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1.
Eur J Ophthalmol ; 32(1): 275-281, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-33158363

RESUMO

PURPOSE: The aim of the present study is to calculate Concavity Shape Index (CSI) in patients with POAG and exfoliative glaucoma (XFG) and correlate CSI with the severity of glaucoma, comparing to control and ocular hypertension (OHT) patients. METHODS: This was a cross­sectional study with 146 eyes/146 subjects: 37 healthy eyes, 23 eyes with OHT and 86 glaucoma eyes (70 with POAG, 16 with XFG). The severity of glaucoma was scored with the Glaucoma Staging System 2 (GSS2). Corvis ST® was used to calculate CSI. RESULTS: Central corneal thickness (CCT) was significantly thinner in POAG (526 ± 40.0 µm) and XFG (520 ± 38.2 µm) than control group (553 ± 28.8 µm). CSI had no significant differences between the groups. XFG had a higher mean of GSS 2 (2.42 ± 1.38) than POAG (1.87 ± 1.55) and OHT (1.87 ± 1.55). OHT had a significantly less deformable cornea than: control (higher A1 length, lower A1 velocity, higher A2 velocity), POAG (higher A1 length, lower A1 velocity, lower deflection amplitude at highest concavity), and XFG group (lower A1 velocity, lower deflection amplitude at highest concavity), which was independent of age and CCT. No significant correlation was found between GSS 2 and CSI. DISCUSSION: OHT patients had stiffer corneas (less deformed by the air puff) when compared to control, POAG or XFG patients. A less deformable cornea could potentially be related to a more resistant LC/peripapillary sclera. As such, this would result in a lesser optic nerve susceptibility to IOP damage.


Assuntos
Glaucoma de Ângulo Aberto , Glaucoma , Hipertensão Ocular , Córnea/diagnóstico por imagem , Estudos Transversais , Glaucoma de Ângulo Aberto/diagnóstico , Humanos , Pressão Intraocular , Hipertensão Ocular/diagnóstico , Tonometria Ocular
2.
Curr Eye Res ; 46(1): 31-34, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32482105

RESUMO

PURPOSE/AIM: Floppy eyelid syndrome (FES) is an ocular manifestation of obstructive sleep apnea (OSA), but no studies have analyzed whether it can be improved by nocturnal continuous positive airway pressure (CPAP) therapy. The aim of this study was to analyze the effect of CPAP on FES by comparing objective measurements before and after 6 months of CPAP therapy. MATERIALS AND METHODS: We conducted a prospective study of 47 patients (74.5% males) with newly diagnosed OSA at a secondary care Portuguese hospital who underwent objective diagnostic testing for FES (upper eyelid eversion for >6 seconds and tarsal conjunctival exposure and upper eyelid laxity ≥1.5 mm). Patients with hyperelastic eyelid or FES were re-evaluated by the same ophthalmologist (blinded to the patients' condition) after 6 months of CPAP therapy. RESULTS: Mean apnea hypopnea index (AHI), analyzed as number of events per hour, was 28.7 ± 18.6 overall and 42.8 ± 20.0 in the supine position. Thirty-four percent of patients had FES. Mean AHI in the supine position was significantly higher in patients with FES (p = .041) and was an independent predictor of FES (p = .034; OR = 0.48). Severe OSA was significantly associated with FES (p = .023). FES resolved in 53.8% of patients after CPAP therapy. Patients with non-reversible FES had more severe OSA and worse airway access according to the Mallampati classification (from class I: visualization of soft palate and entire uvula, to class IV: soft palate not visible). CONCLUSIONS: A higher AHI in the supine position may be predictive of FES. CPAP therapy might reverse FES and patients with non-reversible FES appear to have more severe OSA and a worse airway access.


Assuntos
Pressão Positiva Contínua nas Vias Aéreas , Doenças Palpebrais/fisiopatologia , Apneia Obstrutiva do Sono/fisiopatologia , Apneia Obstrutiva do Sono/terapia , Idoso , Doenças Palpebrais/diagnóstico , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Polissonografia , Estudos Prospectivos , Decúbito Dorsal
4.
Sci Rep ; 9(1): 18972, 2019 12 12.
Artigo em Inglês | MEDLINE | ID: mdl-31831825

