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Papers by Elizabeth Marques

Brazilian Journal of Microbiology, Dec 19, 2019

Stenotrophomonas maltophilia is one of the Gram-negative bacilli most frequently found in the air... more Stenotrophomonas maltophilia is one of the Gram-negative bacilli most frequently found in the airways of cystic fibrosis patients. This opportunistic pathogen is intrinsically multidrug-resistant, and therefore, its treatment presents a challenge. The genetic characterization of S. maltophilia is largely unknown, especially from those strains that colonize/infect the airways of cystic fibrosis patients. This work reports the draft genome sequences of three S. maltophilia isolates recovered from the sputum of a cystic fibrosis pediatric patient in Southeast Brazil. Several resistance- and virulence-related genes were detected. Furthermore, one intact phage and one incomplete prophage region were also identified in all strains. Multilocus sequence typing showed that all strains belonged to a new sequence type (ST264). Interestingly, all S. maltophilia strains were genetically identical, showing persistence for at least 16 months. To our knowledge, this is the first report of S. maltophilia draft genome sequences obtained from a cystic fibrosis pediatric patient in Brazil.

Current Microbiology, May 4, 2021

Burkholderia cepacia complex (Bcc) comprises 24 related species genetically distinct, associated ... more Burkholderia cepacia complex (Bcc) comprises 24 related species genetically distinct, associated with high mortality in cystic fibrosis (CF) patients. Due to a high level of similarity among Bcc species, accurate identification has been problematic, and most conventional and automated phenotypic tests have shown low accuracy. We evaluated accuracy of MALDI-ToF MS decreasing the cut-off score value to distinguish Bcc species compared to recA gene sequencing. A total of 145 Bcc isolates were analyzed. B. vietnamiensis (41.37%), B. cenocepacia IIIA (23.44%), B. multivorans (20%), B. cenocepacia IIIB (11.03%), and B. contaminans (2.75%) among other species were identified by recA sequencing. MALDI-ToF MS identified 100% of Bcc isolates at the genus level and 53.1% at the species level. By decreasing cut-off values for ≥1.70, the correct identification at the species level increased to 74.5%. MALDI-ToF MS proved to be useful at the genus level identification, but it still requires improvements that allow more precise identification, requiring continuous updates and addition of new spectra to its database. A review of interpretative criteria is a field to be explored with a large collection of Bcc species.

Infection, Genetics and Evolution, Aug 1, 2022

Frontiers in Microbiology, May 27, 2022

Due to recent developments in NGS technologies, genome sequencing is generating large volumes of ... more Due to recent developments in NGS technologies, genome sequencing is generating large volumes of new data containing a wealth of biological information. Understanding sequenced genomes in a biologically meaningful way and delineating their functional and metabolic landscapes is a first-level challenge. Considering the global antimicrobial resistance (AMR) problem, investments to expand surveillance and improve existing genome analysis technologies are pressing. In addition, the speed at which new genomic data is generated surpasses our capacity to analyze it with available bioinformatics methods, thus creating a need to develop new, user-friendly and comprehensive analytical tools. To this end, we propose a new web application, CABGen, 1 developed with opensource software. CABGen allows storing, organizing, analyzing, and interpreting bioinformatics data in a friendly, scalable, easy-to-use environment and can process data from bacterial isolates of different species and origins. CABGen has three modules: Upload Sequences, Analyze Sequences, and Verify Results. Functionalities include coverage estimation, species identification, de novo genome assembly, and assembly quality, genome annotation, MLST mapping, searches for genes related to AMR, virulence, and plasmids, and detection of point mutations in specific AMR genes. Visualization tools are also available, greatly facilitating the handling of biological data. The reports include those results that are clinically relevant. To illustrate the use of CABGen, whole-genome shotgun data from 181 bacterial isolates of different species collected in 5 Brazilian regions between 2018 and 2020 were uploaded and submitted to the platform's modules.

