agar. The lectins agglutinating each isolate varied widely depending on culture medium; only five isolates were agglutinated by the same set of lectins regardless of the culture medium used. Lectin typing could be a useful epidemiological tool, but it is necessary to standardise assay conditions (notably culture medium) to enable meaningful comparison of the results produced by different research groups or centres.     

Management of methicillin-resistant Staphylococcus aureus infections

This review addresses selected aspects of the management of severe healthcare-associated infections due to methicillin-resistant Staphylococcus aureus (MRSA), including the limitations of current therapy, potential alternative agents, new therapeutic options, clinical approaches to MRSA bacteraemia/endocarditis and ventilator-associated pneumonia, and strategies to improve outcomes in patients with severe MRSA infections.