RESUMO

The aim is to study risk factors for retinal vein occlusion (RVO), such as thrombophilic and cardiovascular risk factors (CRF). A retrospective consecutive case series of 60 patients with RVO was made, tested for CRF, hyperhomocysteinemia, lupic anticoagulant, antiphospholipid antibody and 5 gene variants: factor V (FV) Leiden (G1691A), factor II (PT G20210A), 5,1-methylenetetra-hydrofolate reductase (MTHFR; 677 C > T and 1298 A > C), plasminogen activator inhibitor 1 (PAI-1; 4 G/5 G). More than 1 CRF were present in 36 patients (60%), which had a significantly higher mean age at diagnosis (66.7 ± 12.9 versus 59.5 ± 13.7 with ≤1 CRF, [t(57) = -2.05, p = 0.045, d = 0.54). Patients with thermolabile MTHFR forms with decreased enzyme activity (T677T or C677T/A1298C) had a significant lower mean age [57.6 ± 15.1; t (58) = 3.32; p = 0.002; d = 0.846] than patients with normal MTHFR enzyme activity (68.5 ± 10.2). Regarding CRF and thermolabile forms of MTHFR, the mean age at diagnosis could be significantly predicted [F(2,56) = 7.18; p = 0.002] by the equation: 64.8 - 10.3 × (thermolabile MTHFR) - 5.31 × ( ≤ 1CRF). Screening of MTHFR polymorphisms may be useful in younger RVO patients, particularly when multiple CRF are absent.


Assuntos
Metilenotetra-Hidrofolato Redutase (NADPH2)/genética , Polimorfismo Genético , Oclusão da Veia Retiniana , Trombofilia , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Oclusão da Veia Retiniana/enzimologia , Oclusão da Veia Retiniana/etiologia , Oclusão da Veia Retiniana/genética , Fatores de Risco , Trombofilia/complicações , Trombofilia/enzimologia , Trombofilia/genética
5.
Food Microbiol ; 80: 1-8, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30704592

RESUMO

Listeria monocytogenes is one of the most important foodborne pathogens due to the high hospitalization and mortality rates associated to an outbreak. Several new molecular methods that accelerate the identification of L. monocytogenes have been developed, however conventional culture-based methods still remain the gold standard. In this work we developed a novel Peptide Nucleic Acid Fluorescence in situ Hybridization (PNA-FISH) method for the specific detection of L. monocytogenes. The method was based on an already existing PNA probe, LmPNA1253, coupled with a novel blocker probe in a 1:2 ratio. The method was optimized for the detection of L. monocytogenes in food samples through an evaluation of several rich and selective enrichment broths. The best outcome was achieved using One Broth Listeria in a two-step enrichment of 24 h plus 18 h. For validation in food samples, ground beef, ground pork, milk, lettuce and cooked shrimp were artificially contaminated with two ranges of inoculum: a low level (0.2-2 CFU/25 g or mL) and a high level (2-10 CFU/25 g or mL). The PNA-FISH method performed well in all types of food matrices, presenting an overall accuracy of ≈99% and a detection limit of 0.5 CFU/25 g or mL of food sample.


Assuntos
Contaminação de Alimentos/análise , Microbiologia de Alimentos/métodos , Hibridização in Situ Fluorescente , Listeria monocytogenes/isolamento & purificação , Animais , Sondas de Ácido Nucleico/genética , Ácidos Nucleicos Peptídicos/genética , Kit de Reagentes para Diagnóstico , Sensibilidade e Especificidade
6.
Future Microbiol ; 12: 1167-1179, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28879777

RESUMO

AIM: This study aims to investigate the morphological transition of Helicobacter pylori during adaptation to water. MATERIALS & METHODS: Different strains were adapted to water. Changes regarding cultivability and cellular morphology were recorded. Expression of 11 genes involved in H. pylori morphological changes was evaluated by real-time PCR. RESULTS: H. pylori presented increased cultivability in water after adaptation. The permanent loss of the spiral shape was observed, but no transition into coccoid form has occurred. Expression levels of genes involved in peptidoglycan assembly of H. pylori 26695 have shown significant changes between adapted and nonadapted strains. CONCLUSION: Adaption to water favors the culturable phenotype and the morphological transition to the rod shape, into a process that implicates the peptidoglycan turnover.


