Substrate specificity profiling of heat-sensitive serine protease from the fungus Onygena corvina.

Proteases catalyze hydrolysis of amide bonds within peptides and proteins, therefore they play crucial functions for organism functioning, but also in industry to facilitate numerous processes. Feather-degrading fungus Onygena corvina (O. corvina) is loaded with numerous proteases that can be utilized for variety of applications. The most active species of these enzymes is heat-sensitive serine protease (NHSSP), from O. corvina fungi and due to its potential applications in industry is an alternative to proteinase K. The uniqueness of NHSSP relies on the ability of NHSSP to hydrolyze peptides at neutral to acidic pH values between 5.0 to 8.5, with an optimum of 6.8 and a temperature activity ranging from 15-50°C making NHSSP exceptionally universal enzyme. Thus, we have performed the in-depth characterization of NHSSP substrate specificity by using a positional scanning substrate combinatorial library (PS-SCL). Afterward, we obtained a set of fluorescent substrates hydrolyzed by NHSSP that served as a leading sequence for the first tailored covalent inhibitor of this enzyme, containing a diphenylphosphonate as a warhead and MeOSuc amine protecting group. Our first inhibitor for NHSSP binds potently with target protease and is a tool for future study of this enzyme functions.

Genetic Load and Adaptive Potential of a Recovered Avian Species that Narrowly Avoided Extinction.

High genetic diversity is a good predictor of long-term population viability, yet some species persevere despite having low genetic diversity. Here we study the genomic erosion of the Seychelles paradise flycatcher (Terpsiphone corvina), a species that narrowly avoided extinction after having declined to 28 individuals in the 1960s. The species recovered unassisted to over 250 individuals in the 1990s and was downlisted from Critically Endangered to Vulnerable in the International Union for the Conservation of Nature Red List in 2020. By comparing historical, prebottleneck (130+ years old) and modern genomes, we uncovered a 10-fold loss of genetic diversity. Highly deleterious mutations were partly purged during the bottleneck, but mildly deleterious mutations accumulated. The genome shows signs of historical inbreeding during the bottleneck in the 1960s, but low levels of recent inbreeding after demographic recovery. Computer simulations suggest that the species long-term small Ne reduced the masked genetic load and made the species more resilient to inbreeding and extinction. However, the reduction in genetic diversity due to the chronically small Ne and the severe bottleneck is likely to have reduced the species adaptive potential to face environmental change, which together with a higher load, compromises its long-term population viability. Thus, small ancestral Ne offers short-term bottleneck resilience but hampers long-term adaptability to environmental shifts. In light of rapid global rates of population decline, our work shows that species can continue to suffer the effect of their decline even after recovery, highlighting the importance of considering genomic erosion and computer modeling in conservation assessments.

Dissecting the effect of soil on plant phenology and berry transcriptional plasticity in two Italian grapevine varieties (Vitis vinifera L.).

Grapevine embodies a fascinating species as regards phenotypic plasticity and genotype-per-environment interactions. The terroir, namely the set of agri-environmental factors to which a variety is subjected, can influence the phenotype at the physiological, molecular, and biochemical level, representing an important phenomenon connected to the typicality of productions. We investigated the determinants of plasticity by conducting a field-experiment where all terroir variables, except soil, were kept as constant as possible. We isolated the effect of soils collected from different areas, on phenology, physiology, and transcriptional responses of skin and flesh of a red and a white variety of great economic value: Corvina and Glera. Molecular results, together with physio-phenological parameters, suggest a specific effect of soil on grapevine plastic response, highlighting a higher transcriptional plasticity of Glera in respect to Corvina and a marked response of skin compared to flesh. Using a novel statistical approach, we identified clusters of plastic genes subjected to the specific influence of soil. These findings could represent an issue of applicative value, posing the basis for targeted agricultural practices to enhance the desired characteristics for any soil/cultivar combination, to improve vineyards management for a better resource usage and to valorize vineyards uniqueness maximizing the terroir-effect.

Replicate contact zones suggest a limited role of plumage in reproductive isolation among subspecies of the variable seedeater (Sporophila corvina).

