Cristiane Silveira de Brito

Nosocomial infections in a Brazilian neonatal intensive care unit: a 4-year surveillance study

INTRODUCTION Report the incidence of nosocomial infections, causative microorganisms, risk factors associated with and antimicrobial susceptibility pattern in the NICU of the Uberlândia University Hospital. METHODS Data were collected through the National Healthcare Safety Network surveillance from January 2006 to December 2009. The patients were followed five times/week from their birth to their discharge or death. RESULTS The study included 1,443 patients, 209 of these developed NIs, totaling 293 NI episodes, principally bloodstream infections (203; 69.3%) and conjunctivitis (52; 17.7%). Device-associated infection rates were as follows: 17.3 primary bloodstream infections per 1,000 central line-days and 3.2 pneumonias per 1000 ventilator-days. The mortality rate in neonates with NI was 11.9%. Mechanical ventilation, total parenteral nutrition, orogastric tube, previous antibiotic therapy, use of CVC and birth weight of 751-1,000g appeared to be associated with a significantly higher risk of NI (p < 0.05). In multiple logistic regression analysis for NI, mechanical ventilation and the use of CVC were independent risk factors (p < 0.05). Coagulase- negative Staphylococcus (CoNS) (36.5%) and Staphylococcus aureus (23.6%) were the most common etiologic agents isolated from cultures. The incidences of oxacillin-resistant CoNS and S. aureus were 81.8% and 25.3%, respectively. CONCLUSIONS Frequent surveillance was very important to evaluate the association of these well-known risk factors with NIs and causative organisms, assisting in drawing the attention of health care professionals to this potent cause of morbidity.

Clinical and Molecular Epidemiology of Multidrug-Resistant P. aeruginosa Carrying aac(6')-Ib-cr, qnrS1 and blaSPM Genes in Brazil.

We described a comprehensive analysis of the molecular epidemiology of multidrug-resistant (MDR) P. aeruginosa. Molecular analysis included typing by Pulsed Field Gel Electrophoresis, identification of genes of interest through PCR-based assays and sequencing of target genes. Case-control study was conducted to better understand the prognostic of patients and the impact of inappropriate therapy in patients with bacteremia, as well as the risk factors of MDR infections. We observed a high rate of MDR isolates (40.7%), and 51.0% of them was independently associated with inappropriate antibiotic therapy. Bacteremia was detected in 66.9% of patients, and prolonged hospital stay was expressive in those resistant to fluoroquinolone. Plasmid-mediated quinolone resistance genes (PMQR), qnrS1 and aac(6’)Ib-cr, were detected in two different nosocomial isolates (5.3%), and the aac(6’)-Ib7 variant was detected at a high frequency (87.5%) in those negative to PMQR. The presence of mutations in gyrA and parC genes was observed in 100% and 85% of selected isolates, respectively. Isolates harboring PMQR genes or mutations in gyrA and parC were not closely related, except in those containing SPM (São Paulo metallo-β-lactamase) clone. In addition, there is no study published in Brazil to date reporting the presence of Pseudomonas aeruginosa isolates harboring both qnrS1 and aac(6’)Ib-cr genes, with alarming frequency of patients with inappropriate therapy.

Multidrug Resistance Related to Biofilm Formation in Acinetobacter baumannii and Klebsiella pneumoniae Clinical Strains from Different Pulsotypes.

The emergence of Acinetobacter baumannii and Klebsiella pneumoniae strains in the hospital environment has been associated with the presence of multiple genetic elements, virulence factors and the ability to form biofilms. This study evaluated the biofilm formation ability of clinical and environmental A. baumannii and K. pneumoniae strains, isolated from various sources and presenting different molecular characteristics, resistance profiles and pulsed-field gel electrophoresis patterns. Fifty-three isolates were recovered from 2009 to 2014 in a Brazilian university hospital. Investigation of biofilm formation was performed for 10 strains of each species assessed by an initial adhesion assay, biofilm cell concentration and biofilm biomass, evaluated by quantitative assays in replicates, in three independent experiments. All strains of A. baumannii were able to attach to polystyrene plates, although two strains adhered to a lesser degree than the control. K. pneumoniae strains showed opposite behaviour, where only three strains adhered significantly when compared to the control. Quantitative evaluation revealed that in five A. baumannii and four K. pneumoniae isolates the biomass production could be characterised as moderate. None of the isolates were strong biofilm producers. Our results demonstrate: (1) biofilm formation is a heterogeneous property amongst A. baumannii and K. pneumoniae clinical strains and it was not associated with certain clonal types; (2) no relationship between multidrug resistance and biofilm production was observed; (3) more virulent K. pneumoniae strains tended to present higher production of biofilm.

Molecular epidemiological survey of the quinolone- and carbapenem-resistant genotype and its association with the type III secretion system in Pseudomonas aeruginosa.

