Lara M. de Almeida

Linezolid Resistance in Brazilian Staphylococcus hominis Strains Is Associated with L3 and 23S rRNA Ribosomal Mutations

Among different clinical staphylococcal strains, linezolid resistance has been mainly mediated by mutations in the central loop of domain V of 23S rRNA, with the G2576T mutation being the most prevalent.…

Identification of Staphylococcus aureus Carrying the mecA Gene in Ready-to-Eat Food Products Sold in Brazil

Since Staphylococcus aureus can cause several types of diseases, the development of antibiotic resistance poses an even greater threat to public health. S. aureus is known to possess the adaptive capability to promptly respond to antibiotics, making it resistant and increasingly difficult to treat; methicillin-resistant strains of S. aureus are a major concern with regard to this species. Previous studies reported the identification of methicillin-resistant S. aureus in food, demonstrating that this can represent a source of S. aureus which may carry the mecA gene. Fifty-seven S. aureus isolates, previously obtained from different types of food, were screened by polymerase chain reaction with specific primers for the mecA gene, which mediates methicillin resistance. Five (9%) isolates showed the presence of mecA gene, demonstrating that food may contain microorganisms possessing resistance genes. This study emphasizes the need to include food as a possible source of S. aureus carrying mecA gene and the need to monitor these products. Moreover, this is the first report of the presence of mecA genes in S. aureus isolated from ready-to-eat food in Brazil and Latin America.

Linezolid resistance in vancomycin-resistant Enterococcus faecalis and Enterococcus faecium isolates in a Brazilian hospital.

Linezolid resistance in vancomycin-resistant enterococci (VRE) strains has been rarely reported, with Enterococcus faecium being the species most commonly associated with these few cases ([1][1], [2][2], [3][3]). Here, we report infections due to linezolid- and vancomycin-resistant Enterococcus (

Novel sequence types (STs) of Staphylococcus aureus isolates causing clinical and subclinical mastitis in flocks of sheep in the northeast of Brazil.

Staphylococcus aureus is one of the most important infectious mastitis causative agents in small ruminants. In order to know the distribution of Staph. aureus strains associated with infectious mastitis in flocks of sheep in the northeast of Brazil and establish whether these clones are related to the strains distributed internationally, this study analysed the genetic diversity of Staph. aureus isolates from cases of clinical and subclinical mastitis in ewes by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). In this research, 135 ewes with mastitis from 31 sheep flocks distributed in 15 districts were examined. Staph. aureus was isolated from sheep milk in 9 (29%) out of 31 herds located in 47% of the districts surveyed. MLST analysis allowed the identification of four STs (ST750, ST1728, ST1729 and ST1730). The last three with their respective novel alleles (glp-220; pta-182 and yqil-180) were recently reported in the Staph. aureus MLST database (http://www.mlst.net). Each novel allele showed only a nucleotide different from those already described. The occurrence of CC133 (ST750 and ST1729) in this study is in agreement with other reports that only a few clones of Staph. aureus seem to be responsible for most cases of mastitis in dairy farms and that some of these clones may have broad geographic distribution. However, the prevalence of CC5 (ST1728 and ST1730)--an important group related to cases of colonization or infection in humans--differs from previous studies by its widespread occurrence and may suggest human contamination followed by selective pressures of the allelic diversifications presented for these STs.

Complete Genome Sequence of Lactococcus lactis Strain AI06, an Endophyte of the Amazonian Açaí Palm

ABSTRACT We report the genome, in a single chromosome, of Lactococcus lactis strain AI06, isolated from the mesocarp of the açaí fruit (Euterpe oleracea) in eastern Amazonia, Brazil. This strain is an endophyte of the açaí palm and also a component of the microbiota of the edible food product.

Comparative analysis of agr groups and virulence genes among subclinical and clinical mastitis Staphylococcus aureus isolates from sheep flocks of the Northeast of Brazil

Staphylococcus aureus is one of the most frequent mastitis causative agents in small ruminants. The expression of most virulence genes of S. aureus is controlled by an accessory gene regulator (agr) locus. This study aimed to ascertain the prevalence of the different agr groups and to evaluate the occurrence of encoding genes for cytotoxin, adhesins and toxins with superantigen activity in S. aureus isolates from milk of ewes with clinical and subclinical mastitis in sheep flocks raised for meat production The agr groups I and II were identified in both cases of clinical and subclinical mastitis. Neither the arg groups III and IV nor negative agr were found. The presence of cflA gene was identified in 100% of the isolates. The frequency of hla and lukE-D genes was high - 77.3 and 82.8%, respectively and all isolates from clinical mastitis presented these genes. The sec gene, either associated to tst gene or not, was identified only in isolates from subclinical mastitis. None of the following genes were identified: bbp, ebpS, cna, fnbB, icaA, icaD, bap, hlg, lukM-lukF-PV and se-a-b-d-e.