After establishing secondary contact, recently diverged populations may remain reproductively isolated or may hybridize to a varying extent depending on factors such as hybrid fitness and the strength of assortative mating. Here, we used genomic and phenotypic data from three independent contact zones between subspecies of the variable seedeater (Sporophila corvina) to examine how coloration and genetic divergence shape patterns of hybridization. We found that differences in plumage coloration are probably maintained by divergent selection across contact zones; however, the degree of plumage differentiation does not match overall patterns of hybridization. Across two parallel contact zones between populations with divergent phenotypes (entirely black vs. pied plumage), populations hybridized extensively across one contact zone but not the other, suggesting that plumage divergence is not sufficient to maintain reproductive isolation. Where subspecies hybridized, hybrid zones were wide and formed by later-generation hybrids, suggesting frequent reproduction and high survivorship for hybrid individuals. Moreover, contemporary gene flow has played an important role in shaping patterns of genetic structure between populations. Replicated contact zones between hybridizing taxa offer a unique opportunity to explore how different factors interact to shape patterns of hybridization. Overall, our results demonstrate that divergence in plumage coloration is important in reducing gene flow but insufficient in maintaining reproductive isolation in this clade, and that other factors such as divergence in song and time since secondary contact may also play an important role in driving patterns of reduced hybridization and gene flow.

Al establecer contacto secundario, las poblaciones que divergieron recientemente pueden permanecer reproductivamente aisladas o pueden hibridarse en distintos grados, dependiendo de factores como la aptitud (fitness) y la fuerza del apareamiento selectivo. Aquí, utilizamos datos genómicos y fenotípicos de tres zonas de contacto independientes entre subespecies del Semillero Variable (Sporophila corvina), para examinar cómo la coloración y la divergencia genética regulan los patrones de hibridación. A través de las zonas de contacto, encontramos que las diferencias en la coloración del plumaje posiblemente se mantienen por selección divergente, pero el grado de diferenciación no coincide con los patrones generales de hibridación. En dos zonas de contacto análogas entre poblaciones con fenotipos divergentes (totalmente negro vs plumaje de varios colores), las poblaciones hibridaron ampliamente en una zona de contacto, pero no en la otra, lo que sugiere que la divergencia del plumaje no es suficiente para mantener el aislamiento reproductivo. Donde las subespecies hibridaron, las zonas híbridas eran amplias y estaban formadas por híbridos de generaciones posteriores, lo que sugiere reproducción frecuente y alta sobrevivencia de los híbridos. Además, el flujo génico ha desempeñado un papel importante en la configuración de patrones de estructura genética entre poblaciones. Las réplicas de zonas de contacto entre taxones que hibridan ofrecen una oportunidad para explorar cómo interactúan diversos factores para dar forma a los patrones de hibridación. En general, nuestros resultados demuestran que la divergencia en la coloración del plumaje es importante para reducir el flujo génico, pero insuficiente para mantener el aislamiento reproductivo en este clado, y que otros factores, como la divergencia en el canto y el tiempo transcurrido desde el contacto secundario, también pueden desempeñar un papel importante en la reducción del flujo génico e hibridación.

Temperature affects organic acid, terpene and stilbene metabolisms in wine grapes during postharvest dehydration.

The partial dehydration of grapes after harvest is a traditional practice in several winegrowing regions that leads to the production of high quality wines. Postharvest dehydration (also known as withering) has a significant impact on the overall metabolism and physiology of the berry, yielding a final product that is richer in sugars, solutes, and aroma compounds. These changes are, at least in part, the result of a stress response, which is controlled at transcriptional level, and are highly dependent on the grape water loss kinetics and the environmental parameters of the facility where grapes are stored to wither. However, it is difficult to separate the effects driven by each single environmental factor from those of the dehydration rate, especially discerning the effect of temperature that greatly affects the water loss kinetics. To define the temperature influence on grape physiology and composition during postharvest dehydration, the withering of the red-skin grape cultivar Corvina (Vitis vinifera) was studied in two conditioned rooms set at distinct temperatures and at varying relative humidity to maintain an equal grape water loss rate. The effect of temperature was also studied by withering the grapes in two unconditioned facilities located in geographic areas with divergent climates. Technological, LC-MS and GC-MS analyses revealed higher levels of organic acids, flavonols, terpenes and cis- and trans-resveratrol in the grapes withered at lower temperature conditions, whereas higher concentrations of oligomeric stilbenes were found in the grapes stored at higher temperatures. Lower expression of the malate dehydrogenase and laccase, while higher expression of the phenylalanine ammonia-lyase, stilbene synthase and terpene synthase genes were detected in the grapes withered at lower temperatures. Our findings provide insights into the importance of the temperature in postharvest withering and its effect on the metabolism of the grapes and on the quality of the derived wines.

