Quantitative and qualitative evaluation of the impact of the G2 enhancer, bead sizes and lysing tubes on the bacterial community composition during DNA extraction from recalcitrant soil core samples based on community sequencing and qPCR

<div><p>Soil DNA extraction encounters numerous challenges that can affect both yield and purity of the recovered DNA. Clay particles lead to reduced DNA extraction efficiency, and PCR inhibitors from the soil matrix can negatively affect downstream analyses when applying DNA sequencing. Further, these effects impede molecular analysis of bacterial community compositions in lower biomass samples, as often observed in deeper soil layers. Many studies avoid these complications by using indirect DNA extraction with prior separation of the cells from the matrix, but such methods introduce other biases that influence the resulting microbial community composition. To address these issues, a direct DNA extraction method was applied in combination with the use of a commercial product, the G2 DNA/RNA Enhancer, marketed as being capable of improving the amount of DNA recovered after the lysis step. The results showed that application of G2 increased DNA yields from the studied clayey soils from layers from 1.00 to 2.20 m. Importantly, the use of G2 did not introduce bias, as it did not result in any significant differences in the biodiversity of the bacterial community measured in terms of alpha and beta diversity and taxonomical composition. Finally, this study considered a set of customised lysing tubes for evaluating possible influences on the DNA yield. Tubes customization included different bead sizes and amounts, along with lysing tubes coming from two suppliers. Results showed that the lysing tubes with mixed beads allowed greater DNA recovery compared to the use of either 0.1 or 1.4 mm beads, irrespective of the tube supplier. These outcomes may help to improve commercial products in DNA/RNA extraction kits, besides raising awareness about the optimal choice of additives, offering opportunities for acquiring a better understanding of topics such as vertical microbial characterisation and environmental DNA recovery in low biomass samples.</p></div

Influence of vintage, geographic location and cultivar on the structure of microbial communities associated with the grapevine rhizosphere in vineyards of San Juan Province, Argentina

Soil microbiomes, as a primary reservoir for plant colonizing fungi and bacteria, play a major role in determining plant productivity and preventing invasion by pathogenic microorganisms. The use of 16S rRNA and ITS high-throughput amplicon sequencing for analysis of complex microbial communities have increased dramatically in recent years, establishing links between wine specificity and, environmental and viticultural factors, which are framed into the elusive terroir concept. Given the diverse and complex role these factors play on microbial soil structuring of agricultural crops, the main aim of this study is to evaluate how external factors, such as vintage, vineyard location, cultivar and soil characteristics, may affect the diversity of the microbial communities present. Additionally, we aim to compare the influence these factors have on the structuring of bacterial and fungal populations associated with Malbec grapevine rhizosphere with that of the more widespread Cabernet Sauvignon grapevine cultivar. Samples were taken from Malbec and Cabernet Sauvignon cultivars from two different vineyards in the San Juan Province of Argentina. Total DNA extracts from the rhizosphere soil samples were sequenced using Illumina’s Miseq technology, targeting the V3-V4 hypervariable 16S rRNA region in prokaryotes and the ITS1 region in yeasts. The major bacterial taxa identified were Proteobacteria, Bacteroidetes and Firmicutes, while the major fungal taxa were Ascomycetes, Basidiomycetes, Mortierellomycetes and a low percentage of Glomeromycetes. Significant differences in microbial community composition were found between vintages and vineyard locations, whose soils showed variances in pH, organic matter, and content of carbon, nitrogen, and absorbable phosphorus.Fil: Oyuela Aguilar, Monica. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Departamento de Ciencias Biológicas; ArgentinaFil: Gobbi, Alex. Universidad de Copenhagen; DinamarcaFil: Browne, Patrick D.. Universidad de Copenhagen; DinamarcaFil: Ellegaard Jensen, Lea. University Aarhus; DinamarcaFil: Hansen, Lars Hestbjerg. Universidad de Copenhagen; DinamarcaFil: Semorile, Liliana. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Microbiología Molecular; ArgentinaFil: Pistorio, Mariano. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; Argentin

Pruning wound protection products induce alterations in the wood mycobiome profile of Grapevines

