耐甲氧西林金黄色葡萄球菌杀白细胞素基因检测

目的 调查临床分离的耐甲氧西林金黄色葡萄球菌(MRSA)杀白细胞毒素基因携带率。方法 收集非重复MRSA菌株83株,按照美国疾病预防控制中心的CA-MRSA 定义分为HA-MRSA和CA-MRSA两组。采用多重PCR法进行SCCmec分型,普通PCR+测序法进行spa分型,普通PCR检测杀白细胞素(PVL)基因。结果 83株MRSA中HA-MRSA、CA-MRSA分别占47.0%、53.0%,SCCmec分型中SCCmecⅠ、Ⅱ、Ⅲ、Ⅳa型和未分型各占1.2%、3.6%、65.1%、28.9%、1.2%,spa分型中83株MRSA共检出15个型,主要分型为t437,t062,t015分别占39.8%, 21.7%,10.8%;PVL阳性的MRSA中HA-MRSA和CA-MRSA分别10.3%、36.4%,两者差异有显著性(P=0.006);33株spa t437中有18株携带PVL基因,阳性率54.5%,50株其他spa分型中仅2株携带PVL基因,阳性率4.0%,两者差异有显著性(P=0.000)。PVL基因阳性的MRSA特征CA-MRSA-Ⅳa-t437 9株,HA-MRSA-Ⅲ-t437 4株,HA-MRSA-Ⅳa-t437 3株;20株PVL基因阳性的MRSA中10株分离自皮肤软组织感染病例,6株分离于耳鼻喉科感染病例,3株分离于呼吸道感染病例,1株分离于败血症病例。结论 CA-MRSA菌株较HA-MRSA菌株的 PVL基因阳性率更高,同时也发现携带有更高毒力的CA-MRSA的克隆已经播散到医院的环境中,引起医院获得性相关感染。     

What is community-associated methicillin-resistant Staphylococcus aureus?

BACKGROUND: A community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infection has been defined as an MRSA infection in a patient who lacks specific risk factors for healthcare exposure. We sought to determine whether the absence or presence of these risk factors still predicts the phenotypic or genotypic characteristics of MRSA strains. METHODS: All clinical MRSA isolates were prospectively collected at the University of Chicago Hospitals from July 2004 through June 2005. Patients were interviewed and/or their medical records were reviewed. Isolates underwent genotyping and susceptibility testing. Data on patients and isolates were stratified in accordance with 8 frequently cited criteria for the identification of CA-MRSA and compared for concordance. RESULTS: Among 616 unique patients from whom MRSA isolates were recovered, 404 (65.6%) had risk factors for healthcare exposure. Of the 404 isolates recovered from these patients, 166 (41.1%) were clindamycin susceptible, 190 (47.0%) carried staphylococcal cassette chromosome mec (SCCmec) type IV, 145 (35.9%) carried the Panton-Valentine leukocidin genes (PVL+), and 162 (40.1%) were identified as sequence type (ST) 8 by multilocus sequence typing (MLST), all of which are characteristics commonly attributed to CA-MRSA strains. CONCLUSIONS: Association with the healthcare environment now has little predictive value for distinguishing patients with infection due to multidrug resistant MRSA isolates from those infected by CA-MRSA isolates, that is, isolates that are clindamycin-susceptible, PVL+, ST8, and/or contain SCCmec type IV. Defining CA-MRSA by the absence of risk factors for healthcare exposure greatly underestimates the burden of epidemic CA-MRSA disease.     

