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Nowadays, consumers demand high quality food products with an extended shelf-life without chemical additives. Edible coatings (EC) containing natural compounds are a promising preservation technology for raw seafood without compromising fresh-like appeal and nutritional content.The aim of this work was to evaluate the effect of Thunnus obesus skin gelatin-based EC containing Codium spp. or Fucus vesiculosus extracts on raw tuna preservation. Three gelatin-based EC (gelatin (5 %) + glycerol (25 %); gelatin (5 %) + Codium spp. (1 %) + glycerol (25 %); gelatin (5 %) + Fucus vesiculosus (1 %) + glycerol (25 %)) were applied directly on the surface of tuna fillets. Functional properties of gelatin and gelatin-based EC containing seaweed extracts were also studied. The gelatin was extracted by an acid-swelling process in the presence/absence of pepsin, followed by subsequent heating/refrigeration, after a pre-treatment with NaOH. The type of acid, temperature and concentration of NaOH greatly influence the process yield. The higher extraction yield was achieved using acetic acid in the presence of pepsin by subsequent refrigeration, especially when skins were previously treated with NaOH (0.2 M). Tuna quality was assessed over 12 days of storage at 4 ± 1 °C in terms of chemical and microbial indices. Results showed that tuna skin gelatin-based EC avoids tuna deterioration. Microbial growth, assessed by total viable counts, and total volatile basic nitrogen were maintained below the maximum limits recommended, contrarily to the control. Additionally, the use of EC increased the stability of red colour during storage.

Objective: To evaluate the possible association between hyperuricemia and cardiorespiratory fitness levels/nutritional profile, grouped into a single variable, in schoolchildren. Method: Cross-sectional study of 2335 students from Elementary schools, aged 7–17 years of both genders, stratified by conglomerates of a municipality in Southern Brazil. Body mass index (BMI) was calculated and cardiorespiratory fitness (CRF) was assessed by the 6-minute run/walk test. The BMI and CRF were grouped into a single variable, considering: (1) low and normal weight/fit; (2) low and normal weight/unfit; (3) overweight-obesity/fit; (4) overweight-obesity/unfit. The Poisson regression (prevalence ratio, PR) was used for the association between hyperuricemia and BMI/CRF ratio with 95% confidence intervals and differences were considered significant when p

Objectives: to evaluate the neuropsychomotor development and the genomic stability associated to folate and blood iron levels in preschool children. Methods: a cross-sectional study in which evaluated the biochemical exams (complete hemogram, serum ferritin, iron and folate), neuropsychomotor development (Denver II Test) and genotoxicity (micronuclei cytome in buccal mucosa cells) of 55 children aging 36-59 months old. Student´s T test, Kruskal-Wallis and Pearson's or Spearman's correlation tests were applied with a significance level of p

Screening of antioxidant potential of dichloromethane and methanolic extracts of twenty-seven seaweeds from the Peniche coast was performed by: total phenolic contents (TPC), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity and oxygen radical absorbance capacity (ORAC). Seaweeds revealing the highest antioxidant activity were screened for cytoprotective potential in MCF-7 cells, including the mitochondrial membrane potential analysis and the caspase-9 activity. High correlation was found between TPC of seaweed extracts and their scavenging capacity on DPPH and peroxyl radicals. The highest antioxidant activity was displayed by the methanolic fraction of brown seaweeds belonging to Fucales, however Ulva compressa presented the highest cytoprotective effect by blunting the apoptosis process. These results suggest that high antioxidant activity may not be directly related with high cytoprotective potential. Thus, seaweeds reveal to be a promising source of compounds with potential against oxidative stress. info:eu-repo/semantics/publishedVersion