Assuntos
Adaptação Fisiológica , Helicobacter pylori/citologia , Helicobacter pylori/genética , Água , Parede Celular , Regulação Bacteriana da Expressão Gênica , Genes Bacterianos/genética , Helicobacter pylori/crescimento & desenvolvimento , Microscopia Eletrônica de Varredura , Peptidoglicano , Fenótipo , RNA Bacteriano , Reação em Cadeia da Polimerase em Tempo Real , Microbiologia da Água
7.
Eur Heart J Acute Cardiovasc Care ; 4(6): 518-27, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25293725

RESUMO

PURPOSE: Conduct a meta-analysis to study the prognostic influence of a previous coronary artery bypass grafting (CABG) in patients admitted for an acute coronary syndrome (ACS). METHODS: A systematic review of the literature was performed using electronic reference databases through January 2013 (MEDLINE, Cochrane Library, Web of Knowledge, Google Scholar and references cited in other studies). Studies in which ACS outcomes with a previous history of CABG were compared with ACS outcomes with no history of previous CABG were considered for inclusion. The main endpoints of interest were mortality and non-fatal acute myocardial infarction. Data was aggregated at three follow-up times using random-effects meta-analysis models. RESULTS: Twenty-four studies were included which provided 387,181 patients for analysis. Previous CABG ACS patients were older, more diabetic and had a more frequent history of a previous myocardial infarction. Pooled in-hospital mortality was higher for the previous CABG ACS patients (OR 1.22 [1.04-1.44], p<0.01, I(2) 88%). The pooled adjusted OR showed no significant differences for the two groups (adjusted OR 1.13 [0.93-1.37], p=0.22, I(2) 92%). Previous CABG ACS patient had a higher pooled 30-day mortality (OR 1.28 [1.05-1.55], p=0.02, I(2) 74%); a higher non-adjusted (OR 1.61 [1.38-1.88], p<0.01, I(2) 70%) and adjusted (adjusted OR 1.37 [1.15-1.65], p<0.01, I(2) 0%) long-term mortality. Both the in-hospital and the long-term re-infarction rates were higher for the previous CABG ACS patients. CONCLUSIONS: According to our data, ACS patients with previous CABG history had a higher risk for short- and long-term adverse events.


Assuntos
Síndrome Coronariana Aguda/diagnóstico , Ponte de Artéria Coronária , Síndrome Coronariana Aguda/complicações , Síndrome Coronariana Aguda/cirurgia , Ponte de Artéria Coronária/estatística & dados numéricos , Feminino , Humanos , Masculino , Prognóstico , Fatores de Risco
8.
Future Microbiol ; 9(10): 1131-42, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25405883

RESUMO

AIM: Developments on synthetic molecules, such as peptide nucleic acid (PNA), make FISH procedures more robust for microbial identification. Fluorochromes use might hinder a broader implementation of PNA-FISH, but colorimetric applications are inexistent so far. METHODS: A biotin-labeled eubacteria probe was used to develop a colorimetric PNA-in situ hybridization (ISH) assay. An enzymatic-conjugate, targeting biotin, was introduced. The procedure was optimized and evaluated regarding sensitivity, specificity and detection limit. RESULTS: RESULTS have shown strong ISH signals. The method was specific, but permeabilization problems were observed for Gram-positive bacteria. Detection limit was 5 × 10(7) CFU/ml, limiting current applications to pre-enriched samples. CONCLUSION: The PNA-ISH procedure described here is a simple alternative to other detection methods, and is also the base for the development of other PNA colorimetric systems.


Assuntos
Bactérias/isolamento & purificação , Técnicas Bacteriológicas/métodos , Colorimetria/métodos , Ácidos Nucleicos Peptídicos , Bactérias/classificação , Bactérias/genética , Sensibilidade e Especificidade
9.
Mem Inst Oswaldo Cruz ; 109(4): 414-9, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25075780

RESUMO

While the influence of water in Helicobacter pylori culturability and membrane integrity has been extensively studied, there are little data concerning the effect of this environment on virulence properties. Therefore, we studied the culturability of water-exposed H. pylori and determined whether there was any relation with the bacterium's ability to adhere, produce functional components of pathogenicity and induce inflammation and alterations in apoptosis in an experimental model of human gastric epithelial cells. H. pylori partially retained the ability to adhere to epithelial cells even after complete loss of culturability. However, the microorganism is no longer effective in eliciting in vitro host cell inflammation and apoptosis, possibly due to the non-functionality of the cag type IV secretion system. These H. pylori-induced host cell responses, which are lost along with culturability, are known to increase epithelial cell turnover and, consequently, could have a deleterious effect on the initial H. pylori colonisation process. The fact that adhesion is maintained by H. pylori to the detriment of other factors involved in later infection stages appears to point to a modulation of the physiology of the pathogen after water exposure and might provide the microorganism with the necessary means to, at least transiently, colonise the human stomach.


