Adaptive changes in geranylgeranyl pyrophosphate synthase gene expression level under ethanol stress conditions in Oenococcus oeni.

Aims: The aim of this study was to investigate the effect of ethanol exposure on the expression level of geranylgeranyl pyrophosphate synthase gene involved in the metabolism of Oenococcus oeni to probe the mechanisms of ethanol tolerance correlated with adaptive changes. Methods and Results: The evaluation of ten potential internal control genes and the comparative study of their stability were performed to select the most stable internal controls for the normalization of expression data. The expression level analysis by qPCR and changes after exposure to ethanol stresses highlighted a significant increase in the presence of higher ethanol concentrations. Conclusions: The analysis of results suggest that O. oeni adjusts the expression of genes to adapt to stress conditions and the high expression level of ggpps would allow a flow of isoprenoid precursors towards the carotenoids and related pathways to stabilize bacterial cell membranes, improving the cell membrane disturbances and preventing cell death induced by ethanol. Significance and Impact of the Study: The involvement of ggpps gene in physiological changes of bacterial behaviour confirmed the exposure to stress requires the activation of defence mechanism to be more tolerant to adverse conditions. Improving the knowledge of stress tolerance and adaptation mechanisms of O. oeni is essential to enhance the efficiency of the malolactic starter in wine and to obtain the development of starters able to survive to direct inoculation with a large benefit for wine technology

Germline variants at SOHLH2 influence multiple myeloma risk

Funding Information: This work was supported by grants from the Knut and Alice Wallenberg Foundation (2012.0193 and 2017.0436), the Swedish Research Council (2017-02023), the Swedish Cancer Society (2017/265), Stiftelsen Borås Forsknings-och Utvecklingsfond mot Cancer, the Nordic Cancer Union (R217-A13329-18-S65), EU-MSCA-COFUND 754299 CanFaster, the Myeloma UK and Cancer Research UK (C1298/A8362), a Jacquelin Forbes-Nixon Fellowship, and Mr. Ralph Stockwell. We thank Ellinor Johnsson and Anna Collin for their assistance. We are indebted to the clinicians and patients who contributed samples. Open access funding provided by Lund University. Publisher Copyright: © 2021, The Author(s).Multiple myeloma (MM) is caused by the uncontrolled, clonal expansion of plasma cells. While there is epidemiological evidence for inherited susceptibility, the molecular basis remains incompletely understood. We report a genome-wide association study totalling 5,320 cases and 422,289 controls from four Nordic populations, and find a novel MM risk variant at SOHLH2 at 13q13.3 (risk allele frequency = 3.5%; odds ratio = 1.38; P = 2.2 × 10−14). This gene encodes a transcription factor involved in gametogenesis that is normally only weakly expressed in plasma cells. The association is represented by 14 variants in linkage disequilibrium. Among these, rs75712673 maps to a genomic region with open chromatin in plasma cells, and upregulates SOHLH2 in this cell type. Moreover, rs75712673 influences transcriptional activity in luciferase assays, and shows a chromatin looping interaction with the SOHLH2 promoter. Our work provides novel insight into MM susceptibility.Peer reviewe

Functional dissection of inherited non-coding variation influencing multiple myeloma risk