Summary The aim of this work was to study the proximate composition and the bioactive profile of Bifurcaria bifurcata. It contains 73.31 ± 0.69% of moisture, 8.57 ± 0.11 g per 100 g dry weight (d.w.) of protein, 5.81 ± 0.14 g per 100 g d.w. of lipid content and 30.15 ± 0.00 g per 100 g d.w. of ash. The polyunsaturated fatty acids were the most abundant fatty acid (FA), accounting for 2426.56 mg per 100 g which represents 41.77% of the total FA. The methanolic fraction showed high quantity of polyphenols (220.01 ± 0.010 phloroglucinol equivalents g−1 extract), DPPH radical reduction capacity (EC50:58.82 μg mL−1) and oxygen radical absorbent capacity (3151.35 ± 119.33 μmol Trolox equivalents g−1 extract). The highest antimicrobial effect was observed against Pseudomonas aeruginosa (11.3 ± 1.5 mm) and Saccharomyces cerevisiae (IC50:17.07 μg mL−1) induced by methanolic and dichloromethane fractions, respectively. Dichloromethane fraction revealed the highest antitumor activity on Caco-2 and HepG-2 cells. Bifurcaria bifurcata can be a promising source of bioactive compounds and functional ingredients.

Objective: The purpose of this study was to measure the prevalence of common mental health disorders among employees of the Family Health Strategy in Santa Cruz do Sul, RS, and to describe sociodemographic and occupational factors associated with these disorders. Methods: This was a cross-sectional, quantitative study, conducted from June to August 2013. In total, 83 employees, who worked at 11 family health units in the city, participated. Participants included doctors, nurses, technicians/nursing assistants, and community health workers. The survey was conducted using a self-administered questionnaire about sociodemographic and occupational issues as well as the matters related to the Self-Reporting Questionnaire. Results: The average age of participants was 37.1±8.9 years. The majority of community health workers were female workers who were married, had children, had a per capita income of up to 2 minimum wages, and had completed high school. The general prevalence of common mental health disorders was 19.7%. Mental health disorders were present in all occupational categories, but were more frequent among male nurses (25%), although this was not statistically significant. Conclusion: This study, in addition to characterizing the population of employees of the Family Health Strategy in Santa Cruz do Sul, RS, in terms of demographic and occupational characteristics, found that psychological distress was present, to varying degrees, in all professional categories evaluated. Given the high prevalence of mental health disorders among these employees, measures are required to ensure the protection and promotion of mental health in this setting.

Methanol, n-hexane and dichloromethane extracts of twelve marine macro-algae (Rhodophyta, Chlorophyta and Heterokontophyta divisions) from Peniche coast (Portugal) were evaluated for their antibacterial and antifungal activity. The antibacterial activity was evaluated by disc diffusion method against Bacillus subtilis (gram positive bacteria) and Escherichia coli (gram negative bacteria). Saccharomyces cerevisiae was used as a model for the antifungal activity by evaluating the growth inhibitory activity of the extracts. The high antibacterial activity was obtained by the Asparagopsis armata methanolic extract (10 mm-0.1 mg/disc), followed by the Sphaerococcus coronopifolius n-hexane extract (8 mm-0.1 mg/disc), and the Asparagopsis armata dichloromethane extract (12 mm-0.3 mg/disc) against Bacillus subtilis. There were no positive results against Escherichia coli. Sphaerococcus coronopifolius revealed high antifungal potential for n-hexane (IC50 = 40.2 µg/ml), dichloromethane (IC50 = 78.9 µg/ml) and methanolic (IC50 = 55.18 µg/ml) extracts against Saccharomyces cerevisiae growth. The antifungal potency of the Sphaerococcus coronopifolius extracts was similar with the standard amphotericin B. Asparagopsis armata and Sphaerococcus coronopifolius reveal to be interesting sources of natural compounds with antimicrobial properties.