Epidemiological and microbiological analysis of community-associated methicillin-resistant Staphylococcus aureus strains isolated from a Japanese hospital

Reports of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections have recently increased in Japan; however, these studies contain limited information on their epidemiology. We performed a single-center study in the Tokyo Medical University Hospital located in Shinjuku, a central area of Tokyo, Japan. From 2,099 MRSA isolates obtained during July 2007 to March 2009, we selected 44 MRSA isolates with a MIC of <2 μg/mL for imipenem. Among 44 isolates, 28 strains had type IV or type V SCCmec, and we classified them as CA-MRSA. We identified only 1 Panton-Valentine leukocidin (PVL)-positive MRSA strain, which belonged to SCCmec type V. The PVL-positive CA-MRSA strain was isolated from a patient with multiple subcutaneous abscesses. The patient had returned to Japan from India; thus, the strain may have been contracted from outside of Japan. Thirteen (46.4%) and 15 strains (53.6%) were isolated from outpatients and inpatients, respectively. The major sites of infection included the respiratory tract (8 strains, 28.6%), skin/soft tissue (4 strains, 14.3%), and nasal cavity (4 strains, 14.3%). It is important to note that the most common site of CA-MRSA infection in inpatients was the respiratory tract; respiratory infections with CA-MRSA frequently cause severe infectious diseases.     

Complex molecular epidemiology of methicillin-resistant staphylococcus aureus isolates from children with cystic fibrosis in the era of epidemic community-associated methicillin-resistant S aureus

BACKGROUND: Limited data exist about the molecular types of methicillin-resistant Staphylococcus aureus (MRSA) strains found in children with cystic fibrosis (CF). We sought to characterize MRSA strains from these patients and compare them with MRSA strains from non-CF pediatric patients. METHODS: All MRSA isolates were collected prospectively at Children's Medical Center in Dallas, TX, and the University of Chicago Comer Children's Hospital in 2004 to 2005. All CF MRSA isolates underwent susceptibility testing, multilocus sequence typing, Panton-Valentine leukocidin gene detection (pvl+), and staphylococcal chromosome cassette mec (SCCmec) typing. RESULTS: A total of 22 of 34 MRSA isolates (64.7%) from patients with CF belonged to clonal complex (CC) 5 and contained SCCmec II, so-called health-care associated MRSA (HA-MRSA) strains. Nine of 34 MRSA strains (26.5%) were CC 8, and contained SCCmec IV, so-called community-associated MRSA (CA-MRSA) strains. The CA-MRSA strains tended to be isolated from newly colonized CF patients. In contrast, CC8 isolates predominated among the non-CF patients (294 of 331 patients; 88.8%). MRSA isolates from children with CF were more likely to be resistant to clindamycin (65% vs 19%, respectively) and ciprofloxacin (62% vs 17%, respectively) compared with strains from non-CF patients (p < 0.001). There was no difference in the rate of pvl+ isolate recovery from children with CF undergoing a surveillance culture (7 of 23 children) compared with those with pulmonary exacerbation (3 of 11 children; p = 1.0). CONCLUSIONS: Both CA-MRSA (CC8) isolates and HA-MRSA (CC5) isolates populate the respiratory tracts of children with CF. HA-MRSA isolates predominated, but CA-MRSA strains predominated among CF patients with newly acquired MRSA strains and among the non-CF patients. The presence of CA-MRSA strains in children with CF was not associated with exacerbation or necrotizing pneumonia.     

Community-associated methicillin-resistant Staphylococcus aureus infections in men who have sex with men: A case series

BACKGROUND: The purpose of the present study was to describe the clinical characteristics and management of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections among a cohort of men who have sex with men. PATIENTS AND METHODS: A retrospective chart review was conducted of patients with culture-proven MRSA at Maple Leaf Medical Clinic (Toronto, Ontario) between November 2004 and December 2005. Cases were identified by individual physicians and by queries in the clinical management system. A standard data collection form was used to record patient demographics, potential risk factors for MRSA and course of illness. When available, antimicrobial sensitivities were recorded. DNA fingerprinting using pulsed-field gel electrophoresis, and genetic analysis for SCCmec typing and detection of the Panton-Valentine leukocidin cytotoxin were performed on six available isolates. RESULTS: Seventeen patients with MRSA infection were identified, 12 (71%) of whom were HIV-positive. The most common clinical presentation was abscess (35%), followed by furuncle (17%), folliculitis (17%), cellulitis (17%) and sinusitis (12%). The majority of MRSA isolates were resistant to ciprofloxacin (92%) and levofloxacin (77%). All isolates were susceptible to trimethoprim-sulfamethoxazole, rifampin, linezolid, gentamicin and clindamycin, while the majority were susceptible to tetracycline (80%). All six isolates tested were SCCmec type IVa-positive and Panton-Valentine leukocidin-positive, and had fingerprint patterns consistent with the CMRSA-10 (USA300) clone. CONCLUSION: The present study describes the clinical presentation and management of CA-MRSA infections occurring in Toronto among men who have sex with men. The infections appear to have been caused by CMRSA-10, which has caused the majority of CA-MRSA outbreaks elsewhere.     