Funding Information: This work was supported by grants from the Knut and Alice Wallenberg Foundation (2012.0193 and 2017.0436), the Swedish Research Council (2017-02023 and 2018-00424), the Swedish Cancer Society (2017/265), the Nordic Cancer Union (R217-A13329-18-S65), Arne and Inga-Britt Lundberg’s Stiftelse (2017-0055), European Research Council (EU-MSCA-COFUND 754299 CanFaster), Myeloma UK and Cancer Research UK (C1298/A8362), The National Institute of Health (R01 DK103794 and R01HL146500), the New York Stem Cell Foundation, a gift from the Lodish Family to Boston Children’s Hospital, and Mr. Ralph Stockwell. We thank Ellinor Johnsson for her assistance between 2011 and 2020. We are indebted to the patients who participated in the study. Publisher Copyright: © 2022, The Author(s).Thousands of non-coding variants have been associated with increased risk of human diseases, yet the causal variants and their mechanisms-of-action remain obscure. In an integrative study combining massively parallel reporter assays (MPRA), expression analyses (eQTL, meQTL, PCHiC) and chromatin accessibility analyses in primary cells (caQTL), we investigate 1,039 variants associated with multiple myeloma (MM). We demonstrate that MM susceptibility is mediated by gene-regulatory changes in plasma cells and B-cells, and identify putative causal variants at six risk loci (SMARCD3, WAC, ELL2, CDCA7L, CEP120, and PREX1). Notably, three of these variants co-localize with significant plasma cell caQTLs, signaling the presence of causal activity at these precise genomic positions in an endogenous chromosomal context in vivo. Our results provide a systematic functional dissection of risk loci for a hematologic malignancy.Peer reviewe

Quantitative expression analysis of geranylgeranyl pyrophophate synthase gene in Oenococcus oeni under different ethanol stresses.

Background Oenococcus (O.) oeni has long been reported as the agent most commonly associated with the MLF and the LAB species most resistant to the presence of ethanol in wine. Objectives The effect of ethanol stress on the expression level of geranylgeranyl pyrophophate synthase gene (GGPPs) was investigated by real-time RT-qPCR to obtain a clearer picture of its function and role in the metabolism of O. oeni. Methods O. oeni cells treated with different ethanol concentrations (7%, 12%, 13%, 15%) and cells grown in absence of ethanol (control) were used in this study. The identification and evaluation of a panel of ten reference genes for real-time PCR normalization was performed. The ten control genes were ranked according to their stability of gene expression measure (M) and the coefficient of variation (V) using geNorm provided by qbase_plus software (Biogazelle). Once stable internal control genes were identified, the expression level of the target GGPPs gene was analyzed by using the geometric mean of copy numbers as normalization factor. Conclusions The quantitative expression gene analysis showed that various changes in the transcription level of the GGPPs gene appeared in response to different ethanol concentrations. Particularly, a significant increase of the expression level in the sample stressed with ethanol 12%, 13% and 15% occurred, while there was no transcriptional increase in the presence of 7% ethanol if compared to the control

Evaluation of Aesculus hippocastanum tincture mother for antioxidant and antimicrobial activities and its effects on protein expression of different bacterial strains

The beneficial health effects of extracts from many types of plants have been known for centuries1 and the search for new natural extracts (such as tincture mother TM), to be used in the food and cosmetics industry, is very important at present2. The aim of this study was to investigate the antioxidant and the antimicrobial activities of Aesculus hippocastanum TM, a fresh plant juice extracted by maceration and solubilised in 65% hydroalcoholic solution. To evaluate the antioxidant, radical scavenging and antiperoxidative activities of extract, FRAP (Ferric Reducing Antioxidant Power), ABTS radical and β-carotene bleaching (BCB) assays were performed. Results indicated the TM possesses elevated activity for radical scavenging and a potent antioxidant ability. The antimicrobial activity and the minimal inhibitory concentration (MIC) were evaluated against selected bacterial strains of significant importance for human health and food production, by using the agar well diffusion assay. A total of thirty-two gram-negative and gram-positive bacteria were employed as screening microorganisms to determine the antimicrobial effect, the action spectrum and the antimicrobial effectiveness of each extract. Results showed that TM provided an high antimicrobial effect against a wide range of bacteria with a MIC <120 μg/ml for 67% of strains. The inhibitory activity showed a moderate efficacy on the growth of 70% of strains in presence of different extract concentrations, while 15% of strains (Hafnia alvei, Enterococcus faecium and Enterococcus gallinarum) presented a count reduction of 2 log cycles in presence of TM at 40 µg/ml concentration. Moreover, the TM effect on protein expression of strains in presence of different MIC was evaluated. Whole-cell protein patterns were analyzed by SDS-PAGE in strains grown in presence/absence of TM and interesting results were obtained. The presence of TM at different concentrations represented a stress condition for strains causing changes in protein profiles. Results of this study underlined important abilities of Aesculus hippocastanum that could be an interesting field for applications within pharmaceutical and food supplement industries. [1]. Russo D, Bonomo MG, Salzano G, Martelli G, Milella L. Nutraceutical properties of Citrus clementina juices. Pharmacol OnLine. 2012; 1:84-93. [2]. Kaushik P, Goyal P. Evaluation of various crude extracts of Zingiber officinale rhizome for potential antibacterial activity: a study in vitro. Adv Microbiol. 2011; 1:7-12

