Robson Souza Leão

Achromobacter xylosoxidans: Characterization of Strains in Brazilian Cystic Fibrosis Patients

ABSTRACT We investigated the possibility of cross-infection among cystic fibrosis patients in two Brazilian reference centers. Achromobacter xylosoxidans isolates (n = 122) were recovered over a 5-year period from 39 patients. Isolates were genetically heterogeneous, but one genotype was present in 56% of the patients, suggesting that cross-infection may have occurred.

Burkholderia cenocepacia, B. multivorans, B. ambifaria and B. vietnamiensis isolates from cystic fibrosis patients have different profiles of exoenzyme production

Knowledge about the virulence mechanisms of species from the Burkholderia cepacia complex (BCC) is still limited. The genomovar heterogeneity and production of different virulence factors are likely to contribute to the variation in the clinical outcome observed in BCC‐infected cystic fibrosis (CF) patients. Therefore, in this study we investigated the genetic polimorphism, the presence of genetic makers associated with virulence and transmissibility in BCC, and the profile of exoenzyme production of 59 BCC isolates obtained from 59 CF patients attending the reference CF centre in Rio de Janeiro, Brazil. The DNA sequence analyses of the recA gene allowed us to identify 40 of these 59 BCC species as being B. cenocepacia, 9 as B. vietnamiensis, 6 as B. multivorans and 4 as B. ambifaria. The assessment of the bacterial genetic polymorphism by PFGE revealed that B. cenocepacia and the B. multivorans isolates belonged to four and two different PFGE profiles with prevalence of two clones, A and B, respectively. All B. vietnamiensis and B. ambifaria belonged to only one PFGE profile (J and E, respectively). None of the isolates exhibited the genetic markers cblA and BCESM, assessed by polymerase chain reaction. In contrast, the profile of enzymatic activity, assessed by phenotypic methods, differed among the BCC species: protease activity was detected only in B. cenocepacia and B. ambifaria isolates, whereas only B. vietnamiensis isolates produced hemolysin. Although the phospholipase C activity was similar among the different species, the level of lipase activity produced by B. multivorans was higher than in the other species. We speculate that the differential characteristics of exoenzyme production may account for the differences in the pathogenic potentials of each BCC species.

ExoU-induced vascular hyperpermeability and platelet activation in the course of experimental Pseudomonas aeruginosa pneumosepsis.

To address the question whether ExoU, a Pseudomonas aeruginosa cytotoxin with phospholipase A2 activity, can induce hemostatic abnormalities during the course of pneumosepsis, mice were instilled i.t. with the ExoU-producing PA103 P. aeruginosa or with a mutant obtained by deletion of the exoU gene. Control animals were instilled with sterile vehicle. To assess the role of ExoU in animal survival, mice were evaluated for 72 h. In all the other experiments, animals were studied at 24 h after infection. PA103-infected mice showed significantly higher mortality rate, lower blood leukocyte concentration, and higher platelet concentration and hematocrit than animals infected with the bacterial mutant, as well as evidences of increased vascular permeability and plasma leakage, which were confirmed by our finding of higher protein concentration in bronchoalveolar lavage fluids and by the Evans blue dye assay. Platelets from PA103-infected mice demonstrated features of activation, assessed by the flow cytometric detection of higher percentage of P-selectin expression and of platelet-derived microparticles as well as by the enzyme immunoassay detection of increased thromboxane A2 concentration in animal plasma. Histopathology of lung and kidney sections from PA103-infected mice exhibited evidences of thrombus formation that were not detected in sections of animals from the other groups. Our results demonstrate the ability of ExoU to induce vascular hyperpermeability, platelet activation, and thrombus formation during P. aeruginosa pneumosepsis, and we speculate that this ability may contribute to the reported poor outcome of patients with severe infection by ExoU-producing P. aeruginosa.

