31 research outputs found
Biofilm formation of Brazilian meticillin-resistant Staphylococcus aureus strains: prevalence of biofilm determinants and clonal profiles
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/publishedVersio
Epidemiologia e fatores de risco associados à colonização por VRE e MRSA em uma unidade de terapia intensiva de adultos
This investigation included a total of 78 VRE-colonized patients and 17 MRSA-colonized
patients through study of the incidence in the period April 2009 to January 2010. We
evaluated the rates of infection/colonization with these phenotypes, risk factors for
colonization, antimicrobial susceptibility profile and characterization of vanA gene in
enterococci strains with high level vancomycin resistance. The identification of S. aureus and
enterococci species was performed by conventional biochemical tests. The vancomycin
minimal inhibitory concentration (MIC) was evaluated by E-test method. The antimicrobial
susceptibility profile and high level aminoglycoside resistance were carried out by discdiffusion.
To assess the genotype enterococcal strains expressing high-level vancomycin
resistance, we used the polymerase chain reaction. Epidemiological data were recorded for all
patients included in the study and were used for the risk factor analysis. A case-control study
was then performed. The cases were defined as only patients with VRE colonization (n=21),
and the controls were those without VRE colonization or infection from any organism
(n=143). A total of 333 patients hospitalized were enrolled in this investigation. Of the 90
patients colonized with Enterococcus spp., 92 samples were isolated. Seventy seven patients
(23.1%) were colonized with VanC VRE and only one patient (0.3%) with VanA-type. Four
of 92 samples were identified as Enterococcus faecium, 11 as Enterococcus faecalis, 26 as
Enterococcus gallinarum and 51 as Enterococcus casseliflavus. The risk factors that were
determined through univariate analysis to be significantly associated with VRE colonization
included nephropathy, diabetes mellitus, prior ICU antibiotic use, vancomycin and
carbapenem use in the ICU. In the multivariate analysis, significant independent risk factors
for VRE colonization were the nephropathy (P < 0.001), prior ICU antibiotic use (P = 0.03)
and carbapenem use (P < 0.001). Our investigation revealed a low frequency of MRSA
colonization (5.1%) with 23.5% of colonized patients progressed to infection by this organism
(P <0.001, OR = 32.1), especially for cases of sepsis (P = 0.01, OR = 20.9). VRE
colonization, particularly the VanC phenotype, was frequent in the ICU and although of little
clinical importance, these microorganisms are considered reservoirs of resistance genes.
There was a correlation between the vancomycin and carbapenems use and VRE colonization,
although the results of multivariate analysis did not demonstrate vancomycin as an
independent risk factor for VRE colonization. We found a low incidence of MRSA in the ICU
and observed a significant relationship between colonization and the development of sepsis by
this microorganism.Conselho Nacional de Desenvolvimento Científico e TecnológicoMestre em Imunologia e Parasitologia AplicadasFoi analisado um total de 78 pacientes colonizados por Enterococcus resistente à vancomicina
(VRE) e 17 pacientes colonizados por Staphylococcus aureus resistente a meticilina (MRSA)
através de estudo de incidência no período de abril de 2009 a janeiro de 2010. Foram
avaliadas as taxas de infecção/colonização por esses fenótipos, fatores de risco para
colonização, perfil de sensibilidade aos antimicrobianos e caracterização do gene vanA em
amostras de VRE com alto nível de resistência à vancomicina. A identificação dos S. aureus e
das espécies de enterococos foi realizada por testes bioquímicos convencionais. A
concentração inibitória mínima (MIC) para vancomicina foi avaliada pelo método de E-test. O
perfil de sensibilidade aos antimicrobianos e a resistência em nível elevado aos
aminoglicosídeos foi realizada por disco difusão. Para avaliar o genótipo das cepas de VRE
foi utilizado o PCR. Dados epidemiológicos foram registrados de todos os pacientes incluídos
nesse estudo e para análise dos fatores de risco para colonização por VRE foi realizado um
estudo caso x controle sendo definidos como caso os pacientes colonizados por VRE sem
infecção por qualquer microrganismo (n=21) e como controle os pacientes sem colonização e
infecção por VRE (n=143). Um total de 333 pacientes hospitalizados foi incluído nessa
investigação. Dos 90 pacientes colonizados por Enterococcus spp., 92 amostras foram
isoladas. Setenta e sete pacientes (23,1%) estavam colonizados com Enterococcus resistentes
à vancomicina do fenótipo VanC e apenas um paciente (0,3%) com VanA. Quatro das 92
amostras foram identificadas como Enterococcus faecium, 11 como Enterococcus faecalis, 26
como Enterococcus gallinarum e 51 como Enterococcus casseliflavus. Os fatores de risco que
foram significativamente associados com a colonização por VRE incluíram nefropatia,
diabetes mellitus, uso de antibiótico prévio à UTI, uso de vancomicina e carbapenêmicos na
unidade sendo o uso de antibiótico prévio à UTI (P = 0,03), uso de carbapenêmicos na
unidade (P < 0,001) e nefropatia (P < 0,001) fatores de risco independentes para colonização.
