Evolution, Molecular ; Methicillin Resistance ; Staphylococcus aureus ; classification ; drug effects ; genetics ; isolation & purification

Child ; Cross Infection ; microbiology ; Disease Outbreaks ; Humans ; Methicillin-Resistant Staphylococcus aureus ; classification ; drug effects ; genetics

OBJECTIVETo detect the enterotoxin genes of Staphylococcus aureus (SA) isolated from clinical specimens and analyze the correlation between enterotoxin genes and drug resistance of SA.

METHODSThe mecA gene and enterotoxin genes A-F of clinical SA isolates were identified by polymerase chain reaction (PCR), and the genes were sequenced to investigate the correlation of these genes to drug resistance.

RESULTSThe detection rate of enterotoxin genes was 100% in 67 methicillin- resistant SA (MRSA), showing no significant difference from the rate in 57 methicillin-sensitive SA (MSSA) (83.5%, χ(2)=0.203, P>0.05). Of the 116 strains carrying enterotoxin genes (93.5%), the detection rates of SEA, SEB, SEC, SED and SEF were 90.5%, 6.9%, 61.3%, 5.2%, 25.9% and 93.5%, respectively, and none of the strains were positive for SEE gene. In these strains, 78 (67.2%) carried 2 or more enterotoxin genes, and the main genotypes were SEA and SEC (33.6%), SEA and SEF (7.8%), and SEA and SEC and SEF (13.8%). Compared with the strains carrying a single enterotoxin gene, those with multiple enterotoxin genes showed a higher drug resistance rate, among which 75% of the SA strains carrying SEA+SEC+SEF were resistant to SXT, significantly higher than the rates of SA carrying SEA (28.6%) and SEA+SEC (38.7%) (P<0.05). The SA strains carrying SEA+SEC+SEF and SEA+SEF showed significantly higher amikacin resistance rates than SA strain carrying SEA (75.0%, 77.0%, 21.5%, respectively, P<0.05).

CONCLUSIONClinical isolates of SA carrying multiple enterotoxin genes have a higher drug resistance rate than those with a single enterotoxin gene, suggesting the the important role of enterotoxin in multidrug resistance.

Drug Resistance, Multiple, Bacterial ; genetics ; Enterotoxins ; genetics ; Humans ; Staphylococcal Infections ; microbiology ; Staphylococcus aureus ; drug effects ; genetics ; isolation & purification

OBJECTIVETo study the antibiotic- and disinfectant-resistance features of and disinfectant-resistant gene distribution in Staphylococcus aureus (Sa) isolated from the urogenital tract of male patients with urogenital tract infection (UTI). total of 152 Sa isolates were collected from the urethral discharge specimens from male UTI patients. The minimum inhibition concentration (MIC) of antimicrobial agents and disinfectants commonly used against Sa were tested by standard ager dilution; the methicillin-resistant Sa (MRSA) isolates detected by cefoxitin disk diffusion and mecA gene amplification; Staphylococcal cassette chromosome mec (SCCmec) genotyping performed by multiplex PCR; the disinfectants gene qac (quaternary ammonium compound) amplified by PCR; and the clonal relatedness of qacA/B-positive MRSA isolates investigated by pulsed-field gel electrophoresis (PFGE).

RESULTSOut of the 152 Sa isolates, 91 (59.9%) were found to be MRSA. SCCmec genotyping showed SCCmec V to be the main type, accounting for 63.7% (58/91), with 8 (8.8%) isolates of SCCmec I, 2 (2.2%) isolates of SCCmec II, 19 (20.9%) isolates of SCCmec III, and 4 (4. 4%) isolates of SCCmec IV. The Sa isolates exhibited high rates of non-susceptibility to penicillin (95.4%) , erythromycin (72.4% ) , ciprofloxacin (42. 8%), and levofloxacin (44.7%), and a fairly high sensitivity to nitrofurantoin, teicoplanin, linezolid, and vancomycin. The MIC in the Sa isolates was 0. 25 -16 microg/ml for chlorhexidine; MIC50 and MIC90 were 2.0 and 4.0 microg/ml respectively for MRSA strains and both 1.0 microg/ml for MSSA strains. Out of the 152 Sa isolates, 72 (47.4%) harbored the qacA/B gene, 6 (3.9%) the smar (qacC + qacD) gene, 9 (5.9%) the qacE delta 1 gene, and 2 (1.3%) the qacH gene, but no qacG and qacJ genes were detected. PFGE analysis showed that the qacA/B-positive MRSA isolates were distributed

CONCLUSIONClinical Sa isolates exhibited varied degrees of resistance to commonly used antibiotics, and in a polyclonal manner. some showed a robust tolerance to chlorhexidine. The main disinfectant-resistant gene is qacA/B. Antimicrobial agents and disinfectants should be used rationally according to clinicians.

