Detection Of Panton-Valentine Leukocidin Research Articles

DETECTION OF PANTON-VALENTINE LEUKOCIDIN (PVL) GENE AGAINST METHICILLIN-RESISTANT STAPHYLOCOCCUS AUREUS (MRSA) IN DIABETIC ULCER PATIENTS

Background: Diabetes Mellitus (DM) patients are susceptible to wound infections brought on by S. aureus bacteria, in dealing with infections caused by bacteria, antibiotic treatment was conducted. Nevertheless, misapplication of antibiotics might lead to drug resistance or MRSA. Panton-Valentine Leukocidin (PVL) is a crucial factor of S. aureus pathogenicity. Skin and soft tissue infections can be caused by the PVL gene. Purpose: To detect PVL gene in ulcer swabs of diabetic mellitus patients against MRSA strains. Method: Employing descriptive quantitative data analysis of PVL gene detection observations on ulcer swab of diabetes mellitus patients using the RT-PCR method. Ulcer swab samples were collected at Dharmahusada and Banyu Urip Diabetes Wound Specialist Hospital, Surabaya, East Java, Indonesia. The research was conducted at the Department of Medical Laboratory Technology, Health Polytechnic Ministry of Health Surabaya, Bacteriology Laboratory for the isolation and identification of MRSA and Molecular Biology Laboratory for the detection of the PVL gene in April–May 2022. Result: From 30 samples of ulcer swabs, 8 samples were positive for MRSA and 22 samples were negative for MRSA. After that, the MRSA positive samples were detected using the RT-PCR method to detect the PVL gene. The results of PVL detection were obtained from 6 samples (75%) positive for detecting the PVL gene and 2 samples (25%) negative for detecting the PVL gene. Conclusion: The results of the study showed that ulcer swabs from diabetes mellitus patients had the PVL gene against MRSA.

Acute Prosthetic Joint Infections with Poor Outcome Caused by Staphylococcus Aureus Strains Producing the Panton-Valentine Leukocidin.

The aim of this study was to investigate whether the presence of Staphylococcus aureus (SA) producing the Panton-Valentine leukocidin (PVL) affects the outcome of Prosthetic Joint Infection (PJI). Patients with acute and chronic PJI sustained by SA were prospectively enrolled at the orthopedic unit of "Casa di Cura Santa Maria Maddalena", from January 2019 to October 2021. PJI diagnosis was reached according to the diagnostic criteria of the International Consensus Meeting on PJI of Philadelphia. Synovial fluid obtained via joint aspirations was collected in order to isolate SA. The detection of PVL was performed via real-time quantitative PCR (RT-qPCR). The outcome assessment was performed using the criteria of the Delphi-based International Multidisciplinary Consensus. Twelve cases of PJI caused by SA were included. Nine (75%) cases were acute PJI treated using debridement, antibiotic and implant retention (DAIR); the remaining three (25%) were chronic PJI treated using two-stage (n = 2) and one-stage revision (n = 1), respectively. The SA strains that tested positive for PVL genes were 5/12 (41.6%,). Treatment failure was documented in three cases of acute PJI treated using DAIR, all supported by SA-PVL strains (p < 0.045). The remaining two cases were chronic PJI treated with a revision arthroplasty (one and two stage, respectively), with a 100% eradication rate in a medium follow-up of 24 months. Although a small case series, our study showed a 100% failure rate in acute PJI, probably caused by SA PVL-producing strains treated conservatively (p < 0.04). In this setting, toxin research should guide radical surgical treatment and targeted antibiotic therapy.

Detection of Panton-Valentine leukocidin and MecA Genes in Staphylococcus aureus isolated from Iraqi Patients.