We evaluated the influence of hesperidin and vitamin C (VitC) on glycemic parameters, lipid profile, and DNA damage in male Wistar rats treated with sucrose overload. Rats were divided into six experimental groups: I-water control; II-sucrose control; III-hesperidin control; IV-VitC control; V-co-treatment of sucrose plus hesperidin; VI-co-treatment of sucrose plus VitC. We measured the levels of triglycerides, total cholesterol, HDL-c, LDL-c, fasting glucose, and glycated hemoglobin (A1C). DNA damage was evaluated in blood and brain cells using the comet assay and the micronucleus test was used to evaluate chromosomal damages in the rat bone marrow. Co-treatment with VitC, but not with hesperidin, normalized the serum glucose. No effect of co-treatments was observed on A1C. The co-treatment with VitC or hesperidin did not influence the lipid profile (p>0.05). Rats co-treated with hesperidin had a significantly lower DNA damage level in blood (p0.05). Hesperidin and VitC showed different effects on sucrose and DNA damage levels. While VitC lowered the serum glucose, hesperidin reduced the DNA damage.

This article belongs to the Special Issue Selected Papers from the 14th International Symposium on Marine Natural Products Surface-associated marine bacteria are an interesting source of new secondary metabolites. The aim of this study was the isolation and identification of epiphytic bacteria from the marine brown alga, Bifurcaria bifurcata, and the evaluation of the antioxidant and antimicrobial activity of bacteria extracts. The identification of epiphytic bacteria was determined by 16S rRNA gene sequencing. Bacteria extracts were obtained with methanol and dichloromethane (1:1) extraction. The antioxidant activity of extracts was performed by quantification of total phenolic content (TPC), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity and oxygen radical absorbance capacity (ORAC). Antimicrobial activities were evaluated against Escherichia coli, Pseudomonas aeruginosa, Bacillus subtilis, Salmonella enteritidis, Staphylococcus aureus, Saccharomyces cerevisiae and Candida albicans. A total of 39 Bifurcaria bifurcata-associated bacteria were isolated and 33 were identified as Vibrio sp. (48.72%), Alteromonas sp. (12.82%), Shewanella sp. (12.26%), Serratia sp. (2.56%), Citricoccus sp. (2.56%), Cellulophaga sp. (2.56%), Ruegeria sp. (2.56%) and Staphylococcus sp. (2.56%). Six (15.38%) of the 39 bacteria Bifurcaria bifurcata-associated bacteria presented less than a 90% Basic Local Alignment Search Tool (BLAST) match, and some of those could be new. The highest antioxidant activity and antimicrobial activity (against B. subtilis) was exhibited by strain 16 (Shewanella sp.). Several strains also presented high antimicrobial activity against S. aureus, mainly belonging to Alteromonas sp. and Vibrio sp. There were no positive results against fungi and Gram-negative bacteria. Bifurcaria bifurcata epiphytic bacteria were revealed to be excellent sources of natural antioxidant and antimicrobial compounds.

Chitin was extracted from Polybius henslowii, a swimming crab, captured in large quantities throughout the Portuguese coast by purse seine vessels as bycatch. After standard chitin extraction procedures, water-soluble chitosan products were obtained via two different methods: (1) N-acetylation with the addition of acetic anhydride and (2) a reaction with hydrogen peroxide. The chemical structure and molecular weight of chitosan derivatives, water-soluble chitosan (WSC) and chitooligosaccharides (COS), were confirmed by Fourier Transform Infrared Spectroscopy (FT-IR) and gel permeation chromatography (GPC). Antioxidant and metal chelation activities were evaluated, and the growth inhibition capacity was tested on four phytopatogens. The chitooligosaccharides from pereopods (pCOS) and shell body parts (sCOS) inhibited all fungal species tested, particularly Cryphonectria parasitica with 84.7% and 85.5%, respectively. Both radical scavenging and antifungal activities proved to be dose-dependent. Chitooligosaccharides with a low molecular weight (2.7, 7.4, and 10.4 Kg&middot, mol&minus, 1) showed the highest activity among all properties tested. These results suggested that chitosan derivatives from P. henslowii raw material could potentially be used against phytopathogens or as ingredient in cosmetics and other products related to oxidative stress.