Coexistence of Panton-Valentine leukocidin-positive and -negative community-associated methicillin-resistant Staphylococcus aureus USA400 sibling strains in a large Canadian health-care region

Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) strains often carry the Panton-Valentine leukocidin (PVL) genes. However, the specific role that PVL plays in the epidemiological features and pathogenesis of CA-MRSA infections has remained undefined and controversial. Conducting a retrospective study on a natural population of MRSA clinical isolates recovered from community and hospital patients in a large Canadian health-care region during a 6-year period, we identified the coexistence of 2 USA400 (a major clonal group of CA-MRSA) sibling strains with and without PVL genes. Polymerase chain reaction and sequence analysis indicated that the PVL-carrying prophage phiSa2mw was present in PVL(+) but absent in PVL(-) USA400 isolates. These strains shared identical genotypic and phenotypic properties and similar clinical characteristics. This study provides direct evidence that PVL genes are not necessarily the key determinants associated with the increasing dissemination of CA-MRSA strains, suggesting that the genomic milieu may play a greater role in this regard.     

Community-associated methicillin-resistant Staphylococcus aureus infective endocarditis with Panton-Valentine leukocidin gene in an injection drug user with HIV infection

Reports of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) isolates carrying Panton-Valentine leukocidin (PVL) gene that causes infective endocarditis in injection drug users (IDUs) with human immunodeficiency virus (HIV) infection are rare in the English language literature. We present a case of CA-MRSA infective endocarditis with bilateral septic lung emboli in a previously healthy 45-year-old IDU. This case suggests that PVL gene-positive CA-MRSA should be considered as a potential pathogen in IDUs with infective endocarditis.     

Two cases of severe necrotizing pneumonia caused by community-acquired methicillin-resistant Staphylococcus aureus]

We present 2 cases with severe necrotizing pneumonia due to community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) infection. The patients were a 89-year-old man and a male student of 15 years of age. Chest X-rays and CT scans demonstrated multiple consolidations with cavitary lesions showing necrotizing pneumonia. MRSA strains were isolated from the sputum cultures on admission in these patients who did not have any established risk factors for MRSA infections such as history of hospitalization, surgery, hemodialysis, the presence of a permanent indwelling catheter or percutaneous medical device, and residence in a long-term care facility. These patients thus satisfied the international criteria for CA-MRSA presented by the Centers for Disease Control and Prevention (CDC). Unfortunately, the first case died of CA-MRSA pneumonia in spite of intensive treatments including anti-MRSA antibiotics. Unlike the severe CA-MRSA cases in western countries, Panton-Valentine leukocidin (PVL) genes were not detected in the present cases, suggesting that factors other than PVL had a significant effect on the severity of necrotizing pneumonia. To the best of our knowledge, this is the first report of severe necrotizing pneumonia caused by CA-MRSA in Japan.     