A high-density integrated map for grapevine based on three mapping populations genotyped by the Vitis18K SNP chip.

KEY MESSAGE: We present a high-density integrated map for grapevine, allowing refinement and improved understanding of the grapevine genome, while demonstrating the applicability of the Vitis18K SNP chip for linkage mapping. The improvement of grapevine through biotechnology requires identification of the molecular bases of target traits by studying marker-trait associations. The Vitis18K SNP chip provides a useful genotyping tool for genome-wide marker analysis. Most linkage maps are based on single mapping populations, but an integrated map can increase marker density and show order conservation. Here we present an integrated map based on three mapping populations. The parents consist of the well-known wine cultivars 'Cabernet Sauvignon', 'Corvina' and 'Rhine Riesling', the lesser-known wine variety 'Deckrot', and a table grape selection, G1-7720. Three high-density population maps with an average inter-locus gap ranging from 0.74 to 0.99 cM were developed. These maps show high correlations (0.9965-0.9971) with the reference assembly, containing only 93 markers with large order discrepancies compared to expected physical positions, of which a third is consistent across multiple populations. Moreover, the genetic data aid the further refinement of the grapevine genome assembly, by anchoring 104 yet unanchored scaffolds. From these population maps, an integrated map was constructed which includes 6697 molecular markers and reduces the inter-locus gap distance to 0.60 cM, resulting in the densest integrated map for grapevine thus far. A small number of discrepancies, mainly of short distance, involve 88 markers that remain conflictual across maps. The integrated map shows similar collinearity to the reference assembly (0.9974) as the single maps. This high-density map increases our understanding of the grapevine genome and provides a useful tool for its further characterization and the dissection of complex traits.

Past and Contemporaneous Otolith Fingerprints Reveal Potential Anthropogenic Interferences and Allows Refinement of the Population Structure of Isopisthus parvipinnis in the South Brazil Bight.

In this study, otolith geochemical signatures (Element:Ca ratios) were used to investigate the long-term spatial shifts of the population structure of Isopisthus parvipinnis, Bigtooth corvina, an economically and ecologically important Brazilian fish species. Two-hundred and ninety-seven juvenile individuals from historical (1975) and contemporary (2018/2019) samples were collected in five subareas [São Paulo: North-NSP, Center-CSP and South-SSP; Paraná (PR) and Santa Catarina (SC)] of the shallow waters off the coast of the South Brazil Bight were analyzed. The main informative single elements were Co:Ca, Cu:Ca, Li:Ca, Mg:Ca, Mn:Ca, Ni:Ca, Na:Ca, and Rb:Ca. Multivariate analysis showed spatial differences in otolith chemical composition over the years. Samples from 1975 presented an overall low reclassification rate (58%), suggesting the existence of two population units: (1) SP + PR; and (2) SC. However, samples from 2018/2019 discriminated four distinct population units with a good overall reclassification (80%): (1) NSP; (2) CSP; (3) SSP + PR; and (4) SC. This spatial differentiation on the geochemical signatures probably reflects the effects of long-term temporal variability in oceanographic conditions, anthropogenic influence, and climate change on this coastal ecosystem. The data also corroborate and refines the population structure scenario of I. parvipinnis recently described using complementary phenotypic tags.

Rapid diversification of the Variable Seedeater superspecies complex despite widespread gene flow.