International audience; The marine environment offers both economic and scientific potential which are relatively untapped from a biotechnological point of view. These environments whilst harsh are ironically fragile and dependent on a harmonious life form balance. Exploitation of natural resources by exhaustive wild harvesting has obvious negative environmental consequences. From a European industry perspective marine organisms are a largely underutilised resource. This is not due to lack of interest but due to a lack of choice the industry faces for cost competitive, sustainable and environmentally conscientious product alternatives. Knowledge of the biotechnological potential of marine organisms together with the development of sustainable systems for their cultivation, processing and utilisation are essential. In 2010, the European Commission recognised this need and funded a collaborative RTD/SME project under the Framework 7-Knowledge Based Bio-Economy (KBBE) Theme 2 Programme ‘Sustainable culture of marine microorganisms, algae and/or invertebrates for high value added products’. The scope of that project entitled ‘Sustainable Production of Biologically Active Molecules of Marine Based Origin’ (BAMMBO) is outlined. Although the Union is a global leader in many technologies, it faces increasing competition from traditional rivals and emerging economies alike and must therefore improve its innovation performance. For this reason innovation is placed at the heart of a European Horizon 2020 Strategy wherein the challenge is to connect economic performance to eco performance. This article provides a synopsis of the research activities of the BAMMBO project as they fit within the wider scope of sustainable environmentally conscientious marine resource exploitation for high-value biomolecules.

Prediabetes (intermediate hyperglycemia) is a high-risk state for diabetes that is defined by higher than normal glycemic levels that are below the level required for a diagnosis of diabetes. Prediabetes is characterized by oxidative stress, yet the associated DNA damage and cytotoxicity remain unknown to date. Therefore, we evaluated the relationship between glycemic alterations, DNA damage and cytotoxicity in the lymphocytes of individuals with pre-diabetes. Fasting plasma glucose (FPG) and glycated hemoglobin (A1C) levels were quantified and used as inclusion criteria. Anthropometric parameters were also evaluated. The cytokinesis-block micronucleus cytome assay (CBMN Cyt) was used to evaluate DNA damage and cytotoxicity. FPG correlated with A1C (r=0.562, p=0.002). Because A1C is the best predictor of diabetes complications, the association between A1C and the evaluated variables was assessed. The waist-hip ratio correlated with A1C (p

O principal escopo é examinar, com base em novas evidências, a evolução das finanças públicas de Minas Gerais na República Velha, com destaque para o lugar do dispêndio com a infraestrutura de transportes. Também se discute a vigência de conflito entre preocupação com o equilíbrio orçamentário e políticas de desenvolvimento. Na primeira seção são analisadas as receitas e despesas estaduais, com ênfase na classificação dos comportamentos assumidos por essas variáveis ao longo do período e na determinação das principais rubricas que as compunham. Também é examinada a evolução da dívida fundada, assim como sua correlação com as despesas públicas. A segunda seção analisa especificamente os gastos com a infraestrutura de transportes, nomeadamente ferrovias e estradas de rodagem. Primeiro o relacionamento entre o crescimento da malha ferroviária e as despesas do Estado com garantias de juros, subvenções e empréstimos a companhias ferroviárias. Em seguida a evolução dos gastos com rodovias. The principal scope of this study is to examine, based on new evidence, the evolution of the public finances of Minas Gerais in the Old Republic, with an emphasis on the role of transportation infrastructure expenditures. Also discussed is the conflict between concern over budgetary equilibrium and development policies. The first section analyzes state revenues and expenses, with emphasis on the classification of the behaviors of these variables over the period and the determination of the main categories that make them up. Also analyzed is the evolution of government funded debt, as well as its correlation with public expenditures. The second section specifically analyzes expenses on transportation infrastructure–in this case, railroads and highways, first by examining the relationship between growth in the railway network and State