A gram-positive bacterium, Staphylococcus aureus, which is widely distributed is considered as a bacterial infection that commonly infects the skin and mucous membranes. Such infections can be the cause of death and illness. In the present study by using reverse transcription-polymerase chain reaction (rt-PCR) the Panton-Valentine leukocidin (PVL) and MecA genes of S. aureus which were isolated from skin and soft tissue infections (SSTIs) in Baghdad, Iraq were investigated. This study included 96 S. aureus isolated from SSTIs and identified by Vitek. The results showed that 61 (63.5%) and 48 (50%) of the isolates were positive for PVL and MecA genes, respectively. This work presented an effective real-time PCR technique for detecting PVL genes alone or in conjunction with MecA. The rt-PCR allows for easier reaction monitoring and eliminates the need for post-PCR processing, saving both resources and time. Moreover, it is ideal for diagnostic applications because of its high sensitivity, simplicity, and specificity. Besides, the rt-PCR has an option to do all the procedures in an automated mode of action.

Irrelevance of Panton-Valentine leukocidin in hidradenitis suppurativa: results from a pilot, observational study

Panton-Valentine leukocidin (PVL) appears to be a virulence factor which, among others, can exacerbate the pathogenicity of Staphylococcus aureus infections, especially inducing severe necrotic, deep-seated skin infections, abscesses, and recurrences. These peculiarities have some overlaps with hidradenitis suppurativa (HS). Our main aim was to assess if S. aureus producing PVL could have some role in influencing clinical features and/or course of HS, specifically in the suppuration and recurrence of lesions. This pilot, mono-centric, observational study included all adult subjects affected with HS consecutively referring to our HS clinic over a 3-month period. Clinically evident suppuration and at least 2 weeks wash out from any antibiotic were the main inclusion criteria. Purulent material from HS skin lesions was collected with swabs in order to isolate micro-organisms, with specific regard to S. aureus. Detection of PVL was performed by real-time quantitative PCR (RT-qPCR). We also analyzed purulent material from suppurative skin lesions other than HS, as a control. Thirty HS patients were included; 29 purulent lesions (96.7%) harbored at least one bacterial species. Five (16.7%) swab samples were positive for S. aureus, none of which was positive for PVL genes. Among the 30 purulent disorders included as controls, 8 (26.3%) were positive for S. aureus; of these, 4 strains (50%) expressed LPV. The study results seem to exclude the pathogenetic involvement of S. aureus producing PVL in HS; as a result, PVL does not seem to represent a potential target in the future development of HS treatments.

Investigation of Nasal Staphylococcus aureus carriage in hospital employees and rapid detection of PVL and mecA genes by RT-PCR

Staphylococcus aureus (S.aureus) can cause serious fatal infections due to several enzymes and toxins that it secretes when it encounters favorable conditions.For this reason, S.aureus is one of the most important pathogens currently causing hospital and community infections.Methicillin-resistant S.aureus(MRSA) strains are resistant to many antibiotics and have the ability to rapidly develop resistance to currently used antibiotics during treatment. MRSA origins cause serious infections with high mortality and also increase the rate of hospital infections that require difficult and longer hospitalization and the use of greater quantities of antibiotics in risk-prone units in hospitals. Another factor that is as important as MRSA in S.aureus pathogenicity is the Panton-Valentine leukocidin(PVL) toxin secreted by S.aureus strains, most commonly isolated from deadly necrotizing pneumonias and community-acquired soft skin tissue infections.PVL is especially observed in community-acquired S.aureus strains. However, studies showed that PVL-positive origins began to spread rapidly to hospital environments. In this study, methicillin-sensitive S.aureus, methicillin-resistant coagulase-negative Staphylococcus (MR-CNS), and Panton-Valentine leukocidine genes were identified by real-time polymerase chain reaction (RT-PCR) by investigating the rate of nasal S.aureus carriers(NSAT) in nasal swab samples taken from hospital staff belonging to the highest risk group for staphylococcal infections, at which screening is aimed. Classical NSAT scans were performed on nasal swap samples taken from 98 healthcare professionals in Hospital. Nasal S.aureus (4 MSSA,13 MRSA) was detected in 17 samples(17.35%). During detection, 48 MR-CNS were detected. In addition, 61 mecA (+),17 nuc (+),1 pvl (+) genes were detected in RT-PCR samples directly from the nasal swabs taken in accordance with the culture method.

Methicillin-resistant Staphylococcus aureus nasal carriage in international medical conference attendees.