Disentangling the evolutionary relationships of rapidly radiating clades is often challenging because of low genetic differentiation and potentially high levels of gene flow among diverging taxa. The genus Sporophila consists of small Neotropical birds that show, in general, relatively low genetic divergence, but particularly high speciation rates and pronounced variation in secondary sexual traits (e.g., plumage color), which can be important in generating premating reproductive isolation. In cases like these, the use of genome-wide sequence data can increase the resolution to uncover a clade's evolutionary history. Here, we used a phylogenomic approach to study the evolutionary history and genetic structure of the Variable Seedeater superspecies complex, which includes S. corvina, S. intermedia, and S. americana. Using ∼25,000 genome-wide single nucleotide polymorphisms (SNPs), we confirmed that the Variable Seedeater superspecies complex is monophyletic. However, a phylogenetic reconstruction based on a mitochondrial marker (ND2) resulted in a discordant tree topology, particularly in the position of Wing-barred Seedeater S. americana, which might be due to a mitochondrial capture event. Our results suggest historical gene flow among lineages, particularly between species with conflicting topologies. Among the four phenotypically variable S. corvina subspecies, our structure analyses identified three main distinct genetic groups (K = 3), and that the entirely black subspecies, S. c. corvina, is derived from within a pied-colored clade. Further, we inferred widespread gene flow across the whole species' distribution, including between subspecies. However, gene flow was about 100 times lower at the geographic boundaries of the entirely black and the pied subspecies, suggesting an important role for plumage divergence in limiting gene flow. Overall, our findings suggest that the early diversification of the Sporophila genus occurred rapidly despite historical gene flow between lineages and that divergence in plumage color possibly influences the extent of gene flow among taxa.

[Determination of 19 chlorophenols in fish by QuEChERS-gas chromatography-mass spectrometry].

With the increasing use of chlorine-containing pesticides, hypochlorous acid disinfection water as well as aquatic product insecticides and fungicides are widely used in the cultivation of fish. This has led to the contamination of fish by chlorophenol compounds. However, currently, there is no standard method for the simultaneous determination of 19 chlorophenol compounds in fish. In this study, the optimum chromatography and mass spectrometry conditions were determined by investigating the instrument parameters. The 19 chlorophenol compounds were well separated using the DB-5MS capillary chromatographic column (30 m×0.25 mm×0.25 µm) with a carrier gas flow rate of 1 mL/min. Under this condition, the chromatographic peak was sharp and symmetric. An analytical method was developed for the simultaneous determination of the 19 chlorophenol compounds in fish using gas chromatography-mass spectrometry coupled with QuEChERS pretreatment. The improved QuEChERS method was used in sample preparation. The 19 chlorophenol compounds were extracted with organic solvents and purified with purifying agents. During the experiment, the effect of the kinds and volumes of the extraction solvent, as well as the types and dosages of the purifying agent, on the recoveries of the 19 chlorophenol compounds were investigated. Moreover, the temperature and time of derivatization, as well as the dosage of the derivatization agent, were optimized. All aforementioned analyses were conducted with the aim of determining the optimal pretreatment method. Finally, the optimized gas chromatography-mass spectrometry conditions were employed for the quantitative determination of 19 chlorophenol compounds in fish samples. Based on the experimental results, the best extraction method was determined to be the one where the extraction agent (10 mL ethyl acetate) was added to 3 g sodium chloride and 5 g anhydrous magnesium sulfate in the test tube, followed by ultrasonication for 15 min. The sample was centrifuged at 4500 r/min for 5 min, and 500 mg C18 was selected as the purifying agent to purify the supernatant. The purified supernatant was blown with nitrogen to less than 1 mL at 45 ℃, and then redissolved with ethyl acetate to 1 mL. Subsequently, the sample solution was passed through a 0.22 µm organic filter membrane, following which 50 µL bis(trimethylsilyl)trifluoroacetamide was added for derivatization at 45 ℃ for 30 min. Lastly, the 19 chlorophenol compounds were determined by gas chromatography-mass spectrometry with an electrospray ionization source and selecting ion monitoring mode. The 19 chlorophenol compounds were then quantitatively analyzed by the external standard method. The compounds showed good linearity in the concentration range of 0.4-10 µg/L, with correlation coefficients (R2) greater than 0.998. The limits of detection and limits of quantification were 0.01-0.05 µg/kg and 0.04-0.16 µg/kg, respectively. Moreover, the average recoveries of the 19 chlorophenol compounds were in the range of 70.6%-115.0% at three spiked levels, and the relative standard deviations were in the range of 2.6%-10.5%. The established method in this study was applied to detect and analyze chlorophenol compounds in actual samples. The experimental results showed that various levels of chlorophenol compounds could be detected in different fishes. Among them, the total amount of chlorophenol compounds detected in the Corvina was 8.74 µg/kg, followed by the Crucian carp at 7.59 µg/kg, and the minimum detected amount in rice fish (1.59 µg/kg). With its simple operation, high sensitivity, and good repeatability, the established method simplifies the pre-treatment of fish samples. It can also meet the requirements for the high-throughput detection of 19 chlorophenol compounds in fish, thereby significantly improving the detection efficiency of chlorophenols. Moreover, the method provides crucial technical support and a theoretical basis for the establishment of feasible detection standards for chlorophenols in China, as well as for the control of residue levels of chlorophenol compounds in fish. The findings have important practical significance to implement management measures during fish breeding and transportation.