Assuntos
Aderência Bacteriana/fisiologia , Células Epiteliais/microbiologia , Helicobacter pylori/patogenicidade , Água , Antígenos de Bactérias/fisiologia , Proteínas de Bactérias/fisiologia , Sistemas de Secreção Bacterianos , Mucosa Gástrica/citologia , Helicobacter pylori/crescimento & desenvolvimento , Interações Hospedeiro-Patógeno , Humanos , Virulência/fisiologia
10.
Mem. Inst. Oswaldo Cruz ; 109(4): 414-419, 03/07/2014. graf
Artigo em Inglês | LILACS | ID: lil-716305

RESUMO

While the influence of water in Helicobacter pylori culturability and membrane integrity has been extensively studied, there are little data concerning the effect of this environment on virulence properties. Therefore, we studied the culturability of water-exposed H. pylori and determined whether there was any relation with the bacterium’s ability to adhere, produce functional components of pathogenicity and induce inflammation and alterations in apoptosis in an experimental model of human gastric epithelial cells. H. pylori partially retained the ability to adhere to epithelial cells even after complete loss of culturability. However, the microorganism is no longer effective in eliciting in vitro host cell inflammation and apoptosis, possibly due to the non-functionality of the cag type IV secretion system. These H. pylori-induced host cell responses, which are lost along with culturability, are known to increase epithelial cell turnover and, consequently, could have a deleterious effect on the initial H. pylori colonisation process. The fact that adhesion is maintained by H. pylori to the detriment of other factors involved in later infection stages appears to point to a modulation of the physiology of the pathogen after water exposure and might provide the microorganism with the necessary means to, at least transiently, colonise the human stomach.


Assuntos
Humanos , Aderência Bacteriana/fisiologia , Células Epiteliais/microbiologia , Helicobacter pylori/patogenicidade , Água , Antígenos de Bactérias/fisiologia , Sistemas de Secreção Bacterianos , Proteínas de Bactérias/fisiologia , Mucosa Gástrica/citologia , Interações Hospedeiro-Patógeno , Helicobacter pylori/crescimento & desenvolvimento , Virulência/fisiologia
11.
J Am Soc Echocardiogr ; 27(5): 463-78, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24656882

RESUMO

The importance of the left atrium in cardiovascular performance has long been acknowledged. Quantitative assessment of left atrial (LA) function is laborious, requiring invasive pressure-volume loops and thus precluding its routine clinical use. In recent years, novel postprocessing imaging methodologies have emerged, providing a complementary approach for the assessment of the left atrium. Atrial strain and strain rate obtained using either Doppler tissue imaging or two-dimensional speckle-tracking echocardiography have proved to be feasible and reproducible techniques to evaluate LA mechanics. It is essential to fully understand the clinical applications, advantages, and limitations of LA strain and strain rate analysis. Furthermore, the technique's prognostic value and utility in therapeutic decisions also need further elucidation. The aim of this review is to provide a critical appraisal of LA mechanics. The authors describe the fundamental concepts and methodology of LA strain and strain rate analysis, the reference values reported with different imaging techniques, and the clinical implications.


Assuntos
Função Atrial , Ecocardiografia Tridimensional/métodos , Técnicas de Imagem por Elasticidade/métodos , Átrios do Coração/diagnóstico por imagem , Átrios do Coração/fisiopatologia , Disfunção Ventricular Esquerda/diagnóstico por imagem , Disfunção Ventricular Esquerda/fisiopatologia , Módulo de Elasticidade , Humanos , Resistência ao Cisalhamento
12.
J Clin Microbiol ; 51(6): 1887-93, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23596234