Fungal pathogens involved in grapevine trunk diseases (GTDs) may infect grapevines throughout their lifetime, from nursery to vineyard, via open wounds in stems, canes or roots. In vineyards, pruning wound protection products (PWPPs) offer the best means to reduce the chance of infection by GTD fungi. However, PWPPs may affect non-target microorganisms that comprise the natural endophytic mycobiome residing in treated canes, disrupting microbial homeostasis and indirectly influencing grapevine health. Using DNA metabarcoding, we characterized the endophytic mycobiome of one-year-old canes of cultivars Cabernet Sauvignon and Syrah in two vineyards in Portugal and Italy and assessed the impact of established and novel PWPPs on the fungal communities of treated canes. Our results reveal a large fungal diversity (176 taxa), and we report multiple genera never detected before in grapevine wood (e.g., Symmetrospora and Akenomyces). We found differences in mycobiome beta diversity when comparing vineyards (p = 0.01) but not cultivars (p > 0.05). When examining PWPP-treated canes, we detected cultivar- and vineyard-dependent alterations in both alpha and beta diversity. In addition, numerous fungal taxa were over- or under-represented when compared to control canes. Among them, Epicoccum sp., a beneficial genus with biological control potential, was negatively affected by selected PWPPs. This study demonstrates that PWPPs induce alterations in the fungal communities of grapevines, requiring an urgent evaluation of their direct and indirect effects on plants health with consideration of factors such as climatic conditions and yearly variations, in order to better advise viticulturists and policy makers.info:eu-repo/semantics/publishedVersio

Effect of training loads on physiological parameters of soccer players

O objetivo desse estudo foi verificar os efeitos de três semanas de treinamentos com intensidades monitoradas sobre a capacidade aeróbia de futebolistas profissionais. Quatorze futebolistas integrantes de equipe da primeira divisão do Campeonato Brasileiro de 2010, foram avaliados pré e pós três semanas de treinamento. O limiar anaeróbio (LAn) foi determinado pelo método bi-segmentado, para isso quatro esforços submáximos de 800 metros com intensidades de 10, 12, 14 e 16 km/h foram aplicados. Trinta três sessões de treinamentos foram quantificadas em zonas de acordo com frequência cardíaca referente ao LAn (FCLAn): Z1 – 10% abaixo, Z2 – 90-100% e Z3 – acima da FCLAn. Durante os treinamentos os jogadores permaneceram 31,17±14,86%, 42,96±14,90% e 25,87±16,67% em Z1, Z2 e Z3, respectivamente. Não foram encontradas diferenças significativas no LAn (pré = 13,29 ± 0,71 km∙h-1 ; pós=12,85 ± 0,90 km∙h-1 ), percepção subjetiva de esforço correspondente ao LAn (pré = 11,53 ± 1,45 u.a; pós=11,23 ± 1,53 u.a) e FCLAn (pré = 166,64 ± 10,69 bpm; pós = 174,50 ± 10,89 bpm), indicando que três semanas de treinamento são insuficiente para gerar adaptações positivas no LAn de futebolistas.The aim of this study was to investigate the effects of three weeks of training with intensity monitored on the aerobic capacity of professional soccer players. Fourteen players, members of a first division Brazilian Championship team in 2010, aged 22.78 ± 3.06 years were evaluated pre and post three weeks of training. The anaerobic threshold intensity LAn was determined by bi-segmented method, for this four submaximal efforts of 800 meters with intensities 10, 12, 14 and 16 km/h were applied. Thirty three training sessions were quantified in zones according to heart rate related to the LAn (FCLAn): Z1 – 10% below, Z2 – 90-100% and Z3 – above the FCLAn. During training participants remained 31.17 ± 14.86%, 42.96% and 25.87 ± 14.90 ± 16.67% in Z1, Z2, and Z3 respectively. There were no significant differences in the LAn (pre = 13,29 ± 0,71 km∙h-1 ; post = 12,85 ± 0,90 km∙h-1 ), perceived exertion (pre = 11,53 ± 1,45 u.a; post = 11,23 ± 1,53 u.a) and FCLAn (pre = 166,64 ± 10,69 bpm; post = 174,50 ± 10,89 bpm) between conditions before and after training, indicating that three weeks of training are insufficient to generate positive changes in soccer players LAn

Effects of different resistance training frequencies on flexibility in older women

