Wine yeast selection in the Iberian Peninsula: Saccharomyces and non-Saccharomyces as drivers of innovation in Spanish and Portuguese wine industries.

Yeast selection for the wine industry in Spain started in 1950 for the understanding of the microbial ecology, and for the selection of optimal strains to improve the performance of alcoholic fermentation and the overall wine quality. This process has been strongly developed over the last 30 years, firstly on Saccharomyces cerevisiae, and, lately, with intense activity on non-Saccharomyces. Several thousand yeast strains have been isolated, identified and tested to select those with better performance and/or specific technological properties. The present review proposes a global survey of this massive ex-situ preservation of eukaryotic microorganisms, a reservoir of biotechnological solutions for the wine sector, overviewing relevant screenings that led to the selection of strains from 12 genera and 22 species of oenological significance. In the first part, the attention goes to the selection programmes related to relevant wine-producing areas (i.e. Douro, Extremadura, Galicia, La Mancha and Uclés, Ribera del Duero, Rioja, Sherry area, and Valencia). In the second part, the focus shifted on specific non-Saccharomyces genera/species selected from different Spanish and Portuguese regions, exploited to enhance particular attributes of the wines. A fil rouge of the dissertation is the design of tailored biotechnological solutions for wines typical of given geographic areas.

Assessment of neural networks and time series analysis to forecast airborne Parietaria pollen presence in the Atlantic coastal regions.

Pollen forecasting models are a useful tool with which to predict episodes of type I allergenic risk and other environmental or biological processes. Parietaria is a wind-pollinated perennial herb that is responsible for many cases of severe pollinosis due to its high pollen production, the long persistence of the pollen grains in the atmosphere and the abundant presence of allergens in their cytoplasm and walls. The aim of this paper is to develop artificial neural networks (ANNs) to predict airborne Parietaria pollen concentrations in the northwestern part of Spain using a 19-year data set (1999-2017). The results show a significant increase in the length of time Parietaria pollen is in the air, as well as significant increases in the annual Parietaria pollen integral and mean daily maximum pollen value in the year. The Neural models show the ability to forecast airborne Parietaria pollen concentrations 1, 2, and 3 days ahead. A developed model with five input variables used to predict concentrations of airborne Parietaria pollen 1 day ahead shows determination coefficients between 0.618 and 0.652.

Fraxinus pollen and allergen concentrations in Ourense (South-western Europe).

In temperate zones of North-Central Europe the sensitization to ash pollen is a recognized problem, also extended to the Northern areas of the Mediterranean basin. Some observations in Switzerland suggest that ash pollen season could be as important as birch pollen period. The allergenic significance of this pollen has been poorly studied in Southern Europe as the amounts of ash pollen are low. Due to the high degree of family relationship with the olive pollen major allergen (backed by a sequence identity of 88%), the Fraxinus pollen could be a significant cause of early respiratory allergy in sensitized people to olive pollen as consequence of cross-reactivity processes. Ash tree flowers in the Northwestern Spain during the winter months. The atmospheric presence of Ole e 1-like proteins (which could be related with the Fra a 1 presence) can be accurately detected using Ole e 1 antibodies. The correlation analysis showed high Spearman correlation coefficients between pollen content and rainfall (R(2)=-0.333, p<0.01) or allergen concentration and maximum temperature (R(2)=-0.271, p<0.01). In addiction CCA analysis showed not significant differences (p<0.05) between the component 1 and 2 variables. PCFA analysis plots showed that the allergen concentrations are related to the presence of the Fraxinus pollen in the air, facilitating the wind speed its submicronic allergen proteins dispersion. In order to forecast the Fraxinus allergy risk periods, two regression equations were developed with Adjusted R(2) values around 0.48-0.49. The t-test for dependent samples shows no significant differences between the observed data and the estimated by the equations. The combination of the airborne pollen content and the allergen quantification must be assessed in the epidemiologic study of allergic respiratory diseases.

Assessment of Quercus flowering trends in NW Spain.