RESUMO

Here, we evaluated a previously established peptide nucleic acid-fluorescence in situ hybridization (PNA-FISH) method as a new diagnostic test for Helicobacter pylori clarithromycin resistance detection in paraffin-embedded gastric biopsy specimens. Both a retrospective study and a prospective cohort study were conducted to evaluate the specificity and sensitivity of a PNA-FISH method to determine H. pylori clarithromycin resistance. In the retrospective study (n = 30 patients), full agreement between PNA-FISH and PCR-sequencing was observed. Compared to the reference method (culture followed by Etest), the specificity and sensitivity of PNA-FISH were 90.9% (95% confidence interval [CI], 57.1% to 99.5%) and 84.2% (95% CI, 59.5% to 95.8%), respectively. In the prospective cohort (n = 93 patients), 21 cases were positive by culture. For the patients harboring clarithromycin-resistant H. pylori, the method showed sensitivity of 80.0% (95% CI, 29.9% to 98.9%) and specificity of 93.8% (95% CI, 67.7% to 99.7%). These values likely represent underestimations, as some of the discrepant results corresponded to patients infected by more than one strain. PNA-FISH appears to be a simple, quick, and accurate method for detecting H. pylori clarithromycin resistance in paraffin-embedded biopsy specimens. It is also the only one of the methods assessed here that allows direct and specific visualization of this microorganism within the biopsy specimens, a characteristic that allowed the observation that cells of different H. pylori strains can subsist in very close proximity in the stomach.


Assuntos
Antibacterianos/farmacologia , Claritromicina/farmacologia , Farmacorresistência Bacteriana , Infecções por Helicobacter/microbiologia , Helicobacter pylori/efeitos dos fármacos , Hibridização in Situ Fluorescente/métodos , Ácidos Nucleicos Peptídicos , Biópsia , Estudos de Coortes , Mucosa Gástrica/microbiologia , Helicobacter pylori/genética , Helicobacter pylori/isolamento & purificação , Humanos , Testes de Sensibilidade Microbiana/métodos , Sondas de Oligonucleotídeos , Estudos Prospectivos , Estudos Retrospectivos , Sensibilidade e Especificidade
13.
BMC Res Notes ; 5: 422, 2012 Aug 08.
Artigo em Inglês | MEDLINE | ID: mdl-22873690

RESUMO

BACKGROUND: Plating methods are still the golden standard in microbiology; however, some studies have shown that these techniques can underestimate the microbial concentrations and diversity. A nutrient shock is one of the mechanisms proposed to explain this phenomenon. In this study, a tentative method to assess nutrient shock effects was tested. FINDINGS: To estimate the extent of nutrient shock effects, two strains isolated from tap water (Sphingomonas capsulata and Methylobacterium sp.) and two culture collection strains (E. coli CECT 434 and Pseudomonas fluorescens ATCC 13525) were exposed both to low and high nutrient conditions for different times and then placed in low nutrient medium (R2A) and rich nutrient medium (TSA).The average improvement (A.I.) of recovery between R2A and TSA for the different times was calculated to more simply assess the difference obtained in culturability between each medium. As expected, A.I. was higher when cells were plated after the exposition to water than when they were recovered from high-nutrient medium showing the existence of a nutrient shock for the diverse bacteria used. S. capsulata was the species most affected by this phenomenon. CONCLUSIONS: This work provides a method to consistently determine the extent of nutrient shock effects on different microorganisms and hence quantify the ability of each species to deal with sudden increases in substrate concentration.


Assuntos
Bactérias/crescimento & desenvolvimento , Técnicas Bacteriológicas/métodos , Meios de Cultura/química , Estresse Fisiológico , Contagem de Colônia Microbiana , Escherichia coli/crescimento & desenvolvimento , Methylobacterium/crescimento & desenvolvimento , Viabilidade Microbiana , Pressão Osmótica , Pseudomonas fluorescens/crescimento & desenvolvimento , Sphingomonas/crescimento & desenvolvimento
14.
BMC Microbiol ; 11: 101, 2011 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-21569555

RESUMO

BACKGROUND: Triple therapy is the gold standard treatment for Helicobacter pylori eradication from the human stomach, but increased resistance to clarithromycin became the main factor of treatment failure. Until now, fastidious culturing methods are generally the method of choice to assess resistance status. In this study, a new genotypic method to detect clarithromycin resistance in clinical samples, based on fluorescent in situ hybridization (FISH) using a set of peptide nucleic acid probes (PNA), is proposed. RESULTS: The set of probes targeting the point mutations responsible for clarithromycin resistance was applied to H. pylori suspensions and showed 100% sensitivity and specificity (95% CI, 79.9-100 and 95% CI, 71.6-100 respectively). This method can also be amenable for application to gastric biopsy samples, as resistance to clarithromycin was also detected when histological slides were tested. CONCLUSIONS: The optimized PNA-FISH based diagnostic method to detect H. pylori clarithromycin resistance shown to be a very sensitive and specific method for the detection of clarithromycin resistance in the H. pylori smears and also proved to be a reliable method for the diagnosis of this pathogen in clinical samples and an alternative to existing plating methods.