This study evaluated the predictors of mortality and the impact of inappropriate therapy on the outcomes of patients with bacteraemia and ventilator-associated pneumonia (VAP). Additionally, we evaluated the correlation of the type III secretion system (TTSS) effector genotype with resistance to carbapenems and fluoroquinolones, mutations in the quinolone resistance-determining regions (QRDRs), metallo-β-lactamase and virulence factors. A retrospective cohort was conducted at a tertiary hospital in patients with multidrug-resistant (MDR) P. aeruginosa bacteraemia (157 patients) and VAP (60 patients). The genes for blaIMP, blaVIM, blaSIM, blaGIM and blaSPM and virulence genes (exoT, exoS, exoY, exoU, lasB, algD and toxA) were detected; sequencing was conducted for QRDR genes on fluoroquinolone-resistant strains. The multivariate analyses showed that the predictors independently associated with death in patients with bacteraemia were cancer and inappropriate therapy. Carbapenem resistance was more frequent among strains causing VAP (53.3 %), and in blood we observed the blaSPM genotype (66.6 %) and blaVIM genotype (33.3 %). The exoS gene was found in all isolates, whilst the frequency was low for exoU (9.4 %). Substitution of threonine to isoleucine at position 83 in gyrA was the most frequent mutation among fluoroquinolone-resistant strains. Our study showed a mutation at position 91 in the parC gene (Glu91Lys) associated with a mutation in gyrA (Thre83Ile) in a strain of extensively drug-resistant P. aeruginosa, with the exoT(+)exoS(+)exoU(+) genotype, that has not yet been described in Brazil to the best of our knowledge. This comprehensive analysis of resistance mechanisms to carbapenem and fluoroquinolones and their association with TTSS virulence genes, covering MDR P. aeruginosa in Brazil, is the largest reported to date.

Etiology and pathogenesis of bloodstream infections associated with the use of long-term central vascular catheter (CVC) in patients who undergone gastrointestinal surgery

CVC is the main factor of risk of bloodstream infections. This study purpose was determining both etiology and pathogenesis of these infections in 80 patients who undergone gastrointestinal surgery and who worn long lasting CVC, in the institution HC-UFU. Cultures were made in nostril, skin of the insertion site, tip and catheter hub, in addition to hemoculture in those suspects of sepsis. The colonization incidence rate of the catheter tip was 12.5/ 1,000 catheter days and the CVC associate infection rate was 3.1/1,000 catheter days. Frequencies of skin, hub and catheter tip colonization were 13.8%, 8.9% and 13.3%, respectively. Coagulase-negative Staphylococci were the most found microorganisms in nostril (74%), skin (45.4%) and hub (75%) and Gram-negative bacilli (50%), followed by S. aureus (25%) the most common ones in catheter tip. Approximately 51% of patients received antibiotics and most of them (53.7%) had therapeutic purpose. The frequency of patients with clinical sepsis was 27.5%. Three cases of bacteremia associated with the use of CVC were detected (3.8%), with S. aureus in two of them and K. pneumoniae in the third one. There was not seen any association of skin and hub colonization with their presence in the catheter tip and in the blood of these patients, but S. aureus was recovered from nostril of those with sepsis by this pathogenic agent. A greater concern is suggested over preventive measures and control of these primary and secondary bloodstream, as well as catheter tip colonization.

Pressure ulcer as a reservoir of multiresistant Gram-negative bacilli: risk factors for colonization and development of bacteremia

The purpose of this study was to identify the risk factors that predispose patients who are hospitalized with pressure ulcers (PUs) colonized by Gram-negative bacilli (GNB) to develop bacteremia. In addition, we also detected main phenotypes of resistance in infected and uninfected PUs. A prospective cohort study was conducted at the Clinical Hospital of the Federal University of Uberlândia including patients with Stage II or greater PUs, colonized or not with GNB, from August 2009 to July 2010. Infected ulcers were defined based on clinical signs and on positive evaluation of smears of wound material translated by a ratio of polymorphonuclear cells to epithelial cells ≥2:1, after Giemsa staining. A total of 60 patients with Stage II PUs were included. Of these 83.3% had PUs colonized and/or infected. The frequency of polymicrobial colonization was 74%. Enterobacteriaceae and GNB non-fermenting bacteria were the most frequent isolates of PUs with 44.0% of multiresistant isolates. Among patients who had infected PUs, six developed bacteremia by the same microorganism with a 100% mortality rate. In addition, PUs in hospitalized patients were major reservoir of multiresistant GNB, also a high-risk population for the development of bacteremia with high mortality rates.