Nossa investigação evidenciou baixa frequência de colonização por MRSA (5,1%) durante o
período estudado com 23,5% dos pacientes colonizados evoluindo com infecção por esse
microrganismo (P < 0,001; OR = 32,1), com destaque para os casos de sepse (P = 0,01; OR =
20,9). A colonização por VRE, predominantemente do fenótipo VanC, foi frequente na UTI e
embora de pouca importância clínica, esses microrganismos são considerados reservatórios de
genes de resistência. Houve correlação positiva entre o uso de vancomicina e carbapenêmicos
e a colonização por VRE. Apesar da baixa colonização por MRSA, observou-se uma relação
entre a colonização e o desenvolvimento de sepse por esse microrganismo
Marcadores genéticos de risco para forte produção de biofilme em cepas clínicas de Staphylococcus aureus resistentes à meticilina e sua associação com o perfil clonal
Methicillin-resistant Staphylococcus aureus (MRSA) is one of the major human pathogens
worldwide and its epidemiology has been the focus of numerous single and multicenter
surveillance studies over the past years. In this study, a phenotypic and genotypic approach were
used to determine the factors that influence adherence and biofilm production of the most
common MRSA SCCmec types, and its relationship with antimicrobial resistance, virulence
genes and the genetic background of S. aureus isolates. The strains used in this study were
selected from a collection of clinical MRSA strains recovered from patients hospitalized in the
Teaching Hospital of the Federal University of Uberlandia, isolated from infections at various
anatomical sites and evaluated for SCCmec type. Fifteen strains carrying different chromosomal
cassettes were selected, five SCCmec II, five SCCmec III and five SCCmec IV, recovered
predominantly from blood (67%), surgical site infections (27%) and pneumonia (6.0%). 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. The genetic relationship between the isolates and a
possible association with the ability to form biofilm were investigated by pulsed field gel
electrophoresis (PFGE). The initial adhesion and biofilm formation were examined by
quantitative assays. To evaluate the association between the hydrophobicity and the ability of
MRSA cell to adhere to an unmodified polystyrene surface, the surface tension and
hydrophobicity of the strains were measured by contact angle technique. There were association
among the values of the electron acceptor parameter, the degree of hydrophobicity and adhesion
ability. SCCmec III and IV strains were less hydrophilic, showed higher values of the electron
acceptor parameter and adhered better than SCCmec II strains. The analysis of biofilm production
showed that SCCmec III strains were characterized as strong biofilm producers; with the average
biomass of biofilm from 0.53 ± 0.12 compared with 0.04 ± 0.04 those non-producers/weak
producers (SCCmec II e IV). The analysis of this study showed five major pulsotypes according
to the PFGE (A-E) with a large genomic diversity observed by the number of subtypes in each
pulsotype. However, biofilm production was related to the dissemination of one specific PFGE
clone (Clone C). The presence of the genes agrI, fnbB and IS256 in clinical MRSA SCCmec III
strains, were considered as genetic risk markers for strong biofilm-formation by an icaindependent
biofilm pathway. This study contributes for the understanding of biofilm production
as a virulence factor potentially involved in the persistence and severity of infections caused by
Staphylococcus aureus belonging to this genotype.Coordenação de Aperfeiçoamento de Pessoal de Nível SuperiorDoutor em Imunologia e Parasitologia AplicadasStaphylococcus aureus resistente à meticilina (MRSA) é um dos principais patógenos humanos
em todo o mundo e sua epidemiologia tem sido foco de numerosos estudos de vigilância
unicêntricos e multicêntricos ao longo dos últimos anos. Neste estudo utilizamos abordagens
fenotípicas e genotípicas para determinar os fatores que influenciam a adesão inicial e produção
de biofilme em cepas clínicas de MRSA carreando os tipos de cassete cromossômico