Usefulness of the restriction-modification test plus staphylococcal cassette chromosome mec types and Panton-Valentine leukocidin encoding phages to identify Staphylococcus aureus methicillin-resistant clones

We studied the usefulness of the restriction-modification (RM) test, staphylococcal cassette chromosome (SCC) mec types, and Panton-Valentine leukocidin (PVL)-encoding phages to identify Staphylococcus aureus methicillin-resistant lineages and to differentiate clones that belong to the same lineage. A total of 108 methicillin-resistant S. aureus (MRSA) strains were characterized by pulsed-field gel electrophoresis (PFGE)--multi-locus sequence typing (MLST)--spa-typing. The RM correctly identified the lineages CC5, CC8, CC30, and CC398, but not CC25 and CC72. The SCCmec and RM combined analysis allowed differentiation between MLST types within the same lineage. Only 5 MRSA strains were PVL-positive. Four PVL-positive USA300 isolates carried elongated-head type PVL-encoding phages, while the sequence type (ST)-30 strain carried an icosahedral-head phage. The combination of the RM test method, SCCmec types, and PVL phage identification could be useful for MRSA typing purposes.     

Staphylococcus aureus skin/soft-tissue infections: the impact of SCCmec type and Panton-Valentine leukocidin

We assessed the role of Panton-Valentine leukocidin (PVL) and SCCmec type in community associated (CA) and healthcare associated (HC) Staphylococcus aureus (SA) skin/soft-tissue infections (STI). We prospectively monitored microbiology results (11 January 2005 to 6 January 2006), screened inpatients with SA in tissue samples or blood, and selected adults with STI. We recorded clinical/microbiological characteristics, and tested saved isolates for PVL genes (real time PCR) and SCCmec type (conventional multiplex PCR). We encountered 204 patients. MRSA strains that accounted for 70.5% CA and 66.0% HC cases, caused more abscesses (55.7% vs 29.7%; p =0.001) and were often PVL-positive (68.9% vs 4.8%; p <0.001). PVL-positive isolates caused more abscesses (72.9% vs 26.5%; p <0.001) but similar bacteremia (7.3% vs 7.1%). SCCmec IVa made up 95.8% of PVL-positive strains and accounted for 69.8% of the abscesses. SCCmec II caused higher mortality (14.8% vs 0-3.1%; p = 0.02). PVL was a predictor of abscesses (p <0.001). Predictors of bacteremia were age > or = 65 y (p =0.004), necrotizing infection (p =0.014), and head/neck location (p =0.05). These findings suggest that SCCmec type and PVL status influence STI manifestations and contribute to MRSA-MSSA differences. PVL is implicated in abscess formation but not bacteremia. Bacteremia is likely related to host condition and/or other virulence factors that were not studied.     

Emergence of community-associated methicillin-resistant Staphylococcus aureus USA 300 clone as a cause of health care-associated infections among patients with prosthetic joint infections

BACKGROUND: Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) has emerged as an important cause of staphylococcal infections, but there have been little data on whether CA-MRSA causes health care-associated infections. METHODS: A case-control study was performed to identify risk factors for prosthetic joint infections (PJI). Antibiograms of isolates associated with PJI were reviewed. Molecular typing of available MRSA isolates was done using pulsed field gel electrophoresis (PFGE). Nares cultures of health care workers who provided care to those orthopedic patients were obtained. RESULTS: Over a 13-month period (January 2003-January 2004), 9.5% of patients with prosthetic hip (THA) or knee (TKA) joint surgery developed PJI (7 TKA and 2 THA). The mean time to development of PJI was 20 days. Five infections were caused by CA-MRSA and 3 by methicillin-susceptible S aureus; one was culture negative. All CA-MRSA isolates had identical antibiograms (resistant to beta-lactams and erythromycin; susceptible to clindamycin, trimethoprim-sulfamethoxazole, rifampin, gentamicin, levofloxacin, and vancomycin). Molecular typing of 2 available CA-MRSA isolates revealed that these were the USA300 clone; these isolates were PVL+ and carried SCCmec IV. CA-MRSA was not recovered from nares cultures from 31 health care workers. In multivariate analysis, TKA (OR, 8.1; 95% CI: 1.3-48.1) and surgery time >180 minutes (OR, 7.4; 95% CI: 1.4-39.6) were associated with PJI. CONCLUSION: We have demonstrated that the CA-MRSA USA300 clone is no longer just a cause of community-acquired infections but has also emerged as a cause of health care-associated infections, causing PJI at our institution.     