Carriage of methicillin-resistant Staphylococcus aureus (MRSA) is associated with its transmission. International travels and massive gatherings may accelerate such transmission. MRSA carriage was surveyed among the attendees of two international medical conferences held in Taipei in 2010. A total of 209 attendees from 23 countries were recruited. Nasal specimens were collected from each volunteer and subjected to polymerase chain reaction (PCR) detection for MRSA. Molecular analysis, including pulsed-field gel electrophoresis, multilocus sequence typing (MLST), typing of staphylococcal cassette chromosome mec (SCCmec) and staphylococcal protein A (spa) genes, and detection of Panton-Valentine leukocidin (PVL) and sasX genes, was performed. MRSA carriage was detected in 10 (4.8%) attendees from Vietnam (3/8, 37.5%), Korea (2/6, 33.3%), Japan (2/41, 4.9%), Philippines (2/52, 3.8%), and Bangladesh (1/4, 25.0%). The proportion of MRSA colonizers was significantly higher in the local hospital group compared to those from the other groups (3/17 vs. 7/192, p<0.05). Six MRSA isolates were available for molecular analysis. They all carried a type IV SCCmec gene. Five pulsotypes were identified; four genotypes, respectively, were identified by MLST and spa typing. None of the isolates carried either PVL or sasX genes. None of common molecular characteristics was shared by isolates from different countries. Most of these isolates were local endemic community clone in each country. As healthcare workers, a certain proportion of international medical conference attendees harbored MRSA in their nares, mostly local endemic community clones in each country, which has the potential of spread among attendees.

Prevalence of methicillin-resistant Staphylococcus aureus (MRSA) infection and the molecular characteristics of MRSA bacteraemia over a two-year period in a tertiary teaching hospital in Malaysia

BackgroundMethicillin-resistant Staphylococcus aureus (MRSA) is an established pathogen that causes hospital- and community-acquired infections worldwide. The prevalence rate of MRSA infections were reported to be the highest in Asia. As there is limited epidemiological study being done in Malaysia, this study aimed to determine the prevalence of MRSA infection and the molecular characteristics of MRSA bacteraemia.MethodsTwo hundred and nine MRSA strains from year 2011 to 2012 were collected from a tertiary teaching hospital in Malaysia. The strains were characterized by antimicrobial susceptibility testing, staphylococcal cassette chromosome mec (SCCmec) typing, detection of Panton-Valentine leukocidin (PVL) gene, multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). Patient’s demographic and clinical data were collected and correlated with molecular data by statistical analysis.ResultsMale gender and patient >50 years of age (p < 0.0001) were significantly associated with the increased risk of MRSA acquisition. Fifty-nine percent of MRSA strains were HA-MRSA that carried SCCmec type II, III, IV and V while 31% were CA-MRSA strains with SCCmec III, IV and V. The prevalence of PVL gene among 2011 MRSA strains was 5.3% and no PVL gene was detected in 2012 MRSA strains. All of the strains were sensitive to vancomycin. However, vancomycin MIC creep phenomenon was demonstrated by the increased number of MRSA strains with MIC ≥1.5 μg/mL (p = 0.008) between 2011 and 2012. Skin disease (p = 0.034) and SCCmec type III (p = 0.0001) were found to be significantly associated with high vancomycin MIC. Forty-four percent of MRSA strains from blood, were further subtyped by MLST and PFGE. Most of the bacteraemia cases were primary bacteraemia and the common comorbidities were diabetes, hypertension and chronic kidney disease. The predominant pulsotype was pulsotype C exhibited by SCCmec III-ST239. This is a first study in Malaysia that reported the occurrence of MRSA clones such as SCCmec V-ST5, untypeable-ST508, SCCmec IV-ST1 and SCCmec IV-ST1137.ConclusionsSCCmec type III remained predominant among the MRSA strains in this hospital. The occurrence of SCCmec IV and V among hospital strains and the presence of SCCmec III in CA-MRSA strains are increasing. MRSA strains causing bacteraemia over the two-year study period were found to be genetically diverse.

