Publications by authors named "Ana Azevedo"

372 Publications

Factors associated with Leishmania infection in dogs and geospatial analysis in the Sertão of Paraíba, Northeast Brazil.

Rev Bras Parasitol Vet 2021 28;30(2):e023620. Epub 2021 May 28.

Laboratório de Biologia Molecular do Semiárido, Programa de Pós-graduação em Ciência e Saúde Animal, Universidade Federal de Campina Grande - UFCG, Patos, PB, Brasil.

Visceral leishmaniasis (VL) is a zoonosis with a worldwide distribution that has a major impact on public health. The aim of this study was to verify the prevalence of canine infection by Leishmania infantum, the factors associated with the infection and its spatial distribution in the municipality of Mãe D'Água, in the Sertão region of Paraíba State, Northeast Brazil. Blood samples were collected from 150 dogs for diagnosis by the DPP®, ELISA-S7®, ELISA-EIE® and qPCR assays. The prevalence was calculated considering the positivity in at least two tests. SaTScan® was used for spatial analysis. The prevalence of canine infection with Leishmania was 18.6% (28/150), with the rural area being identified as a risk factor (Odds Ratio (OR) = 2.93). The permanence of the dog loose during the night (OR = 0.33) and deworming (OR = 0.30) were identified as protective factors. A risk cluster was formed in the northern region of the urban area. Mãe D'Água showed a pattern of active transmission in the rural area, but VL control measures also need to be carried out in the urban area to prevent human cases and the spread of the disease in the risk zone.
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http://dx.doi.org/10.1590/S1984-29612021047DOI Listing
June 2021

Genomic Selection in Tropical Forage Grasses: Current Status and Future Applications.

Front Plant Sci 2021 30;12:665195. Epub 2021 Apr 30.

Embrapa Gado de Leite, Juiz de Fora, Brazil.

The world population is expected to be larger and wealthier over the next few decades and will require more animal products, such as milk and beef. Tropical regions have great potential to meet this growing global demand, where pasturelands play a major role in supporting increased animal production. Better forage is required in consonance with improved sustainability as the planted area should not increase and larger areas cultivated with one or a few forage species should be avoided. Although, conventional tropical forage breeding has successfully released well-adapted and high-yielding cultivars over the last few decades, genetic gains from these programs have been low in view of the growing food demand worldwide. To guarantee their future impact on livestock production, breeding programs should leverage genotyping, phenotyping, and envirotyping strategies to increase genetic gains. Genomic selection (GS) and genome-wide association studies play a primary role in this process, with the advantage of increasing genetic gain due to greater selection accuracy, reduced cycle time, and increased number of individuals that can be evaluated. This strategy provides solutions to bottlenecks faced by conventional breeding methods, including long breeding cycles and difficulties to evaluate complex traits. Initial results from implementing GS in tropical forage grasses (TFGs) are promising with notable improvements over phenotypic selection alone. However, the practical impact of GS in TFG breeding programs remains unclear. The development of appropriately sized training populations is essential for the evaluation and validation of selection markers based on estimated breeding values. Large panels of single-nucleotide polymorphism markers in different tropical forage species are required for multiple application targets at a reduced cost. In this context, this review highlights the current challenges, achievements, availability, and development of genomic resources and statistical methods for the implementation of GS in TFGs. Additionally, the prediction accuracies from recent experiments and the potential to harness diversity from genebanks are discussed. Although, GS in TFGs is still incipient, the advances in genomic tools and statistical models will speed up its implementation in the foreseeable future. All TFG breeding programs should be prepared for these changes.
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http://dx.doi.org/10.3389/fpls.2021.665195DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8120112PMC
April 2021

Thyroid Hormones within the Normal Range and Cardiac Function in the General Population: The EPIPorto Study.

Eur Thyroid J 2021 Apr 8;10(2):150-160. Epub 2020 Jul 8.

EPIUnit - Instituto de Saúde Pública, Universidade do Porto, Porto, Portugal.

Background: Hypothyroidism and hyperthyroidism are associated with marked changes in cardiac structure and function. However, the association of thyroid function within the normal range with cardiac structure and function in the general population remains uncertain.

Methods: Eight hundred thirty-five subjects aged ≥45 years from the EPIPorto cohort (evaluation between 2006 and 2008) were cross-sectionally analyzed. We excluded participants with TSH, free T4 (FT4), or free T3 (FT3) outside of the reference range or with self-reported cardiovascular or thyroid disease. Cardiac structure and function were evaluated by echocardiography. We used linear regression models unadjusted and adjusted for sex and age (model 1), and sex, age, BMI, diabetes, hypertension, and smoking (model 2). Nonlinear associations were assessed using restricted cubic splines.

Results: The mean age was 61.5 years (SD 10.5); 61.1% of the patients were women. In the adjusted model 2, heart rate was positively associated with FT3; diastolic blood pressure was positively associated with TSH; LV end-diastolic and end-systolic volumes were inversely associated with TSH, and ejection fraction was nonlinearly associated with FT3, with higher ejection fractions near the limits of the reference range. Left ventricle (LV) posterior wall thickness was nonlinearly associated with FT4 in the adjusted model 1, with a greater thickness near the limits of the reference range. Regarding diastolic function, no significant associations were observed in adjusted models.

Conclusions: Thyroid function within the reference range was associated with heart rate, blood pressure, cardiac structure, and function. Increasing thyroid function (lower TSH, higher FT4, or higher FT3) was associated with a higher heart rate, a lower diastolic blood pressure, and larger LV volumes. LV wall thickness and ejection fraction had a U-shaped association with thyroid hormones.
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http://dx.doi.org/10.1159/000508407DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8077596PMC
April 2021

Manufacturing of bacteriophages for therapeutic applications.