O diabetes mellitus (DM) inclui um grupo de doenças metabólicas resultante de disfunção na secreção e/ou ação da insulina. O diagnóstico de DM é definido com base na avaliação da glicemia de jejum(GJ) e hemoglobina glicada (A1C) que também informa a persistência da hiperglicemia. Entretanto novas tecnologias analíticas estão sempre sob constante investigação. A espectroscopia de absorção molecular no infravermelho com Transformada de Fourier (FT-IR) associada à análise multivariada tem surgido como uma alternativa nesse contexto. O presente estudo buscou avaliar a FT-IR, através da técnica de reflectância difusa, para quantificar glicose e A1C em um grupo de indivíduos adultos. O estudo foi composto por 46 pacientes de 32 a 59 anos, com valores GJ de 71,67 a 135 mg/dL e de A1C de 5,3 a 7,2 %. O modelo PLS-DRIFTS para GJ o apresentou RMSECV=1,37 mg/dL e R²=0,995 e para A1C, RMSECV=0,06% R²=0,993 sem exclusão de amostras e usando a faixa de 4000-2401, 2300-600 cm-1. Dessa forma foi possível concluir que FT-IR associada a análise multivariada se mostrou adequado para predizer os parâmetros bioquímicos de glicose e A1C de jejum com um erro mínimo, atendendo os requisitos analíticos da ANVISA para quantificação destes parâmetros.

Hepatic ischemia-reperfusion injury has a significant impact on liver resection and transplantation. Many strategies have been developed to reduce the effects of ischemia-reperfusion injury, including pharmacologic and ischemic preconditioning; however, studies comparing these two methods are lacking.An experimental study was performed in a swine model. Eighteen swine were randomly assigned to three different groups: an ischemic preconditioning (IschPC) group, a pharmacologic preconditioning (PharmPC) group, and a control group. All animals underwent a 40-min liver ischemia, followed by 40 min of reperfusion. The IschPC group received a short period of ischemia (10 min) and a short period of reperfusion (15 min) before prolonged ischemia. The PharmPC group received inhaled sevoflurane for 30 min before prolonged ischemia. The control group did not receive any intervention before prolonged ischemia. Blood samples and liver tissue were obtained after ischemic and reperfusion periods. Injury was evaluated by measure of DNA damage (using COMET assay) and serum biochemical markers (transaminases, alkaline phosphatase, amylase, bilirubin, and C-reactive protein [CRP]).No significant difference was found in serum biochemical markers, except for the C-reactive protein level that was lower in the PharmPC group than in the control group soon after hepatic ischemia. Soon after prolonged ischemia, DNA damage index, both in blood samples and in liver tissue samples, was similar among the groups. However, an increase in DNA damage after reperfusion was higher in the control group than in the PharmPC group (P 0.05). The increase in DNA damage in the IschPC group was half of that observed in the control, but this difference was not statistically significant.Our results suggest an early protective effect of PharmPC (lower levels of C-reactive protein soon after ischemia). The protective effect observed after reperfusion was higher with PharmPC than with ischemic preconditioning. The simultaneous use of both methods could potentiate protection for ischemia-reperfusion.

Antioxidants play an important role as Reactive Oxygen Species (ROS) chelating agents and, therefore, the screening for potent antioxidants from natural sources as potential protective agents is of great relevance. The main aim of this study was to obtain antioxidant-enriched fractions from the common seaweed Fucus spiralis and evaluate their activity and efficiency in protecting human cells (MCF-7 cells) on an oxidative stress condition induced by H₂O₂. Five fractions, F1-F5, were obtained by reversed-phase vacuum liquid chromatography. F3, F4 and F5 revealed the highest phlorotannin content, also showing the strongest antioxidant effects. The cell death induced by H₂O₂ was reduced by all fractions following the potency order F4 > F2 > F3 > F5 > F1. Only fraction F4 completely inhibited the H₂O₂ effect. To understand the possible mechanisms of action of these fractions, the cellular production of H₂O₂, the mitochondrial membrane potential and the caspase 9 activity were studied. Fractions F3 and F4 presented the highest reduction on H₂O₂ cell production. All fractions decreased both caspase-9 activity and cell membrane depolarization (except F1). Taken all together, the edible F. spiralis reveal that they provide protection against oxidative stress induced by H₂O₂ on the human MCF-7 cellular model, probably acting as upstream blockers of apoptosis. info:eu-repo/semantics/publishedVersion