Molecular Characteristic and Virulence Gene Profiles of Community-Associated Methicillin-Resistant Staphylococcus aureus Isolates from Pediatric Patients in Shanghai, China.

Staphylococcus aureus is a globally important human pathogen, especially among children and immunocompromised patients. The emergence and spread of community-associated methicillin-resistant S. aureus (CA-MRSA) has become a serious public health problem worldwide. The aim of this study was to investigate the prevalence, molecular characteristics and virulence profiles of CA-MRSA infections from pediatric patients in a university hospital in Shanghai, China. A total of 80 CA-MRSA isolates were collected from July 2012 to December 2013 in Shanghai Children's Medical Center and analyzed by multilocus sequence typing, staphylococcus chromosomal cassette mec (SCCmec) typing, and spa typing. The detection of Panton-Valentine Leukocidin (pvl), superantigenic and exfoliative toxins, and adhesin genes was also performed. Overall, 16 distinct sequence types (STs) were identified among the 80 isolates. Among them, ST59 was found to be the most prevalent, followed by ST398 (11.3%, 9/80) and ST88 (8.8%, 7/80). SCCmec types IV and V were observed, at 60 and 40%, respectively. Thirty spa types were identified, spa t437 (23.8%) was the most predominant type. All 80 isolates exhibited carriage of at least four virulence genes. Thirty-four (42.5%, 34/80) isolates harbored ≥10 tested virulence genes. Adhesion genes were present in most of the MRSA isolates, including the following: icaA (100%), clfA (100%), sdrC (95%), and sdrE (63.8%). The prevalence of pvl gene was 20%, and multidrug resistance was observed in 36% of all strains. In addition, ST59-MRSA-IV with t437 accounted for 21.3% of occurrences, making it the most prevalent clone. Isolates that were carriers of toxin genes, and hla (100%) and hlg (87.5%) were the most frequent. In conclusion, simultaneous carriage of multiple virulence genes and genetically considerable diversity were very common among CA-MRSA from pediatric patients in Shanghai. ST59-MRSA-IV with t437 was still the most predominant type. The combination of virulence gene profiles and antibiotic resistance may help ST59 to be successfully spread among children.

Characterization of Staphylococcus aureus from Human Immunodeficiency Virus (HIV) patients in Accra, Ghana.

The aims of this study were to: a) determine the nasal carriage prevalence of Stahpylococcus aureus among HIV patients, b) to characterize S. aureus strains isolated. Characterization of S. aureus isolates was done by antibiotyping, spa typing, and detection of Panton-Valentine leukocidin (PVL) genes. S. aureus isolated (10/124; 8%) belonged to spa types t084 (n = 3), t10828 (n = 2), t311, t304, t774, t645, and t091. The isolates were resistant to penicillin (100%), tetracycline (40%), rifampicin (10%), fucidic acid (10%), norfloxacin (10%), erythromycin (10%), and sulfamethoxazole trimethoprim (10%). Multidrug resistance (MDR) was detected in 30% of the isolates. The finding of MDR S. aureus among HIV-positive patients suggests that surveillance of antimicrobial resistant S. aureus among this patient group could be considered as an infection control measure in the hospital.

Molecular Detection of Panton-Valentine Leukocidin (PVL) Toxins in Clinical Isolates of Staphylococcus aureus from Maitama District Hospital, Abuja, Nigeria

