A unique SaeS allele overrides cell-density dependent expression of saeR and lukSF-PV in the ST30-SCCmecIV lineage of CA-MRSA.

ST30 (CC30)-SCCmec IV (USA1100) is one of the most common community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) lineages. ST30 isolates typically carry lukSF-PV genes encoding the Panton-Valentine leukocidin (PVL) and are responsible for outbreaks of invasive infections worldwide. In this study, twenty CC30 isolates were analyzed. All were very susceptible to non-ß-lactam antimicrobials, 18/20 harbored the lukSF-PV genes, only 1/20 exhibited agr-rnaIII dysfunction, and the majority was not able to form biofilm on inert surfaces. Analysis of lukSF-PV temporal regulation revealed that opposite to other CA-MRSA isolates, these genes were more highly expressed in early log phase than in stationary phase. This inverted lukSF-PV temporal expression was associated with a similar pattern of saeRS expression in the ST30 isolates, namely high level expression in log phase and reduced expression in stationary phase. Reduced saeRS expression in stationary phase was associated with low expression levels of the sae regulators, agr and agr-upregulator sarA, which activate the stationary phase sae-P1 promoter and overexpression of agr-RNAIII restored the levels of saeR and lukSF-PV trancripts in stationary phase. Altered SaeRS activity in the ST30 isolates was attributed to amino acid substitutions (N227S, E268K and S351T) in the HTPase_c domain of SaeS (termed SaeS(SKT)). Complementation of a USA300 saeS mutant with the saeS(SKT) and saeS alleles under the direction of the log phase sae-P3 promoter revealed that saeR and lukSF-PV transcription levels were more significantly activated by saeS(SKT) than saeS. In summary our data identify a unique saeS allele (saeS(SKT)) which appears to override cell-density dependent SaeR and PVL expression in ST30 CA-MRSA isolates. Further studies to determine the contribution of saeS(SKT) allele to the pathogenesis of infections caused by ST30 isolates are merited.

The antimicrobial susceptibility, biofilm formation and genotypic profiles of Staphylococcus haemolyticus from bloodstream infections.

We analysed the antimicrobial susceptibility, biofilm formation and genotypic profiles of 27 isolates of Staphylococcus haemolyticus obtained from the blood of 19 patients admitted to a hospital in Rio de Janeiro, Brazil. Our analysis revealed a clinical significance of 36.8% and a multi-resistance rate of 92.6% among these isolates. All but one isolate carried the mecA gene. The staphylococcal cassette chromosome mec type I was the most prevalent mec element detected (67%). Nevertheless, the isolates showed clonal diversity based on pulsed-field gel electrophoresis analysis. The ability to form biofilms was detected in 66% of the isolates studied. Surprisingly, no icaAD genes were found among the biofilm-producing isolates.

First report in South America of companion animal colonization by the USA1100 clone of community-acquired meticillin-resistant Staphylococcus aureus (ST30) and by the European clone of methicillin-resistant Staphylococcus pseudintermedius (ST71).

BACKGROUND: Methicillin-resistant staphylococci can colonize and cause diseases in companion animals. Unfortunately, few molecular studies have been carried out in Brazil and other countries with the aim of characterizing these isolates. Consequently, little is known about the potential role of companion animals in transmitting these resistant bacteria to humans. In this work we searched for mecA gene among Staphylococcus isolates obtained from nasal microbiota of 130 healthy dogs and cats attended in a veterinary clinic located in the west region of Rio de Janeiro. The isolates recovered were identified to the species level and characterized using molecular tools. RESULTS: A community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) isolate related to USA1100 (Southwest Pacific clone) and susceptible to all non-ß-lactams was detected in a cat (1.7%, 1/60). Another coagulase-positive isolate harboring mecA was recovered from a dog (1.4%, 1/70) and identified as Staphylococcus pseudintermedius (MRSP) related to the European clone (ST71). The two isolates of Staphylococcus conhii subsp. urealyticus (1.4%, 1/70 dogs and 1.7%, 1/60 cats), similarly to the MRSP isolate, also presented high-level multiresistance. The majority of the methicillin-resistant coagulase-negative staphylococci recovered were Staphylococcus saprophyticus (5.7%, 4/70 dogs and 6.7%, 4/60 cats) and all clustered into the same PFGE type. CONCLUSIONS: This work demonstrates that mecA-harboring Staphylococcus isolates are common members of the nasal microbiota of the healthy companion animals studied (9.2%, 12/130 animals), including some high-level multiresistant isolates of S. pseudintermedius and S. conhii subsp. urealyticus. The detection, for the first time in South America, of USA1100-related CA-MRSA and of ST71 MRSP (European clone), colonizing companion animals, is of concern. Both S. pseudintermedius and S. aureus are important agents of infections for animals. The USA1100 CA-MRSA is a causative of severe and disseminated diseases in healthy children and adults. Additionally, MRSP is a nosocomial pathogen in veterinarian settings. It had already been demonstrated that the virulent ST71 MRSP is geographically spread over Europe and USA, with potential for zoonotic infections.

agr RNAIII divergently regulates glucose-induced biofilm formation in clinical isolates of Staphylococcus aureus.

Staphylococcus aureus is an important nosocomial and community-acquired pathogen. Hospital infections are frequently complicated by the ability of bacteria to form biofilms on different surfaces. The development of bacterial films on medical indwelling devices, such as prostheses, often requires surgical procedures to remove the contaminated implant. Indeed, biofilm formation on central endovenous catheters is a major cause of primary bacteraemia in hospitals. The modulation of virulence factors in S. aureus is orchestrated by a number of global regulators including agr RNAIII. To improve our understanding of the role of the agr quorum-sensing system in biofilm formation by S. aureus, we constructed a number of agr-null mutants, derived from contemporary clinical isolates. Analysis of these mutants indicates that agr has a significant impact on biofilm development for most of the isolates tested. Our data show that RNAIII can control both biofilm formation and accumulation. The agr effect included both up- and downregulation of biofilms, even for isolates within the same lineage, corroborating the hypothesis that the mechanisms involved in S. aureus biofilms are complex and probably multifactorial.

Commensal isolates of methicillin-resistant Staphylococcus epidermidis are also well equipped to produce biofilm on polystyrene surfaces.

OBJECTIVES: To study biofilm production and to detect icaAD, atlE and aap genes in 137 isolates of methicillin-resistant Staphylococcus epidermidis (MRSE) obtained from healthy individuals from the community (35 isolates), from hospitalized patients at the Antônio Pedro University Hospital (25 isolates) and from individuals from a home-care system (HCS; 77 isolates). METHODS: Biofilm production was determined in vitro using polystyrene inert surfaces. icaAD, atlE and aap genes were detected using PCR. Hybridization experiments were also carried out to confirm some PCR results. Antimicrobial susceptibility testing was carried out using the NCCLS methods. RESULTS: Although many of the commensal MRSE isolates produced biofilms, the percentage of biofilm producers was significantly higher (P = 0.0107) among hospital isolates (76%) than among isolates from the community (60%) and from the HCS (57%). An association was observed between multiresistance and biofilm production for isolates obtained from healthy individuals from the community and from household contacts from the HCS (P < 0.0001). The concomitant presence of the ica operon and atlE and aap genes was associated with the strong biofilm-producer phenotype (P < 0.0001). CONCLUSION: Because many of the commensal MRSE isolates obtained from nares produced biofilms and carried icaAD, aap and atlE genes, biofilms or such genetic elements should not be used as markers for clinical significance. The biofilm environment seems to increase genetic exchanges and hence may contribute to multiresistance phenotypes.