The risk of developing cardiovascular disease (CVD) is related to lifestyle (e.g. diet, physical activity and smoking) as well as to genetic factors. This study aimed at evaluating the association between CVD risk factors and DNA damage levels in children and adolescents. Anthropometry, diet and serum CVD risk factors were evaluated by standard procedures. DNA damage levels were accessed by the comet assay (Single cell gel electrophoresis; SCGE) and cytokinesis-blocked micronucleus (CBMN) assays in leukocytes. A total of 34 children and adolescents selected from a population sample were divided into three groups according to their level of CVD risk. Moderate and high CVD risk subjects showed significantly higher body fat and serum CVD risk markers than low risk subjects (P

This study evaluated the association between primary DNA damage and chromosomal damage with iron intake and red blood cell parameters of iron status in a sample of healthy children and adolescents from a low-socioeconomic community.The level of primary DNA damage was assessed using an alkaline comet assay and the level of chromosomal damage was assessed using the cytokinesis-block micronucleus assay. A automated complete blood count was used to evaluate red blood cell status. The intake of iron was measured using a food-recall questionnaire.According to hemoglobin levels, only 1 of the 30 subjects evaluated was anemic. Nevertheless, 43% of the sampled subjects showed decreased mean corpuscular volume in addition to an increased amount of primary DNA damage (P0.05). Mean corpuscular volume was negatively correlated with primary DNA damage (r = -0.429, P = 0.020) but not with chromosomal damage. The association between iron and primary DNA damage showed a U-shaped curve, indicating that an intake of approximately 15 mg of iron per day (up to two-fold of the dietary recommended intake) could minimize primary DNA damage in this age group. The frequency of micronuclei and nucleoplasmic bridges, indicators of chromosomal breakage/loss and chromosomal end-fusions, respectively, showed a negative correlation with iron intake. These results indicate that an intake of iron15 mg/d could increase genomic stability in binucleated lymphocytes of the same group.An intake of iron ≥ 15 mg/d can decrease DNA damage in young subjects.

During childhood and adolescence, physical inactivity, excess weight, and poor nutrition are risk factors for chronic diseases, especially obesity, hypertension, and diabetes mellitus. Early intervention can prevent the development of these complications.To determine the presence of cardiovascular risk (obesity and hypertension) in schoolchildren and its potential interactions with cardio-respiratory fitness.This was a cross-sectional study conducted in a stratified cluster sample of 1,666 schoolchildren, aged between 7 and 17 years, 873 (52.4%) of them male and 793 (47.6%) of them female. The following variables were evaluated: systolic blood pressure (SBP), diastolic blood pressure (DBP), body mass index (BMI), body fat percentage (BF %), and cardio-respiratory fitness. SBP and DBP were correlated with waist circumference (WC), waist-hip ratio (WHR), sum of skin folds (SigmaSF), and cardio-respiratory fitness.A BMI assessment of the students showed that 26.7% of them were overweight or obese, and 35.9% had body fat percentage over moderately high. As to blood pressure, we found that 13.9% and 12.1% of the students were borderline or hypertensive, for SBP and DBP, respectively. There was an association among hypertension, obesity, and cardio-respiratory fitness. There was a significant correlation of SBP and DBP with all variables, and also a weak to moderate correlation with age, weight, height, BMI, and waist circumference.The presence of hypertension associated with obesity and its effects on cardio-respiratory fitness stress the importance of recommending, since childhood, a more active and healthy lifestyle.