Biotechnol Adv 2021 Jul-Aug;49:107758. Epub 2021 Apr 22.

iBB - Institute for Bioengineering and Biosciences, Department of Bioengineering, Instituto Superior Técnico, Universidade de Lisboa, Av. Rovisco Pais, 1, 1049-001 Lisbon, Portugal. Electronic address:

Bacteriophages, or simply phages, are the most abundant biological entities on Earth. One of the most interesting characteristics of these viruses, which infect and use bacteria as their host organisms, is their high level of specificity. Since their discovery, phages became a tool for the comprehension of basic molecular biology and originated applications in a variety of areas such as agriculture, biotechnology, food safety, veterinary, pollution remediation and wastewater treatment. In particular, phages offer a solution to one of the major problems in public health nowadays, i.e. the emergence of multidrug-resistant bacteria. In these situations, the use of virulent phages as therapeutic agents offers an alternative to the classic, antibiotic-based strategies. The development of phage therapies should be accompanied by the improvement of phage biomanufacturing processes, both at laboratory and industrial scales. In this review, we first present some historical and general aspects related with the discovery, usage and biology of phages and provide a brief overview of the most relevant phage therapy applications. Then, we showcase current processes used for the production and purification of phages and future alternatives in development. On the production side, key factors such as the bacterial physiological state, the conditions of phage infection and the operation parameters are described alongside with the different operation modes, from batch to semi-continuous and continuous. Traditional purification methods used in the initial phage isolation steps are then described followed by the presentation of current state-of-the-art purification approaches. Continuous purification of phages is finally presented as a future biomanufacturing trend.
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http://dx.doi.org/10.1016/j.biotechadv.2021.107758DOI Listing
April 2021

Neuro-COVID frequency and short-term outcome in the Northern Portuguese population.

Eur J Neurol 2021 Apr 21. Epub 2021 Apr 21.

Neurology Department, Centro Hospitalar Universitário do Porto-Hospital de Santo António, Porto, Portugal.

Background And Purpose: COVID-19-related acute neurological phenotypes are being increasingly recognised, with neurological complications reported in more than 30% of hospitalised patients. However, multicentric studies providing a population-based perspective are lacking.

Methods: We conducted a retrospective multicentric study at five hospitals in Northern Portugal, representing 45.1% of all hospitalised patients in this region, between 1 March and 30 June 2020.

Results: Among 1261 hospitalised COVID-19 patients, 457 (36.2%) presented neurological manifestations, corresponding to a rate of 357 per 1000 in the North Region. Patients with neurologic manifestations were younger (68.0 vs. 71.2 years, p = 0.002), and the most frequent neurological symptoms were headache (13.4%), delirium (10.1%), and impairment of consciousness (9.7%). Acute well-defined central nervous system (CNS) involvement was found in 19.1% of patients, corresponding to a rate of 217 per 1000 hospitalised patients in the whole region. Assuming that all patients with severe neurological events were hospitalised, we extrapolated our results to all COVID-19 patients in the region, estimating that 116 will have a severe neurological event, corresponding to a rate of nine per 1000 (95% CI = 7-11). Overall case fatality in patients presenting neurological manifestations was 19.8%, increasing to 32.6% among those with acute well-defined CNS involvement.

Conclusions: We characterised the population of hospitalised COVID-19 patients in Northern Portugal and found that neurological symptoms are common and associated with a high degree of disability at discharge. CNS involvement with criteria for in-hospital admission was observed in a significant proportion of patients. This knowledge provides the tools for adequate health planning and for improving COVID-19 multidisciplinary patient care.
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http://dx.doi.org/10.1111/ene.14874DOI Listing
April 2021

Development of an automated yeast-based spectrophotometric method for toxicity screening: Application to ionic liquids, GUMBOS, and deep eutectic solvents.

Chemosphere 2021 Aug 18;277:130227. Epub 2021 Mar 18.

LAQV, REQUIMTE, Departamento de Ciências Químicas, Faculdade de Farmácia, Universidade do Porto, Porto, Portugal. Electronic address:

Saccharomyces cerevisiae has been used as a eukaryotic model organism for studying the toxic effects of various compounds. In this context, an automated spectrophotometric method based on the enzymatic reduction of methylene blue dye to a colorless product by living yeast cells was implemented in a sequential injection analysis system. Loss of yeast viability/impaired metabolic activity was monitored by an increase in optical density at 664 nm. To prove the usefulness of this approach, the toxicity of ILs (ionic liquids), GUMBOS (group of uniform materials based on organic salts), and DESs (deep eutectic solvents) was examined. Differences obtained between IC values confirmed the impact of structural elements on each compounds' toxicity. While DESs appeared to be less toxic than ILs, GUMBOS were found to be among the most toxic compounds to yeast cells and thus can be viewed as promising antimicrobial candidates. The automated methodology showed satisfactory repeatability and reproducibility (RSD < 9%), which is in good agreement with Green Chemistry principles. In fact, the method required consumption of only 40 μL of reagents and produced less than 2 mL of effluents per cycle. Thus, the developed assay can be used as an alternative tool for toxicity screening.
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http://dx.doi.org/10.1016/j.chemosphere.2021.130227DOI Listing
August 2021

mRNA vaccines manufacturing: Challenges and bottlenecks.

Vaccine 2021 04 24;39(16):2190-2200. Epub 2021 Mar 24.