Infective Endocarditis from Furuncle with Meningitis Complication Caused by Methicillin-resistant Staphylococcus aureus

Infective endocarditis (IE) may be acquired in the community as community-acquired (CA) IE or in the healthcare setting. In Japan, cases of CA-methicillin-resistant Staphylococcus aureus (MRSA) infection as skin infection have been increasing. CA-MRSA strains, including the USA300 clone, have higher pathogenicity and are more destructive to tissue than healthcare-associated MRSA strains because of the toxins they produce, including arginine-catabolic mobile element (ACME) and Panton-Valentine leukocidin (PVL). However, only a few IE cases induced by USA300 have been reported. We herein report a 64-year-old man who developed CA-IE from a furuncle caused by USA300 MRSA producing PVL and ACME, which resulted in complications of meningitis.     

Panton-Valentine leukocidin is not a virulence determinant in murine models of community-associated methicillin-resistant Staphylococcus aureus disease

Increases in the incidence and severity of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections have spawned efforts to define unique virulence properties among prevalent strains. Panton-Valentine leukocidin (PVL), a pore-forming cytotoxin, has garnered attention because of its epidemiologic association with CA-MRSA. Using both the clinical isolate LAC, which is representative of the epidemic USA300 strain, and its isogenic PVL-negative strain in murine models of staphylococcal skin infection and pneumonia, we expanded upon recent studies by assessing the contribution of PVL in the genetic background of BALB/c mice. The data presented in this report support the observation that PVL does not contribute to the pathogenesis of staphylococcal infection of mice.     

Community acquired MRSA infections in a paediatric population in Greece

We investigated the characteristics of 20 community acquired methicillin resistant Staphylococcus aureus (MRSA) strains isolated in a paediatric hospital in Athens. Eighteen of these, all isolated from skin and soft tissue infections, carried the Panton-Valentine leukocidin (PVL) determinants. They all were found resistant to fusidic acid, tetracycline and kanamycin, and displayed a PFGE pattern identical to that of the well-described ST80 CA-MRSA clone circulating in various European countries.     

Genetic characterization of Staphylococcus aureus isolates carrying Panton-Valentine leukocidin genes in Bangladesh

To assess the spread and genetic characteristics of Panton-Valentine leukocidin (PVL) gene-carrying Staphylococcus aureus in Bangladesh, we investigated 59 strains (49 isolates from clinical specimens and 10 isolates colonized in the nasal cavities of medical staff), including 26 methicillin-resistant S. aureus (MRSA) strains. The PVL gene was detected only in methicillin-susceptible S. aureus (MSSA) strains (7 clinical strains and 2 colonizing strains). PVL gene-positive MSSA strains were found to belong to coagulase serotypes III or VI and were classified into sequence types ST88 (CC88), ST772, and ST573 (CC1) by multilocus sequence typing, and agr types 2 or 3. These types were different from those determined for MRSA (coagulase serotypes I and IV, ST240 and ST361, and agr type 1). PVL gene-positive MSSA possessed a larger number of virulence factor genes than MRSA, although they were susceptible to more antimicrobials. These findings suggest that the PVL gene is distributed to limited populations of S. aureus clones with specific genetic traits that are distinct from MRSA in Bangladesh, but genetically close to CA-MRSA clones in the CC1 lineage reported in the United States and European countries.     