estafilocócico (SCCmec) mais frequentes nos hospitais brasileiros, e sua relação com resistência,
genes de virulência e perfil clonal. As cepas de MRSA utilizadas neste estudo foram selecionadas
a partir de uma coleção de amostras clínicas recuperadas de pacientes internados no Hospital de
Clínicas da Universidade Federal de Uberlândia, isoladas de infecções em diversos sítios
anatômicos. Foram selecionadas para o estudo quinze cepas carreando diferentes cassetes
cromossômicos, cinco SCCmec II, cinco SCCmec III e cinco SCCmec IV, recuperadas
predominantemente de sangue (67%), sítio cirúrgico (27%) e pneumonia (6,0%). O tipo de
SCCmec, o grupo agr e a presença de genes de virulência (bbp, clfA, icaA, icaD, fnbB, bap, sasC
e IS256) foram avaliados por PCR. A relação genética entre as amostras e a possível associação
com a capacidade de formação de biofilme foram investigadas por eletroforese em gel de campo
pulsado (PFGE). A adesão inicial e a capacidade de formação de biofilme foram examinadas por
ensaios quantitativos. Adicionalmente, a associação entre a hidrofobicidade e a capacidade da
célula de MRSA aderir a uma superfície de poliestireno não modificada, foi avaliada através da
medida dos ângulos de contato. Houve associação entre o grau de hidrofobicidade e a capacidade
de adesão. Cepas clínicas de MRSA carreando SCCmec III e IV foram menos hidrofílicas,
apresentaram valores mais altos de tensão interfacial do componente aceptor de elétrons e
aderiram melhor do que as cepas SCCmecII. As análises da produção de biofilme mostraram que
cepas carreando SCCmec III foram caracterizadas como fortemente produtoras de biofilme, com
a média da biomassa do biofilme de 0,53 ± 0,12 em comparação com 0,04 ± 0,04 daquelas nãoprodutoras/
fraco produtoras (SCCmec II e SCCmec IV). A tipagem molecular por PFGE
evidenciou cinco pulsotipos (A-E) com uma grande diversidade clonal observada pelo número de
subtipos em cada pulsotipo. Entretanto, a produção de biofilme esteve relacionada a
disseminação de um clone específico (Clone C). Os genes agrI, fnbB e IS256 em cepas clínicas
de MRSA carreando SCCmec III, foram considerados marcadores genéticos de risco para forte
produção de biofilme por uma via independente de ica. Nosso estudo contribui para a
compreensão da produção de biofilme como um fator de virulência, potencialmente envolvido na
severidade e persistência de infecções causadas por S. aureus pertencentes a este genótipo
The nares as a CA-MRSA reservoir in the healthy elderly
Abstract: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
Spread of multidrug-resistant Acinetobacter baumannii and Pseudomonas aeruginosa clones in patients with ventilator-associated pneumonia in an adult intensive care unit at a university hospital
Background:In Brazil, ventilator-associated pneumonia (VAP) caused by carbapenem resis- tant Acinetobacter baumanniiand Pseudomonas aeruginosaisolates 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. aeruginosaVAP in an adult intensive care unit (ICU).Methods: It was conducted a cohort study of patients with VAP caused by carbapenem resistant A. baumanniiand P'. aeruginosaduring 14 months in an adult ICU. Genomic studies were used to investigate the clonal relatedness of carbapenem resistant OXA-23-producing A. baumanniiand P. aeruginosaclinical 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. baumanniiVAP and hemodialysis as independent variable associated with P. aeruginosaVAP. All carbapenem resistant clinical and environmental isolates of A. baumanniiwere OXA-23 producers. No MBL-producer P. aeruginosawas detected. Molecular typing revealed a polyclonal pattern; however, clone A (clinical) and H (surface) were the most frequent among isolates of A. baumanniitested, with a greater pattern of resistance than other isolates. In P. aeruginosathe most frequent clone I was multi-sensitive.Conclusion: These findings suggest the requirement of constant monitoring of these microor- ganisms in order to control the spread of these clones in the hospital environment