Department of Biochemical Engineering, University College London, Bernard Katz Building, Gordon Street, London WC1H 0AH, United Kingdom. Electronic address:

Vaccines are one of the most important tools in public health and play an important role in infectious diseases control. Owing to its precision, safe profile and flexible manufacturing, mRNA vaccines are reaching the stoplight as a new alternative to conventional vaccines. In fact, mRNA vaccines were the technology of choice for many companies to combat the Covid-19 pandemic, and it was the first technology to be approved in both United States and in Europe Union as a prophylactic treatment. Additionally, mRNA vaccines are being studied in the clinic to treat a number of diseases including cancer, HIV, influenza and even genetic disorders. The increased demand for mRNA vaccines requires a technology platform and cost-effective manufacturing process with a well-defined product characterisation. Large scale production of mRNA vaccines consists in a 1 or 2-step in vitro reaction followed by a purification platform with multiple steps that can include Dnase digestion, precipitation, chromatography or tangential flow filtration. In this review we describe the current state-of-art of mRNA vaccines, focusing on the challenges and bottlenecks of manufacturing that need to be addressed to turn this new vaccination technology into an effective, fast and cost-effective response to emerging health crises.
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http://dx.doi.org/10.1016/j.vaccine.2021.03.038DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7987532PMC
April 2021

First-trimester miscarriage: A histopathological classification proposal.

Heliyon 2021 Mar 8;7(3):e06359. Epub 2021 Mar 8.

School of Medicine, University of Minho, Surgical Sciences Domain Research, Campus de Gualtar, 4710-057, Braga, Portugal.

Background: Histopathology of first-trimester abortion products may be useful in document an intrauterine pregnancy, identifying an important pathology affecting the mother or the embryo and diagnosing conditions that are likely to recur in future pregnancies or that explain the adverse fetal outcome. Relevant information provided by histology is essential to determine the cause and to guide the patients with early pregnancy failure.

Aims: Histopathological classification proposal in first-trimester miscarriage.

Methods: Published pathological criteria in first-trimester abortion specimens were collected, standardized and focused into a comprehensive diagnosis. The idea was to create a comprehensive classification related to major pathophysiological processes. Thus, the histological criteria were grouped into 7 categories: i. Changes suggesting aneuploidy (CSA) or metabolic storage disease; ii. Embryo anomaly (EA); iii. Multifactorial (MF) causes; iv. Maternal causes (MC); v. Gestational trophoblastic disease, such as hydatidiform mole (HM) and non neoplastic lesions and neoplasms; vi. Ectopic pregnancy; vii. Other. So, a 6-years retrospective study of first-trimester spontaneous miscarriage were reviewed. Two groups were created: i. Study group include specimens with pathological diagnosis; ii. Control group incorporate specimens with pathological diagnosis and additional genetic study in order to validate pathological criteria.

Results: Pathological criteria concordance between inter-observers was generally good, with an excellent correlation in EA and HM categories. Despite greater inter-observer disagreement in the CSA and MC categories the correlation with the genetic results was very positive.

Conclusion: A standardized, reproducible and biologically comprehensive histopathological classification may improve fetal follow-up and couple's management.
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http://dx.doi.org/10.1016/j.heliyon.2021.e06359DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7969342PMC
March 2021

A Scoping Review of Process Indicators for Measuring Quality of Care in Glaucoma.

J Glaucoma 2021 05;30(5):e198-e204

Surgery and Physiology and Cardiovascular R&D Center, Faculty of Medicine, Universidade do Porto.

Prcis: There are no standardized process quality indicators (QIs) in glaucoma care. Although they can be inferred from guidelines and trials, they should be designed and standardized to allow better assessment of the quality of care.

Purpose: QIs are crucial for assessing the performance of any health care system. To allow efficiency, effectiveness, and patient-centeredness, there is a need for prompt acquisition of up-to-date information. Among the available QIs, process indicators have the highest sensitivity to frequent changes and could better reflect the implementation outcomes of novel ideas and technology. This study aimed to map the available information regarding process QIs in glaucoma care, identify the current development stage of these indicators, and systematically synthesize them.

Materials And Methods: We performed a scoping review of 4 electronic bibliographic databases for studies reporting on process QIs in glaucoma. We retrieved 7502 references and created a domain list reflecting the core idea underlying each indicator.

Results: We summarized information from 18 documents and listed 20 domains. The most mentioned domains were follow-up, optic nerve head assessment, visual field test, and intraocular pressure. Indicators regarding the quality of life assessment, patient assistance, or presence of written protocols were less frequently mentioned.

Conclusions: There are notable variations among process QIs in glaucoma and significant heterogeneity in their descriptions in published studies. Although novel indicators can be inferred from guidelines and trials, they should be designed and standardized for better assessment of performance in health systems to improve their quality.
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http://dx.doi.org/10.1097/IJG.0000000000001825DOI Listing
May 2021

Beyond Val30Met transthyretin (TTR): variants associated with age-at-onset in hereditary ATTRv amyloidosis.

Amyloid 2021 Jun 18;28(2):100-106. Epub 2021 Jan 18.

UnIGENe, IBMC - Institute for Molecular and Cell Biology, i3S - Instituto de Investigação e Inovação em Saúde, Universidade do Porto, Porto, Portugal.

Objectives: V30M in transthyretin () gene is causative for hereditary ATTRv amyloidosis (familial amyloid polyneuropathy). ATTRv amyloidosis shows a wide variation in age-at-onset (AO) between clusters, families, and among generations. We aim at identifying genetic modifiers of disease onset that may contribute to this variability in Portuguese patients by identifying other variants in , beyond the ATTRv amyloidosis causing variant that could play a regulatory role in its expression level.

Methods: We analysed DNA samples of 330 ATTRV30M carriers (299 patients, 31 aged-asymptomatic carriers aged >40 years) from 120 families currently under follow-up. A generalised estimating equation analysis (GEE) was used to take into account non-independency of AO between relatives. An intensive analysis was performed in order to understand a possible regulation of gene expression.

Results: We found 11 rare variants in the promoter, coding and intron/exon boundaries of the gene associated with the onset of symptoms before and after age 40 years, namely 2 novel ones and a tandem CA-dinucleotide repeat. Furthermore, of the 4 common variants found, one was significantly associated with AO and may influence the constitutive splicing of TTR pre-mRNA. The seven ATTRV30M/V30M homozygous do not carry any of the variants identified in this study, including the common ones. analysis disclosed significant alterations in the mechanism of splicing, transcription factors and miRNAs binding.