Disinfectants ; pharmacology ; Drug Resistance, Bacterial ; genetics ; Genotype ; Humans ; Male ; Staphylococcus aureus ; drug effects ; genetics ; Urinary Tract Infections ; microbiology

Anti-Bacterial Agents ; pharmacology ; Humans ; Methicillin ; pharmacology ; Methicillin Resistance ; genetics ; Sequence Homology ; Staphylococcus aureus ; drug effects ; genetics

OBJECTIVETo analyze multilocus sequence typing (MLST) of methicillin-resistant Staphylococcus aureus (MRSA) strains in 2000 and 2005, and get a primary knowledge of MLST Characterization of MRSA.

METHODSSequence analysis was conducted on seven allelic genes of 29 methicillin-resistant Staphylococcus aureus strains and 2 methicillin-sensitive Staphylococcus aureus (MSSA) strains and the allelic profiles were gained from internet database.

RESULTSAll 12 MRSA strains in 2000 were sequence type (ST) 239 and 10 MRSA strains in 2005 were ST239, while 7 MRSA strains in 2005 were new types, ST5 (41.18%, 7/17). ST6 and ST630 were allelic profiles of 2 MSSA strains. ST239 was the most prevalent allelic profile (75.86%, 22/29), while ST5 was the second prevalent allelic profile (24.14%, 7/29) among all isolates.

CONCLUSIONST239 and ST5 are the most prevalent MRSA clones in this research. MRSA strains have different allelic profile from MSSA strains. MLST might provide an unambiguous method for assigning MRSA and MSSA isolates to known clones or assigning them as novel clones via the internet. Further studies need to be taken by increasing strains.

Bacterial Typing Techniques ; Base Sequence ; DNA, Bacterial ; genetics ; Genes, Bacterial ; Methicillin-Resistant Staphylococcus aureus ; classification ; drug effects ; genetics

BACKGROUNDIncreasing prevalence of Staphylococcus aureus (S. aureus), particularly methicillin-resistant S. aureus (MRSA) has been reported in China. In this study, we investigated the drug resistance characteristic, genetic background, and molecular epidemiological characteristic of S. aureus in Changsha.

METHODSBetween January 2006 and December 2008, 293 clinical isolates of S. aureus were collected from 11 hospitals in Changsha and identified by the Vitek-2 system. All the isolates were verified as MRSA by PCR amplification of both femA and mecA genes. K-B disk method was used to test drug sensitivity of S. aureus to antibiotics. Pulsed-field gel electrophoresis (PFGE) was performed for genotypic and homologous analysis of 115 isolates randomly selected from the original 293 clinical S. aureus isolates.

RESULTSS. aureus was highly resistant to penicillin, ampicillin, erythromycin, and clindamycin with resistant rates of 96.6%, 96.6%, 77.1%, and 67.2% respectively. All the isolates were susceptible to tecoplanin, vancomycin, and linezolid. MRSA accounted for 64.8% (190/293) of all the S. aureus strains. The 115 S. aureus isolates were clustered into 39 PFGE types by PFGE typing, with 13 predominant patterns (designated types A to M) accounting for 89 isolates. The most prevalent PFGE type was type A (n = 56, 48.7%) and 100.0% of type A strains were MRSA. PFGE type A included 13 subtypes, and the most prevalent subtype was subtype A1 (46.4%, 26/56). Strains with PFGE type A were isolated from eight hospitals (8/11), and both subtypes A1 and A4 strains were isolated in a university hospital.

CONCLUSIONSClinical isolates of S. aureus in Changsha were resistant to multiple traditional antibiotics. There was an outbreak of PFGE type A MRSA in this area and the A1 subtype was the predominant epidemic clone. Dissemination of the same clone was an important reason for the wide spread of MRSA.

Ampicillin ; pharmacology ; Anti-Bacterial Agents ; pharmacology ; China ; Clindamycin ; pharmacology ; Electrophoresis, Gel, Pulsed-Field ; Erythromycin ; pharmacology ; Humans ; Methicillin-Resistant Staphylococcus aureus ; drug effects ; genetics ; Microbial Sensitivity Tests ; Penicillins ; pharmacology ; Staphylococcus aureus ; drug effects ; genetics ; Vancomycin ; metabolism

