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The rapid increased multidrug resistance in Klebsiella pneumoniae has led to a renewed interest in polymyxin antibiotics, such as colistin, as antibiotics of last resort, not least in low/middle in...

Introduction: Infections, mainly bloodstream infections (BSI), acquired in the intensive care unit (ICU) are associated with significant rates of morbidity and mortality, especially when caused by pathogens. The objectives the study were to determine risk factors and mortality rates in 30 days for patients with BSI caused by microorganisms resistant versus microorganisms susceptible to antimicrobials. Methods: A case-vs-control study was carried out, in which patients were identified with BSI acquired in the ICU. Control patients were paired, considering: age, acute mean severity classification (ASIS), and chronic disease scores, Charlson's Comorbidity Index (CHARLSON) both ≥ 3. Results: Retrospective cohort study of 531 patients, 254 with bloodstream infection (BSI), and 277 uninfected controls during hospital stay. Of those infected, 181 presented multidrug-resistant isolates (MDR) and 73, isolates susceptible to antibiotics. The univariate analysis showed statistically significant frequencies for BSIs, when compared with controls, of isolates resistant to multiple drugs and susceptible. There was also significance in the mortality rate among patients with resistant and susceptible pathogens (40.3% vs 34.2%, P

This study used whole-genome sequencing to analyze the first case of NDM-1-producing Acinetobacter baumannii belonging to the novel sequence type 1465/CC216 recovered in Brazil. The study identified an unusual plasmid carrying blaNDM-1 gene, in which some genes of the Tn125 transposon were lost. Besides, on the chromosome, the strain reported here presented blaOXA-106 gene, a variant of blaOXA-51 gene, and blaADC-25 with ISAba1 upstream. The isolation of new STs of A. baumannii carrying blaNDM-1 genes elicits our concerns about the possible spread of these genes among clinically relevant bacteria.

Bloodstream infection (BSI) remains the most frequent adverse event among premature infants worldwide, associated with increased hospital stay length and costs, poor outcomes, and even death. The aim of this study was to investigate the origin of catheter-related bloodstream infections (CR-BSIs) by Staphylococcus epidermidis in critical neonates, using molecular biology techniques. This was an observational study comprising neonates presenting BSI using central venous catheters (CVCs) for over 24 hours. Skin cultures obtained from the catheter insertion site, CVC hub, and neonate nasal and intestinal mucosa, were performed weekly from 48 hours from the beginning of the invasive procedure, as well as from the tip of the CVC after its withdrawal. Quantitative cultures were also performed of the tip with modifications and the “roll-plate” technique. Isolates identification were obtained using a VITEK II automated system. Molecular subtyping by pulsed-field gel electrophoresis (PFGE) was used to determine the origin of all BSIs. Among 19 primary BSI caused by S. epidermidis, BSI origin was defined in only 21.1% of the cases, where one was characterized as definite intraluminal, two as definite extraluminal, and one as translocation. Origin was indeterminate in 78.9% of the cases analyzed by PFGE. A total of 27 different genotype profiles were obtained. The spread of a prevalent clone in the unit (clone A) was detected in 28.6% of the samples. Most BSIs related to catheter caused by S. epidermidis were not able to prove the origin of most microorganisms present in neonate blood, demonstrating the difficulty in determining it in this population of patients. The lack of a better explanation of the origin of CR-BSI in neonates limits the direction of specific intervention measures.

The dissemination of antimicrobial resistance genes and the bacterium that harbor them have increasingly become a public concern, especially in low- and middle-income countries. The present study used whole-genome sequencing to analyze 10 KPC-2-producing Klebsiella pneumoniae isolates obtained from clinical specimens originated from Brazilian hospitals. The study documents a relevant "snapshot" of the presence of class 1 integrons in 90% of the strains presenting different gene cassettes (dfrA30, dfrA15, dfrA12, dfrA14, aadA1, aadA2, and aac(6')Iq), associated or not with transposons. Two strains presented nonclassical integron (lacking the normal 3'conserved segment). In general, most strains showed a complex resistome, characterizing them as highly resistant. Integrons, a genetically stable and efficient system, confer to bacteria as highly adaptive and low cost evolution potential to bacteria, even more serious when associated with high-risk clones, indicating an urgent need for control and prevention strategies to avoid the spread of resistance determinants in Brazil. Despite this, although the class 1 integron identified in the KPC-2-producing K. pneumoniae clones is important, our findings suggest that other elements probably have a greater impact on the spread of antimicrobial resistance, since many of these important genes were not related to this cassette.

