The regulator LdhR and the d-lactate dehydrogenase LdhA of Burkholderia multivorans play a role in carbon overflow and in planktonic cellular aggregates formation

This paper version is the accepted manuscript posted online 21 July 2017. It has peer-review. This publication hasn't any creative commons license associated. The deposited article version contains attached the supplementary materials within the pdf.LysR-type transcriptional regulators (LTTR) are the most commonly found regulators in Burkholderia cepacia complex, comprising opportunistic pathogens causing chronic respiratory infections in cystic fibrosis (CF) patients. Despite LTTRs being global regulators of pathogenicity in several bacteria, few have been characterized in Burkholderia Here, we showed that gene ldhR of B. multivorans encoding a LTTR is co-transcribed with ldhA encoding a d-lactate dehydrogenase, and evaluate their implication in virulence traits like exopolysaccharide (EPS) synthesis and biofilm formation. Comparison of wild-type (WT) and its isogenic ΔldhR mutant grown in medium with 2% d-glucose revealed a negative impact on EPS biosynthesis and on cells' viability in the presence of LdhR. Loss of viability in WT cells was caused by intracellular acidification as consequence of cumulative organic acids secretion including d-lactate, this last one absent from the ΔldhR mutant supernatant. Furthermore, LdhR is implicated in the formation of planktonic cellular aggregates. WT cell aggregates reached 1000 μm after 24 hours in liquid cultures; in contrast to ΔldhR mutant aggregates that never grew more than 60 μm. Overexpression of d-lactate dehydrogenase LdhA in the ΔldhR mutant partially restored formed aggregates size, suggesting a role for fermentation inside aggregates. Similar results were obtained for surface-attached biofilms, with WT cells producing more biofilm. A systematic evaluation of planktonic aggregates in Burkholderia CF clinical isolates showed aggregates in 40 out of 74. As CF patients' lung environment is microaerophilic and bacteria are found as free aggregates/biofilms, LdhR and LdhA might have central roles in adaptation to this environment.IMPORTANCE Cystic fibrosis patients often suffer from chronic respiratory infections caused by several microorganisms. Among them are the Burkholderia cepacia complex bacteria which cause progressive deterioration of lung function and, in some patients, might develop into fatal necrotizing pneumoniae with bacteremia, known as "cepacia syndrome". Burkholderia pathogenesis is multifactorial since they express several virulence factors, form biofilms, and are highly resistant to antimicrobial compounds, making their eradication from the CF patients' airways very difficult. As Burkholderia is commonly found in the CF lungs in the form of cell aggregates and biofilms, the need to investigate the mechanisms of cellular aggregation is obvious. In this study we demonstrate the importance of a d-lactate dehydrogenase and a regulator, in regulating carbon overflow, cellular aggregates and surface-attached biofilm formation. This not only enhances our understanding of Burkholderia pathogenesis, but can also lead to the development of drugs against these proteins to circumvent biofilm formation.Instituto Gulbenkian de Ciência (Gene Expression Unit); Programa Operacional 833 Regional de Lisboa 2020 grant: (LISBOA-01-0145-FEDER-007317); Fundação para a Ciência e a Tecnologia grants: (PTDC/QUI-BIQ/118260/2010, UID/BIO/04565/2013, SFRH/BPD/86475/2012).info:eu-repo/semantics/acceptedVersio

Essential oils as antibacterial agents against food-borne pathogens: are they really as useful as they are claimed to be ?

Original articleMost studies evaluating the use of essential oils (EO) as antibacterial agents focus mainly on minimal inhibitory concentrations (MIC) rather than minimal bactericidal concentrations (MBC). In this work, we compared MICs and MBCs of EO from condiment plants commonly used in Mediterranean Europe, namely Origanum vulgare, Salvia lavandulaefolia, Salvia officinalis, Salvia sclarea and Rosmarinus officinalis, aiming to evaluate their application as disinfecting agents in minimally processed produce. Outbreaks-related pathogens such as Listeria monocytogenes, Pseudomonas aeruginosa and Yarrowia lipolytica were used. Results showed that all EO were able to reduce bacterial growth in all bacterial strains tested, particularly O. vulgare. However, fewer EO exhibited bactericidal activities, and were only effective against one or two bacterial strains, hence eliminating the possibility to use them as broad range disinfectants. Furthermore, the necessary concentrations were too high for food application. Hence, our work suggests the need to evaluate MBC rather than MIC and questions EO usefulness in controlling undesired microorganisms. Overall, and despite the large volume of data published on EO, results obtained were not very encouraging for a realistic application on produce and question the viability of EOs as disinfecting agents in foodinfo:eu-repo/semantics/publishedVersio

β-globin haplotypes in normal and hemoglobinopathic individuals from Reconcavo Baiano, State of Bahia, Brazil