BACKGROUND: Susceptibility testing of Staphylococcus aureus often requires cumbersome supplementary tests. MicroScan Pos Breakpoint Combo Panel Type 28 (PBC28) (Siemens, USA) includes cefoxitin screening to detect methicillin-resistant Staphylococcus aureus (MRSA), inducible clindamycin resistance detection (ICD), and determination of low-range minimum inhibitory concentration of vancomycin (0.5-16 microgram/mL). The purpose of this study was to evaluate the performance of PBC28 in comparison with that of Pos Combo Type 1A (PC1A) (Siemens). METHODS: From December 2009 to March 2010, 500 non-duplicate clinical isolates of S. aureus were tested with PC1A and PBC28. Categorical agreements (CA) between the interpretations of the 2 panels were estimated. The presence of the mecA gene was determined by PCR, and double-disk diffusion test (D-test) was performed on the isolates resistant to erythromycin but susceptible or intermediately resistant to clindamycin. Ninety-six isolates representing various vancomycin minimum inhibitory concentrations (MICs) were tested in parallel with repeat PBC28, broth macrodilution, and epsilometer test (E test). RESULTS: The CA was 99.3% with a very major error (VME) of 0.2%, major error (ME) of 0.1%, and minor error (mE) of 0.4% in total. PBC28 showed 100% CA for 1 isolate with vancomycin MIC of 4 microgram/mL and 35 isolates (7.0%) with MIC of 2 microgram/mL. However, only 15, 27, and 35 isolates with vancomycin MIC of 2 microgram/mL showed 100% CA in repeat PBC28, broth macrodilution, and E test, respectively. PC1A and PBC28 detected all 314 mecA-positive isolates. Among the 63 isolates tested with the D-test, 58 (92.1%) were positive, and the results were 100% concordant with those of ICD. CONCLUSIONS: PBC28 can be appropriate susceptibility testing of S. aureus, including MRSA detection and ICD. However, the lower-range vancomycin MIC test was not reproducible enough to reliably differentiate MIC of 2 microgram/mL from MIC< or =1 microgram/mL.
Anti-Bacterial Agents/*pharmacology ; Bacterial Proteins/genetics ; Cefoxitin/*pharmacology ; Clindamycin/*pharmacology ; Drug Resistance, Bacterial ; Methicillin-Resistant Staphylococcus aureus/genetics/isolation & purification ; *Microbial Sensitivity Tests ; Reagent Kits, Diagnostic ; Sensitivity and Specificity ; Staphylococcus aureus/*drug effects/genetics/isolation & purification ; Vancomycin/*pharmacology

The timely detection of blood-borne pathogens is one of the most important functions of the microbiology laboratory. Recently, methicillin-resistant staphylococci have become the most important pathogens seen by the laboratory. The purpose of this study was to evaluate Staphy agar, a novel screening medium, for the detection methicillin-resistant Staphylococcus aureus, S. epidermidis, or other coagulase-negative staphylococci (CNS) from positive blood cultures showing Gram-positive cocci in clusters. Eighty-six blood cultures that yielded Gram-positive cocci in clusters were included in this study. The organisms were finally identified by the Vitek system, and oxacillin resistance was confirmed by polymerase chain reaction (PCR)-based mecA gene detection. The identification and oxacillin resistance of all S. aureus strains showed complete agreement with the Vitek and PCR results. The presumptive detection of S. epidermidis and other CNS were consistent with the Vitek system in 94.7%, and the screening of oxacillin resistance was consistent with the result of PCR in 92.1% of 38 strains. The Staphy agar method is reliable and rapid for differentiating Gram-positive cocci in clusters in blood and for determining their methicillin resistance.
*Bacterial Proteins ; Carrier Proteins/genetics ; Drug Resistance, Microbial ; *Hexosyltransferases ; Muramoylpentapeptide Carboxypeptidase/genetics ; Oxacillin/*pharmacology ; Penicillin-Binding Proteins ; *Peptidyl Transferases ; Staphylococcus aureus/*drug effects/genetics ; Staphylococcus epidermidis/*drug effects/genetics

Emergence and spread of low-level mupirocin resistance in staphylococci have been increasingly reported in recent years. The aim of this study was to characterize missense mutations within the chromosomal isoleucyl-tRNA synthetase gene (ileS) among clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) with low-level mupirocin resistance. A total of 20 isolates of MRSA with low-level mupirocin resistance (minimal inhibitory concentration, 16-64 microgram/mL) were collected from 79 patients in intensive care units for six months. The isolates were analyzed for isoleucyl-tRNA synthetase (IleS) mutations that might affect the binding of mupirocin to the three-dimensional structure of the S. aureus IleS enzyme. All isolates with low-level mupirocin resistance contained the known V588F mutation affecting the Rossman fold, and some of them additionally had previously unidentified mutations such as P187F, K226T, F227L, Q612H, or V767D. Interestingly, Q612H was a novel mutation that was involved in stabilizing the conformation of the catalytic loop containing the KMSKS motif. In conclusion, this study confirms that molecular heterogeneity in ileS gene is common among clinical MRSA isolates with low-level mupirocin resistance, and further study on clinical mutants is needed to understand the structural basis of low-level mupirocin resistance.
Staphylococcus aureus/drug effects/*genetics ; *Mutation, Missense ; Mupirocin/*pharmacology ; Methicillin Resistance ; Isoleucine-tRNA Ligase/*genetics ; Intensive Care Units ; Humans ; Electrophoresis, Gel, Pulsed-Field ; Drug Resistance, Bacterial