Comparison of methicillin-resistant Staphylococcus aureus strains isolated in 2003 and 2008 with an emergence of multidrug resistant ST22: SCCmec IV clone in a tertiary hospital, Malaysia.

BACKGROUND/PURPOSE: Infections caused by methicillin-resistant Staphylococcus aureus (MRSA) continue to be a problem for clinicians worldwide. The objective of this study was to determine the changes in antibiograms of MRSA and their genotypic characteristics. METHODS: The antibiograms of 162 MRSA isolates (52 from 2003 and 110 from 2008) from a tertiary hospital were analyzed by antimicrobial susceptibility tests, the Panton-Valentine leukocidin (PVL) and staphylococcal cassette chromosome mec (SCCmec) types were determined by polymerase chain reaction, and genetic relatedness by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). RESULTS: All the isolates were sensitive to vancomycin. Resistance to ciprofloxacin, clindamycin, erythromycin, and gentamicin remained high throughout the study period, although a small decrease was observed in 2008 for ciprofloxacin (96% to 90%) and gentamicin (90% to 83%). Similarly, a slight decrease in resistance toward fusidic acid (10% to 9%), linezolid (2% to 1%), rifampicin (8% to 4%), and teicoplanin (4% to 0%) was observed between 2003 and 2008. In contrast, there was a significant increase (p < 0.05) in resistance rates toward trimethoprim-sulfamethoxazole, netilmicin, and tetracycline between 2003 and 2008. Ninety-six percent of the isolates from both 2003 and 2008 were multidrug resistant. Three SCCmec types (SCCmec type III, 90%; SCCmec type IV, 9%; SCCmec V, 1%) were observed. SCCmec type IV (n = 15) and pvl gene (n = 3) were detected in 2008 isolates but not in 2003 isolates. Most of the SCCmec type IV isolates (12 of 15) belonged to sequence type 22 (ST22) and were resistant to erythromycin and ciprofloxacin, with 11 being multidrug resistant. Most of the isolates were genetically related (F > 0.8) as determined by PFGE. Some isolates from 6 years apart shared similar PFGE profiles, indicating the persistence of a particular genotype. Five STs (ST239, ST772, ST22, ST6, and ST1178) were identified among the 2008 isolates but only one ST (ST239) was observed in 2003 isolates. CONCLUSION: Vancomycin remains the most active agent in vitro against S. aureus infection followed by linezolid and teicoplanin. The prevalence of resistance to fluoroquinolones, aminoglycosides (netilmicin), and tetracyclines had increased over the years. The Malaysian multidrug-resistant MRSA isolates were mostly SCCmec type III and ST239, although SCCmec type IV: ST22 is gaining importance. There was a correlation between resistotypes and PFGE profiles.

Temporal changes in the genotypes of methicillin-resistant Staphylococcus aureus strains isolated from a tertiary Malaysian hospital based on MLST, spa, and mec-associated dru typing.

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the main bacterial pathogens responsible for nosocomial infections leading to pneumonia, bloodstream, skin, and soft tissue infections. The objective of this study was to investigate the genomic changes of MRSA in a tertiary hospital between the years 2003, 2004, 2007, and 2008. One hundred fifty-four MRSA strains were characterized by multilocus sequence typing (MLST), spa, and mec-associated dru typing. Among the 154 strains, 29 different dru, 15 spa, and 8 MLST types were identified. Seven sequence types (STs) (ST239, ST22, ST5, ST6, ST80, ST573, and ST241) were identified among 2007-08 strains, although only 2 STs (ST239 and ST20) were observed among 2003 strains. Clones ST239-t037-dt13g, ST22-t032-(dt10a and dt10aw), and 28 other MRSA clones being introduced in 2007-2008 have replaced the ST239-t037 (dt13d, 14h, 13i, 13l, 13m, 15m, 15l, and 11al) clones present in 2003. The predominant MLST clone, ST239 (90.3%), was further distinguished into 7 different spa types and 26 different dru types, including 17 novel dru types. Maximum parsimony tree based on dru repeats revealed that 10 dru types (dt11am, dt13j, dt15n, dt13q, dt13n, dt13p, dt13f, dt13ao, dt12j, dt7v) shared the same MLST-spa types with dt13d, suggesting that these MRSA clones might have evolved from ST239-t037-dt13d. In conclusion, our data showed that the ST239-t037-dt13d clone and other MRSA clones in 2003 were replaced by ST239-t037-dt13g and other new emerging spa and dru types.

Characterisation of the Virulence Factors and Genetic Types of Methicillin Susceptible Staphylococcus aureus from Patients and Healthy Individuals.