Conclusions: Variants within the promoter region may modify disease expressivity and variants in the 3'UTR can impact the efficacy of novel therapeutic interventions. Importantly, the putative mechanisms of regulation of gene expression within the gene deserve to be better explored, in order to be used in the future as potential therapeutical targets.
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http://dx.doi.org/10.1080/13506129.2020.1857236DOI Listing
June 2021

Definition of baseline renal function in hospitalised patients with multiple preadmission measures of serum creatinine-A joint modelling approach.

Nephrology (Carlton) 2020 Dec 7;25(12):897-905. Epub 2020 Sep 7.

EPIUnit - Instituto de Saúde Pública, Universidade do Porto, Porto, Portugal.

Aim: With the use of a joint model (JM) we investigated how different definitions of baseline serum creatinine (SCr) would affect the association between an acute increase in inpatients' SCr and 30-day mortality and whether this effect depends on premorbid SCr trajectory.

Methods: This was a retrospective study including adult patients admitted to a tertiary acute-care hospital in Porto, Portugal, between January 1, 2013, and December 31, 2015, who had at least two preadmission ambulatory and two inpatients SCr measurements. The baseline SCr was defined as the lowest (-min), the most recent (-last) or the median (-medi) value over the preadmission period. The JM combined a linear mixed model for repeated inpatient SCr relative to baseline value and a Cox proportional survival model. Preadmission SCr courses were identified using linear regression and subsequently clustered based upon a patient-specific slope. Preadmission SCr trajectories were described as median SCr courses within clusters.

Results: SCr trajectories were: "Stable" (78.0% of patients), "Decreasing" (11.3%) and "Increasing" (10.7%). Overall, an increase in inpatient SCr by 50% relative to baseline SCr-min raised the risk of 30-day mortality by 74%; the estimate was not different from hazard ratio (HR) obtained for SCr-last (1.78) and SCr-medi (1.71). We found no differences in HR across preadmission trajectories.

Conclusion: The increased risk of death associated with an abrupt rise in inpatient SCr depends neither on the definition of baseline SCr nor patients' SCr trajectory before hospitalisation. Preadmission SCr-medi value may be the least biased estimate of the baseline renal function.
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http://dx.doi.org/10.1111/nep.13768DOI Listing
December 2020

Skeletal Anomalies in Senegalese Sole ( Kaup) Fed with Different Commercial Enriched A Study in Postlarvae and Juveniles.

Animals (Basel) 2020 Dec 24;11(1). Epub 2020 Dec 24.

Department of Anatomy, Animal Production and Clinical Veterinary Sciences, Faculty of Veterinary Science, University of Santiago de Compostela, Campus Universitario S/N, 27002 Lugo, Spain.

The high incidence of skeletal anomalies in Senegalese sole () still constitutes a bottleneck constraining its production. There are diverse commercially available products for the enrichment of live preys, but few reports of their influence on skeletogenesis in Senegalese sole. This study evaluated the presence of vertebral anomalies in postlarvae and juvenile Senegalese sole fed with spp. metanauplii enriched with four commercial products (EA, EB, EC, and ED) in a fish farm. The most frequent alterations consisted of deformations of the neural/haemal arches and spines and fusions and deformations of hypurals, epural, or parhypural. The correspondence analysis ordered fish from each age in separated semiaxis, indicating the presence of different anomaly patterns for the two sampled stages. The results showed only very light changes in the frequency of vertebral abnormalities among tested enrichment products, i.e., individuals from EC and EA lots displayed less vertebral body anomalies and/or vertebral column deviations at 31 and 105 days after hatching, respectively. The existence of a large shared malformation pattern in all the experimental groups leads to impute to the rearing conditions as the main driving factor of the onset of such group of anomalies, probably masking some dietary effect.
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http://dx.doi.org/10.3390/ani11010022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7823604PMC
December 2020

Estimation of Missing Baseline Serum Creatinine for Acute Kidney Injury Diagnosis in Hospitalized Patients.

Nephron 2021 18;145(2):123-132. Epub 2020 Dec 18.

EPIUnit - Instituto de Saúde Pública, Universidade do Porto, Porto, Portugal.

Introduction: In hospitalized patients, information on preadmission kidney function is often missing, impeding timely and accurate acute kidney injury (AKI) detection and affecting results of AKI-related studies.

Objective: In this study, we provided estimates of preadmission serum creatinine (SCr), based on a multivariate linear regression (Model 1) and random forest model (Model 2) built with different parametrizations. Their accuracy for AKI diagnosis was compared with the accuracy of commonly used surrogate methods: (i) SCr at hospital admission (first SCr) and (ii) SCr back-calculated from the assumed estimated glomerular filtration rate of 75 mL/min/1.73 m2 (eGFR 75).

Methods: From 44,670 unique adult admissions to a tertiary referral centre between 2013 and 2015, we analysed 8,540 patients with preadmission SCr available. To control for differences in characteristics of patients with and without SCr, we used an inverse probability weighting technique.

Results: Estimates of SCr were likely to be higher than true preadmission SCr in a low Cr concentration and undervalued in high concentrations although for Model 2 Complete-SCr these differences were smallest. The true cumulative incidence of AKI was 14.8%. Model 2 Complete-SCr had the best agreement for AKI diagnosis (kappa 0.811, 95% CI 0.787-0.835), while surrogate methods resulted in the lowest agreement: (kappa 0.553, 0.516-0.590) and (0.648, 0.620-0.676) for first SCr and eGFR 75, respectively.

Conclusions: Multivariable imputation of preadmission SCr, taking into account elementary admission data, improved accuracy in AKI diagnosis over commonly used surrogate methods. Random forest-based models can serve as an effective tool in research.
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http://dx.doi.org/10.1159/000512080DOI Listing
December 2020

Conversion methods for modified Jaffe reaction assays of serum creatinine.