This paper sought to chart airborne Quercus pollen counts over the last 20 years in the region of Galicia (NW Spain) with a view to detecting the possible influence of climate change on the Quercus airborne pollen season (APS). Pollen data from Ourense, Santiago de Compostela, Vigo and Lugo were used. The Quercus airborne pollen season was characterized in terms of the following parameters: pollen season start and end dates, peak pollen count, pollen season length and pollen index. Several methods, dates and threshold temperatures for determining the chill and heat requirements needed to trigger flowering were applied. A diverse APS onset timing sequence was observed for the four cities as Quercus flowers few days in advance in Vigo. The variations observed could be related to differences in the meteorological conditions or the thermal requirements needed for flowering. Thermal requirements differed depending on local climate conditions in the study cities: the lowest values for chilling accumulation were recorded in Vigo and the highest in Lugo, whereas the lowest heat accumulation was achieved in Vigo. Differences in APS trends between cities may reflect variations in weather-related trends. A significant trend towards rising Quercus pollen indices and higher maximum daily mean pollen counts was observed in Ourense, linked to the more marked temperature increase across southern Galicia. A non-uniform trend towards increased temperatures was noted over the study period, particularly in late summer and early autumn in all four study cities. Additionally, an increase in spring temperatures was observed in south-western Galicia.

Impact of an enhanced screening program on the detection of non-AIDS neoplasias in patients with human immunodeficiency virus infection.

BACKGROUND: The incidence of non-AIDS defining cancer (NADC) is higher in people living with HIV (PLWH) than in the general population, and it is already one of the leading causes of death in the HIV-infected population. It is estimated that the situation will be aggravated by the progressive aging of PLWH. Early diagnosis through intensive cancer screening may improve the ability for therapeutic interventions and could be critical in reducing mortality, but it might also increase expenditure and harms associated with adverse events. The aim of this study is to evaluate an enhanced screening program for early diagnosis of cancer in PLWH compared to standard practice. The specific objectives are (1) to compare the frequency of cancer diagnosed at an early stage, (2) to analyze safety of the enhanced program: adverse events and unnecessary interventions, (3) to analyze the cost-utility of the program, and (4) to estimate the overall and site-specific incidence of NADC in PLWH. METHODS: We will conduct a multicenter, non-blinded, randomized, controlled trial, comparing two parallel arms: conventional vs enhanced screening. Data will be recorded in an electronic data collection notebook. Conventional intervention group will follow the standard of care screening in the participating centers, according to the European AIDS Clinical Society recommendations, and the enhanced intervention group will follow an expanded screening aimed to early detection of lung, liver, anal, cervical, breast, prostate, colorectal, and skin cancer. The trial will be conducted within the framework of the Spanish AIDS Research Network Cohort (CoRIS). DISCUSSION: The trial will evaluate the efficacy, safety, and efficiency of an enhanced screening program for the early diagnosis of cancer in HIV patients compared to standard of care practice. The information provided will be relevant since there are currently no studies on expanded cancer screening strategies in patients with HIV, and available data estimating cost effectiveness or cost-utility of such as programs are scarce. An enhanced program for NADC screening in patients with HIV could lead to early diagnosis and improve the prognosis of these patients, with an acceptable rate of unnecessary interventions, but it is critical to demonstrate that the benefits clearly outweigh the harms, before the strategy could be implemented. TRIAL REGISTRATION: ClinicalTrials.gov NCT04735445. Registered on 25 June 2019.

Secondary Outcomes of the Ole e 1 Proteins Involved in Pollen Tube Development: Impact on Allergies.

Ole e 1 protein is involved in olive fertilization mechanisms controlling pollen tube development. Similarly to the process by which pollen grains hydrated and form a pollen tube upon arrival at the female gametophyte, when pollen grains fall on the nasal mucosa the expression of Ole e 1 protein induce allergic reaction in sensitive individuals. The research was conducted in Ourense (North-western Spain), during the 2009-2018 period. Ole e 1 protein was collected using a Cyclone Sampler and processed with the ELISA methodology. Airborne Olea pollen were monitored using a Hirst type volumetric sampler. Allergy risk episodes identified by pollen concentrations were detected in five of the 10 studied years, all with moderate risk. Actual risk episodes of allergy increased when the combination of pollen and Ole e 1 concentrations were considered. Moderate risk episodes were detected during 9 years and high-risk episodes during 3 years. In addition, some years of low annual pollen concentrations recorded high total amounts of Ole e 1. During the years with lower pollen production, the tree increases the synthesis of Ole e 1 to ensure proper pollen tube elongation in order to complete a successful fertilization. This fact could justify higher sensitization rates in years in which a lower pollen production is expected. The present method contributes to the determination of the real exposure to Ole e 1 allergen evaluating the role of this protein as an aeroallergen for sensitized population. The allergen content in the atmosphere should be considered to enhance the prevention of pollinosis clinical symptomatology and the reduction of medicine consumption.

Epidemiology and clinical course of hepatitis A in Cantabria before and after the epidemic outbreak of June 2016. / Epidemiología y curso clínico de la hepatitis A en Cantabria antes y después del brote epidémico de junio 2016.