A rising incidence of extended-spectrum beta-lactamase-producing Enterobacteriacae has caused an increased use of carbapenems and subsequent spread of carbapenem-resistant strains. Two recent artic...

The rapid increased multidrug resistance in

Objective: To establish a baseline of knowledge regarding about inappropriate therapy, virulence and resistance in a cohort of patients infected with S. aureus. Methods: Retrospective cohort study in tertiary-care university hospital was employed to evaluate the risk factors and the impact of inappropriate therapy among patients with Staphylococcus aureus infections, resistance and virulence. To assess the presence of the genes was performed PCR. Results: Patients with MRSA were older and hospitalized 17 days longer than those with MSSA infection, which were in ICU with a bloodstream infection. 50.0% received inadequate antibiotic therapy and we found virulence factors associated with MRSA (mecA, LukS, fnbB and clfA genes). Conclusion: These data show that surveillance studies related to Staphylococcus aureus infections remain essential to identify resistance and inform policy on resistance.

It appears inevitable that severe acute respiratory syndrome coronavirus 2 will continue to spread. Although we still have limited information on the epidemiology of this virus, there have been multiple reports of superspreading events (SSEs), which are associated with both explosive growth early in an outbreak and sustained transmission in later stages. Although SSEs appear to be difficult to predict and therefore difficult to prevent, core public health actions can prevent and reduce the number and impact of SSEs. To prevent and control of SSEs, speed is essential. Prevention and mitigation of SSEs depends, first and foremost, on quickly recognizing and understanding these events, particularly within healthcare settings. Better understanding transmission dynamics associated with SSEs, identifying and mitigating high-risk settings, strict adherence to healthcare infection prevention and control measures, and timely implementation of nonpharmaceutical interventions can help prevent and control severe acute respiratory syndrome coronavirus 2, as well as future infectious disease outbreaks.

Carbapenemase-producing organisms are pandemic and a significant threat to public health. We investigated the clonal relatedness of colistin-resistant Klebsiella pneumoniae strains producing KPC-type carbapenemase (KPC-KP) causing subsequent infections or colonization. Moreover, we aimed to gain insight into the ability of biofilm production in K. pneumoniae strains producing carbapenemase. Twenty-two consecutive KPC-KP and one KPC-negative strain was identified from an adult intensive care unit in Brazil. Seventy-five percent of isolates that harbored the blaKPC gene exhibited genetic relatedness by pulsed-field gel electrophoresis, and none presented the plasmid-mediated mcr-1 and blaNDM genes. This study showed that the majority of repeated KPC infections in adults were caused by a clone that caused the previous infections/colonizations even after a long period of time and illustrates the capacity of multiple clones producing biofilms to coexist in the same patient at the same time, becoming a reservoir of KPC-KP in the hospital environment.

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.

Plasmid-mediated quinolone resistance (PMQR) determinants combined with mutations in quinolone resistance-determining regions (QRDRs) and clonal dissemination were investigated in 40 fluoroquinolone-resistant Klebsiella pneumoniae and Escherichia coli 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.