Pathogenic Staphylococcus aureus is the most frequently isolated Gram positive bacterium from clinical specimens; and it is among the leading cause of infection in man. S. aureus has gained significant interest in recent years as an important nosocomial pathogen – owing to its multidrug resistant nature which is associated to several virulence factors of the organism including Panton-valentine leukocidin (PVL). PVL is part of the toxins produced by pathogenic S. aureus – which help the organism to exacerbate their pathogenicity/virulence in the phase of an infection. This study evaluated the prevalence of PVL-positive S. aureus from clinical specimens – owing to the dearth of information on this subject matter in Nigeria. Out of the 118 non-consecutive S. aureus isolates employed for this study, only 56 isolates were biochemically confirmed as pathogenic S. aureus. The antibiogram showed that the S. aureus isolates were most susceptible to gentamicin, vancomycin, ciprofloxacin, erythromycin and linezolid. However, they were highly resistant to the cephamycin, cefoxitin (82.1%). The S. aureus isolates also showed reduced susceptibility to tigecycline (71.4%), clindamycin (66.1%), chloramphenicol (48.2%), amoxicillin-clavulanic acid (53.6%) and sulphamethoxazole-trimethoprim (48.2%). The prevalence of PVL genes in this study was 10.7%. Only 6 isolates of S. aureus (10.7%) were confirmed by PCR to harbour the PVL genes; and these S. aureus isolates were from wound samples, abscess and urine samples. The occurrence of pathogenic S. aureus harbouring drug resistant genes such as PVL genes in the hospital environment pose serious health and therapeutic challenges especially in choosing antimicrobial therapy for treatment. S. aureus isolates with PVL genes could also disseminate with high propensity within the hospital environment; and this could result in the outbreak of nosocomial infections. Continues antibiotic stewardship in our hospitals will help in the control and prevention of the emergence and dissemination of drug-resistant microbes in both the community and hospital environment.

(404) - Clinical and Molecular Epidemiologic Characterization of Methicillin-Resistant Staphylococcus aureus (MRSA) Infections Occurring Early After Lung Transplant

(404) - Clinical and Molecular Epidemiologic Characterization of Methicillin-Resistant Staphylococcus aureus (MRSA) Infections Occurring Early After Lung Transplant

Continued Emergence of USA300 Methicillin-ResistantStaphylococcus aureusin the United States: Results from a Nationwide Surveillance Study

The epidemiology of methicillin-resistant Staphylococcus aureus (MRSA) is changing, with USA300 emerging first in community and then in healthcare settings. We performed nationwide surveillance to assess recent trends in the molecular epidemiology of MRSA. One hundred consecutive unique clinically significant S. aureus isolates were recovered from patients at each of 43 US centers between July 1, 2011, and December 31, 2011. Susceptibility testing, pulsed-field gel electrophoresis (PFGE), staphylococcal protein A gene (spa) and staphylococcal cassette chromosome mec typing, and Panton-Valentine leukocidin detection were performed on all MRSA isolates. Of 4,131 isolates collected, 2,093 (51%) were MRSA. Specimen sources of MRSA isolates included wound or abscess (54%), blood (24%), lower respiratory tract (11%), and other sterile site (10%). Thirty percent were isolated more than 48 hours after hospital admission (ie, were associated with nosocomial acquisition of infection). USA300 was the most common PFGE type (1,269 isolates; 61%), overall and in all regions, followed by USA100 (368 isolates; 18%). Among 173 spa types found, the most common were t008 (51%) and t002 (18%); no other spa type accounted for more than 2% of isolates. One strain type (USA300/t008/IV) constituted almost half of all MRSA isolates (1,005 isolates; 48%) and was the most common at all body sites, causing 37% of MRSA bloodstream infections (BSIs) and 38% of nosocomial MRSA infections. Multidrug-resistant phenotypes were found among 34 USA300 isolates (3%) from 18 states. The USA300 PFGE type continues to advance nationwide. A single strain type (USA300/t008/IV) predominates in all regions and infection sites and is now more common than USA100 as a cause of MRSA BSI and nosocomial infections. Although most USA300 retain typical susceptibility profiles, multidrug-resistant phenotypes are emerging.