Bioinformatics based discovery of new keratinases in protease family M36.

Keratinases are proteases that can catalyze the degradation of insoluble keratinous biomass. Keratinases in protease family M36 (MEROPS database) are endo-acting proteases. In total, 687 proteases are classified in family M36. In the present study, new keratinolytic enzymes were identified in protease family M36 using the bioinformatics tool Conserved Unique Peptide Patterns (CUPP). Via CUPP, M36 family members were classified into 11 groups, with CUPP group 1 containing the three currently known and sequenced family M36 keratinases (derived from the fungi Fusarium oxysporum, Microsporum canis and Onygena corvina) as well as an additional 71 uncharacterized M36 proteases. In order to assess the relevance of CUPP group 1 categorization to keratinolytic function, four uncharacterized M36 proteases and the known keratinase from F. oxysporum (in CUPP group 1) were selected for recombinant expression and keratinolytic activity assessment. The four hitherto unknown M36 proteases were from Phaeosphaeria nodorum, Aspergillus clavatus, Pseudogymnoascus pannorum and Nectria haematococca, and represent four different fungal taxonomical classes. The genes encoding the selected M36 proteases were individually expressed in Pichia pastoris and all proteases displayed keratinase activity on keratin azure. Additionally, the activity on different keratinase substrates, optimal reaction conditions and thermal stability were determined for the two most active new keratinases. The results validate the applicability of CUPP for function-based discovery of non-characterized keratinases and present new robust keratinases for potential use in keratin upgrading.

Volatile Compounds in Monovarietal Wines of Two Amarone Della Valpolicella Terroirs: Chemical and Sensory Impact of Grape Variety and Origin, Yeast Strain and Spontaneous Fermentation.

Aroma profiles of withered Corvina and Corvinone wines from two different Valpolicella terroirs were investigated in relationship to yeast strain and use of spontaneous fermentation. The results indicated that volatile chemical differences between wines were mainly driven by grape origin, which was associated with distinctive compositional profiles. Wine content in terpenes, norisoprenoids, benzenoids and C6 alcohols, as well as some fermentative esters, were indeed significantly affected by grape origin. Conversely, yeast strain influence was mainly associated with fermentation-derived esters. Sensory analysis, besides confirming the major role of grape origin as driver of wine differentiation, indicated that spontaneous fermentations reduced the sensory differences associated with grape origin and variety, mainly due to high content of acetic acid and ethyl acetate.

Effects of Traditional and Modern Post-Harvest Withering Processes on the Composition of the Vitis v. Corvina Grape and the Sensory Profile of Amarone Wines.

In the Valpolicella area (Verona, Italy) Vitis vinifera cv. Corvina is the main grape variety used to produce Amarone wine. Before starting the winemaking process, the Corvina grapes are stored in a withering (i.e., dehydrating) warehouse until about 30% of the berry weight is lost (WL). This practice is performed to concentrate the metabolites in the berry and enrich the Amarone wine in aroma and antioxidant compounds. In compliance with the guidelines and strict Amarone protocol set by the Consorzio of Amarone Valpolicella, withering must be carried out by setting the grapes in a suitable environment, either under controlled relative air humidity (RH) conditions and wind speed (WS)-no temperature modification is to be applied-or, following the traditional methods, in non-controlled environmental conditions. In general, the two processes have different dehydration kinetics due to the different conditions in terms of temperature, RH, and WS, which affect the accumulation of sugars and organic acids and the biosynthesis of secondary metabolites such as stilbenes and glycoside aroma precursors. For this study, the two grape-withering processes were carried out under controlled (C) and non-controlled (NC) conditions, and the final compositions of the Corvina dried grapes were compared also to evaluate the effects on the organoleptic characteristics of Amarone wine. The findings highlighted differences between the two processes mainly in terms of the secondary metabolites of the dried grapes, which affect the organoleptic characteristics of Amarone wine. Indeed, by the sensory evaluation, wines produced by adopting the NC process were found more harmonious, elegant, and balanced. Finally, we can state how using a traditional system, grapes were characterised by higher levels of VOCs (volatile compounds), whilst wines had a higher and appreciable complexity and finesse.