This study used whole-genome sequencing (WGS) and PFGE to analysis KPC-2-producing Klebsiella pneumoniae strains from clinical specimens collected in Brazilian hospitals. The study identifies the emergence of a novel small IncX3 plasmid (pKPB11), 12,757-bp in length, in a high-risk K. pneumoniae ST11/CG258 lineage, a successful clonal group in Brazil, carrying the bla

The dissemination of antimicrobial resistance genes and the bacterium that harbor them have increasingly become a public concern, especially in low- and middle-income countries. The present study used whole-genome sequencing to analyze 10 KPC-2-producing

Background Carbapenem-resistant Acinetobacter baumannii (CR-Ab) has become a worrying health care problem, mainly in developing countries, such as Brazil. The objective was to investigate the prevalence and prognostic factors for CR-Ab infections at a Brazilian university hospital and examine the impact of inappropriate antimicrobial therapy on patient outcome. Methods A retrospective study on hospitalized patients with CR-Ab infections was carried out from January 2013 to December 2017. An epidemiologic analysis was carried out to determine the frequency of infections, the epidemiologic indicators by year, the risk factors for 30-day mortality, and the impact of inappropriate therapy. Results A total of 489 patients were included in the study. A rate of 0.7 per 1,000 patient-day CR-Ab infections was observed, mostly in the lungs (54.7%), and predominantly in the adult intensive care unit. The occurrence of infections by CR-Ab per 1,000 patient-days in November 2014 exceeded the established control limit, confirming an outbreak. Conclusions The prevalence of CR-Ab increased in the investigated hospital, passing to an endemic pathogen with a direct impact on mortality and the control of these strains.

INTRODUCTION: The present study aimed to determine the incidence of health care-associated infections (HCAIs) and identify the main resistant microorganisms in intensive care unit (ICU) patients in a Brazilian university hospital. METHODS: A retrospective cohort study was conducted in a Brazilian teaching hospital between 2012 and 2014. RESULTS: Overall, 81.2% of the infections were acquired in the ICU. The most common resistant pathogenic phenotypes in all-site and bloodstream infections were oxacillin-resistant coagulase-negative staphylococci and carbapenem-resistant Acinetobacter spp. (89.9% and 87.4%; 80.6% and 70.0%), respectively. CONCLUSIONS: There is an urgent need to focus on HCAIs in ICUs in Brazil.

Biofilms plays an important role in medical-device-related infections. This study aimed to determine the factors that influence adherence and biofilm production, as well as the relationship between strong biofilm production and genetic determinants in clinical isolates of meticillin-resistant Staphylococcus aureus (MRSA). Fifteen strains carrying different chromosomal cassettes recovered from hospitalized patients were selected; five SCCmecII, five SCCmecIII and five SCCmecIV. The SCCmec type, agr group and the presence of the virulence genes (bbp, clfA, icaA, icaD, fnbB, bap, sasC and IS256) were assessed by PCR. PFGE and multilocus sequence typing (MLST) techniques were also performed. The initial adhesion and biofilm formation were examined by quantitative assays. The surface tension and hydrophobicity of the strains were measured by the contact angle technique to evaluate the association between these parameters and adhesion ability. SCCmecIII and IV strains were less hydrophilic, with a high value for the electron acceptor parameter and higher adhesion in comparison with SCCmecII strains. Only SCCmecIII strains could be characterized as strong biofilm producers. The PFGE showed five major pulsotypes (AE); however, biofilm production was related to the dissemination of one specific PFGE clone (C) belonging to MLST ST239 (Brazilian epidemic clonal complex). The genes agrI, fnbB and IS256 in SCCmecIII strains were considered as genetic determinants associated with strong biofilm-formation by an ica-independent biofilm pathway. This study contributes to the understanding of biofilm production as an aggravating factor potentially involved in the persistence and severity of infections caused by multidrug-resistant MRSA belonging to this genotype., We thank FAPEMIG (Fundação de Amparo à Pesquisa de Minas Gerais, proceeding APQ 01398-11) and CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, PDSE proceeding 8952/11-6) for the financial support and scholarships. We also thank Dr Teruyo Ito, Juntendo University, Japan, and Dr Elsa Masae Mamizuka, Universidade de São Paulo, Brazil, for kindly providing the control strains used in this study., info:eu-repo/semantics/publishedVersion