Five restriction site polymorphisms in the β-globin gene cluster (HincII-5‘ ε, HindIII-G γ, HindIII-A γ, HincII- ψβ1 and HincII-3‘ ψβ1) were analyzed in three populations (n = 114) from Reconcavo Baiano, State of Bahia, Brazil. The groups included two urban populations from the towns of Cachoeira and Maragojipe and one rural Afro-descendant population, known as the “quilombo community”, from Cachoeira municipality. The number of haplotypes found in the populations ranged from 10 to 13, which indicated higher diversity than in the parental populations. The haplotypes 2 (+ - - - -), 3 (- - - - +), 4 (- + - - +) and 6 (- + + - +) on the βA chromosomes were the most common, and two haplotypes, 9 (- + + + +) and 14 (+ + - - +), were found exclusively in the Maragojipe population. The other haplotypes (1, 5, 9, 11, 12, 13, 14 and 16) had lower frequencies. Restriction site analysis and the derived haplotypes indicated homogeneity among the populations. Thirty-two individuals with hemoglobinopathies (17 sickle cell disease, 12 HbSC disease and 3 HbCC disease) were also analyzed. The haplotype frequencies of these patients differed significantly from those of the general population. In the sickle cell disease subgroup, the predominant haplotypes were BEN (Benin) and CAR (Central African Republic), with frequencies of 52.9% and 32.4%, respectively. The high frequency of the BEN haplotype agreed with the historical origin of the afro-descendant population in the state of Bahia. However, this frequency differed from that of Salvador, the state capital, where the CAR and BEN haplotypes have similar frequencies, probably as a consequence of domestic slave trade and subsequent internal migrations to other regions of Brazil

Anti-angiogenic effects of pterogynidine alkaloid isolated from Alchornea glandulosa

<p>Abstract</p> <p>Background</p> <p>Angiogenesis, a complex multistep process that comprehends proliferation, migration and anastomosis of endothelial cells (EC), has a major role in the development of pathologic conditions such as inflammatory diseases, tumor growth and metastasis. Brazilian flora, the most diverse in the world, is an interesting spot to prospect for new chemical leads, being an important source of new anticancer drugs. Plant-derived alkaloids have traditionally been of interest due to their pronounced physiological activities. We investigated the anti-angiogenic potential of the naturally occurring guanidine alkaloid pterogynidine (Pt) isolated from the Brazilian plant <it>Alchornea glandulosa</it>. The purpose of this study was to examine which features of the angiogenic process could be disturbed by Pt.</p> <p>Methods</p> <p>Human umbilical vein endothelial cells (HUVEC) were incubated with 8 μM Pt and cell viability, proliferation, apoptosis, invasion and capillary-like structures formation were addressed. Nuclear factor κB (NFκB), a transcription factor implicated in these processes, was also evaluated in HUVEC incubated with Pt. Quantifications were expressed as mean ± SD of five independent experiments and one-way analysis of variance (ANOVA) followed by the Dunnet test was used.</p> <p>Results</p> <p>A significant decrease in proliferation and invasion capacity and an effective increase in apoptosis as assessed by bromodeoxyuridine (BrdU), double-chamber and terminal transferase dUTP nick end labeling (TUNEL) assay, respectively, have been found. Pt also led to a drastic reduction in the number of capillary-like structures formation when HUVEC were cultured on growth factor reduced-Matrigel (GFR-Matrigel) coated plates. In addition, incubation of HUVEC with Pt resulted in reduced NFκB activity.</p> <p>Conclusion</p> <p>These findings emphasize the potential use of Pt against pathological situations where angiogenesis is stimulated as tumor development.</p

Quinze anos de ecografia intravascular coronária emintervenc ̧ão coronária percutânea em Portugal

INTRODUCTION: Coronary intravascular ultrasound (IVUS) is increasingly important in catheterization laboratories due to its positive prognostic impact. This study aims to characterize the use of IVUS in percutaneous coronary intervention (PCI) in Portugal. METHODS: A retrospective observational study was performed based on the Portuguese Registry on Interventional Cardiology of the Portuguese Society of Cardiology. The clinical and angiographic profiles of patients who underwent PCI between 2002 and 2016, the percentage of IVUS use, and the coronary arteries assessed were characterized. RESULTS: A total of 118 706 PCIs were included, in which IVUS was used in 2266 (1.9%). Over time, use of IVUS changed from none in 2002 to generally increasing use from 2003 (0.1%) to 2016 (2.4%). The age of patients in whom coronary IVUS was used was similar to that of patients in whom IVUS was not used, but in the former group there were fewer male patients, and a higher prevalence of cardiovascular risk factors (hypertension, hypercholesterolemia and diabetes), previous myocardial infarction, previous PCI, multivessel coronary disease, C-type or bifurcated coronary lesions, and in-stent restenosis. IVUS was used in 54.8% of elective PCIs and in 19.15% of PCIs of the left main coronary artery. CONCLUSION: Coronary IVUS has been increasingly used in Portugal since 2003. It is used preferentially in elective PCIs, and in patients with higher cardiovascular risk, with more complex coronary lesions and lesions of the left main coronary artery.info:eu-repo/semantics/publishedVersio