Assuntos
Antibacterianos/farmacologia , Claritromicina/farmacologia , Farmacorresistência Bacteriana , Helicobacter pylori/efeitos dos fármacos , Helicobacter pylori/genética , Hibridização in Situ Fluorescente/métodos , Ácidos Nucleicos Peptídicos , Genótipo , Humanos , Testes de Sensibilidade Microbiana/métodos , Tipagem Molecular , Sondas de Oligonucleotídeos/genética , Sensibilidade e Especificidade
15.
PLoS One ; 6(3): e14786, 2011 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-21479268

RESUMO

BACKGROUND: Our current understanding of biofilms indicates that these structures are typically composed of many different microbial species. However, the lack of reliable techniques for the discrimination of each population has meant that studies focusing on multi-species biofilms are scarce and typically generate qualitative rather than quantitative data. METHODOLOGY/PRINCIPAL FINDINGS: We employ peptide nucleic acid fluorescence in situ hybridization (PNA FISH) methods to quantify and visualize mixed biofilm populations. As a case study, we present the characterization of Salmonella enterica/Listeria monocytogenes/Escherichia coli single, dual and tri-species biofilms in seven different support materials. Ex-situ, we were able to monitor quantitatively the populations of ∼56 mixed species biofilms up to 48 h, regardless of the support material. In situ, a correct quantification remained more elusive, but a qualitative understanding of biofilm structure and composition is clearly possible by confocal laser scanning microscopy (CLSM) at least up to 192 h. Combining the data obtained from PNA FISH/CLSM with data from other established techniques and from calculated microbial parameters, we were able to develop a model for this tri-species biofilm. The higher growth rate and exopolymer production ability of E. coli probably led this microorganism to outcompete the other two [average cell numbers (cells/cm(2)) for 48 h biofilm: E. coli 2,1 × 10(8) (± 2,4 × 10(7)); L. monocytogenes 6,8 × 10(7) (± 9,4 × 10(6)); and S. enterica 1,4 × 10(6) (± 4,1 × 10(5))]. This overgrowth was confirmed by CSLM, with two well-defined layers being easily identified: the top one with E. coli, and the bottom one with mixed regions of L. monocytogenes and S. enterica. SIGNIFICANCE: While PNA FISH has been described previously for the qualitative study of biofilm populations, the present investigation demonstrates that it can also be used for the accurate quantification and spatial distribution of species in polymicrobial communities. Thus, it facilitates the understanding of interspecies interactions and how these are affected by changes in the surrounding environment.


Assuntos
Bactérias/genética , Bactérias/isolamento & purificação , Biofilmes/crescimento & desenvolvimento , Hibridização in Situ Fluorescente/métodos , Ácidos Nucleicos Peptídicos/química , Bactérias/citologia , Bactérias/crescimento & desenvolvimento , Indóis , Microscopia de Fluorescência , Reprodutibilidade dos Testes , Especificidade da Espécie
16.
BMC Microbiol ; 11: 57, 2011 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-21418578

RESUMO

BACKGROUND: It is well established that Legionella pneumophila is a waterborne pathogen; by contrast, the mode of Helicobacter pylori transmission remains unknown but water seems to play an important role. This work aims to study the influence of five microorganisms isolated from drinking water biofilms on the survival and integration of both of these pathogens into biofilms. RESULTS: Firstly, both pathogens were studied for auto- and co-aggregation with the species isolated from drinking water; subsequently the formation of mono and dual-species biofilms by L. pneumophila or H. pylori with the same microorganisms was investigated. Neither auto- nor co-aggregation was observed between the microorganisms tested. For biofilm studies, sessile cells were quantified in terms of total cells by SYTO 9 staining, viable L. pneumophila or H. pylori cells were quantified using 16 S rRNA-specific peptide nucleic acid (PNA) probes and cultivable cells by standard culture techniques. Acidovorax sp. and Sphingomonas sp. appeared to have an antagonistic effect on L. pneumophila cultivability but not on the viability (as assessed by rRNA content using the PNA probe), possibly leading to the formation of viable but noncultivable (VBNC) cells, whereas Mycobacterium chelonae increased the cultivability of this pathogen. The results obtained for H. pylori showed that M. chelonae and Sphingomonas sp. help this pathogen to maintain cultivability for at least 24 hours. CONCLUSIONS: It appears that M. chelonae may have an important role in the survival of both pathogens in drinking water. This work also suggests that the presence of some microorganisms can decrease the cultivability of L. pneumophila but not the viability which indicates that the presence of autochthonous microorganisms can lead to misleading results when the safety of water is assessed by cultivable methods alone.