Comparison of collagen characteristic from the skin and swim bladder of Gulf corvina (Cynoscion othonopterus).

Collagens extracted from different tissues and fish species display different physicochemical properties, thus novel sources require characterization. Gulf corvina (Cynoscion othonopterus) is processed industrially for food. Of the by-products, the swim bladder is used for fish maw, but other tissues are treated as waste. In the present study, pepsin-soluble collagen from Gulf corvina skin and swim bladder was extracted and characterized. Skin produced a higher collagen yield (82 ± 1.53 %) than swim bladder (69 ± 1.60 %). Both collagens exhibited electrophoresis bands corresponding to ([α1(I)]2α2(I)) and ß chains, all characteristic of type I collagen. Spectra analysis showed the collagens to maintain their triple-helix structure. The skin collagen had a lower denaturation temperature (29.8 °C) than the swim bladder collagen (32.5 °C), due to its relatively low imino acid content (168 vs. 190 /1000 residues, respectively). Both collagens were highly soluble in acidic pH ranges; Zeta potential values were 5.5 for the skin collagen and 6.2 for the swim bladder collagen. Gulf corvina skin and swim bladder are excellent sources of type I collagen with similar physicochemical properties.

Diversity and foraging patterns of bees on flowers of Cucurbita pepo (Cucurbitaceae) in Costa Rica

Abstract Introduction: The species and functional diversity of pollinators are important components for the reproduction of cultivated plants. More information is necessary about this diversity and its geographical variation in crops such as Cucurbita pepo, an important crop in global agriculture. Objective: To describe the taxonomic diversity, geographic variation and foraging patterns of the community of bees that visit C. pepo crops in Costa Rica. Methods: Squash fields were visited at 11 locations within three geographic regions of the country, where the groups of bees and their relative frequency were determined. Through video recordings, information was obtained on their behavior at two locations. Results: A minimum of 27 species belonging to 19 genera and 2 families of bees were found. Three species were dominant in 10 localities (Eucera limitaris, Apis mellifera and Trigona corvina). Altitude reduces bee diversity due to the dominance of Bombus ephipiatus in high regions. Two genera of halictids (Megalopta and Caenaugochlora) that are rarely reported in this crop were frequently observed. Trigona bees dominated among the flowers later in the morning, lacerating nectary holes to facilitate nectar collection. Conclusions: Squash fields in Costa Rica are visited by a highly diverse bee community, which may ensure pollination via complementarity in the face of spatial or seasonal changes in environmental conditions.

Resumen Introducción: Dada la importancia del componente diversidad para la polinización de plantas cultivadas, es necesario obtener más información de esta diversidad y su variación geográfica en cultivos como Cucurbita pepo, uno de los cultivos más importantes de la agricultura centroamericana. Objetivo: Describir la diversidad y la variación geográfica de la comunidad de abejas que visitan este cultivo en Costa Rica, y algunos aspectos de sus patrones de búsqueda de alimento. Métodos: Se visitaron cultivos de C. pepo en 11 localidades dentro de tres regiones geográficas de Costa Rica, donde se determinó los grupos de abejas y su frecuencia relativa. Por medio de grabaciones de video se registró el comportamiento de cada grupo en dos localidades. Resultados: Fueron encontradas un mínimo de 27 especies pertenecientes a 19 géneros y 2 familias de abejas. Tres especies son dominantes en 10 localidades (Eucera limitaris, Apis mellifera y Trigona corvina). La altitud reduce la diversidad de abejas debido a la dominancia de Bombus ephipiatus en regiones altas. Se observaron dos géneros de halíctidos (Megalopta y Caenaugochlora) no previamente reportados en este cultivo. Abejas Trigona dominan las flores en horarios más tardíos de la mañana, donde algunas veces muerden los orificios de los nectarios para facilitar la recolecta de néctar. Conclusiones: La diversidad de abejas que visitan C. pepo en Costa Rica parece asegurar su polinización ante cambios espaciales o estacionales en condiciones ambientales.

Diversity of Italian red wines: A study by enological parameters, color, and phenolic indices.

