| Staphylococcus aureus is not only a pathogen of nosocomial infection, but also acommon cause of skin and soft tissue infections in the community. In the pre-antibiotic era,the consequences of infections caused by S. aureus were very dangerous. In1941, the fatalityrate in Portuguese cases suffered by S. aureus was high as82%. Since1940s, the powerfulpenicillin has been developed and applied in clinic, and the infections caused by S. aureuswere significantly decreased. With the widespread use of penicillin, some S. aureus strainsthat produce penicillinase, a hydrolysis enzyme to degrade the beta-lactam ring of penicillin,were emerged. To combat penicillinase, a semi-synthetic penicillin termed methicillin(methicilhn) was introduced to clinic in1959. However, only two years later (1961), Jevonsreported the discovery of the first case of methicillin-resistant S. aureus (MRSA) infection inthe United Kingdom. From1970s, MRSA has been spread to worldwide and become a majorglobal pathogen in the late1980s, resulting in increased morbidity and mortality, increasedhealth care costs, brought serious economic losses and adverse effects to not only hospitals,but also the communities. At present, MRSA has been ranked the first epidemic pathogen forthe hospital infections, and the morbidity was up to80%in the United States. Every year, it isestimated that about100,000people were hospitalized due to MRSA infection in the UnitedStates.MRSA that emerged in hospitals were designed as hospital-acquired MRSA(HA-MRSA),which usually affect elders or patients with more serious illness, skin wounds (such asbedsores),etc. However, recent studies showed that the proportion of infections caused by newtypes of MRSA in the community was increasing. This newly popular MRSA was firstdiscovered in New York in1997, occured in the environment of the community, and often served as risk factors for health people. The community-onset MRSA, also known ascommunity-acquired MRSA (CA-MRSA) is distinct from HA-MRSA and refers to the MRSAstrains isolated from outpatients or hospitalized patients within48-72hours admission. Todistinguish CA-MRSA and HA-MRSR based on simple clinical documonents of patients maybe not accurate.More and more scientists would like to identify CA-MRSA and HA-MRSAby molecular biology data of the isolates. Usually, SCCmec types I, II and III mobile elementsare carried by HA-MRSA isolates. These SCCmec elements are relatively larger (35~70kb)and carry multiple resistance genes (likly to be multidrug resistant strains). SCCmec types IVand V are more likely to spread in CA-MRSA strains and relatively shorter (20~40kb) withfewer other resistance genes in addition to mecA.One way to control the spread of MRSA is by determining the genotypic characteristicsof MRSA clones and the genetic relatedness of strains in different geographic regions.Severalmolecular methods have been developed to type MRSA isolates, including pulsed-field gelelectrophoresis (PFGE), multilocus sequence typing (MLST), and typing based on uniquefragments of the genetic loci SCCmec and the polymorphisms of X region encodingstaphylococcal protein A (spa). With these typing methods, several MRSA clones have beenidentified worldwide, including New York/Japan (ST5-SCCmecII), Hungarian (ST239-SCCmecIII) and USA300(ST8-SCCmecIV) clones. MRSA isolates also harbour numerousdeterminants associated with antibiotic resistance. The association of MRSA antimicrobialresistance profiles (ARPs) with their molecular characteristics can provide useful informationfor the clinical treatment of MRSA infections.MRSA was originally detected in China in the1970s, and the proportion of MRSAamong S. aureus isolates increased to approximately20%after1980, which continued toincrease at this level until2000. In the21stcentury, MRSA prevalence rapidly increased,comprising60%of S. aureus in hospital isolates in2008. Two major clones,ST239-MRSA-SCCmec type III and ST5-MRSA-SCCmec type II, were shown to beprevalent in China during2005and2006. To determine the dynamic status of predominantMRSA types in China,517S. aureus isolates, with309MRSA isolates collected, from9teaching hospitals in6cities were subjected to antimicrobial susceptibility testing andmolecular typing in this study. The correlation between the MRSA molecular types and theirantibiotic resistance phenotypes was also investigated. The main results are as following. 1. MRSA detection. To confirm the MRSA from S. aureus isolates, mecA and femB genesof the517clinical strains were respectively detected by multiplex polymerase chain reaction(PCR). The result showed that59.8%(309/517) of the collected S. aureus isolates wereMRSA.2. Molecular Typing. i) SCCmec typing. SCCmec typing of309MRSA isolates wasperformed using four unique and specific pairs of primers for SCCmec types I, II, III, IV and V.The results demonstrated that the most common SCCmec type was type III, which was present in178isolates (57.6%,178/309). SCCmec type II was the second most predominant type,comprising22.0%(68/309) of the isolates. The third predominant type was SCCmec type IV,19of309(6.2%) isolates belonged to SCCmec type I,13isolates (4.2%) belonged to SCCmec typeV and four isolates (4/309;1.3%) were defined as NT (nontypable). ii) spa typing. The X regionof each MRSA isolate was amplified by PCR. The amplified products were sequenced andanalysed based on the spa database Web site (http://www.ridom.de/spaserver), and each isolatewas assigned a specific spa type. The309MRSA bacteria can be divided into31spa genotypes,with the dominant type for type t030(90,29.1%), type t002(49,15.9%) and t037(48,15.5%).iii)MLST. MRSA isolates were subjected to MLST analysis by PCR amplification andsequencing of the seven housekeeping genes (arc, aroE, glpF, gmk, pta, tpi and yqiL). Allelicprofiles and sequence types (STs) were assigned according to the MLST Web site(http://saureus.mlst.net/). Fifteen STs and4new STs were found. ST239was the mostpredominant ST (51.5%,159/309), ST5was determined to be the second most common ST(25.2%,78/309), and ST59comprised10.7%of the tested isolates and was the third mostcommon ST. iv) PFGE. The SmaI-digested genomic DNA of each MRSA isolate wasseparated by PFGE. The PFGE patterns were analysed with BioNumerics version6.6(Applied Maths, Belgium) according to the unweighted pair-group matching analysisclustering algorithm. The309MRSA is divided into23cluster groups, named A-W, including17major cluster groups,6small cluster groups and37sporadic strains. Type E, type I, types Land M were the major types, which accounted for9.7%(30/309),10.0%(31/309),12.9%(40/309) and7.8%(24/309),respectively. v) PVL detection. A PCR for pvl genes wasperformed. The presence of lukF-PV and lukS-PV genes encoding the components of the PVLtoxin was positive in24.6%(76/309) isolates.3. Antimicrobial susceptibility testing. Antibiogram testing of the MRSA isolates was performed by Vitek microbial analysis instrument, according to the guidelines set by Clinicaland Laboratory Standards Institute (CLSI). Exclusion of intermediate resistance,97.6%of theMRSA isolates were resistant to oxacillin,100%to penicillin,99.4%to ampicillin,77.3%togentamicin,80.2%to ciprofloxacin,80.9%to clindamycin,86.2%to erythromycin and81.7%to levofloxacin. The results also showed that74.8%(231/309) of the MRSA were multidrugresistant (MDR) isolatesIn conclusion, among the isolates collected, three major clones were found prevalent inChina, ST239-MRSA-III-t030, ST239-MRSA-III-t037and ST5-MRSA-II-t002. These threeclones were associated with two characteristic resistance profiles, namely, gentamicin/ciprofloxacin/rifampicin/levofloxacin for the first clone and gentamicin/ciprofloxacin/clindamycin/erythromycin/tetracycline/levofloxacin/bactrim for the latter two. Severalgeographically unique minor (such as ST59-MRSA-IV-t437) clones were also identified. |
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