Fabienne Antunes Ferreira

First Report of Infection with Community-Acquired Methicillin-Resistant Staphylococcus aureus in South America

ABSTRACT Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) has recently emerged in the southwestern Pacific, North America, and Europe. These S. aureus isolates frequently shared some genetic characteristics, including the SCCmec type IV and lukS-lukF genes. In this paper we show that typical CA-MRSA isolates have spread to South America (Brazil).

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

BackgroundA 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.ResultsThe 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.ConclusionThe 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.

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.

Complete Genome Sequence of a Variant of the Methicillin-Resistant Staphylococcus aureus ST239 Lineage, Strain BMB9393, Displaying Superior Ability To Accumulate ica-Independent Biofilm

ABSTRACT Biofilm is considered an important virulence factor in nosocomial infections. Herein, we report the complete genome sequence of a variant of methicillin-resistant Staphylococcus aureus, strain BMB9393, which is highly disseminated in Brazil. This strain belongs to the lineage ST239 and displays increased ability to accumulate ica-independent biofilm and to invade human epithelial cells.

Comparison of in vitro and in vivo systems to study ica-independent Staphylococcus aureus biofilms.

The ability of Staphylococcus aureus to form biofilms is considered an important factor in the pathogenesis of central venous catheter-related bacteremia and infections associated with the use of medical prostheses. Different methods have been described for assessing staphylococcal biofilms, but few comparative studies have been attempted to evaluate these techniques; especially related to ica-independent biofilm formation/accumulation. In this study we compared some in vitro and in vivo techniques to evaluate ica-independent biofilms produced by methicillin-resistant S. aureus. We observed that biofilms formed on human fibronectin-covered surfaces were about three times higher than those produced on inert polystyrene surfaces. However, despite the difference in absolute values, a linear correlation was detected between these two models. We also found that biofilms formed on polystyrene or polyurethane surfaces treated with human serum were easily detachable during washing and staining processes. The mouse model of subcutaneous foreign body showed good correlation with the in vitro techniques using either inert polystyrene or solid-phase fibronectin. Thus, our data showed that the microtiter-plate-based spectrophotometric assay is an appropriate method for preliminary biofilm investigations, mainly when a large number of isolates, mutants or systems need to be tested.

The multifaceted resources and microevolution of the successful human and animal pathogen methicillin-resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most important bacterial pathogens based on its incidence and the severity of its associated infections. In addition, severe MRSA infections can occur in hospitalised patients or healthy individuals from the community. Studies have shown the infiltration of MRSA isolates of community origin into hospitals and variants of hospital-associated MRSA have caused infections in the community. These rapid epidemiological changes represent a challenge for the molecular characterisation of such bacteria as a hospital or community-acquired pathogen. To efficiently control the spread of MRSA, it is important to promptly detect the mecA gene, which is the determinant of methicillin resistance, using a polymerase chain reaction-based test or other rapidly and accurate methods that detect the mecA product penicillin-binding protein (PBP)2a or PBP2’. The recent emergence of MRSA isolates that harbour a mecA allotype, i.e., the mecC gene, infecting animals and humans has raised an additional and significant issue regarding MRSA laboratory detection. Antimicrobial drugs for MRSA therapy are becoming depleted and vancomycin is still the main choice in many cases. In this review, we present an overview of MRSA infections in community and healthcare settings with focus on recent changes in the global epidemiology, with special reference to the MRSA picture in Brazil.

An evaluation of matrix-assisted laser desorption ionization time-of-flight mass spectrometry for the identification of Staphylococcus pseudintermedius isolates from canine infections

It has been proposed, based on taxonomic and molecular studies, that all canine isolates belonging to Staphylococcus intermedius group (SIG) should be renamed Staphylococcus pseudintermedius. However, isolates of SIG and other coagulase-positive staphylococci share many phenotypic characteristics, which could lead to misidentification. The accuracy of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) for identifying S. pseudintermedius isolates obtained from canine infections was evaluated, using a polymerase chain reaction (PCR)-based identification as the gold standard. In addition, MALDI-TOF MS was compared with conventional biochemical tests. A central problem was the incorrect identification of S. pseudintermedius isolates as S. intermedius by either MALDI-TOF MS or biochemical identification. From the 49 S. pseudintermedius isolates identified by the molecular method, only 21 could be assigned to this species by the biochemical approach and only 12 by MALDI-TOF MS. The 6 S. aureus isolates were correctly identified by all 3 techniques. However, using biochemical tests, 9 S. pseudintermedius were mistakenly classified as S. aureus, indicating a reduced specificity relative to the MALDI-TOF MS system. Analysis with the MALDI-TOF MS platform allowed rapid and accurate identification of the 49 isolates to the S. intermedius group but the approach was very limited in identifying S. pseudintermedius isolates, as only 12 of 49 isolates were correctly identified, a sensitivity of 0.24 (95% confidence interval: 0.13–0.39).

The influence of different factors including fnbA and mecA expression on biofilm formed by MRSA clinical isolates with different genetic backgrounds

Biofilm formation is considered an important virulence factor in implanted device-associated infections caused by methicillin-resistant Staphylococcus aureus (MRSA). Recent studies demonstrated that the ica-independent biofilms produced by MRSA are multifactorial. Despite the recent progress achieved in this field, the bacterial factors associated with biofilm formation/accumulation and regulation among clinical MRSA isolates remain largely unknown. In this study, using MRSA isolates from diverse multilocus sequence typing (MLST) clonal complexes that produce different amounts of biofilm, and a number of phenotypic and molecular approaches, we investigated the correlation between biofilm-associated factors and the ability of the bacteria to accumulate biofilm.

Comparison of different methods for detecting methicillin resistance in MRSA isolates belonging to international lineages commonly isolated in the American continent

The aim of the present paper was to compare different methods for detecting methicillin resistance in Staphylococcus aureus. Among the isolates analyzed, 52 belonged to MRSA international lineages commonly detected in the American continent and 14 to sporadic MRSA clones. Both 30 μg‐cefoxitin disk and PBP2a had 100% sensibility/specificity when the low‐level heterogeneous isolates were tested and, thus, are highly recommended.

Complete genome sequence of the MRSA isolate HC1335 from ST239 lineage displaying a truncated AgrC histidine kinase receptor

Methicillin-resistant Staphylococcus aureus (MRSA) is still one of the most important hospital pathogen globally. The multiresistant isolates of the ST239-SCCmecIII lineage are spread over large geographic regions, colonizing and infecting hospital patients in virtually all continents. The balance between fitness (adaptability) and virulence potential is likely to represent an important issue in the clonal shift dynamics leading the success of some specific MRSA clones over another. The accessory gene regulator (agr) is the master quorum sensing system of staphylococci playing a role in the global regulation of key virulence factors. Consequently, agr inactivation in S. aureus may represent a significant mechanism of genetic variability in the adaptation of this healthcare-associated pathogen. We report here the complete genome sequence of the methicillin-resistant S. aureus, isolate HC1335, a variant of the ST239 lineage, which presents a natural insertion of an IS256 transposase element in the agrC gene encoding AgrC histidine kinase receptor.