Escherichia coli producing KPC-2 carbapenemase: first report in Brazil☆

Carbapenem resistance in Escherichia coli isolates is rare; resistance has usually been attributed to the presence of an AmpC β-lactamase in association with the loss of porins (Poirel et al., 2004). Klebsiella pneumoniae carbapenemase (KPC)-producing E. coli have been described in a few cases, involving KPC-2 and KPC-3 acquisition in the United States (Bratu et al., 2007; Deshpande et al., 2006; Hong et al, 2005) and KPC-2 in Israel (Navon-Venezia et al., 2006) and China (Cai et al., 2008). KPC enzymes have recently been described in Brazil in Klebsiella pneumoniae (Monteiro et al., 2009; Pavez et al., 2009; Peirano et al., 2009) and Enterobacter cloacae (Zavascki et al., 2009). This study is the first reported description of the presence of KPC-2 in E. coli isolates in Brazil. Since May 2008, in the University Hospital Pedro Ernesto, a 600-bed teaching hospital located in Rio de Janeiro City, Rio de Janeiro State, has isolated a KPC-2– producing K. pneumoniae clone from different patients hospitalized in different wards (personal communication). In an effort to curb this trend, we have begun routine screening of all Enterobacteriaceae isolates with carbapenem resistance or reduced carbapenem susceptibility for presence of carbapenemases by modified Hodge test (Anderson et al., 2007). From August 2008 to May 2009, we recovered 4 carbapenemase-positive E. coli isolates from postoperative fluid collection in nephrectomy (1 isolate), urine (1 isolate), and blood (2 isolates) from patients with different clinical backgrounds hospitalized in different wards. The isolates showed polymerase chain reactionpositive results by using blaKPC primers (Peirano et al., 2009), and amplicons sequencing revealed 100% of sequence identity with blaKPC-2. The main microbiologic findings for isolates are summarized in Table 1. According to the antimicrobial susceptibility testing by disk diffusion, all isolates exhibited resistance or reduced

First report of Paenibacillus cineris from a patient with cystic fibrosis

We describe the recovery of Paenibacillus cineris from a Brazilian cystic fibrosis (CF) patient. Paenibacillus spp. are mainly environmental organisms. Although the role that these species play in lung disease is not known, we highlight the importance of careful attention to unusual bacteria from respiratory secretions of CF patients.

Panton-Valentine leukocidin (PVL) gene carriage among Staphylococcus aureus strains with SCCmec types I, III, IV, and V recovered from cystic fibrosis pediatric patients in Brazil

The prevalence of methicillin-resistant Staphylococcus aureus (MRSA) is increasing in patients with cystic fibrosis (CF). We report a molecular characterization, antimicrobial resistance, and Panton-Valentine leukocidin (PVL) toxin gene detection of MRSA strains from 28 Brazilian pediatric CF patients (1 strain per patient). A significant proportion (50%) of MRSA SCCmec IV isolates was observed. Nearly half of MRSA strains harboring the PVL genes distributed in all SCCmec types detected. Multilocus sequence typing (MLST) analyses showed majority (57.1%) of the isolates belonged to known epidemic lineages, such as UK/EMRSA-3, Pediatric/USA 800, Southwest Pacific clone, and Brazilian/Hungarian clone. To our knowledge, this is the first Brazilian study of molecular epidemiology based on MLST and SCCmec typing and the first description of PVL genes in MRSA from CF patients.

KPC-2 Carbapenemase-producing Klebsiella pneumoniae isolates from patients with Cystic Fibrosis

Bacteria producing Klebsiella pneumoniae carbapenemases (KPCs) are rapidly emerging as a cause of multidrug-resistant infections worldwide. KPCs enzyme are plasmid-borne and can accumulate and transfer resistance determinants to other classes of antibiotics.We report two cases of KPC-2 carbapenemase-producing Klebsiella pneumoniae isolates from Cystic Fibrosis patients.

KPC‐2 producing Klebsiella pneumoniae and Escherichia coli co‐infection in a catheter‐related infection

We describe the first report of simultaneous blood infection with KPC-2 producing Klebsiella pneumoniae and Escherichia coli in a Brazilian patient. We highlight the importance of implementing efficient infection control measures to limit the spread of these phenotypes in a hospital setting.

Influence of biofilm formation in the susceptibility of Pseudomonas aeruginosa from Brazilian patients with cystic fibrosis.

Ferreira AG, Leão RS, Carvalho‐Assef APD, Folescu TW, Barth AL, Marques EA. Influence of biofilm formation in the susceptibility of Pseudomonas aeruginosa from Brazilian patients with cystic fibrosis. APMIS 2010; 118: 606–12.

Comparison of the worldwide transmissible Pseudomonas aeruginosa with isolates from brazilian cystic fibrosis patients

Cross-infection with Pseudomonas aeruginosa among cystic fibrosis (CF) patients is a rare occurrence. However, the emergence of transmissible strains has been reported between unrelated individuals. We analyzed the genetic relationship among P. aeruginosa isolates from Brazilian CF patients and transmissible clones which are worldwide spread. The data does not indicate the presence of closely related variant clones.