Porto Biomed J 2020 May-Jun;5(3):e72. Epub 2020 Jun 4.

EPIUnit-Instituto de Saúde Pública, Universidade do Porto.

Background: Modifications in Jaffe serum creatinine (sCr) assays question the suitability of the results for direct comparison.

Methods: sCr in adult in-patients was routinely measured either by SRM 909-standardized/noncompensated (method A) or isotope dilution mass spectrometry traceable/compensated method (reference). We converted values by method A into values by the reference using a formula provided by the manufacturer [Beckman Coulter (BC)] and traditional equating methods.

Results: The BC-based conversion and linear equating resulted in underestimated sCr values, whereas equipercentile equating (EE) provided sCr with not significantly different distribution from the reference values. Proportions of patients with renal impairment did not differ between the reference and EE-converted sCr, as opposed to BC-recalculated values. Three percent of patients were classified into better renal function category when applying BC versus EE conversion.

Conclusions: Equipercentile equation was a more accurate method for recalculation of sCr obtained from different Jaffe reaction assays than the linear equating or the BC linear formula. This study emphasizes the importance of the derivation sample specificity when applying research results to other real-world populations.
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http://dx.doi.org/10.1097/j.pbj.0000000000000072DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7722398PMC
June 2020

Bio-Based Nanoparticles as a Carrier of β-Carotene: Production, Characterisation and In Vitro Gastrointestinal Digestion.

Molecules 2020 Sep 30;25(19). Epub 2020 Sep 30.

INL-International Iberian Nanotechnology Laboratory, 4715-330 Braga, Portugal.

β-carotene loaded bio-based nanoparticles (NPs) were produced by the solvent-displacement method using two polymers: zein and ethylcellulose. The production of NPs was optimised through an experimental design and characterised in terms of average size and polydispersity index. The processing conditions that allowed to obtain NPs (<100 nm) were used for β-carotene encapsulation. Then β-carotene loaded NPs were characterised in terms of zeta potential and encapsulation efficiency. Transmission electron microscopy, Fourier transform infrared spectroscopy and X-ray diffraction analysis were performed for further morphological and chemical characterisation. In the end, a static in vitro digestion following the INFOGEST protocol was performed and the bioaccessibility of β-carotene encapsulated in both NPs was determined. Results show that the best conditions for a size-controlled production with a narrow size distribution are lower polymer concentrations and higher antisolvent concentrations. The encapsulation of β-carotene in ethylcellulose NPs resulted in nanoparticles with a mean average size of 60 ± 9 nm and encapsulation efficiency of 74 ± 2%. β-carotene loaded zein-based NPs resulted in a mean size of 83 ± 8 nm and encapsulation efficiency of 93 ± 4%. Results obtained from the in vitro digestion showed that β-carotene bioaccessibility when encapsulated in zein NPs is 37 ± 1%, which is higher than the value of 8.3 ± 0.1% obtained for the ethylcellulose NPs.
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http://dx.doi.org/10.3390/molecules25194497DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7582681PMC
September 2020

GUMBOS and nanoGUMBOS in chemical and biological analysis: A review.

Anal Chim Acta 2020 Oct 9;1133:180-198. Epub 2020 Jul 9.

LAQV, REQUIMTE, Departamento de Ciências Químicas, Laboratório de Química Aplicada, Faculdade de Farmácia, Universidade do Porto, Rua Jorge Viterbo Ferreira, 228, 4050-313, Porto, Portugal. Electronic address:

GUMBOS (group of uniform materials based on organic salts) is a novel class of materials that exhibits similar features to those of ionic liquids, but have melting points between 25 and 250 °C. GUMBOS can be easily converted into nanomaterials (nanoGUMBOS), with advantages of working at nanoscale. Due to the huge number of possible cation-anion combinations, these materials can be multifunctional and designed for a specific task. This review highlights the possibility of fine-tuning GUMBOS physical and chemical properties in view of changing their ionic counterparts. Their outstanding potential for analytical applications is shown through recent developments in areas such as sensing, and solid-phase extraction. Available methods for synthesis of nanoGUMBOS, and their different outcomes in shapes and optical properties are described, with pros and cons being outlined. Finally, an analysis is made of opportunities and challenges faced by this class of organic ionic materials.
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http://dx.doi.org/10.1016/j.aca.2020.06.028DOI Listing
October 2020

Purification of Plasmid DNA by Multimodal Chromatography.

Methods Mol Biol 2021 ;2197:193-205

Department of Bioengineering, Institute for Bioengineering and Biosciences, Instituto Superior Técnico, Universidade de Lisboa, Lisbon, Portugal.

Multimodal (MM) chromatography can be described as a chromatographic method that uses more than one mode of interaction between the target molecule and the ligand to achieve a particular separation. Owing to its advantages over traditional chromatography, such as higher selectivity and capacity, its application for the purification of biomolecules with therapeutic interest has been widely studied. The potential of MM chromatography for the purification of plasmid DNA has been demonstrated. In this chapter, a downstream process for the purification of supercoiled plasmid DNA using MM chromatography with two different ligands-Capto™ adhere and PPA HyperCell™-is described. In both the cases, the purification process yields a high purity and highly homogeneous sc plasmid product.
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http://dx.doi.org/10.1007/978-1-0716-0872-2_10DOI Listing
March 2021

Primary Purification of Plasmid DNA Using Differential Isopropanol Precipitation.

Methods Mol Biol 2021 ;2197:151-165

Department of Bioengineering, Institute for Bioengineering and Biosciences, Instituto Superior Técnico, Universidade de Lisboa, Lisbon, Portugal.

