In vitro production of panton-valentine leukocidin among strains of methicillin-resistant Staphylococcus aureus causing diverse infections.
ABSTRACT Community-acquired methicillin-resistant Staphylococcus aureus strains have recently been associated with severe necrotizing infections. Greater than 75% of these strains carry the genes for Panton-Valentine leukocidin (PVL), suggesting that this toxin may mediate these severe infections. However, to date, studies have not provided evidence of toxin production.
Twenty-nine community-acquired methicillin-resistant Staphylococcus aureus and 2 community-acquired methicillin-susceptible S. aureus strains were collected from patients with infections of varying severity. Strains were analyzed for the presence of lukF-PV and SCCmecA type. PVL production in lukF-PV gene-positive strains was measured by ELISA, and the amount produced was analyzed relative to severity of infection.
Only 2 of the 31 strains tested, 1 methicillin-resistant Staphylococcus aureus abscess isolate and 1 nasal carriage methicillin-susceptible S. aureus isolate, were lukF-PV negative. All methicillin-resistant Staphylococcus aureus strains were SCCmec type IV. PVL was produced by all strains harboring lukF-PV, although a marked strain-to-strain variation was observed. Twenty-six (90%) of 29 strains produced 50-350 ng/mL of PVL; the remaining strains produced PVL in excess of 500 ng/mL. The quantity of PVL produced in vitro did not correlate with severity of infection.
Although PVL likely plays an important role in the pathogenesis of these infections, its mere presence is not solely responsible for the increased severity. Factors that up-regulate toxin synthesis in vivo could contribute to more-severe disease and worse outcomes in patients with community-acquired methicillin-resistant Staphylococcus aureus infection.
Article: Variations in amount of TSST-1 produced by clinical methicillin resistant Staphylococcus aureus (MRSA) isolates and allelic variation in accessory gene regulator (agr) locus.[show abstract] [hide abstract]
ABSTRACT: Staphylococcus aureus (S. aureus) is an important pathogen associated with both nosocomial and community-acquired infections and its pathogenicity is attributed to its potential to produce virulence factors. Since the amount of toxin produced is related to virulence, evaluating toxin production should be useful for controlling S. aureus infection. We previously found that some strains produce relatively large amounts of TSST-1; however, no reports have described the amount of TSST-1 produced by clinical isolates. Amounts of TSST-1 produced by clinical methicillin resistant S. aureus (MRSA) isolates were measured by Western blotting. We determined their accessory gene regulator (agr) class by PCR and investigated whether TSST-1 production correlates with variations in the class and structure of the agr. We found that 75% of surveyed MRSA isolates (n = 152) possessed the tst gene and that 96.7% belonged to agr class 2. The concentrations of TSST-1 secreted into culture supernatants by 34 strains measured by Western blotting differed 170-fold. Sequencing the entire agr locus (n = 9) revealed that some had allelic variations regardless of the amount of TSST-1 produced whereas sequencing the sar, sigma factor B and the tst promoter region revealed no significant changes. The amounts of TSST-1 produced by clinical MRSA isolates varied. The present results suggest that TSST-1 production is not directly associated with the agr structure, but is instead controlled by unknown transcriptional/translational regulatory systems, or synthesized by multiple regulatory mechanisms that are interlinked in a complex manner.BMC Microbiology 04/2009; 9:52. · 3.04 Impact Factor
Article: Rapid detection of Staphylococcus aureus Panton-Valentine leukocidin in clinical specimens by enzyme-linked immunosorbent assay and immunochromatographic tests.[show abstract] [hide abstract]
ABSTRACT: Staphylococcus aureus strains producing Panton-Valentine leukocidin (PVL) have been epidemiologically linked to specific human infections. To evaluate immunological tests that may be used to diagnose infections with PVL-producing strains, we prospectively collected pus, respiratory tract specimens, and joint fluid specimens from which S. aureus had been isolated in clinical laboratories in six countries. An enzyme-linked immunosorbent assay (ELISA) and an immunochromatographic test (ICT) targeting LukS-PV were performed directly with clinical samples for the detection of PVL. The same tests were applied to S. aureus culture supernatants. The corresponding S. aureus isolates were characterized by PCR for the presence of the PVL locus (lukS-PV and lukF-PV) and the mecA gene. A total of 185 samples from 144 skin infections, 23 bone and joint infections, and 18 lower respiratory tract infections were analyzed. By PCR, 72/185 S. aureus isolates were PVL locus positive (PVL(+)); 28 of these were also mecA positive. PVL was detected in the supernatants of all PVL(+) strains by both ELISA and an ICT, while no signal was observed with PVL-negative strains. The PVL concentrations in human clinical samples that grew PVL(+) strains ranged from 0 to 399 microg/ml by ELISA. By the use of 0.015 microg/ml of PVL as a cutoff value, PVL was detected in 65/72 (90%) of the clinical samples by ELISA. The sensitivity and specificity of the ELISA test were 90% and 100%, respectively. By the ICT, PVL was detected in 57/72 (79%) of the samples, and the sensitivity and specificity of ICT were 79% and 100%, respectively. PVL is expressed by S. aureus during human infection, and a PVL-specific ELISA and ICT could be reliable tests for the diagnosis of infections caused by PVL-producing strains.Journal of clinical microbiology 04/2010; 48(4):1384-90. · 4.16 Impact Factor
Article: Community-associated methicillin-resistant Staphylococcus aureus infections in a pediatric intensive care unit.[show abstract] [hide abstract]
ABSTRACT: Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infection is an increasing problem worldwide. In developing countries, there is little data on CA-MRSA infection in children. This study reviewed the clinical features and outcomes of children admitted in a Tunisian pediatric intensive care unit with severe CA-MRSA infections. Retrospective chart review of patients coded for CA-MRSA over 10 years. There were 14 (0.32% of all admissions) patients identified with severe CA-MRSA infections. The median age was three months (range, 0.5-156 months). All patients had pulmonary involvement. Six children (42.8%) developed septic shock. Two (14.3%) patients had multifocal infection with deep venous thrombosis. Two (14.3%) patients died. Severe CA-MRSA pneumonia dominated presentation. The mortality of CA-MRSA infection in our series is lower than that previously reported.The Journal of Infection in Developing Countries 01/2011; 5(8):587-91. · 1.19 Impact Factor