An extensive survey was conducted on 110 Italian monovarietal red wines from a single vintage to determine their standard compositional, color, and phenolic characteristics, analysing more than 35 parameters evaluated through methods commonly used in the wine industry. 'Primitivo' achieved the highest average alcohol strength (15.4% v/v) and dry extract values, while 'Cannonau' showed the lowest total acidity. 'Corvina' had the lowest phenolic content (1065 mg/L by Folin-Ciocalteu assay), remarkably different from the highest found in 'Sagrantino' (3578 mg/L), the latter being also the richest variety in both proanthocyanidins and vanillin-reactive flavanols. 'Teroldego' wines were the richest in both total and monomeric anthocyanins (702 and 315 mg/L, respectively), followed by 'Aglianico' and 'Raboso Piave', while 'Corvina', 'Nebbiolo', and 'Nerello Mascalese' were the poorest. 'Montepulciano' and 'Sangiovese' showed intermediate values for the majority of the parameters analyzed. A multivariate PCA-DA approach allowed achieving both a classification of the different wines as well as the discrimination of 'Sangiovese' wines produced in two regions (Emilia Romagna and Toscana) that returned a 42-66% success rate depending on the zone considered. Taking into account the number and diversity of the wines analyzed, a correlation study helped in better understanding the underlying relations between the most common and widespread analytical techniques for phenolic and color determinations.

A new maximum size record of the shi drum (Umbrina cirrosa Linnaeus, 1758) for Aegean Sea / Um novo registro de tamanho máximo do corvina (Umbrina cirrosa Linnaeus, 1758) para o Mar Egeu

The present study has been conducted to find out new findings on maximum length and weight values of Umbrina cirrosa in the Bay of Saros (Northern Aegean Sea, Turkey). On September, 11, 2016, a single specimen of Umbrina cirrosa with 68.8 cm total length and 2600.00 g total weight was caught by handline at a depth of 20 m. The provable size is the second largest length ever reported in the all seas of the world.

O presente estudo foi realizado para descobrir novas descobertas sobre os valores máximos de comprimento e peso de Umbrina cirrosa na Baía de Saros (Mar Egeu do Norte, Turquia). Em 11 de setembro de 2016, um único espécime de Umbrina cirrosa com 68.8 cm de comprimento total e 2600.00 g de peso total foi capturado por linha de mão a uma profundidade de 20 m. O tamanho provável é o segundo maior comprimento já registrado em todos os mares do mundo.

Elucidations on the structures of some putative flavonoids identified in postharvest withered grapes (Vitis vinifera L.) by quadrupole time-of-flight mass spectrometry.

Grape dehydration is an oenological process used for production of high-quality reinforced and sweet wines. High-resolution mass spectrometry (ultrahigh-performance liquid chromatography/quadrupole time-of-flight [UHPLC/QTOF]) was used to deepen the characterization of some flavonoids previously proposed in Corvina and Raboso Piave withered grapes. By performing a targeted data analysis workflow and orthogonal identification approaches (tandem mass spectrometry [MS/MS]), in silico fragmentation, and calculation of putative retention time), elucidation on the structures of six compounds previously proposed was achieved (taxifolin-pentoside, two tetrahydroxyflavanone-hexoside derivatives, a tetrahydroxy-dimethoxyflavanone-hexoside derivative, a pentahydroxyflavone, and peonidin-O-pentoside); and the structures of four putative new grape flavonoids were characterized (dihydromyricetin-O-hexoside, taxifolin-di-O-hexoside, isorhamnetin, and a pinoquercetin isomer). Findings enlarge the panorama of flavonoids in grape and of their possible biosynthetic pathways.

The Effect of Soil on the Biochemical Plasticity of Berry Skin in Two Italian Grapevine (V. vinifera L.) Cultivars.

Grapevine represents a particularly interesting species as concerns phenotypic plasticity, considering that the terroir, meaning the contribution of the geography, geology, and climate of a certain place, together with the agronomical practices utilized, may deeply influence the berry phenotype at the physiological, molecular, and biochemical levels. This phenomenon leads to the production of wines that, although produced from the same variety, exhibit different enological profiles and represents an issue of increasing interest from both a biological and an economic point of view. The main objective of the present study was to deepen the understanding of phenotypic plasticity in grapevine, trying to dissect the role of one its important components - the soil - by investigating the singular effect that different physico-chemical soil properties can produce in terms of berry plasticity at the phenological, physiological, and biochemical levels in a red and a white variety of great economic importance in Italy and overseas: Corvina and Glera. The results indicated a genotype-dependent response to the soil factor, with higher biochemical plasticity in Corvina with respect to Glera and suggested a key role of specific soil properties, including the skeleton, texture, and mineral composition, on the metabolite profile of berry skin.