This study used whole-genome sequencing (WGS) and PFGE to analysis KPC-2-producing Klebsiella pneumoniae strains from clinical specimens collected in Brazilian hospitals. The study identifies the emergence of a novel small IncX3 plasmid (pKPB11), 12,757-bp in length, in a high-risk K. pneumoniae ST11/CG258 lineage, a successful clonal group in Brazil, carrying the blaKPC-2 gene on a non-Tn4401 genetic element (NTEKPC-Ic). Comparative analysis of the pKPB11 showed that this plasmid reduced its size, losing part of its conjugation apparatus. The pKPB11 was also compared to another strain sequenced in this study (KPC89) that had the hybrid IncX3-IncU plasmid (pKP89), of approximately 45 kb in length, similarly carrying the blaKPC-2 gene on NTEKPC-Ic. To the best of our knowledge, pKPB11 is the first example of small IncX3 plasmid found in a high-risk KPC-2-producing K. pneumoniae ST11/CG258.

Summary Background Healthcare-associated infection (HAI) represents a major problem for patient safety worldwide. Aim To provide an up-to-date picture of the extent, aetiology, risk factors and patterns of infections in intensive care units (ICUs) in 28 Brazilian hospitals of different sizes. Methods A one-day point prevalence survey in 2016 enrolled the ICUs of hospitals from the 12 meso-regions in Minas Gerais state, south-east Brazil. Hospitals were classified as university or non-university hospitals. All patients with >48 h of admission to the study ICUs at the time of the survey were included. Findings In total, 303 patients were studied; of these, 155 (51.2%) were infected and 123 (79.4%) had at least one ICU-acquired infection. The most common ICU-acquired infections were pneumonia (53.0%) and bloodstream infection (27.6%). One hundred and nineteen bacterial isolates were cultured; the most common were Acinetobacter baumannii (27.1%), Pseudomonas aeruginosa (27.1%) and Staphylococcus aureus (39.0%). According to type of infection, the most common pathogens were P. aeruginosa (30.4%) in pneumonia, coagulase-negative staphylococci (23.4%) and Enterobacteriaceae (23.4%) in bloodstream infections, and Enterobacteriaceae (47.6%) in urinary tract infections. Conclusion This study found that the overall prevalence of ICU-acquired infections in surveyed Brazilian hospitals was higher than that reported in most European countries and the USA. A greater proportion of infections were caused by non-fermenting Gram-negative bacteria. These observations, along with a high rate of antimicrobial use, illustrate the urgent need for HAIs to be a priority in the public health agenda of Brazil.

Background: In Brazil, ventilator-associated pneumonia (VAP) caused by carbapenem resistant Acinetobacter baumannii and Pseudomonas aeruginosa isolates are associated with significant mortality, morbidity and costs. Studies on the clonal relatedness of these isolates could lay the foundation for effective infection prevention and control programs. Objectives: We sought to study the epidemiological and molecular characteristics of A. baumannii vs. P. aeruginosa VAP in an adult intensive care unit (ICU). Methods: It was conducted a cohort study of patients with VAP caused by carbapenem resistant A. baumannii and P. aeruginosa during 14 months in an adult ICU. Genomic studies were used to investigate the clonal relatedness of carbapenem resistant OXA-23-producing A. baumannii and P. aeruginosa clinical isolates. The risk factors for acquisition of VAP were also evaluated. Clinical isolates were collected for analysis as were samples from the environment and were typed using pulsed field gel electrophoresis. Results: Multivariate logistic regression analysis identified trauma diagnosed at admission and inappropriate antimicrobial therapy as independent variables associated with the development of A. baumannii VAP and hemodialysis as independent variable associated with P. aeruginosa VAP. All carbapenem resistant clinical and environmental isolates of A. baumannii were OXA-23 producers. No MBL-producer P. aeruginosa was detected. Molecular typing revealed a polyclonal pattern; however, clone A (clinical) and H (surface) were the most frequent among isolates of A. baumannii tested, with a greater pattern of resistance than other isolates. In P. aeruginosa the most frequent clone I was multi-sensitive. Conclusion: These findings suggest the requirement of constant monitoring of these microorganisms in order to control the spread of these clones in the hospital environment. Keywords: Acinetobacter baumannii, Intensive care unit, Molecular epidemiology, Multidrug-resistant