Methicillin sensitive Staphylococcus aureus is an important bacterial pathogen associated with hospital- and community-acquired infections leading to endocarditis, skin tissue infection and pneumonia. The objective of this study was to determine both the genetic characteristics of methicillin-sensitive S. aureus (MSSA) strains, and the occurrence of virulence factors produced by S. aureus strains isolated from UMMC and healthy students in the University from year 2009. Out of 429 nasal swab samples, 67 were MSSA. The prevalence of 21 different virulence genes among 67 Malaysian clinical and community MSSA strains was determined by PCR, and their genetic features were assessed by PCR-RFLP of coa gene, agr types, spa typing and PFGE. The five predominant virulence genes were ica (79 %), efb and fnbA (61 % each), sdrE (57 %) and hlg (45 %). Toxin genes (enterotoxin, etd and pvl) were significantly more common (P < 0.05) in clinical strains compared to community strains. Three agr genotypes were observed: agr type I (45 %), agr type III (25 %) and agr type II (19 %). All 67 MSSA strains were distinguished into 26 profiles by PCR-RFLP of coa, 55 pulsotypes and 21 spa types. Four novel spa types (t7312, t7581, t7582 and t7583) were observed. In conclusion, different virulence profiles were observed in MSSA strains in Malaysia where toxin genes were more prevalent among clinical strains. No correlation between DNA profiles (coa-RFLP, PFGE and spa) and virulotypes was observed. The Malaysian MSSA strains from clinical and community sources were genetically diverse and heterogeneous.

Antimicrobial susceptibility profiling and genomic diversity of multidrug-resistant Acinetobacter baumannii isolates from a teaching hospital in Malaysia.

The resistance phenotypes and genomic diversity of 185 Acinetobacter baumannii isolates obtained from the intensive care unit (ICU) of a local teaching hospital in Kuala Lumpur from 2006 to 2009 were determined using antimicrobial susceptibility testing and pulsed-field gel electrophoresis (PFGE). Antibiogram analyses showed that the isolates were fully resistant to ß-lactam antimicrobials and had high resistance rates to the other antimicrobial agents tested. However, the isolates were susceptible to polymyxin B. Resistance to cefoperazone/sulbactam was only detected in strains isolated from 2007 to 2009. Some environmental isolates and an isolate from the hands of a healthcare worker (HCW) had identical resistance profiles and PFGE profiles that were closely related to patient isolates. Cluster analyses based on the PFGE profiles showed there was a persistent clone of endemic isolates in the ICU environment. The transmission route from HCWs to fomites to patients, which caused a long-term infection in the ICU of the University Malaya Medical Centre, was observed in this study. These data provide a better understanding of A. baumannii epidemiology within the hospital and the possible transmission routes. Knowledge of changes in the resistance rates of A. baumannii in our local hospital will improve antimicrobial therapy.

Macrorestriction analysis and antimicrobial susceptibility profiling of Salmonella enterica at a University Teaching Hospital, Kuala Lumpur.

The genetic diversity and antimicrobial resistance rates of clinical Salmonella isolates (2007-2008) at the University of Malaya Medical Centre, Kuala Lumpur, were investigated and the genetic diversity of the isolates was determined by pulsed-field gel electrophoresis (PFGE) and repetitive extragenic palindromic (REP)-PCR. XbaI-PFGE analysis generated 57 profiles (Dice coefficient, F=0.08-1.00), whereas REP-PCR using the REP primer generated only 35 (F=0.34-1.00). PFGE was therefore the more discriminative and reproducible method for assessing the genetic diversity of salmonellae. The antibiograms of 78 Salmonella isolates were assessed against 19 antimicrobials using the disk diffusion method. Twenty serotypes were identified, with the most common being S. Enteritidis (18%) followed by S. Typhimurium (14%), S. Paratyphi B var Java (9%), S. Weltevreden (9%), and S. Corvallis (9%). A total of 38 resistant profiles were defined, with 53.8% of the isolates being resistant to three or more antimicrobials. The highest resistance rates were observed for cephalothin (55.1%), tetracycline (47.4%), and nalidixic acid (35.9%). The presence of multidrug-resistant Salmonella strains is a cause for concern as it may limit the treatment of severe salmonellosis. One multidrug-resistant S. Enteritidis strain was a putative extended-spectrum beta-lactamase producer, based on a double disk diffusion analysis, and was resistant to ceftriaxone (MIC>32 microg/mL). The data generated by this study will contribute towards epidemiological monitoring and investigations of Salmonella infections in Malaysia.

Prevalence of mupirocin resistance in methicillin-resistant Staphylococcus aureus strains isolated from a Malaysian hospital.

Mupirocin is used topically to treat skin infection caused by methicillin-resistant Staphylococcus aureus (MRSA). One hundred eighty-eight strains (isolated in 2003, 2004, 2007, and 2008) were tested for mupirocin susceptibility using disk diffusion method and minimum inhibitory concentration (MIC). Mupirocin resistance was detected in 10 (5%) strains with 2 of them showing MIC of 256 mg/l. PCR detection using gene-specific primers showed that all 10 mupirocin-resistant strains harbored ileS2 gene whereas mupA gene was detected in 2 mupirocin-resistant strains with MIC of 256 mg/l. Amplification of agr grouping and SCCmec typing showed that all 10 strains were agr group I and SCCmec type III. Sequence analysis of region X of the spa gene yielded 4 distinct spa types (t037, t363, t421, and t6405) which were clonally related. In conclusion, the rate of mupirocin resistance in Malaysia is still low but is much higher than previous reports in Malaysia.