High-Density Livestock Production and Molecularly Characterized MRSA Infections in Pennsylvania

Background: European studies suggest that living near high-density livestock production increases the risk of sequence type (ST) 398 methicillin-resistant Staphylococcus aureus (MRSA) colonization. To our knowledge, no studies have evaluated associations between livestock production and human infection by other strain types.Objectives: We evaluated associations between MRSA molecular subgroups and high-density livestock production.Methods: We conducted a yearlong 2012 prospective study on a stratified random sample of patients with culture-confirmed MRSA infection; we oversampled patients from the Geisinger Health System with exposure to high-density livestock production in Pennsylvania. Isolates were characterized using S. aureus protein A (spa) typing and detection of Panton-Valentine leukocidin (PVL) and scn genes. We compared patients with one of two specific MRSA strains with patients with all other strains of MRSA isolates, using logistic regression that accounted for the sampling design, for two different exposure models: one based on the location of the animals (livestock model) and the other on crop field application of manure (crop field model).Results: Of 196 MRSA isolates, we identified 30 spa types, 47 PVL-negative and 15 scn-negative isolates, and no ST398 MRSA. Compared with quartiles 1–3 combined, the highest quartiles of swine livestock and dairy/veal crop field exposures were positively associated with community-onset-PVL-negative MRSA (CO-PVL-negative MRSA vs. all other MRSA), with adjusted odds ratios of 4.24 (95% CI: 1.60, 11.25) and 4.88 (95% CI: 1.40, 17.00), respectively. The association with CO-PVL-negative MRSA infection increased across quartiles of dairy/veal livestock exposure (trend p = 0.05).Conclusions: Our findings suggest that other MRSA strains, beyond ST398, may be involved in livestock-associated MRSA infection in the United States.Citation: Casey JA, Shopsin B, Cosgrove SE, Nachman KE, Curriero FC, Rose HR, Schwartz BS. 2014. High-density livestock production and molecularly characterized MRSA infections in Pennsylvania. Environ Health Perspect 122:464–470; http://dx.doi.org/10.1289/ehp.1307370

Emergence of methicillin-resistant Staphylococcus aureus USA300 genotype as a major cause of late-onset nosocomial pneumonia in intensive care patients in the USA

To compare demographic and clinical characteristics, and methicillin-resistant Staphylococcus aureus (MRSA) strain characteristics, in patients with early-onset (EO) and late-onset (LO) MRSA nosocomial pneumonia. This was a retrospective analysis of data from a multicenter observational study of nosocomial pneumonia patients admitted between November 2008 and July 2010. Laboratory analyses performed on MRSA isolates included confirmation of antimicrobial susceptibility and heteroresistance to vancomycin, USA typing, staphylococcal cassette chromosome (SCC) mec typing, and detection of Panton-Valentine leukocidin (PVL) genes. We identified 134 patients; 42 (31%) had EO MRSA pneumonia and 92 (69%) had LO MRSA pneumonia. The patients in the LO group were more likely to have risk factors for multidrug-resistant pathogens (98% vs. 76%, p<0.001). The MRSA USA300 strain was found with equal frequency in the EO and LO groups. Likewise, both groups had similar frequencies of isolates exhibiting PVL and SCCmec type IV. Our findings provide further evidence of the continued migration of community-associated MRSA into the healthcare setting in the USA. MRSA USA300 genotype has emerged as a significant cause of LO nosocomial pneumonia in intensive care units. Appropriate anti-MRSA antimicrobial therapy should be considered for both EO and LO hospital-acquired pneumonia and ventilator-associated pneumonia.

In Vitro Susceptibilities and Molecular Analysis of Vancomycin-Intermediate and Vancomycin-Resistant Staphylococcus aureus Isolates

There is increasing frequency of vancomycin-intermediate and -resistant Staphylococcus aureus (VISA and VRSA) isolates identified in clinical practice. There are limited reports evaluating susceptibility patterns and molecular characteristics of these strains. Laboratory analysis was performed on 13 VRSA and 33 VISA isolates, including susceptibility testing by broth microdilution, detection of Panton-Valentine leukocidin (PVL) genes, arginine catabolic mobile element (ACME), and staphylococcal cassette chromosome mec typing using polymerase chain reaction. Strain typing using pulsed-field gel electrophoresis (PFGE) was performed on VRSA isolates. Telavancin, linezolid, tigecycline, and minocycline were active against >90% of VISA isolates, while >90% of VRSA isolates were susceptible to ceftaroline, daptomycin, linezolid, minocyline, tigecycline, rifampin, and trimethoprim/sulfamethoxazole. There were no VISA or VRSA isolates that carried PVL genes or ACME, and most strains (69.8%) were staphylococcal cassette chromosome mec type II. VRSA isolates were predominantly related to USA100 (53.8%) and none were related to USA300 or USA400. A large number of available antimicrobial agents retain very good in vitro activity against VRSA and VISA isolates. The present isolates appear to be derived from healthcare-associated strains based on the absence of features associated with community-associated strains, and VRSA isolates are polyclonal by PFGE.