In this study, the distribution and prevalence of extended-spectrum beta-lactamases belonging to the TEM, SHV and CTX-M families were estimated among samples of Escherichia coli and Klebsiella spp. at the university hospital of Santa Maria, Rio Grande do Sul. Over a 14-month period, 90 microorganisms were selected as likely ESBL producers. The isolates were subjected to confirmatory phenotype tests for the presence of ESBL. Through investigating the respective genes using the polymerase chain reaction, the ESBL types present in each microorganism were then determined. Fifty-five samples (61.1%) were confirmed as ESBL-positive by means of the combined disc method, and 57 (63.3%) were found to be ESBL producers by means of the double disc method. From the polymerase chain reaction, ESBLs of TEM and SHV types were more frequently present in Klebsiella pneumoniae, while ESBL of CTX-M type was more frequently present in Klebsiella oxytoca.

Cryptococcus neoformans is a pathogenic fungus that causes life-threatening meningoencephalitis in immunocompromised patients (HIV-positive patients), and lymphoproliferative disorders in patients subjected to organ transplantation and other immunosuppressive therapies. This fungus is commonly found in soil and avian excreta, mainly from pigeon and turkey. We describe the isolation and characterization of 17 clinical and 10 environmental (pigeon excreta) isolates from the Brazilian state Rio Grande do Sul. We analyzed capsule formation, carbon assimilation pattern, canavanine-glycine-bromothymol blue (CGB) reaction, and nitrate and urease tests, as well as susceptibility to antifungal drugs. The genetic variability among C. neoformans isolates was studied using randomly amplified polymorphic DNA (RAPD) analysis. Eight of 22 arbitrary polymerase chain reaction primers used confirmed genetic polymorphism among the environmental isolates tested, suggesting that it remains feasible to use RAPD analysis as a typing method. Three of the selected primers yielded 10 molecular subclasses. The majority of the clinical isolates were assigned to the molecular subclass F. The RAPD data obtained reinforce the developing consensus about the population structure of this fungus.

In the present study, seasonal changes in semen quality of Lusitano stallions were evaluated to establish the best time for semen cryopreservation. Seasonal changes on testicular size, seminal characteristics and semen freezability were therefore evaluated by optical microscopy and flow cytometry. Five Lusitano stallions belonging to National Stud Farm, “Coudelaria Nacional” were used in the study. Semen was collected in four periods defined as spring (March/April), summer (June/July), autumn (September/October) and winter (December/February). Ejaculates were collected once a week for 4 weeks, and cryopreserved in 0.5-ml plastic straws. Before and after each period of semen collection, testicles of all stallions were measured. For evaluation, one straw per ejaculate, per stallion, and per season was thawed at 37 �C for 30 s and microscopic evaluation (motility) was performed. A flow cytometer (Becton Dickinson, San Jose, CA, USA) was used to evaluate sperm viability and acrosomal integrity by using the combination of fluorescent probes propidium iodide (PI) and SYBR-14 (LIVE/DEAD Sperm viability Kit (L-7011), Molecular Probes, Eugene, Oregon) for sperm viability. In addition, acrosome integrity was evaluated using the fluorescein labelled lectin from the peanut plant, Arachis hypogea (FITC-PNA, Sigma Chemical, St. Louis, MO) and propidium iodide (PI) probe. Results were evaluated by the Cell-Quest software (Becton Dickinson, San Jose, CA, USA) and statistical differences by a one-way ANOVA, followed by post hoc comparison tests. The results indicated significant seasonal differences in testicular size and sperm production (p�0.05). Ultrasonographic testicular volume was maximal in March (234.03±15.8 cm3), decreased to a minimum in October and June (203.5±17.0 cm3, 168.6±10.0 cm3, respectively) and started to increase again in January (190.8±8.8 cm3). A significant positive correlation between testicular volume and sperm production (r = 0.66; P� 0.01) was demonstrated, although minimum testicular volume was reached in October while minimum sperm production was registered in January. Viability in frozen-thawed semen from ejaculates collected in autumn was 37.73±2.20% and in summer 40.99±4.08%; the results differ significantly from other seasons: winter 54.26±2.10% and spring 46.71±2.32% (p�0.05). The high number of dead spermatozoa after the freezing‐thawing process in autumn/winter most likely indicates that seasonally related changes in seminal plasma composition may affect semen preservation. Nevertheless, no significant differences were observed for acrosome integrity in different periods of the experiment. An inter-stallion variation was observed for testicular size and sperm production, but testicular size, besides sperm production, was positively correlated in all individual stallions. At the time of semen collection, sperm concentrations determined by photometry showed that both testis size and sperm production of housed Lusitano stallions were affected by season. Testicular size and function were maximal in the spring, but testicular volume was smallest in October, while sperm production continued to decrease till January. The results indicate that in our climatic/latitude conditions, it is preferable to freeze semen during winter/spring period.

