Agnes Marie Sá Figueiredo

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).

The Predominant Variant of the Brazilian Epidemic Clonal Complex of Methicillin-Resistant Staphylococcus aureus Has an Enhanced Ability to Produce Biofilm and to Adhere to and Invade Airway Epithelial Cells

Methicillin-resistant Staphylococcus aureus (MRSA) has emerged as a therapeutic problem. In the present study, the molecular characterization by pulsed-field gel electrophoresis of MRSA isolates collected from a university hospital revealed that the predominant variant of the Brazilian epidemic clonal complex (BECC) was responsible for the increase in the incidence of MRSA strains, which reached 28% in 1998. It was verified that this predominant variant of the BECC displayed an enhanced ability to produce biofilm on inert polystyrene surfaces and to adhere to and invade epithelial airway cells. These results indicate that MRSA strains belonging to the BECC have evolved advantageous properties that might play a role in their predominance as international nosocomial pathogens.

Isolation and molecular characterization of methicillin- resistant coagulase-negative staphylococci from nasal flora of healthy humans at three community institutions in Rio de Janeiro City

We describe the isolation and molecular characterization of methicillin-resistant coagulase-negative staphylococci (MRCNS) from the nasal flora of healthy humans from three institutions located in Rio de Janeiro City. Swabs were obtained from the nares of students attending a non-residential public school and adults from two military quarters. Isolates of staphylococci were tested for the presence of the mecA gene by hybridization with a specific probe. S. epidermidis was the most frequent MRCNS (38 of the total 45 CNS isolated). Twenty-five percent of nasal staphylococcal carriers studied were colonized with MRCNS. Pulsed-field gel electrophoresis (PFGE) of SmaI-digested genomic DNA was carried out to study the clonality of the methicillin-resistant S. epidermidis (MRSE) isolates. In addition to cross-colonization among individuals belonging to the same institution, familial cross-colonization appeared to contribute to the spread of the methicillin-resistant isolates among two inter-communicable institutions. Indeed, the wide genomic diversity among the MRSE flora suggests that the spread of the mecA gene among these isolates might also have occurred via horizontal transmission. Despite the limited number of institutions analysed, it is reasonable to conclude that our data do not represent a situation unique to the three organizations but may reflect other communities in Rio with respect to transmission of MRCNS.

Spread of the Brazilian epidemic clone of a multiresistant MRSA in two cities in Argentina.

Methicillin-resistant Staphylococcus aureus (MRSA) is recognised as an important cause of nosocomial infection. The spread of some MRSA epidemic clones is well documented. In Brazil, and more recently in Portugal, a considerable number of hospital infections has been caused by a unique multiresistant MRSA clone designated as the Brazilian epidemic clone. This paper describes the spread of this clone in hospitals in two cities in Argentina.

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.

Isolation of methicillin-resistant coagulase-negative staphylococci from patients undergoing continuous ambulatory peritoneal dialysis (CAPD) and comparison of different molecular techniques for discriminating isolates of Staphylococcus epidermidis

Coagulase-negative staphylococci (CNS) have emerged as an important pathogen in nosocomial infections. About 80%-90% of CNS isolates associated with hospital infections are methicillin-resistant coagulase-negative staphylococci (MRCNS). The aims of this study were to screen for MRCNS isolates in the flora of a small population of patients undergoing continuous ambulatory peritoneal dialysis (CAPD) and to evaluate the discriminatory power of different molecular methods: pulsed-field gel electrophoresis (PFGE), mecA location, ClaI/mecA polymorphism and arbitrarily primed polymerase chain reaction (AP-PCR) for characterizing isolates of methicillin-resistant Staphylococcus epidermidis (MRSE). Seventy-nine CNS isolates were recovered from the 11 CAPD patients studied. Using a methicillin screening agar and a DNA specific mecA probe we verified that 30 of the 79 (38%) CNS isolates were resistant to methicillin (MRCNS). Twenty-two of the 30 MRCNS (73%) were MRSE, 7 (23%) methicillin-resistant S. haemolyticus (MRSH(ae)) and 1 (3%) methicillin-resistant S. hominis (MRSH(om)). All patients analyzed carried MRCNS in their flora, in one or more sites. Since CAPD patients have high risk for developing peritonitis, the colonization of these patients with MRCNS might represent an additional problem, due to the therapeutic restrictions imposed by these multiresistant isolates. A wide genetic diversity was verified when the PFGE of the MRSE isolates was analyzed. The 22 MRSE isolates displayed a total of 15 PFGE different patterns (11 PFGE types and 4 subtypes). The location of mecA in the SmaI-fragmented genome DNA did not bring any additional advantage for epidemiologic characterization of the isolates. The ClaI/mecA polymorphism was able to correctly discriminate 12 from the 15 PFGE patterns. In addition, the DNA of 20 MRSE isolates were used for AP-PCR typing. These isolates belonged to 14 PFGE patterns (11 types and 3 subtypes) and displayed 15 genotypes (for the association of PFGE, mecA location and ClaI/mecA polymorphism). A total of 17 different amplification patterns was verified using the primer 1. Only for 2 genotypes, strains having identical genetic backgrounds were further discriminated by AP-PCR (2 of 15 genotypes (87%) for AP-PCR and 1 of 15 genotypes for PFGE; (93%). Concluding, our results indicated that the AP-PCR can be an alternative and useful tool for monitoring and genotyping MRSE colonization and also to molecular characterizing MRSE outbreaks in hospitals.