A method for the intermediate recovery of plasmid DNA (pDNA) from alkaline lysates is described that comprises differential isopropanol precipitation steps. In a first low-cut precipitation, a smaller amount of isopropanol (20% v/v) is used so that only high molecular weight RNA precipitates. After solid liquid separation, a high-cut precipitation is performed by bringing isopropanol concentration to 70% v/v to precipitate pDNA. Tests made with lysates show that the differential precipitation increases purity threefold compared to the conventional one-step precipitation at 70% v/v without affecting pDNA recovery (>80%).
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http://dx.doi.org/10.1007/978-1-0716-0872-2_8DOI Listing
March 2021

Correlation between the Spread of COVID-19 and the Interest in Personal Protective Measures in Poland and Portugal.

Healthcare (Basel) 2020 Jul 9;8(3). Epub 2020 Jul 9.

CEOS.PP, Porto Accounting and Business School, Polytechnic Institute of Porto, 4200-465 Porto, Portugal.

The pandemic of the coronavirus disease 2019 (COVID-19), has gained extensive coverage in public media and global news, generated international and national communication campaigns to educate the communities worldwide and raised the attention of everyone. The coronavirus has caused viral pneumonia in tens of thousands of people around the world, and the COVID-19 outbreak changed most countries' routines and concerns and transformed social behaviour. This study explores the potential use of Google Trends (GT) in monitoring interest in the COVID-19 outbreak and, specifically, in personal protective equipment and hand hygiene, since these have been promoted by official health care bodies as two of the most protective measures. GT was chosen as a source of reverse engineering data, given the interest in the topic and the novelty of the research. Current data on COVID-19 are retrieved from GT using keywords in two languages-Portuguese and Polish. The geographical settings for GT are two countries: Poland and Portugal. The period under analysis is 20 January 2020, when the first cases outside China were known, to 15 June 2020. The results show that there is a correlation between the spread of COVID-19 and the search for personal protective equipment and hand hygiene and that GT can help, to a certain extent, understand people's concerns, behaviour and reactions to sanitary problems and protection recommendations.
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http://dx.doi.org/10.3390/healthcare8030203DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7551869PMC
July 2020

Patient and system delays in the treatment of acute coronary syndrome.

Rev Port Cardiol (Engl Ed) 2020 Mar 6;39(3):123-131. Epub 2020 May 6.

EPIUnit - Instituto de Saúde Pública, Universidade do Porto, Porto, Portugal; Departamento de Ciências da Saúde Pública e Forenses e Educação Médica, Faculdade de Medicina da Universidade do Porto, Porto, Portugal.

Introduction: Early reperfusion for patients with ST-segment elevation myocardial infarction (STEMI) is indicated by the European Society of Cardiology, while a timely invasive strategy is recommended for patients with high-risk and intermediate-risk non-ST-elevation acute coronary syndromes (NSTE-ACS). This study aims to assess patient and system delays according to diagnosis and risk profile, and to identify predictors of prolonged delay.

Methods: We assembled a cohort of patients (n=939) consecutively admitted to the cardiology department of two hospitals, one in the metropolitan area of Porto and one in the north-east region of Portugal, between August 2013 and December 2014.

Results: The proportion of patients with time from symptom onset to first medical contact (FMC) ≥120 min was highest among high-risk NSTE-ACS (57.7%), followed by intermediate-risk NSTE-ACS (52.1%) and STEMI (43.3%). Regardless of diagnosis and risk stratification, use of own transportation and inability to interpret cardiac symptoms correctly were associated with prolonged delays. Regarding system delays, we found that 78.0% of patients with STEMI and 65.8% of patients with high-risk NSTE-ACS were treated in a timeframe exceeding the recommended limits. Admission to a non-percutaneous coronary intervention-capable hospital, admission on weekends and complications at admission were associated with prolonged delays to treatment.

Conclusions: Due to both patient and system delays, a large proportion of STEMI and high-risk NSTE-ACS patients still fail to have access to timely reperfusion.
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http://dx.doi.org/10.1016/j.repc.2019.07.007DOI Listing
March 2020

Anything but Conventional Chromatography Approaches in Bioseparation.

Biotechnol J 2020 Aug 11;15(8):e1900274. Epub 2020 May 11.

Department of Biotechnology, Delft University of Technology, Van der Maasweg 9, Delft, 2629 HZ, The Netherlands.

While packed bed chromatography, known as conventional chromatography, has been serving the biopharmaceutical industry for decades as the bioseparation method of choice, alternative approaches are likely to take an increasing leading role in the next few years. The high number of new biological drugs under development, and the need to make biopharmaceuticals widely accessible, has been driving the academia and industry in the quest of anything but conventional chromatography approaches. In this perspective paper, these alternative approaches are discussed in view of current and future challenges in the downstream processing field.
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http://dx.doi.org/10.1002/biot.201900274DOI Listing
August 2020

Development of microsatellite markers for Lippia alba and related Lippia species.

Mol Biol Rep 2020 Jun 23;47(6):4911-4915. Epub 2020 Apr 23.

Departamento de Biologia, Instituto de Ciências Biológicas, Universidade Federal de Juiz de Fora (UFJF), Juiz de Fora, Minas Gerais, 36036-900, Brazil.

Microsatellite primers were developed in Lippia alba complex to better understanding the origins and evolution of the species. We sought to increase the numbers of available simple sequence repeat (SSR) markers. We performed low-coverage (~ twofold) genomic DNA sequencing of a diploid accession and generated a de novo assembly comprising 175,572 contigs. Sixteen SSR loci were selected and of these 13 SSR loci were successfully amplified in 20 L. alba tetraploid accessions and in 12 other Lippia species. Only one SSR locus was monomorphic, whereas 12 loci were polymorphic, yielding one to nine alleles. The heterozygosity was similar among markers, with values of 0.274-0.485; the polymorphism information content values varied from 0.237 to 0.367. These markers were successfully amplified in related species with 85% of transferability on average. Thus, we demonstrate the utility of including a de novo assembly step to obtain SSR markers from low-coverage genomic datasets.
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http://dx.doi.org/10.1007/s11033-020-05445-zDOI Listing
June 2020

Persuasive antimicrobial stewardship intervention in the context of a KPC outbreak: a controlled interrupted time series analysis.