BACKGROUND AND OBJECTIVES: Since June 2016, there have been outbreaks of hepatitis A in various European countries, mainly affecting men who have sex with men (MSM). The aim of this study was to assess their clinical and epidemiological impact in Cantabria, Spain. MATERIAL AND METHODS: We retrospectively collected all cases of hepatitis A diagnosed in Cantabria between January 2013 and September 2018. We compared 2 periods: January 2013-May 2016 and June 2016-September 2018. RESULTS: A total of 156 cases were diagnosed, observing an increase in the incidence starting in October 2016. With regard to 2013-2016, we observed a higher proportion of men (50.0% vs. 84.5%; p=.012) with a predominance of the homosexual orientation (80.6%) and a higher rate of sexual transmission (0% vs. 48.3%; p=.061) for the patients in the 2016-2018 period. From the clinical standpoint, all cases of severe hepatitis occurred during this latter period. CONCLUSIONS: Our results reaffirm the high clinical and epidemiological impact of the epidemic outbreak in Cantabria and emphasizes the need for optimising the current prevention measures against hepatitis A.

Use of benznidazole to treat chronic Chagas' disease: a systematic review with a meta-analysis.

BACKGROUND AND OBJECTIVES: The recent significant increase in the number of immigrants entering the European Union from South and Central America means that chronic Chagas' disease is an increasingly frequent diagnosis among immigrants in Europe. Our objectives were to evaluate published evidence on the treatment of chronic Chagas' disease with benznidazole and on the potential benefits of this drug in the chronic phase of the disease. METHODS: We performed a systematic review and meta-analysis by means of an electronic search of the published literature, with no language restrictions, until October 2008. We included studies on chronically infected patients of any age who were in the indeterminate phase or had visceral involvement and for whom treatment with benznidazole was compared with placebo or no treatment. The primary endpoint was response to therapy (whether serological, parasitological or clinical), as it was measured in each of the studies included. Clinical response to therapy was also analysed. RESULTS: We identified 696 studies, from which we chose 9: 3 clinical trials and 6 observational studies. Compared with placebo or no treatment, benznidazole increases 18-fold the probability of a response to therapy [global odds ratio (OR), 18.8; 95% confidence interval (CI), 5.2-68.3]. This effect was mainly observed in clinical trials (OR, 70.8; 95% CI, 16-314), whereas in observational studies it was much less marked (OR, 7.8; 95% CI, 2.1-28.9), and even less so when only observational studies in adults were considered (OR, 6.3; 95% CI, 1.6-24.7). Patients treated with benznidazole had a significantly lower risk of clinical events (OR, 0.29; 95% CI, 0.16-0.53). Up to 18% of patients discontinued treatment due to toxicity (cutaneous reactions followed by gastrointestinal disturbances); this was less common in children than in adults. CONCLUSIONS: Analysis of available information reveals that the efficacy of treatment in late chronic infection is doubtful. Although data generally point to a beneficial effect, this could be marginal. This uncertainty is largely the result of differences in the study populations, endpoints and follow-up periods, and the fact that almost all of the information on treatment in the late chronic phase comes from non-randomized studies.

Application of PCR-TTGE and PCR-RFLP for intraspecific and interspecific characterization of the genus Saccharomyces using actin gene (ACT1) primers.

In this work the actin gene was used to establish phylogenetic relationships of wider and more diffuse species of the genus Saccharomyces in food ecology by temporal temperature gradient electrophoresis (TTGE) and amplified restriction fragment length polymorphism (RFLP) analysis. Results for DNA RFLP analysis varied considerably, and some enzymes showed a high intra- and interspecific power; however, comparison of experimental results with those provided by the National Center for Biotechnology Information database disclosed a number of interesting variations. Only some experimental results matched the theoretical ones. A theoretical study of melting temperatures using available information from partial sequences of the actin gene was done. Several Saccharomyces species and strains could be distinguished using different TTGE melting points. Some degree of discrimination was achieved under different conditions, in that the Saccharomyces strains tested were separated into groups like the results obtained by PCR-RFLP.

Assessment of the potential real pollen related allergenic load on the atmosphere of Porto city.