Plasmid-mediated quinolone resistance (PMQR) determinants combined with mutations in quinolone resistance-determining regions (QRDRs) and clonal dissemination were investigated in 40 fluoroquinolone-resistant

FAPEMIG - Fundação de Amparo a Pesquisa do Estado de Minas Gerais Klebsiella pneumoniae é um agente etiológico frequente em infecções relacionadas à assistência à saúde no Brasil e possui uma grande capacidade de desenvolver ou adquirir resistência aos antimicrobianos. Este estudo investigou a disseminação clonal e os tipos de sequência (STs) predominantes em amostras clínicas de K. pneumoniae apresentando genes blaCTX-M e blaKPC, bem como a presença de genes para fatores de virulência e a capacidade de produção de biofilmes. Adicionalmente, também foi examinado o impacto das resistências aos carbapenêmicos e polimixina no fitness bacteriano de K. pneumoniae produtoras de carbapenemase (KPC). Para as avaliações epidemiológicas foram selecionadas randomicamente 60 amostras não-duplicadas de K. pneumoniae, sendo 30 amostras de infecção K. pneumoniae sensível aos carbapenêmicos (KpSC), 20 infecções e 10 colonizações por K. pneumoniae resistente aos carbapenêmicos (KpRC). As amostras foram selecionadas a partir de uma coleção recuperada de pacientes internados no Hospital de Clínicas da Universidade Federal de Uberlândia (HC-UFU) no período de junho de 2009 a julho de 2015. Para investigação dos genes de resistência blaCTX-M e blaKPC e genes de virulência fimH, fimA, wabG, iucC, rmpA, ecpA, mrkD e khe, as amostras foram avaliadas por reação em cadeia da polimerase (PCR). A confirmação de blaKPC-2 foi realizada por sequenciamento genético. Os pulsotipos, STs e complexos clonais foram determinados por Pulsed Field Gel Electrophoresis (PFGE) e Multilocus Sequence Typing (MLST). A adesão inicial e a formação de biofilme foram examinadas por ensaios quantitativos, e os resultados confirmados por microscopia eletrônica de varredura. Para avaliação do fitness foram utilizados experimentos de competição em pares, in vitro, utilizando três amostras de K. pneumoniae: uma resistente aos carbapenêmicos, contendo 5 dos 8 genes de virulência avaliados, uma resistente aos carbapenêmicos e polimixina, menos virulenta, e a amostra ATCC 10031 multissensível. Diferenças importantes foram encontradas na caracterização dos grupos KpSC e KpRC, destacando-se o elevado uso prévio de β-lactâmicos (53%), carbapenêmicos (73,3%) e polimixina B (43,3%) naqueles infectados por amostras resistentes. A terapia inadequada embora tenha sido prevalente em 56,7% dos casos, foi bem maior (70%) entre pacientes com infecções por KpRC. No total, 75% das amostras foram caracterizadas como multirresistentes. Além disso, foi observado alto consumo em DDD de cefepime, ceftriaxona e carbapenêmicos, com tendência ascendente entre o início e o final do período estudado. Em relação à presença de genes de resistência, 80% das amostras apresentaram o gene blaCTX-M e 100% das amostras com resistência aos carbapenêmicos possuíam o gene blaKPC. Os genes de virulência foram detectados com frequências elevadas em ambos os grupos, sem relação com a resistência aos carbapenêmicos. Entretanto, entre as amostras de KpRC, os genes fimH, fimA e wabG foram predominantes (83%). Observou-se disseminação policlonal das amostras com predominância dos STs 11 e 340, pertencentes ao complexo clonal 258. Todas as amostras de K. pneumoniae avaliadas foram capazes de aderir a uma superfície de poliestireno não modificada. No entanto, a análise estatística evidenciou que três amostras apresentaram taxas de adesão notavelmente mais altas do que as demais (p