Analysis of different molecular methods for typing methicillin-resistant Staphylococcus aureus isolates belonging to the Brazilian epidemic clone

The extensive geographic spread of MRSA isolates belonging to the Brazilian epidemic clone (BEC) limited the value of pulsed-field gel electrophoresis (PFGE) in epidemiological studies of outbreaks caused by these strains. Thus, the discriminatory power of eight different molecular methods was evaluated in an attempt to establish a methodology for genotyping BEC isolates involved in intra-hospital outbreaks. BEC isolates from five hospitals in Teresina City, Piaui State were genotyped by conventional electrophoresis or PFGE of Cla I- or Sma I-digested genomic DNA hybridised with specific labelled mecA, Tn554, IS257 and IS256 probes. The combination of PFGE with Cla I/mecA, Cla I/Tn554, Cla I/IS257, Sma I/mecA and Sma I/IS257 probe-fingerprinting techniques provided a very poor discriminatory power for BEC strains. Although Cla I/IS256 fingerprinting discriminated 17 different polymorphisms among the isolates displaying PFGE A1 pattern, this strategy was not reproducible. In contrast, the combination of PFGE and Sma I/IS256 polymorphisms differentiated BEC isolates into nine stable polymorphisms. Thus combination of PFGE and hybridisation with IS256 probe may be recommended as a useful means of typing BEC strains involved in intra-hospital infections.

Emergence of multiresistant variants of the community-acquired methicillin-resistant Staphylococcus aureus lineage ST1-SCCmecIV in 2 hospitals in Rio de Janeiro, Brazil.

Usually, community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) is susceptible to a variety of non-beta-lactam drugs. These isolates commonly display SCCmecIV and are associated with community-acquired infections. More recently, CA-MRSA has been isolated from health-care-associated diseases. We characterized MRSA isolates from 2 hospitals in Rio de Janeiro area to assess the entry of new lineages. The isolates were primary genotyped using a combination of molecular typing methods including SCCmec, restriction modification test, and Panton-Valentine leukocidin (PVL) detection. Pulsed-field gel electrophoresis was carried out for representatives of each lineages found. Disk diffusion test was performed as recommended by the Clinical and Laboratory Standards Institute. SCCmecIV was the predominant cassette mec detected. The most frequent MRSA lineage, a PVL nonproducer, was allocated in the CC1-SCCmecIV. It was found that 56% of these isolates were resistant to 3 or more non-beta-lactam drugs. Multilocus sequence typing of a representative of the CC1 isolates supported our finds that multiresistant variants of a CA-MRSA lineage (ST1-SCCmecIV) emerged in this city.

Molecular Characterization of Methicillin Resistant Staphylococcus aureus Strains Isolated from Intensive Care Units in Iran: ST22-SCCmec IV/t790 Emerges as the Major Clone.

Introduction The emergence of methicillin-resistant Staphylococcus aureus (MRSA) in different patient populations is a major public health concern. This study determined the prevalence and distribution of circulating molecular types of MRSA in hospitalized patients in ICU of hospitals in Tehran. Materials and Methods A total of 70 MRSA isolates were collected from patients in eight hospitals. Antimicrobial resistance patterns were determined using the disk diffusion method. The presence of toxin encoding genes and the vancomycin resistance gene were determined by PCR. The MRSA isolates were further analyzed using multi-locus sequence, spa, SCCmec, and agr typing. Results The MRSA prevalence was 93.3%. Antimicrobial susceptibility testing revealed a high resistance rate (97.1%) to ampicillin and penicillin. The rate of resistance to the majority of antibiotics tested was 30% to 71.4%. Two isolates belonging to the ST22-SCCmec IV/t790 clone (MIC ≥ 8 μg/ml) had intermediate resistance to vancomycin. The majority of MRSA isolates (24.3%) were associated with the ST22-SCCmec IV/t790 clone; the other MRSA clones were ST859-SCCmec IV/t969 (18.6%), ST239-SCCmec III/t037 (17.1%), and ST291-SCCmec IV/t030 (8.6%). Conclusions The circulating MRSA strains in Iranian hospitals were genetically diverse with a relatively high prevalence of the ST22-SCCmec IV/t790 clone. These findings support the need for future surveillance studies on MRSA to better elucidate the distribution of existing MRSA clones and detect emergence of new MRSA clones.