The knowledge of the allergen content in the atmosphere is a useful tool to stablish the risk allergy warnings for the sensitive people. In Portugal the main airborne allergenic pollen come from trees (such as Betula or Olea), grasses or weeds (mainly Urticaceae). The present study sought the quantification of the Bet v 1, Ole e 1, Lol p1 and Par j1-2 aeroallergen concentration as well as how weather variables influence in the pollen and allergen concentration in Porto city. Aerobiological study was carried out by a Hirst-type volumetric sampler for pollen collection and a Burkard Cyclone sampler for the aeroallergens. A regression analysis between pollen and allergens was conducted for the identification the allergenic risk days. High Pollen Allergen Potency in the atmosphere was observed considering the low levels of airborne pollen detected. A significant and positive correlation has been obtained between pollen and aeroallergen values with the temperatures whereas the correlation was negative with relative humidity, rainfall and wind speed. Back trajectory methodology was applied in order to analyse the discordances between pollen and allergen maximum concentrations. The analysis showed that when the pollen and allergen peaks were registered on the same day, air masses always comes from the continent. However, when the peaks do not coincide, the air mass comes from the continent in the case of the pollen peak and from the sea for the allergen peak. This behaviour can be a consequence of the high humidity in the air masses from the sea, which can benefit the allergen release from pollen grains. In our study it was observed that the available traditional information for allergenic Type I patients, corresponding to the amount of pollen grains in the bioaerosol, do not accurately identify the real allergenic load in the air.

Oleaceae cross-reactions as potential pollinosis cause in urban areas.

It is worth noting the allergological problems induced by a not accurate design of the ornamental vegetation in the parks and streets of the cities. Usually, in the Oleaceae family, only the olive pollen is considered an important aeroallergen but other species of the family could be an important source of airborne pollen allergens. Pollen from Fraxinus, Olea and Ligustrum and its main aeroallergens were sampled in the atmosphere of an urban area in North-Western Spain during 2011. The allergen bioaerosol content was quantified by using specific 2-site ELISA and Ole e 1 antibodies. The Fra e 1 and Lig v 1 allergens were detected by means Ole e 1 antibodies. This fact demonstrates the cross-reactivity between the main allergens of Fraxinus, Olea and Ligustrum, plants widely species used as ornamental in the cities. Therefore, the urban allergenic people sensitized to Olea pollen could present allergenic reactions during the winter (due to ash pollen allergens), the spring (caused by olive pollen allergens) and the early summer (triggered by the privet flowering). As a consequence, sensitivity to the pollen of one species may favour development of sensitivity to all three species as consequence of the priming effect. The combination of pollen count and the allergen quantification must be assessed in the epidemiologic study of allergic respiratory diseases.

Airborne castanea pollen forecasting model for ecological and allergological implementation.

Castanea sativa Miller belongs to the natural vegetation of many European deciduous forests prompting impacts in the forestry, ecology, allergological and chestnut food industry fields. The study of the Castanea flowering represents an important tool for evaluating the ecological conservation of North-Western Spain woodland and the possible changes in the chestnut distribution due to recent climatic change. The Castanea pollen production and dispersal capacity may cause hypersensitivity reactions in the sensitive human population due to the relationship between patients with chestnut pollen allergy and a potential cross reactivity risk with other pollens or plant foods. In addition to Castanea pollen's importance as a pollinosis agent, its study is also essential in North-Western Spain due to the economic impact of the industry around the chestnut tree cultivation and its beekeeping interest. The aim of this research is to develop an Artificial Neural Networks for predict the Castanea pollen concentration in the atmosphere of the North-West Spain area by means a 20years data set. It was detected an increasing trend of the total annual Castanea pollen concentrations in the atmosphere during the study period. The Artificial Neural Networks (ANNs) implemented in this study show a great ability to predict Castanea pollen concentration one, two and three days ahead. The model to predict the Castanea pollen concentration one day ahead shows a high linear correlation coefficient of 0.784 (individual ANN) and 0.738 (multiple ANN). The results obtained improved those obtained by the classical methodology used to predict the airborne pollen concentrations such as time series analysis or other models based on the correlation of pollen levels with meteorological variables.

Engineering of an oenological Saccharomyces cerevisiae strain with pectinolytic activity and its effect on wine.

A pectinolytic industrial yeast strain of Saccharomyces cerevisiae was generated containing the S. cerevisiae endopolygalacturonase gene (PGU1) constitutively expressed under the control of the 3-phosphoglycerate kinase gene (PGK1) promoter. The new strain contains DNA derived exclusively from yeast and expresses a high polygalacturonic acid hydrolyzing activity. Yeast transformation was carried out by an integrative process targeting a dispensable upstream region of the acetolactate synthase locus (ILV2), which determines sulfometuron methyl resistance. Microvinification assays were performed on white and red musts with the transformed UCLMS-1M strain and with the same strain untransformed. It was found that the changes in the pectic polysaccharide contents did not directly affect the taste or flavor of the wine. From the data reported, it is deduced that the chief advantage of using the modified strain is that it improves the yield of must/wine extraction, while it also positively affects some variables relating to appearance.