Clonal Dissemination of Linezolid-Resistant Staphylococcus haemolyticus Exhibiting the G2576T Mutation in the 23S rRNA Gene in a Tertiary Care Hospital in Brazil

Linezolid, the first oxazolidinone class agent to be introduced clinically, has broad activity against many important multidrug-resistant Gram-positive pathogens, including methicillinresistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE). Linezolid inhibits protein synthesis by interacting with 23S rRNA in the 50S ribosomal subunit, and it should interfere with the binding of aminoacyl-tRNA to the ribosomal A site in the bacterial ribosome (5). Although linezolid resistance is mediated by the cfr-encoded product (8) or by mutations in ribosomal proteins (6, 9), mutations in the central loop of domain V of 23S rRNA have been reported as the main mechanism associated with the expression of linezolid resistance among clinical staphylococcal strains (2, 3, 11). In this study, we report the first clonal dissemination of linezolid-resistant Staphylococcus haemolyticus strains with the G2576T mutation in 23S rRNA in Brazil. From March 2008 to January 2010, nine S. haemolyticus strains exhibiting high-level resistance to linezolid (MICs of 32 to 128 g/ml) were isolated from blood and catheter cultures from different inpatients in a tertiary care hospital. In this institution, linezolid was introduced in November 2000 and is currently the drug of choice for the treatment of VRE infections, which are highly prevalent. Identification testing was performed by the Vitek-2 system (bioMérieux, St. Louis, MO), and the susceptibility of bacterial strains was tested using disk diffusion and agar dilution methods according to the Clinical and Laboratory Standards Institute (CLSI) (1). Etests (AB Biodisk, Solna, Sweden) were used for oxacillin and vancomycin. The chromosomal DNA was obtained using the technique of phenol-chloroform extraction. The presence of the cfr gene and mutations in the domain V region of 23S rRNA and the rplC, rplD, and rplV genes were investigated as described previously (2, 4, 9, 10). The PCR products were sequenced and aligned with the corresponding nucleotide sequence from linezolid-susceptible S. haemolyticus strain JCSC1435 (GenBank accession no. AP006716.1). Pulsed-field gel electrophoresis (PFGE) typing was performed according to the method of Struelens et al. (12) with a modification. Bacterial DNA was digested with the restriction enzyme FastDigest SmaI (Fermentas Life Sciences, Canada). The G2576T mutation in the domain V region of 23S rRNA was identified in all strains except the linezolid-susceptible S. haemolyticus control strain, which was obtained from the same hospital. On the other hand, all strains showed the wild-type L3, L4, and L22 ribosomal proteins when compared to the linezolid-susceptible S. haemolyticus strain JCSC1435. Furthermore, the cfr gene was not identified in any isolate (Table 1). Cases of linezolid resistance in S. haemolyticus are still extremely rare and associated with the G2576T mutation in domain V of 23S rRNA (7, 11, 13). In our institution, linezolid-resistant S. haemolyticus strains with the G2576T mutation have been disseminated since 2008 and represent 1% of all S. haemolyticus isolates obtained during the period of this study, whereas linezolid-resistant Staphylococcus epidermidis strains represent 0.04% of total isolates of this species (2). No linezolid-resistant Staphylococcus aureus strain has been identified so far. Because these strains were collected over a 2-year period, it is possible to infer that these strains belong to an endemic clone of S. haemolyticus that has probably spread in our institution and become linezolid resistant under selective pressure. Although linezolid is potentially active against S. haemolyticus strains, reports of clinical S. haemolyticus

Environmental dissemination of vanA-containing Enterococcus faecium strains belonging to hospital-associated clonal lineages

Department of Clinical Analysis, School of Pharmacy, Universidade de São Paulo, São Paulo, Brazil; Bacteriology Branch, Instituto Adolfo Lutz, São Paulo, Brazil; Environmental Company of São Paulo State (CETESB), São Paulo, Brazil; Department of Infectious Diseases, Faculty of Medicine, Universidade de São Paulo, São Paulo, Brazil; Department of Microbiology, Institute of Biomedical Sciences, Universidade de São Paulo, São Paulo, Brazil

Molecular mechanisms of membrane impermeability in clinical isolates of Enterobacteriaceae exposed to imipenem selective pressure

Intrinsic mechanisms leading to carbapenem-induced membrane impermeability and multidrug resistance are poorly understood in clinical isolates of Enterobacteriaceae. In this study, molecular behaviours during the establishment of membrane impermeability in members of the Enterobacteriaceae family under imipenem selective pressure were investigated. Clinical isolates (n = 22) exhibiting susceptibility to multiple antibiotics or characterised as extended-spectrum β-lactamase (ESBL)- or AmpC-producers were submitted to progressive passages on Mueller-Hinton agar plates containing subclinical concentrations of imipenem [0.5 × the minimum inhibitory concentration (MIC)]. Changes in outer membrane permeability were evaluated by determination of antimicrobial MICs, sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE), and gene expression analysis related to membrane permeability (i.e. omp35-like, omp36-like and acrA) and regulatory mechanisms (i.e. marA and ompR) by quantitative reverse transcription PCR. Following imipenem induction, 73% of isolates showed increased carbapenem MICs by ≥2 doubling dilutions. At an early stage of treatment, imipenem modulated the expression of porins and efflux pump genes, represented by a reduction of 78% in omp36-like and a two-fold increase in acrA expression. Transcriptional factors marA and ompR were also affected by imipenem induction, increasing mRNA expression by 14- and 4-fold, respectively. High marA expression levels were associated with higher values of acrA expression. These results suggest that imipenem is an important factor in the development of an adaptive response to carbapenems by regulating key genes involved in the control of efflux pumps and porins, which could lead to a multidrug-resistant profile in clinical isolates, contributing to possible treatment failure.

Complete Genome Sequence of Staphylococcus aureus FCFHV36, a Methicillin-Resistant Strain Heterogeneously Resistant to Vancomycin

ABSTRACT We report here the sequence of the entire chromosome of Staphylococcus aureus strain FCFHV36, a methicillin-resistant strain heterogeneously intermediate to vancomycin, bearing a type II staphylococcal chromosome cassette mec element (SCCmec), belonging to multilocus sequence type (MLST) 105, and isolated from a vertebra of a patient with osteomyelitis.