Two different chitosan polymers were obtained from chitin extracted from the crab Polybius henslowii through different methods: (1) water-soluble chitosan (WSC) through N-acetylation with addition of acetic anhydride and (2) low molecular-weight water-soluble chitosan (LWSC) by the use of H2O2. WSC and LWSC polymers were characterized by FTIR and GPC analysis and investigated for antibacterial activity against a range of Gram-negative bacteria (Escherichia coli 1103, Escherichia coli 10503, Salmonella sp.) and Gram-positive bacteria (Staphylococcus aureus, Bacillus subtilis and Lactobacillus planctarum). This study is a contribution to economic valorization of Polybius by-catch and sustainable development of west coast Portuguese fisheries communities. Financial support was obtained from European Fisheries Fund – Axis 4 managed by Fishery Local Action Group Oeste-Portugal.

Este estudo foi realizado com os primeiros isolados ambientais de C. neoformans sorotipo D, obtidos no Rio Grande Susceptibility of C. neoformans to heat and antifungal agents 5 do Sul. Objetivando-se avaliar a susceptibilidade a agentes antifúngicos de forma mais rigorosa, utilizou-se a técnica de referência proposta pelo NCCLS, Caldo Yeast Nitrogen Base (YNB) proposto por Ghannoum et al., Antibiotic medium 3, caldo YNB adequado à metodologia do NCCLS e o E-test. Os resultados indicaram que todos os isolados foram sensíveis à anfotericina B (0,0625 -0,5 μg/mL), fluconazol (0,125 – 4,0 μg/ mL), itraconazol (0,031 – 0,25 μg/ml) e fluorocitosina (0,125-4,0 μg/mL) através das técnicas empregadas. Nos testes de termotolerância (47ºC/30 min), observou-se que as culturas de C. neoformans sorotipo D são mais sensíveis do que as de C. neoformans sorotipo A. In this work we studied the susceptibility to heat and antifungal agents of the first strains of environmental Cryptococcus neoformans var neoformans (serotype D) isolated in the state of Rio Grande do Sul, Brazil. In order to achieve a rigorous analysis, we employed the methodology recommended by NCCLS, Yeast Nitrogen Base (YNB) proposed by Ghannoum et al (YNB-1), Antibiotic medium 3 (AM3) indicated by others, YNB adjusted to the NCCLS methodology (YNB-2) and Etest. Our results indicate that all strains were susceptible to amphotericin B (0.0625 – 0.5 μg/mL), fluconazole (0.125 – 8.0 μg/mL), itraconazole (0.031 – 0.25 μg/mL) and flucytosine (0.125 – 4.0 μg/mL). The C. neoformans serotype D strains were more susceptible to heat (47ºC / 30 min) than C. neoformans serotype A.