Modification of kinetic parameters of glycogen phosphorylase from mantle tissue of Mytilus galloprovincialis by a phosphorylation mechanism.

Initial rate and affinity studies on mantle Mytilus phosphorylase a were carried out in order to find possible differences in its kinetic properties with respect to phosphorylase b. Phosphorylase a was not stimulated for any AMP concentrations. Michaelis constants (Km) are 0.05 mg/ml glycogen, 1.15 mM inorganic phosphate and 1.50 mM glucose-1-phosphate. The Kms for the substrates, in the direction of glycogen breakdown, are enhanced by non-saturating concentrations of cosubstrate, without reducing the apparent maximum velocity. First order and hyperbolic kinetics and values of the allosteric constant smaller than 2 were observed. These results suggest a catalytic mechanism different to that shown for mantle Mytilus phosphorylase b.

Somatostatin inhibits VIP- and isoproterenol-stimulated cyclic AMP accumulation in rat prostatic epithelial cells.

The dual regulation of cyclic AMP accumulation was studied in rat prostatic epithelial cells incubated with somatostatin, vasoactive intestinal peptide (VIP), and the beta-adrenergic agent isoproterenol. Somatostatin noncompetitively inhibited the stimulatory effect of VIP and isoproterenol, but it did not alter basal cyclic AMP levels. In addition to the multifactorial regulation of the cyclic AMP system in rat prostatic epithelium, these results suggest that somatostatin may play a physiological role at this level.

Evaluation of polygalacturonase activity in Saccharomyces cerevisiae wine strains.

A total of 61 S. cerevisiae strains, 60 of them isolated from wine ecosystems, were evaluated for the presence of the gene encoding endopolygalacturonase (PGU1) and for polygalacturonase (PG) activity. Nine strains lack the gene PGU1 and did not exhibit PG activity on plate assays. Of the 52 strains showing an amplified band corresponding to the size of PGU1 gene, only 36 degraded polygalacturonic acid (PGA) and 17 did not degrade it at any of the pH values used. The coding region of the PGU1 gene (ORF YJR153w) was not present in some PG activity negative strains. The S. cerevisiae UCLMS-39 strain was selected for its specific activity at different pHs, temperatures and oenological parameters. The temperature and pH optima were 50 degrees C and 3.5-5.5 respectively and it was only affected by ethanol. The PGU1 gene was cloned and sequenced. The production of a biologically functional endoPG in S. cerevisiae UCLMS-39 brings us a step closer to improving the qualities of outstanding enological yeasts naturally lacking PG activity.

Yeasts present during wine fermentation: comparative analysis of conventional plating and PCR-TTGE.

Yeasts isolated from must before and during fermentation at a wine cellar of La Mancha region in Spain were characterised using Polymerase Chain Reaction / Restriction Fragments Lengths Polymorphism and Polymerase Chain Reaction / Temporal Temperature Gradient Gel Electrophoresis. S. cerevisiae strains were differentiated using mtDNA restriction analysis. Direct PCR-TTGE was also used to study biodiversity during wine fermentation, and revealed the variations in the population. It was observed that isolation by conventional plating may afford a skewed view of the strains taking part in wine fermentation.

Beta-glucosidase activity in a Saccharomyces cerevisiae wine strain.

Beta-glucosidase activity contributes to aroma formation during the winemaking process. This study investigated whether beta-glucosidase activity was expressed by wild Saccharomyces cerevisiae strains and by a laboratory strain. beta-Glucosidase activity was assayed on several culture media and under various growth conditions. The highest activities were obtained in Yeast Extract Peptone medium, but activity was also detected using grape juice as the growth medium, although a 25% drop activity was observed when anaerobic conditions were employed. A number of parameters affecting beta-glucosidase activity were evaluated. Optimal conditions for activity were pH 4 and a temperature of 40-50 degrees C. The results showed beta-glucosidase activity to be present during the process of winemaking, although different from the optimal conditions.

Higher levels of serum copper in schizophrenic patients treated with depot neuroleptics.

The findings of previous research on the status of trace elements in patients with schizophrenia have been controversial. We studied 62 outpatients with a DSM-IV diagnosis of schizophrenia, and compared them with sex- and age-matched healthy controls. Serum copper levels were significantly higher in schizophrenic patients (mean 117.4 microg/dl; S.D. 23.4) than in healthy controls (105.6+/-27.9). Those patients on treatment with depot neuroleptics had higher copper levels. Zinc levels did not differ between patients and healthy controls. Altered levels of trace elements in schizophrenic patients may be a consequence of antipsychotic treatment.