Assuntos
Biofilmes , Helicobacter pylori/crescimento & desenvolvimento , Legionella pneumophila/crescimento & desenvolvimento , Microbiologia da Água , Abastecimento de Água , Antibiose , Contagem de Colônia Microbiana , Comamonadaceae/crescimento & desenvolvimento , Helicobacter pylori/genética , Helicobacter pylori/isolamento & purificação , Legionella pneumophila/genética , Legionella pneumophila/isolamento & purificação , Viabilidade Microbiana , Mycobacterium chelonae/crescimento & desenvolvimento , RNA Ribossômico 16S/genética , Sphingomonas/crescimento & desenvolvimento
17.
Int J Pediatr Otorhinolaryngol ; 74(7): 807-11, 2010 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-20452684

RESUMO

OBJECTIVE: The transmission of the gastric pathogen Helicobacter pylori involves the oral route. Molecular techniques have allowed the detection of H. pylori DNA in samples of the oral cavity, although culture of H. pylori from these type of samples has been sporadic. Studies have tried to demonstrate the presence of H. pylori in adenotonsillar tissue, with contradictory results. Our aim was to clarify whether the adenotonsillar tissue may constitute an extra gastric reservoir for H. pylori. METHODS: Sixty-two children proposed for adenoidectomy or tonsillectomy were enrolled. A total of 101 surgical specimens, 55 adenoid and 46 tonsils, were obtained. Patients were characterized for the presence of anti-H. pylori antibodies by serology. On each surgical sample rapid urease test, immunohistochemistry, fluorescence in situ hybridization (FISH) with a peptide nucleic acid probe for H. pylori, and polymerase chain reaction-DNA hybridization assay (PCR-DEIA) directed to the vacA gene of H. pylori were performed. RESULTS: Thirty-nine percent of the individuals had anti-H. pylori antibodies. Rapid urease test was positive in samples of three patients, all with positive serology. Immunohistochemistry was positive in samples of two patients, all with negative serology. All rapid urease test or immunohistochemistry positive cases were negative by FISH. All samples tested were negative when PCR-DEIA for H. pylori detection was used directly in adenotonsillar specimens. CONCLUSIONS: The adenotonsillar tissue does not constitute an extra gastric reservoir for H. pylori infection, at least a permanent one, in this population of children. Moreover, techniques currently used for detecting gastric H. pylori colonization are not adequate to evaluate infection of the adenotonsillar tissues.


Assuntos
Tonsila Faríngea/microbiologia , Infecções por Helicobacter/diagnóstico , Tonsila Palatina/microbiologia , Adenoidectomia , Anticorpos Antibacterianos/sangue , Criança , DNA Bacteriano/genética , Feminino , Helicobacter pylori/imunologia , Humanos , Imuno-Histoquímica , Hibridização in Situ Fluorescente , Masculino , Sondas de Ácido Nucleico , Reação em Cadeia da Polimerase , Tonsilectomia , Urease/análise
18.
Biotechnol Lett ; 31(4): 477-85, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19116693

RESUMO

In nature, the biofilm mode of life is of great importance in the cell cycle for many microorganisms. Perhaps because of biofilm complexity and variability, the characterization of a given microbial system, in terms of biofilm formation potential, structure and associated physiological activity, in a large-scale, standardized and systematic manner has been hindered by the absence of high-throughput methods. This outlook is now starting to change as new methods involving the utilization of microtiter-plates and automated spectrophotometry and microscopy systems are being developed to perform large-scale testing of microbial biofilms. Here, we evaluate if the time is ripe to start an integrated omics approach, i.e., the generation and interrogation of large datasets, to biofilms--"biofomics". This omics approach would bring much needed insight into how biofilm formation ability is affected by a number of environmental, physiological and mutational factors and how these factors interplay between themselves in a standardized manner. This could then lead to the creation of a database where biofilm signatures are identified and interrogated. Nevertheless, and before embarking on such an enterprise, the selection of a versatile, robust, high-throughput biofilm growing device and of appropriate methods for biofilm analysis will have to be performed. Whether such device and analytical methods are already available, particularly for complex heterotrophic biofilms is, however, very debatable.