Antimicrobial susceptibility profiles of meticillin-susceptible and -resistant Staphylococcus aureus: focus on daptomycin minimum inhibitory concentrations at a tertiary care centre in Mumbai, India

In this study, daptomycin minimum inhibitory concentrations (MICs) for Staphylococcus aureus isolates were determined using Etest strips and were correlated with staphylococcal cassette chromosome mec (SCC mec) types and Panton–Valentine leukocidin (PVL) gene positivity. In total, 60 meticillin-resistant S. aureus (MRSA) and 60 meticillin-susceptible S. aureus (MSSA) isolates from clinical samples were subjected to antimicrobial susceptibility testing by disk diffusion according to Clinical and Laboratory Standards Institute guidelines, polymerase chain reaction (PCR) detection of PVL and mecA genes, and SCC mec typing. Daptomycin MICs were determined using Etest strips. The mecA gene was present in all MRSA isolates and was absent in 59 MSSA isolates. The PVL gene was present in 41 MRSA isolates and 25 MSSA isolates. Amongst the MRSA isolates, 10 were SCC mec type III, 27 were SCC mec IV and 23 were SCC mec V. Moreover, 26 SCC mec IV and 15 SCC mec V isolates were PVL-positive. All SCC mec III isolates were multidrug-resistant and PVL-negative. Daptomycin MICs ranged from 0.047 μg/mL to 1 μg/mL for MRSA and from 0.19 μg/mL to 1 μg/mL for MSSA. All MRSA and MSSA isolates were susceptible to daptomycin. Although SCC mec III MRSA and PVL-positive MSSA were resistant to more antimicrobial classes than SCC mec IV and V MRSA and PVL-negative MSSA, there does not appear to be a significant correlation between SCC mec types, PVL positivity and daptomycin MICs. MICs were not as low as expected for a newly introduced drug.

Usefulness of antimicrobial resistance pattern for detecting PVL- or TSST-1-producing meticillin-resistant Staphylococcus aureus in a French university hospital

Several recent reports have suggested that community-associated meticillin-resistant Staphylococcus aureus (MRSA) clones, particularly those harbouring genes for Panton-Valentine leukocidin (PVL) or toxic shock syndrome toxin 1 (TSST-1), are increasingly responsible for infections in hospitals. Here, a retrospective study was carried out to investigate whether antimicrobial resistance patterns could be used to detect these pathogens in a French university hospital. Isolates were characterized by antimicrobial susceptibility testing, PCR profiling (PVL genes and tst), PFGE typing and multilocus sequence typing. Demographic and clinical data were collected from all patients. For PVL-positive MRSA, the typical antimicrobial resistance pattern (susceptible to fluoroquinolones, non-susceptible to fusidic acid, kanamycin resistant and susceptible to gentamicin and tobramycin) had a sensitivity of 77.8 % and a positive predictive value (PPV) of 100 %. For tst-positive MRSA, the antimicrobial resistance pattern (susceptible to fluoroquinolones and non-susceptible to fusidic acid) had a sensitivity of 100 % and a PPV of 72.4 %. These results suggest that phenotypic rules based on antimicrobial resistance patterns are potentially useful for the detection of PVL- and tst-positive MRSA isolates.