Community-onset methicillin-resistant Staphylococcus aureus associated with antibiotic use and the cytotoxin Panton-Valentine leukocidin during a furunculosis outbreak in rural Alaska

BACKGROUND: Community-onset methicillin-resistant Staphylococcus aureus (CO-MRSA) reports are increasing, and infections often involve soft tissue. During a CO-MRSA skin infection outbreak in Alaska, we assessed risk factors for disease and whether a virulence factor, Panton-Valentine leukocidin (PVL), could account for the high rates of MRSA skin infection in this region. METHODS: We conducted S. aureus surveillance in the outbreak region and a case-control study in 1 community, comparing 34 case patients with MRSA skin infection with 94 control subjects. An assessment of traditional saunas was performed. S. aureus isolates from regional surveillance were screened for PVL genes by use of polymerase chain reaction, and isolate relatedness was determined by use of pulsed-field gel electrophoresis (PFGE). RESULTS: Case patients received more antibiotic courses during the 12 months before the outbreak than did control subjects (median, 4 vs. 2 courses; P=.01) and were more likely to use MRSA-colonized saunas than were control subjects (44% vs. 13%; age-adjusted odds ratio, 4.6; 95% confidence interval, 1.7-12). The PVL genes were present in 110 (97%) of 113 MRSA isolates, compared with 0 of 81 methicillin-susceptible S. aureus isolates (P<.001). The majority of MRSA isolates were closely related by PFGE. CONCLUSION: Selective antibiotic pressure for drug-resistant strains carrying PVL may have led to the emergence and spread of CO-MRSA in rural Alaska.     

Comparative analysis of USA300 virulence determinants in a rabbit model of skin and soft tissue infection

Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections are frequently associated with strains harboring genes encoding Panton-Valentine leukocidin (PVL). The role of PVL in the success of the epidemic CA-MRSA strain USA300 remains unknown. Here we developed a skin and soft tissue infection model in rabbits to test the hypothesis that PVL contributes to USA300 pathogenesis and compare it with well-established virulence determinants: alpha-hemolysin (Hla), phenol-soluble modulin-alpha peptides (PSMα), and accessory gene regulator (Agr). The data indicate that Hla, PSMα, and Agr contribute to the pathogenesis of USA300 skin infections in rabbits, whereas a role for PVL could not be detected.     

Involvement of Panton-Valentine leukocidin-producing Staphylococcus aureus in primary skin infections and pneumonia.

Panton-Valentine leukocidin (PVL) is a cytotoxin that causes leukocyte destruction and tissue necrosis. It is produced by fewer than 5% of Staphylococcus aureus strains. A collection of 172 S. aureus strains were screened for PVL genes by polymerase chain reaction amplification. PVL genes were detected in 93% of strains associated with furunculosis and in 85% of those associated with severe necrotic hemorrhagic pneumonia (all community-acquired). They were detected in 55% of cellulitis strains, 50% of cutaneous abscess strains, 23% of osteomyelitis strains, and 13% of finger-pulp-infection strains. PVL genes were not detected in strains responsible for other infections, such as infective endocarditis, mediastinitis, hospital-acquired pneumonia, urinary tract infection, and enterocolitis, or in those associated with toxic-shock syndrome. It thus appears that PVL is mainly associated with necrotic lesions involving the skin or mucosa.     

Is Panton-Valentine leukocidin the major virulence determinant in community-associated methicillin-resistant Staphylococcus aureus disease?

Methicillin-resistant Staphylococcus aureus (MRSA) remains a major problem in hospitals, and it is now spreading in the community. A single toxin, Panton-Valentine leukocidin (PVL), has been linked by epidemiological studies to community-associated MRSA (CA-MRSA) disease. However, the role that PVL plays in the pathogenesis of CA-MRSA has not been tested directly. To that end, we used mouse infection models to compare the virulence of PVL-positive with that of PVL-negative CA-MRSA representing the leading disease-causing strains. Unexpectedly, strains lacking PVL were as virulent in mouse sepsis and abscess models as those containing the leukotoxin. Isogenic PVL-negative (lukS/F-PV knockout) strains of USA300 and USA400 were as lethal as wild-type strains in a sepsis model, and they caused comparable skin disease. Moreover, lysis of human neutrophils and pathogen survival after phagocytosis were similar between wild-type and mutant strains. Although the toxin may be a highly linked epidemiological marker for CA-MRSA strains, we conclude that PVL is not the major virulence determinant of CA-MRSA.