Antimicrob Resist Infect Control 2020 04 21;9(1):55. Epub 2020 Apr 21.

EPIUnit, Instituto de Saúde Pública da Universidade do Porto, Porto, Portugal.

Introduction: Antimicrobial resistance is a major public health threat. Antimicrobial stewardship (AMS) is one of the key strategies to overcome resistance, but robust evidence on the effect of specific interventions is lacking. We report an interrupted time series (ITS) analysis of a persuasive AMS intervention implemented during a KPC producing Klebsiella pneumoniae outbreak.

Methods: A controlled ITS for carbapenem consumption, total antibiotic consumption and antibiotic-free days, between January 2012 and May 2018 was performed, using segmented regression analysis. The AMS intervention was implemented in the Vascular Surgery ward starting on April 2016 in the context of a KPC outbreak. The General Surgery ward was taken as a control group. Data were aggregated by month for both wards, including 51 pre-intervention and 26 intervention points.

Results: The AMS intervention produced a level change in carbapenem consumption of - 11.14 DDDs/100 patient-days accompanied by a decreasing trend of total antibiotic consumption and stable rate of antibiotic-free days in Vascular Surgery ward. These differences were not apparent in the control group. No differences in mortality or readmission rates between pre-intervention and intervention periods were noticed in any of the groups.

Conclusion: Persuasive AMS interventions on top of previously implemented restrictive interventions can reduce carbapenem consumption without increasing total antibiotic consumption. Starting persuasive AMS interventions in an outbreak setting does not compromise the sustainability of the intervention.
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http://dx.doi.org/10.1186/s13756-020-00718-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7175563PMC
April 2020

Induction of Synthetic Polyploids and Assessment of Genomic Stability in .

Front Plant Sci 2020 26;11:292. Epub 2020 Mar 26.

Department of Biology, Federal University of Juiz de Fora, Juiz de Fora, Brazil.

Polyploidy is widely recognized as a major evolutionary force in plants and has been reported in the genus (Verbenaceae). , the most studied species, has been documented as a polyploid complex involving at least four ploidal levels. presents remarkable chemical and genetic variation and represents a model for understanding genome organization. Although the economic and medicinal importance of the species has been widely described, no established polyploid induction protocol has been reported so far. Here, we describe the production of synthetic polyploid plants of using colchicine. The ploidal levels were estimated by flow cytometry and chromosome counting. In addition, FISH and molecular markers approaches were used to confirm the stability of the synthetic polyploids. The major component of the essential oils was estimated by GCMS to compare with the natural individuals. Tetraploids and triploids were produced providing new opportunities for investigating medicinal, pharmacological, and economic applications as well as addressing intrinsic questions involved in the polyploidization process in tropical plants.
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http://dx.doi.org/10.3389/fpls.2020.00292DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7113378PMC
March 2020

Serum creatinine trajectories in real-world hospitalized patients: clinical context and short-term mortality.

J Investig Med 2020 04 15;68(4):870-881. Epub 2020 Mar 15.

EPIUnit, Instituto de Saúde Pública da Universidade do Porto, Porto, Portugal.

Fluctuations in serum creatinine (SCr) during hospitalization may provide additional prognostic value beyond baseline renal function. This study aimed to identify groups of patients with distinct creatinine trajectories over hospital stay and assess them in terms of clinical characteristics and short-term mortality. This retrospective study included 35 853 unique adult admissions to a tertiary referral center between January 2012 and January 2016 with at least three SCr measurements within the first 9 days of stay. Individual SCr courses were determined using linear regression or linear-splines model and grouped into clusters. SCr trajectories were described as median SCr courses within clusters. Almost half of the patients presented with changing, mainly declining SCr concentration during hospitalization. In comparison to patients with an increase in SCr, those with a significant decline were younger, more often admitted via the emergency department, more often required a higher level of care, had fewer comorbidities and the more pronounced the fall in SCr, the greater the observed difference. Regardless of baseline renal function, an increase in SCr was related to the highest in-hospital mortality risk among compared clusters. Also, patients with normal renal function at admission followed by decreasing SCr were at higher risk of inpatient death, but lower 90-day postdischarge mortality than patients with a stable SCr. Acute changes in inpatient SCr convey important prognostic information and can only be interpreted by looking at their evolution over time. Recognizing underlying causes and providing adequate care is crucial for improving adverse prognosis.
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http://dx.doi.org/10.1136/jim-2019-001185DOI Listing
April 2020

Genetic relationships and polyploid origins in the Lippia alba complex.

Am J Bot 2020 03 1;107(3):466-476. Epub 2020 Mar 1.

Department of Biology, Federal University of Juiz de Fora, Juiz de Fora, MG, 36036-241, Brazil.

Premise: Plant genomes vary in size and complexity due in part to polyploidization. Latitudinal analyses of polyploidy are biased toward floras of temperate regions, with much less research done in the tropics. Lippia alba has been described as a tropical polyploid complex with diploid, triploid, tetraploid, and hexaploid accessions. However, no data regarding relationships among the ploidal levels and their origins have been reported. Our goals are to clarify the relationships among accessions of Lippia alba and the origins of each ploidal level.

Methods: We investigated 98 samples representing all five geographical regions of Brazil and all ploidal levels using microsatellite (SSR) allelic variation and DNA sequences of ITS and trnL-F. Nine morphological structures were analyzed from 33 herbarium samples, and the chemical compounds of 78 accessions were analyzed by GC-MS.