The assessment of risk factors for the nosocomial acquisition of colonization and infection by vancomycin-resistant Enterococcus faecium (VREfm) is often problematic due to scarce data on antibiotic use. A 30-month prospective cohort study was conducted to characterize VREfm strains isolated during an outbreak and endemic period, identifying the risk factors, antibiotic consumption, and prevalence of virulence determinants.The study was conducted in a tertiary care hospital. A representative number (171 patients) of isolates that were classified as resistant to high-level vancomycin (minimum inhibitory concentration (MIC) ≥ 256 μg/ml) were investigated.Among 171 colonized patients, 22 (12.9%) developed VRE infection. All VREfm isolates harboured vanA genes. Genes codifying virulence factors such as enterococcal surface protein (esp), aggregation substance 1 (asa1), and gelatinase (gelE) were detected in the VREfm studied. All patients infected with VRE had previously been colonized and became infected on average 14 days after colonization. Only previous use of aminoglycosides was a risk factor independently associated with VRE infection; however, glycopeptide consumption in defined daily doses (DDD) per 1000 patient-days was associated with the presence of this microorganism. The monthly colonization pressure ranged from 0.004% to 1.32% during the 30-month study period.We found a high incidence of VRE in a tertiary care hospital, independently associated with the prior use of aminoglycosides and the administration of glycopeptides.

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.

Summary Background Carbapenem-resistant Enterobacteriaceae (CRE), especially those that produce Klebsiella pneumoniae carbapenemase (KPC) and are associated with colistin resistance, pose a severe health threat due to the limited treatment options. Aim To describe two outbreaks of KPC-producing K. pneumoniae in an adult intensive care unit (AICU) in Brazil. In May 2015, 14 patients had colistin-susceptible KPC-producing strains (ColS-KPC), and in July 2015, nine patients had colistin-resistant KPC-producing strains (ColR-KPC). Methods Between September 2014 and August 2015, we performed surveillance at a university hospital and all CRE were tested for bla KPC genes. Clonality was investigated by pulsed-field gel electrophoresis. Resistance to colistin was confirmed by broth microdilution method. Consumption of carbapenems and colistin was expressed as defined daily doses. Findings In all, 111 patients with CRE were identified during the surveillance period; K. pneumoniae was the major isolate (77.13%). The two outbreaks were identified when infection rates (KPC per 1000 patient-days) exceeded the background level. Rates of carbapenem and colistin consumption were high. Control measures (bedside alcohol gel, contact precautions, regular rectal swabs) did not curtail the outbreaks. Mortality rates were 42.9% and 44.4% for ColS-KPC- and ColR-KPC-infected patients, respectively. After the death of four infected patients with ColR-KPC, the unit was closed to new admissions. Conclusion Our experience demonstrates the serious risks presented by KPC, and especially ColR-KPC, in Brazilian AICUs. Selective pressure from excessive antibiotic use and transmission on healthcare workers' hands were likely the major factors in transmission.

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 (Sao 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.

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.

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.

The bacterial factors associated with bacteremia by multidrug-resistant and extensively drug-resistant P. aeruginosa, including overexpression of efflux pumps, AmpC overproduction, and loss/alteration of the OprD porin in isolates that are non-Metallo-β-Lactamase producing were analyzed in a retrospective study. Molecular analyses included strain typing by Pulsed Field Gel Electrophoresis and identification of key genes via qualitative and quantitative PCR-based assays. Previous use of carbapenems and tracheostomy was independently associated with the development of bacteremia by extensively drug-resistant and multidrug-resistant strains of P. aeruginosa. A high consumption of antimicrobials was observed, and 75.0% of the isolates contained amplicons with the blaSPM-1 and blaVIM genes, with a cross-dissemination of SPM-1. Of the 47 non-Metallo-β-Lactamase isolates, none had another type of carbapenemase. However, the isolates exhibited high rates of hyperproduction of AmpC, loss of the OprD porin (71.4%) and the presence of MexABOprM (57.1%) and MexXY (64.3%). This study suggests that in non-Metallo-β-Lactamase isolates, the association of intrinsic resistance mechanisms could contributes to the expression of multidrug-resistant/extensively drug-resistant phenotypes.