Methicillin-resistant Staphylococcus aureus in hospitals in Tbilisi, the Republic of Georgia, are variants of the Brazilian clone

The purpose of this study was to characterise methicillin-resistant Staphylococcus aureus (MRSA) isolates from the Republic of Georgia, part of the former Soviet Union. Thirty-two non-duplicate MRSA isolates were collected in the period from May 2006 to February 2007. The patient data were analysed and the isolates were characterised by staphylococcal protein A (spa) typing, staphylococcal chromosome cassette mec (SCCmec) typing, multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE) and the detection of Panton-Valentine leukocidin (PVL) genes. Only two closely related spa types were found; 29 isolates were of spa type 459 and three were t030. The spa types belonged to sequence type (ST) 239, clonal complex (CC) 8. All isolates were multiresistant, PVL-negative and harboured SCCmec type IIIA. Based on the molecular findings and PFGE, the isolates most closely resembled the pandemic Brazilian clone (ST239-IIIA).

Rapid Increase of Genetically Diverse Methicillin-ResistantStaphylococcus aureus,Copenhagen, Denmark

In Copenhagen, methicillin-resistant Staphylococcus aureus (MRSA) accounted for <15 isolates per year during 1980-2002. However, since 2003 an epidemic increase has been observed, with 33 MRSA cases in 2003 and 110 in 2004. We analyzed these 143 cases epidemiologically and characterized isolates by pulsed-field gel electrophoresis, Staphylococcus protein A (spa) typing, multilocus sequence typing, staphylococcal chromosome cassette (SCC) mec typing, and detection of Panton-Valentine leukocidin (PVL) genes. Seventy-one percent of cases were community-onset MRSA (CO-MRSA); of these, 36% had no identified risk factors. We identified 29 spa types (t) and 16 sequence types (STs) belonging to 8 clonal complexes and 3 ST singletons. The most common clonal types were t024/ST8-IV, t019/ST30-IV, t044/ST80-IV, and t008/ST8-IV (USA300). A total of 86% of isolates harbored SCCmec IV, and 44% had PVL. Skin and soft tissue infections dominated. CO-MRSA with diverse genetic backgrounds is rapidly emerging in a low MRSA prevalence area.

StaphPlex System for Rapid and Simultaneous Identification of Antibiotic Resistance Determinants and Panton-Valentine Leukocidin Detection of Staphylococci from Positive Blood Cultures

Phenotypic methods take several days for identification and antimicrobial susceptibility testing of staphylococcal isolates after gram-positive cocci in clusters (GPCC) are observed in positive blood cultures. We developed and validated a StaphPlex system that amplifies and detects 18 gene targets simultaneously in 1 reaction for species-level identification of staphylococci, detection of genes encoding Panton-Valentine leukocidin (PVL), and antimicrobial resistance determinants of staphylococci. The StaphPlex system was compared to phenotypic methods for organism identification and antimicrobial resistance detection for positive blood culture specimens in which GPCC were observed. Among a total of 360 GPCC specimens, 273 (75.8%), 37 (10.3%), 37 (10.3%), 1 (0.3%), 3 (0.8%), and 9 (2.5%) were identified by StaphPlex as coagulase-negative Staphylococcus (CoNS), methicillin-resistant Staphylococcus aureus (MRSA), methicillin-susceptible S. aureus (MSSA), or mixed infections of CoNS and MRSA, CoNS and MSSA, or nonstaphylococci, respectively, with an overall accuracy of 91.7%. The 277 CoNS-containing specimens were further identified to the species level as containing 203 (73.3%) Staphylococcus epidermidis isolates, 10 (3.6%) Staphylococcus haemolyticus isolates, 27 (9.7%) Staphylococcus hominis isolates, 1 (0.4%) Staphylococcus lugdunensis isolate, and 36 (13.0%) other CoNS isolates, with an overall accuracy of 80.1% compared to an API STAPH test and CDC reference identification. Numerous very major errors were noticed when detection of aacA, ermA, ermC, tetM, and tetK was used to predict in vitro antimicrobial resistance, but relatively few major errors were observed when the absence of these genes was used to predict susceptibility. The StaphPlex system demonstrated 100% sensitivity and specificity, ranging from 95.5% to 100.0% when used for staphylococcal cassette chromosome mec typing and PVL detection. StaphPlex provides simultaneous staphylococcal identification and detection of PVL and antimicrobial resistance determinants within 5 h, significantly shortening the time needed for phenotypic identification and antimicrobial susceptibility testing.