Assuntos
Biofilmes , Pesquisa Biomédica/tendências , Biologia de Sistemas/tendências
19.
Water Res ; 43(1): 229-37, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-18977505

RESUMO

The study of biofilm ecology and interactions might help to improve our understanding of their resistance mechanisms to control strategies. Concerns that the diversity of the biofilm communities can affect disinfection efficacy have led us to examine the effect of two antimicrobial agents on two important spoilage bacteria. Studies were conducted on single and dual species biofilms of Bacillus cereus and Pseudomonas fluorescens. Biofilms were formed on a stainless steel rotating device, in a bioreactor, at a constant Reynolds number of agitation (Re(A)). Biofilm phenotypic characterization showed significant differences, mainly in the metabolic activity and both extracellular proteins and polysaccharides content. Cetyl trimethyl ammonium bromide (CTAB) and glutaraldehyde (GLUT) solutions in conjunction with increasing Re(A) were used to treat biofilms in order to assess their ability to kill and remove biofilms. B. cereus and P. fluorescens biofilms were stratified in a layered structure with each layer having differential tolerance to chemical and mechanical stresses. Dual species biofilms and P. fluorescens single biofilms had both the highest resistance to removal when pre-treated with CTAB and GLUT, respectively. B. cereus biofilms were the most affected by hydrodynamic disturbance and the most susceptible to antimicrobials. Dual biofilms were more resistant to antimicrobials than each single species biofilm, with a significant proportion of the population remaining in a viable state after exposure to CTAB or GLUT. Moreover, the species association increased the proportion of viable cells of both bacteria, comparatively to the single species scenarios, enhancing each other's survival to antimicrobials and the biofilm shear stress stability.


Assuntos
Biofilmes/efeitos dos fármacos , Compostos de Cetrimônio/farmacologia , Glutaral/farmacologia , Bacillus cereus/citologia , Bacillus cereus/efeitos dos fármacos , Bacillus cereus/fisiologia , Aderência Bacteriana/efeitos dos fármacos , Reatores Biológicos , Cetrimônio , Viabilidade Microbiana/efeitos dos fármacos , Fenótipo , Pseudomonas fluorescens/citologia , Pseudomonas fluorescens/efeitos dos fármacos , Pseudomonas fluorescens/fisiologia , Especificidade da Espécie
20.
Med Chem ; 4(6): 616-23, 2008 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-18991747

RESUMO

The present work examines the potential of sesquiterpenoids to sensitize Escherichia coli and Staphylococcus aureus, and modulate their susceptibility to the standard antibiotics ciprofloxacin, erythromycin, gentamicin and vancomycin. It was tested samples of three sesquiterpenoids: guaiazulene, nerolidol (racemic mixture of the cis and trans isomers) and germacrene D enriched natural extract. Experiments were conducted aiming to assess the antimicrobial effects of the antibiotic-sesquiterpenoid combination on bacterial growth inhibition, by the disc diffusion assay and the minimum inhibitory concentration (MIC) assessment, the bactericidal effects, the post-antibiotic effect (PAE) and the effect on membrane permeability. The data related with the antimicrobial activity evidenced, through the disc diffusion assay, an antibiotic S. aureus antimicrobial activity enhancement by sesquiterpenoids presence. The MIC value for E. coli decreased significantly by sesquiterpenoids combination with ciprofloxacin, erythromycin and gentamicin, and for S. aureus, with all four selected antibiotics. This combination also increased the PAE, with the exception of guaiazulene, which seemed to quench antibiotic antimicrobial action. A moderate correlation between antimicrobial action and impairment of cell membrane function was detected for germacrene D enriched extract, and nerolidol, as single treatments and in combination with antibiotic, while a poor correlation was obtained for guaiazulene. This study provides basis for the evaluation of sesquiterpenoids as alternative or possible synergistic compounds for current antimicrobial chemotherapeutics, showing the practical utility of natural derived products to increase the susceptibility of E. coli and S. aureus.


Assuntos
Antibacterianos/farmacologia , Escherichia coli/efeitos dos fármacos , Sesquiterpenos/farmacologia , Staphylococcus aureus/efeitos dos fármacos , Membrana Celular/efeitos dos fármacos , Membrana Celular/ultraestrutura , Corantes , Testes de Sensibilidade Microbiana , Propídio
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