Impact of agr dysfunction on virulence profiles and infections associated with a novel methicillin-resistant Staphylococcus aureus (MRSA) variant of the lineage ST1-SCCmec IV.

BACKGROUND: A novel variant of the ST1-SCCmecIV methicillin-resistant Staphylococcus aureus (MRSA) lineage, mostly associated with nosocomial bloodstream infections (BSI), has emerged in Rio de Janeiro. Bacterial biofilm has been considered a major virulence factor in central venous catheter-associated BSI. The mechanisms involved in biofilm formation/accumulation are multifactorial and complex. Studies have suggested that biofilm production was affected in vitro and vivo for agr-null mutants of S. aureus. RESULTS: The impact of naturally occurring inhibition of agr signaling on virulence profiles and infections associated with the ST1 variant was investigated. agr dysfunction was detected in a significant percentage (13%) of the isolates with concomitant increase in biofilm accumulation in vitro and in vivo, and enhanced ability to adhere to and invade airway cells. The biofilm formed by these ST1 isolates was ica-independent and proteinaceous in nature. In fact, the improved colonization properties were paralleled by an increased expression of the biofilm-associated genes fnbA, spa and sasG. The transcription of sarA, a positive regulator of agr, was two-times reduced for the agr-dysfunctional MRSA. Remarkably, the agr inhibition was genetically stable. Indeed, agr-dysfunctional isolates succeed to colonize and cause both acute and chronic infections in hospitalized patients, and also to effectively accumulate biofilm in a mouse subcutaneous catheter implant model. CONCLUSION: The ability of agr-dysfunctional isolates to cause infections in humans and to form biofilm in the animal model suggests that therapeutic approaches based on agr-inactivation strategies are unlikely to be effective in controlling human-device infections caused by ST1 isolates. The increased biofilm accumulation associated with the acquisition of multiple antimicrobial resistant traits might have influenced (at least in part) the expansion of this USA400 related clone in our hospitals.

Heteroduplex mobility assay and single-stranded conformation polymorphism analysis as methodologies for detecting variants of human erythroviruses.

Variant samples from the three genotypes of erythroviruses have already been detected using sequencing as methodology for analysis. This study aimed to investigate the efficacy of single-stranded conformation polymorphism (SSCP) analysis and heteroduplex mobility assay (HMA) as methodologies to detect human erythrovirus variants, using their VP1 unique region sequences. Clinical samples and plasmids of PVBAUA, A6, LaLi, V9Gh3051, and D91.1 erythrovirus variants as prototypes of the three genotypes were used. SSCP analysis was able to distinguish all divergences among the plasmids, including the two mutation points between LaLi and A6 plasmids that led to distinct electrophoresis mobility patterns. Although HMA analysis was unabled to detect two mutation points between LaLi and A6, it enabled the differentiation among all other plasmids that revealed specific electrophoresis patterns, with high-enough sensibility to detect 1.5% nucleotide substitutions. When 57 clinical samples were analyzed, 33 of them presented an identical pattern to PVBAUA by HMA and SSCP analyses, two of them were sequenced and presented an identical sequence in relation to PVBAUA. Another pattern was found for 21 samples. Among these, two samples were sequenced, revealing one mutation point in relation to PVBAUA, while each one of the three remaining samples presented a distinct pattern, showing two or three mutations in relation to PVBAUA by sequencing. HMA and SSCP analyses were suggested as methodologies suited for detecting genetic mutations of human erythroviruses in developing countries because of their practicability and minor costs for reagents and equipment.