Results: Genetic distance analysis, the alignment block pattern, as well as RAxML and Bayesian trees showed that accessions grouped by ploidal level. The triploids form a well-defined group that originated from a single group of diploids. The tetraploids and hexaploid grouped together in SSR and trnL-F analyses. The recovered groups agree with chemical data and morphology.

Conclusions: The accessions grouped by ploidal level. Only one origin of triploids from a single group of diploids was observed. The tetraploid origin is uncertain; however, it appears to have contributed to the origin of the hexaploid. This framework reveals linkages among the ploidal levels, providing new insights into the evolution of a polyploid complex of tropical plants.
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http://dx.doi.org/10.1002/ajb2.1443DOI Listing
March 2020

Coexpression and Transcriptome analyses identify active Apomixis-related genes in Paspalum notatum leaves.

BMC Genomics 2020 Jan 28;21(1):78. Epub 2020 Jan 28.

Centro de Biologia Molecular e Engenharia Genética (CBMEG), Universidade Estadual de Campinas (UNICAMP), Campinas, SP, Brazil.

Background: Paspalum notatum exhibits both sexual and apomictic cytotypes and, thus, is considered a good model for studies of apomixis because it facilitates comparative approaches. In this work, transcriptome sequencing was used to compare contrasting P. notatum cytotypes to identify differential expression patterns and candidate genes involved in the regulation of expression of this trait.

Results: We built a comprehensive transcriptome using leaf and inflorescence from apomictic tetraploids and sexual diploids/tetraploids and a coexpression network based on pairwise correlations between transcript expression profiles. We identified genes exclusively expressed in each cytotype and genes differentially expressed between pairs of cytotypes. Gene Ontology enrichment analyses were performed to better interpret the data. We de novo assembled 114,306 reference transcripts. In total, 536 candidate genes possibly associated with apomixis were detected through statistical analyses of the differential expression data, and several interacting genes potentially linked to the apomixis-controlling region, genes that have already been reported in the literature, and their neighbors were transcriptionally related in the coexpression network.

Conclusions: Apomixis is a highly desirable trait in modern agriculture due to the maintenance of the characteristics of the mother plant in the progeny. The reference transcriptome, candidate genes and their coexpression network identified in this work represent rich resources for future grass breeding programs.
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http://dx.doi.org/10.1186/s12864-020-6518-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6986084PMC
January 2020

Effect of Two Different Pose Estimation Approaches on Lower Extremity Biomechanics in Professional Dancers.

Annu Int Conf IEEE Eng Med Biol Soc 2019 Jul;2019:2947-2950

Different algorithms can be used to estimate the pose of musculoskeletal models in biomechanical studies. Visual 3D uses segment optimization whereas OpenSim uses global optimization. Thus, our purpose was to study whether the two approaches would influence the estimation of lower extremity biomechanical parameters. Marker trajectories and ground reaction forces of 6 professional dancers were collected during a single-leg forward jump-landing. The same data set was processed using both approaches. Our findings suggested that the sagittal knee and ankle angles and moments were highly comparable between the two approaches. The ankle sagittal angle and moment showed the lowest offset. On the other hand, the choice of a kinematic model was likely to affect the hip, more evident in the frontal and transverse planes. This may be due to different factors such as the pelvis and femur positions or larger amount of soft tissue in the thigh.
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http://dx.doi.org/10.1109/EMBC.2019.8857683DOI Listing
July 2019

Metabolic syndrome severity score is associated with diastolic dysfunction and low-grade inflammation in a community-based cohort.

Eur J Prev Cardiol 2020 12 17;27(19):2330-2333. Epub 2019 Dec 17.

Serviço de Cardiologia, Centro Hospitalar de Vila Nova de Gaia/Espinho, Vila Nova de Gaia, Portugal.

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http://dx.doi.org/10.1177/2047487319895400DOI Listing
December 2020

Unraveling candidate genes underlying biomass digestibility in elephant grass (Cenchrus purpureus).

BMC Plant Biol 2019 Dec 10;19(1):548. Epub 2019 Dec 10.

Embrapa Gado de Leite, Juiz de Fora, MG, 36038-330, Brazil.

Background: Elephant grass [Cenchrus purpureus (Schumach.) Morrone] is used for bioenergy and animal feed. In order to identify candidate genes that could be exploited for marker-assisted selection in elephant grass, this study aimed to investigate changes in predictive accuracy using genomic relationship information and simple sequence repeats for eight traits (height, green biomass, dry biomass, acid and neutral detergent fiber, lignin content, biomass digestibility, and dry matter concentration) linked to bioenergetics and animal feeding.

Results: We used single-step, genome-based best linear unbiased prediction and genome association methods to investigate changes in predictive accuracy and find candidate genes using genomic relationship information. Genetic variability (p < 0.05) was detected for most of the traits evaluated. In general, the overall means for the traits varied widely over the cuttings, which was corroborated by a significant genotype by cutting interaction. Knowing the genomic relationships increased the predictive accuracy of the biomass quality traits. We found that one marker (M28_161) was significantly associated with high values of biomass digestibility. The marker had moderate linkage disequilibrium with another marker (M35_202) that, in general, was detected in genotypes with low values of biomass digestibility. In silico analysis revealed that both markers have orthologous regions in other C4 grasses such as Setaria viridis, Panicum hallii, and Panicum virgatum, and these regions are located close to candidate genes involved in the biosynthesis of cell wall molecules (xyloglucan and lignin), which support their association with biomass digestibility.

Conclusions: The markers and candidate genes identified here are useful for breeding programs aimed at changing biomass digestibility in elephant grass. These markers can be used in marker-assisted selection to grow elephant grass cultivars for different uses, e.g., bioenergy production, bio-based products, co-products, bioactive compounds, and animal feed.
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http://dx.doi.org/10.1186/s12870-019-2180-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6905061PMC
December 2019