Objective The main purpose of the investigation reported here was to analyze the effect of resistance training (RT) performed at different weekly frequencies on flexibility in older women. Participants and methods Fifty-three older women (≥60 years old) were randomly assigned to perform RT either two (n=28; group “G2x”), or three (n=25; group “G3x”) times per week. The RT program comprised eight exercises in which the participants performed one set of 10–15 repetitions maximum for a period of 12 weeks. Anthropometric, body-composition, and flexibility measurements were made at baseline and post-study. The flexibility measurements were obtained by a fleximeter. Results A significant group-by-time interaction (P\u3c0.01) was observed for frontal hip flexion, in which G3x showed a higher increase than G2x (+12.8% and +3.0%, respectively). Both groups increased flexibility in cervical extension (G2x=+19.1%, G3x=+20.0%), right hip flexion (G2x=+14.6%, G3x=+15.9%), and left hip flexion (G2x=+25.7%, G3x=+19.2%), with no statistical difference between groups. No statistically significant differences were noted for the increase in skeletal muscle mass between training three versus two times a week (+7.4% vs +4.4%, respectively). Conclusion Twelve weeks of RT improves the flexibility of different joint movements in older women, and the higher frequency induces greater increases for frontal hip flexion

Characterizing 1-year development of cervical cord atrophy across different MS phenotypes: A voxel-wise, multicentre analysis

Background: Spatio-temporal evolution of cord atrophy in multiple sclerosis (MS) has not been investigated yet. Objective: To evaluate voxel-wise distribution and 1-year changes of cervical cord atrophy in a multicentre MS cohort. Methods: Baseline and 1-year 3D T1-weighted cervical cord scans and clinical evaluations of 54 healthy controls (HC) and 113 MS patients (14 clinically isolated syndromes (CIS), 77 relapsing-remitting (RR), 22 progressive (P)) were used to investigate voxel-wise cord volume loss in patients versus HC, 1-year volume changes and clinical correlations (SPM12). Results: MS patients exhibited baseline cord atrophy versus HC at anterior and posterior/lateral C1/C2 and C4–C6 (p < 0.05, corrected). While CIS patients showed baseline volume increase at C4 versus HC (p < 0.001, uncorrected), RRMS exhibited posterior/lateral C1/C2 atrophy versus CIS, and PMS showed widespread cord atrophy versus RRMS (p < 0.05, corrected). At 1 year, 13 patients had clinically worsened. Cord atrophy progressed in MS, driven by RRMS, at posterior/lateral C2 and C3–C6 (p < 0.05, corrected). CIS patients showed no volume changes, while PMS showed circumscribed atrophy progression. Baseline cord atrophy at posterior/lateral C1/C2 and C3–C6 correlated with concomitant and 1-year disability (r = −0.40/–0.62, p < 0.05, corrected). Conclusions: Voxel-wise analysis characterized spinal cord neurodegeneration over 1 year across MS phenotypes and helped to explain baseline and 1-year disability

Whole-metagenome-sequencing-based community profiles of Vitis vinifera L. cv. Corvina berries withered in two post-harvest conditions

Vitis vinifera L. cv. Corvina grape forms the basis for the production of unique wines, such as Amarone, whose distinctive sensory features are strongly linked to the post-harvest grape withering process. Indeed, this process increases sugar concentration and changes must characteristics. While microorganisms involved in must fermentation have been widely investigated, few data are available on the microbiota of withered grapes. Thus, in this paper, a whole metagenome sequencing (WMS) approach was used to analyse the microbial consortium associated with Corvina berries at the end of the withering process performed in two different conditions (“traditional withering,” TW or “accelerated withering,” AW), and to unveil whether changes of drying parameters could have an impact on microbial diversity. Samples of healthy undamaged berries were collected and washed, to recover microorganisms from the surface and avoid contamination with grapevine genetic material. Isolated DNA was sequenced and the data obtained were analyzed with several bioinformatics methods. The eukaryotic community was mainly composed by members of the phylum Ascomycota, including Eurotiomycetes, Sordariomycetes, and Dothideomycetes. Moreover, the distribution of the genera Aspergillus and Penicillium (class Eurotiomycetes) varied between the withered berry samples. Instead, Botryotinia, Saccharomyces, and other wine technologically useful microorganisms were relatively scarce in both samples. For prokaryotes, 25 phyla were identified, nine of which were common to both conditions. Environmental bacteria belonging to the class Gammaproteobacteria were dominant and, in particular, the TW sample was characterized by members of the family Pseudomonadaceae, while members of the family Enterobacteriaceae dominated the AW sample, in addition to Sphyngobacteria and Clostridia. Finally, the binning procedure discovered 15 putative genomes which dominated the microbial community of the two samples, and included representatives of genera Erwinia, Pantoea, Pseudomonas, Clostridium, Paenibacillus, and of orders Lactobacillales and Actinomycetales. These results provide insights into the microbial consortium of Corvina withered berries and reveal relevant variations attributable to post-harvest withering conditions, underling how WMS could open novel perspectives in the knowledge and management of the withering process of Corvina, with an impact on the winemaking of important Italian wines