Frontiers in Medicine, Mar 15, 2021

Multidrug-resistant microorganisms are a well-known global problem, and gram-negative bacilli are... more Multidrug-resistant microorganisms are a well-known global problem, and gram-negative bacilli are top-ranking. When these pathogens are associated with bloodstream infections (BSI), outcomes become even worse. Here we applied whole-genome sequencing to access information about clonal distribution, resistance mechanism diversity and other molecular aspects of gram-negative bacilli (GNB) isolated from bloodstream infections in Brazil. It was possible to highlight international high-risk clones circulating in the Brazilian territory, such as CC258 for Klebsiella pneumoniae, ST79 for Acinetobacter baumannii and ST233 for Pseudomonas aeruginosa. Important associations can be made such as a negative correlation between CRISPR-Cas and K. pneumoniae CC258, while the genes bla TEM, bla KPC and bla CTX−M are highly associated with this clone. Specific relationships between A. baumannii clones and bla OXA−51 variants were also observed. All P. aeruginosa ST233 isolates showed the genes bla VIM and bla OXA486. In addition, some trends could be identified, where a new P. aeruginosa MDR clone (ST3079), a novel A. baumannii clonal profile circulating in Brazil (ST848), and important resistance associations in the form of bla VIM−2 and bla IMP−56 being found together in one ST233 strain, stand out. Such findings may help to develop approaches to deal with BSI and even other nosocomial infections caused by these important GNB.

Brazilian Journal of Microbiology, 2020

The application of next-generation sequencing tools revealed that the cystic fibrosis respiratory... more The application of next-generation sequencing tools revealed that the cystic fibrosis respiratory tract is a polymicrobial environment. We have characterized the airway bacterial microbiota of five adult patients with cystic fibrosis during a 14-month period by 16S rRNA tag sequencing using the Illumina technology. Microbial diversity, estimated by the Shannon index, varied among patient samples collected throughout the follow-up period. The beta diversity analysis revealed that the composition of the airway microbiota was highly specific for each patient, showing little variation among the samples of each patient analyzed over time. The composition of the bacterial microbiota did not reveal any emerging pathogen predictor of pulmonary disease in cystic fibrosis or of its unfavorable clinical progress, except for unveiling the presence of anaerobic microorganisms, even without any established clinical association. Our results could potentialy help us to translate and develop strategies in response to the pathobiology of this disease, particularly because it represents an innovative approach for CF centers in Brazil.

Revista Brasileira de Ortopedia, 2022

Resumo Objetivo Estimar a frequência da colonização por Staphylococcus aureus e as bactérias não ... more Resumo Objetivo Estimar a frequência da colonização por Staphylococcus aureus e as bactérias não suscetíveis à cefalosporina, em pacientes com fratura proximal do fêmur durante a internação pré-operatória. Métodos Avaliação da prevalência e incidência em 63 pacientes hospitalizados ao longo de um ano. O tempo médio de internação pré-tratamento foi de 12 dias. As amostras foram coletadas das narinas, pele da virilha e mucosa anal, durante a internação prévia ao tratamento e testadas pela técnica de disco-difusão. Resultados A incidência da colonização hospitalar e a prevalência de resultados positivos foram de 14,3% e 44,4% para Staphylococcus aureus; 3,2% e 6,4% para S. aureus resistente à meticilina; 28,6% e 85,7% para Staphylococcus coagulase-negativo resistente à meticilina; 28,6% e 61,9% para Enterobacteriaceae não suscetível à cefazolina (KFNSE); e 20,6% e 28,6% para Enterobacteriaceae não suscetível à cefuroxima (CXNSE). Além da duração do período de internação pré-tratamento,...

Bacteria producing Klebsiella pneumoniae carbapenemases (KPCs) are rapidly emerging as a cause of... more 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.