Metazoan parasites of Plagioscion squamosissimus (Osteichthyes: Sciaenidae) of two rivers from the eastern Amazon (Brazil).

Plagioscion squamosissimus, commonly known as "white hake" or "corvina", is among the most commercially important fish in the Amazon and host to a great diversity of endoparasites. The aim of the present study was to compare the communities and infracommunities of metazoan parasites that infect P. squamosissimus in two rivers from the eastern Brazilian Amazon. A total of 75 specimens of P. squamosissimus were collected from Lago Grande do Curuái and the mouth of the Tapajós River. Morphological analysis revealed the presence of 16 parasite species, three myxozoans (Myxobolus sp., Ceratomyxa sp., Henneguya sp.), two trematodes (Austrodiplostomum compactum, Digenea gen. sp. (metacercariae), two monogeneans (Diplectanum sp., Euryhaliotrema sp.), three nematodes (Procamallanus (S.) sp., Anisakis sp., Pseudoproleptus sp. (larva), two acanthocephalans (Rhadinorhynchus plagioscionis and Neoechinorhynchus sp.), one Cestoda Ptychobothriidae gen. sp. and three crustaceans (Therodamas sp., Ergasilus sp., Dolops sp.). Six new records of parasites of P. squamosissimus were made, of which three were myxozoan species, one nematode species and two crustacean species. There were differences in the component communities of parasites of both rivers studied and these were discussed.

An alternative for proteinase K-heat-sensitive protease from fungus Onygena corvina for biotechnology: cloning, engineering, expression, characterization and special application for protein sequencing.

BACKGROUND: A neutral, heat-sensitive serine protease (NHSSP) originating from the feather-degrading fungus Onygena corvina (O. corvina) was described and defined as an alkaline serine protease of the subtilisin type S8 family, exhibiting an enzymatic activity at neutral pH. Generally, broad specificity proteases, such as proteinase K or trypsin, have found numerous applications in research and biotechnology. RESULTS: We report the cloning and expression in the yeast PichiaPink™ system, as well as purification, and characterization of the NHSSP. Recombinant, His6-tagged NHSSP was efficiently expressed from an optimized, synthetic gene and purified using a simple protocol based on ammonium sulfate fractionation and hydrophobic interaction chromatography. The enzyme shows atypical C-terminal processing, the coded preproprotein undergoes signal peptide removal and maturation through the clipping of a propeptide section and 10 amino acids (aa) from the C-terminus, including the His6-tag. The deletion variant has been constructed, devoid of the C-terminal ORF segment, thus eliminating the need for C-terminal processing. Both NHSSP variants exhibit very similar enzymatic characteristics. The purified enzymes were characterized to determine the optimal proteolytic conditions. We revealed that the mature NHSSP is reproducibly active over a wide pH range from neutral to mild acidic (pH of 5.0 to 8.5), with an optimum at pH 6.8, and at temperatures of 15 to 50 °C with an optimum at 38-42 °C. Interestingly, we demonstrated that the protease can be fully deactivated by a moderate increase in temperature of about 15 °C from the optimum to over 50 °C. The protease was partially sensitive to serine protease inhibitors, and not inhibited by chelating or reducing agents and detergents. SDS induced autolysis of NHSSP, which points to a high stimulation of its proteolytic activity. CONCLUSIONS: The NHSSP was produced as a recombinant protein with high efficiency. Compared to proteinase K, the most common serine protease used, NHSSP shows an approx. twofold higher specific activity. Protein sequencing can be a valuable technical application for the protease. The protein coverage is significantly higher in comparison to trypsin and reaches about 84-100% for ß-lactoglobulin (BLG), antibody (mAb) light and heavy chains. Furthermore, the option to perform digestions at neutral to slightly acidic pH-values down to pH 5.0 avoids modification of peptides, e.g. due to deamidation.