Hypervirulence and biofilm production in KPC-2-producing Klebsiella pneumoniae CG258 isolated in Brazil

In this study, we describe the frequency of virulence genes in Klebsiella pneumoniae carbapenemase-2-producing Klebsiella pneumoniae (KPC-KP), including hypervirulent (hv) and hypermucoviscous (hm) strains by whole-genome sequencing. We also evaluate the capacity for biofilm formation by using phenotypic techniques. The occurrence of several virulence genes (fimABCDEFGHIK, mrkABCDFHJ, ecpA, wabG, entB, ugE, irp1, irp2, traT, iutA and ureADE) and a high frequency of hvhmKPC-KP isolates was found. Most hospital-associated lineages of KPC-KP belong to the international clonal group 258 (CG258). Biofilm formation was a constant feature among 90.9 % of KPC-KP strains. This report suggests a close relationship between ST437 and weak biofilm production, given that all weakly biofilm-producing strains belonged to this sequence type. This also supports the dissemination of KPC-KP containing numerous virulence determinants belonging to the biofilm-producing CG258 type in Brazil, including hv and hm strains. These factors allow this pathogen to cause infections, leading to its rapid expansion and persistence in hospital settings.

The nares as a CA-MRSA reservoir in the healthy elderly.

INTRODUCTION The frequency of methicillin-resistant Staphylococcus aureus (MRSA) has increased in the community. This study evaluated the prevalence of MRSA and community-acquired (CA)-MRSA in 120 healthy elderly. METHODS The MRSA were evaluated for the presence of the IS256, mecA, agr, icaA, icaD, fnbB , and pvl genes with PCR. RESULTS Frequency of S. aureus and MRSA colonization was 17.8% and 19%, respectively. CA-MRSA isolate showed SCC mec IV, fnbB+ , and icaD+ . CONCLUSIONS CA-MRSA was detected, with genotype determined as SCC mec type IV/IS256/ fnbB+ / icaA / icaD+ / bbp-/agr2 / bap / pvl, characterizing this population as a possible reservoir of this organism in the community.

Molecular characterization and clonal dynamics of nosocomial blaOXA-23 producing XDR Acinetobacter baumannii

The emergence of infections associated to new antimicrobial resistance in Acinetobacter baumannii (Ab) genotypes represents a major challenge. In this context, this study aimed to determine the diversity of resistance mechanisms and investigate clonal dissemination and predominant sequence types (STs) in multidrug-resistant Ab strains of clinical (tracheal aspirate, n = 17) and environmental (surface, n = 6) origins. Additionally, the major clones found in clinical (A) and environmental (H) strains had their complete genomes sequenced. All strains were submitted to polymerase chain reactions (PCR) for the detection of the ISAba1/blaOXA-51-like and ISAba1/blaOXA-23-like genes, while the expression of genes encoding the carO porin, AdeABC (adeB), AdeFGH (adeG), and AdeIJK (adeJ) efflux pumps was determined by real time PCR (qPCR). Most of the strains were characterized as extensively drug-resistant (XDR) with high minimal inhibitory concentrations (MICs) detected for tigecycline and carbapenems. Associations between ISAba1/OXA-51 and ISAba1/OXA-23 were observed in 91.3% and 52.2% of the strains, respectively. Only the adeB gene was considered hyper-expressed. Furthermore, most of the strains analyzed by the MuLtilocus Sequence-Typing (MLST) were found to belong to the clonal complex 113 (CC113). In addition, a new ST, ST1399, belonging to CC229, was also discovered herein. Strains analyzed by whole genome sequencing presented resistance genes linked to multidrug-resistance phenotypes and confirmed the presence of Tn2008, which provides high levels carbapenem-resistance.

High frequency of the combined presence of QRDR mutations and PMQR determinants in multidrug-resistant Klebsiella pneumoniae and Escherichia coli isolates from nosocomial and community-acquired infections

Plasmid-mediated quinolone resistance (PMQR) determinants combined with mutations in quinolone resistance-determining regions (QRDRs) and clonal dissemination were investigated in 40 fluoroquinolone-resistant Klebsiellapneumoniae and Escherichiacoli isolates from nosocomial and community-acquired infections. We observed nucleotide substitutions in gyrA (Ser83Ile, Val37Leu, Lys154Arg, Ser171Ala, Ser19Asn, Ile198Val, Ser83Tyr, Ser83Leu, Asp87Asn and Asp87Gly) and parC genes (Ser80Ile, Glu84Lys, Ala129Ser, Val141Ala and Glu84Gly). Two novel substitutions were detected in the gyrA gene (Val37Leu and Ile198Val). The presence of PMQR genes predominated in community isolates (55.5 %). In addition to the frequent presence of the class 1 integron in isolates from community-acquired infections, the genetic similarity results obtained by PFGE showed high genomic diversity. This study suggests that management of multidrug-resistant Enterobacteriaceae isolates from the community are a possible source of genetic mobile elements that carry genes that confer resistance to fluoroquinolones. More attention should be paid to the surveillance of community-acquired infections.