New Microbes and New Infections, 2018

We describe the first detection of a KPC-2-and QnrB-producing Enterobacter cloacae from a patient... more We describe the first detection of a KPC-2-and QnrB-producing Enterobacter cloacae from a patient with cystic fibrosis. The bla KPC-2 and qnrB-1 genes were located in a 79.8-kb plasmid. The presence of bla KPC-2 and qnrB-1 genes was determined by PCR and sequencing. Mobilization of plasmid containing bla KPC2 gene was assayed by conjugation.

Revista Hospital Universitário Pedro Ernesto, 2013

Journal of Medical Microbiology, 1989

The tracheobronchial secretions from patients with cystic fibrosis often contain high amounts of ... more The tracheobronchial secretions from patients with cystic fibrosis often contain high amounts of free proteases. To evaluate whether human leucocyte elastase (HLE) can favour the persistence of bacterial airways infection, we exposed the frog palate mucosa to HLE and then to radiolabelled Pseudomonas aeruginosa and followed the sequence of events by scanning electronmicroscopy. In response to HLE there was a marked outpouring of mucus and a desquamation of the epithelium. P. aeruginosa was shown to adhere to recently secreted granules of mucus and to the exposed submucosal underlying connective tissues. For the eight different bacterial strains studied, a significative adherence to HLE-injured mucosa was observed only in strains that possessed internal haemagglutinating activity. Neither the presence of fimbriae, nor of the mucoid exopolysaccharide, nor of the bacterial surface haemagglutinating activity could be related to adherence of P. aeruginosa to the injured mucosa. These results support the hypothesis that HLE enhances bacterial infection of the respiratory mucosa both by inducing mucus hypersecretion and by exposing receptors to the microbial adhesins. It is also suggested that P. aeruginosa internal lectins may be implicated in adherence to host tissues.

Journal of Cystic Fibrosis, 2007

Aim of study: to evaluate the spectrum of pulmonary infection among Romanian CF children. Materia... more Aim of study: to evaluate the spectrum of pulmonary infection among Romanian CF children. Materials and Methods: study lot included 93 children, median age at diagnosis = 11.8 yrs, ranging from 3 month to 18 yr. Patients were classified in 4 age groups, as follow: under 1 years old (n = 18), 1−3 yrs (n = 34), 3−7 yrs (n = 7), 7−18 years old (n = 33). We have analyzed microbiological results obtained from: sputum samples, throat swabs or pharingotracheal aspirates, for five years ago. The pursued pathogens were: P. aeruginosa, S. aureus, MRSA, E. coli, Klebsiella sp, S. maltophilia, Aspergilus fumigatus, Candida albicans. We could not determine Burkholderia cepacia complex. Results: Table 1-Pathogens prevalence in CF children. Among study lot 19.35% (18 patients) had infection with 2 or more pathogens. Conclusions: P. aeruginosa is the most frequent pathogen in our lot study and its prevalence is rising with age, while S. aureus predominates in babies. A.fumigatus has is a significant pathogen, also in older children. An important percentage of mixed infection was registered, especially in delayed diagnosed children.

Cellular Microbiology, 2005

As Pseudomonas aeruginosa ExoU possesses two functional blocks of homology to calcium-independent... more As Pseudomonas aeruginosa ExoU possesses two functional blocks of homology to calcium-independent (iPLA(2)) and cytosolic phospholipase A(2) (cPLA(2)), we addressed the question whether it would exhibit a proinflammatory activity by enhancing the synthesis of eicosanoids by host organisms. Endothelial cells from the HMEC-1 line infected with the ExoU-producing PA103 strain exhibited a potent release of arachidonic acid (AA) that could be significantly inhibited by methyl arachidonyl fluorophosphonate (MAFP), a specific PLA(2) inhibitor, as well as significant amounts of the cyclooxygenase (COX)-derived prostaglandins PGE(2) and PGI(2). Cells infected with an isogenic mutant defective in ExoU synthesis did not differ from non-infected cells in the AA release and produced prostanoids in significantly lower concentrations. Infection by PA103 induced a marked inflammatory response in two different in vivo experimental models. Inoculation of the parental bacteria into mice footpads led to an early increase in the infected limb volume that could be significantly reduced by inhibitors of both COX and lipoxygenase (ibuprofen and NDGA respectively). In an experimental respiratory infection model, bronchoalveolar lavage (BAL) from mice instilled with 10(4) cfu of PA103 exhibited a marked influx of inflammatory cells and PGE(2) release that could be significantly reduced by indomethacin, a non-selective COX inhibitor. Our results suggest that ExoU may contribute to P. aeruginosa pathogenesis by inducing an eicosanoid-mediated inflammatory response of host organisms.

Epidemiology and Infection, 2017

Acinetobacter spp. are important healthcare pathogens, being closely linked to antibiotic resista... more Acinetobacter spp. are important healthcare pathogens, being closely linked to antibiotic resistance and outbreaks worldwide. Although such species are rarely observed in patients with cystic fibrosis (CF), we describe the characteristics of 53 strains of Acinetobacter spp. isolated from the sputum of 39 Brazilian patients with CF. The species distribution was A. baumannii (n = 29), A. pittii (n = 13), A. nosocomialis (n = 8), A. seifertii (n = 1), A. soli (n = 1) and A. variabilis (n = 1) determined by partial rpoB gene sequencing. Sixteen strains (10 A. baumannii, 3 A. pittii and 3 A. nosocomialis) were multidrug-resistant (MDR) by disk diffusion test (30%) and eight MDR carbapenem-resistant A. baumannii strains harboured the blaOXA-23-like oxacillinase gene. Thirty-three sequence types (STs) were identified by multilocus sequence typing of which eight were novel (A. baumannii: 843, 844, 845, 847, 848; A. pitti: 643; A. nosocomialis: 862 and A. seifertii: 846); six STs (2 A. bauma...

Epidemiology and Infection, Nov 22, 2016

Achromobacter spp. are opportunistic pathogens increasingly recovered from adult patients with cy... more Achromobacter spp. are opportunistic pathogens increasingly recovered from adult patients with cystic fibrosis (CF). We report the characterization of 122 Achromobacter spp. isolates recovered from 39 CF patients by multilocus sequence typing, virulence traits, and susceptibility to antimicrobials. Two species, A. xylosoxidans (77%) and A. ruhlandii (23%) were identified. All isolates showed a similar biofilm formation ability, and a positive swimming phenotype. By contrast, 4•3% and 44•4% of A. xylosoxidans and A. ruhlandii, respectively, exhibited a negative swarming phenotype, making the swimming and swarming abilities of A. xylosoxidans significantly higher than those of A. ruhlandii. A. xylosoxidans isolates from an outbreak clone also exhibited significantly higher motility. Both species were generally susceptible to ceftazidime, ciprofloxacin, imipenem and trimethoprim/sulphamethoxazole and there was no significant difference in susceptibility between isolates from chronic or sporadic infection. However, A. xylosoxidans isolates from chronic and sporadic cases were significantly more resistant to imipenem and ceftazidime than isolates of the outbreak clone.

APMIS

Burkholderia cenocepacia complex is associated with high transmissibility, virulence, and poor pr... more Burkholderia cenocepacia complex is associated with high transmissibility, virulence, and poor prognosis in cystic fibrosis (CF) patients. However, extrapulmonary infections are rare. We investigated the genome of a B. cenocepacia IIIA isolated from a liver abscess in a Brazilian CF patient and compared it to strain J2315. The whole genome was sequenced, and contigs were annotated by Rapid Annotation using Subsystem Technology. The Pathosystems Resource Integration Center was used to map antimicrobial and virulence genes. The genomic island (GIs) analysis was performed using two prediction methods, and the presence of putative plasmids and insertion sequences (ISs) was investigated. The isolate was confirmed as B. cenocepacia IIIA to ST‐28 (ET12 lineage). A total of 64 genes for antimicrobial resistance and 47 genes related to virulence were identified. Among the virulence factors, there was a predominance of factors related to the invasion mechanism, to the flagellar biosynthesis protein, and to the RNA polymerase sigma factor for flagellar operon (cdpA). Two IS families (IS3 and IS5) and only one plasmid were found. On average 56 GIs were predicted by at least one of the methods applied. Comparative analysis showed resistance mechanisms and virulence factors revealing invasive determinants used by B. cenocepacia IIIA (ET12) in the process of disease spread to other infection sites (extrapulmonary) of highly virulent strains in CF patients.

Infection, 2014

Despite the recent advances in diagnosis and treatment, mortality rates due to infective endocard... more Despite the recent advances in diagnosis and treatment, mortality rates due to infective endocarditis (IE) remain high if not aggressively treated with antibiotics, whether or not associated with surgery. Data on the prevalence, epidemiology and etiology of IE from developing countries remain scarce. The aim of this observational, prospective cohort study was to report a 5-year experience of IE at two teaching hospitals in Rio de Janeiro, Brazil. Demographical, anamnestic and microbiological characteristics of 71 IE patients were evaluated during the period of January 2009 to March 2013. The mean age of the IE patients was 49.8 ± 2.4 years, of which 41 (57.7%) were males. The median time between the onset of symptoms and diagnosis of IE was 35.8 ± 4.8 days. A total of 31 (43.6%) cases of community-acquired infective endocarditis (CAIE) and 40 (56.3%) cases of healthcare-acquired infective endocarditis (HAIE) were observed. Staphylococcus aureus (30%) was the predominant cause of IE....

Microbiology and Immunology, 2001

Bacteria of Stenotrophomonas maltophilia have been isolated with increasing frequency from the ai... more Bacteria of Stenotrophomonas maltophilia have been isolated with increasing frequency from the airways of cystic fibrosis (CF) patients, usually following P. aeruginosa infections, but their adherence to human epithelial respiratory cells has never been investigated. In this study, various S. maltophilia strains were seen to adhere to epithelial respiratory cells in vitro, mainly along intercellular junctions. Bacteria could also enter into host cells, as determined by the gentamicin exclusion assay and transmission electron microscopy. Cells co-incubated with P. aeruginosa and S. maltophilia exhibited a significantly decreased adherence of these latter bacteria. No decrease in S. maltophilia adherence was observed when co-infection was carried out with heat-killed P. aeruginosa or when respiratory cells were first incubated with P. aeruginosa, before incubation with S. maltophilia. Our data suggest that P. aeruginosa infections do not account for the increased prevalence of S. maltophilia in CF patient airways, that thermolabile products from P. aeruginosa can control the adherence of S. maltophilia to respiratory cells and also that these two bacteria do not compete for cell receptors.

BMC pulmonary medicine, Jan 8, 2015

Pulmonary deterioration after B.cepacia complex (BCC) colonization has a heterogeneous pattern. T... more Pulmonary deterioration after B.cepacia complex (BCC) colonization has a heterogeneous pattern. The aim was to investigate the clinical outcome of BCC colonization in CF patients chronically colonized with P. aeruginosa. CF patients chronically colonized with P. aeruginosa were divided into three groups: intermittent (I), chronic (II) and no colonization (III) with BCC. Body mass index (BMI) percentile and spirometric parameters were analyzed at three different times in each group. Fifty-six patients chronically colonized with P. aeruginosa were included. Of these, 27 also had evidence of BCC colonization (13 intermittent and 14 chronic). BMI percentile was significantly lower among patients chronically colonized by both P. aeruginosa and BCC. Mean values of FEV1 and FVC % were also significantly lower in these patients, both at the time of chronic BCC colonization and 24 months forward. Chronic BCC colonization is associated with significant loss of lung function. Lower BMI might b...