Assessment of the genetic polymorphism and physiological characterization of indigenous Oenococcus oeni strains isolated from Aglianico del Vulture red wine

The aim of this study was a reliable intra-species discrimination and strain biodiversity in Oenococcus oeni populations of two different Aglianico wineries by molecular, biochemical, and physiological characterization. Pulsed field gel electrophoresis (PFGE) analysis revealed a high polymorphism related to the origin (winery) of strains, while differential display PCR (DD-PCR) allowed a further discrimination of strains from the same winery. Moreover, the heterogeneity of these natural populations was investigated by capillary electrophoresis and enzymatic assays. A variability related to a different surface charge distribution was observed among strains, linked to their origin. Malolactic activity study evidenced strain-specific differences in malic acid degradation, and then, only the presence of L(-)-malic acid in the medium induced the mle gene. This study provided evidences on the importance of intra-species biodiversity ofmalolactic bacterial populations in wine ecosystems, as eachwine possess peculiar winemaking conditions and physical–chemical properties which make specific the bacterial survival and growth. This study highlighted a great biodiversity among O. oeni strains that can be also winery specific. Such biodiversity within a certain winery and winemaking area is important for selecting malolactic starters, and strain-specific trait identification is especially important to match individual strains to specific industrial process

CAPILLARY ELECTROPHORESIS ANALYSIS OF STRESSED Oenococcus oeni

The aim of this work was to study the electrophoretic behaviour of Oenococcus oeni (O. oeni) under the effect of ethanol stress. The bacterial outer surface of O. oeni is rich of ionizable groups so the changes in the charging rates in the optimal and stressed conditions could be evaluated by capillary electrophoresis. As a first attempt, it was necessary to optimize the electrophoretic conditions for the identification and efficient separation of this microorganism by capillary electrophoresis. After this preliminary study, the electropherograms show significant differences between cells stressed by ethanol and cells growth in optimal condition. Interestingly, it was observed a substantial difference in electrophoretic profile among different O. oeni strains. The experimental results confirmed the power of capillary electrophoresis for microbial analysis (characterization and separation of microorganisms) since permitting rapid, easy and highly sensitive microbial analysis at low costs for several biological samples

Flow cytometry and capillary electrophoresis analyses in ethanol-stressed Oenococcus oeni strains and changes assessment of membrane fatty acid composition

Aims: This study aimed to investigate the dynamics and physiological heterogeneity of Oenococcus oeni under different conditions, cell membrane fluidity and permeability variations, and assessment of changes in cell surface charging rates. Methods and Results: Flow cytometry, membrane fatty acid analysis and capillary electrophoresis were performed to study ethanol-induced variations. Different physiological states were assessed, revealing cell subpopulations able to adapt and withstand to environmental stress, in order to recover their functionality. Moreover, total results demonstrated changes in cell surface and membrane fatty acid redistribution with a saturation degree and an unsaturated/saturated fatty acid ratio fairly steady in control and in different ethanol stresses. Conclusions: This study revealed a great variability among O. oeni strains and the importance to investigate the mechanisms by a multiparametric approach based on the structural and physiological bacterial adjustments in different stresses tolerance. Significance and Impact of the Study: Intermediate physiological state assessment in O. oeni with recovery possibility could be an important criterion for potential starter culture application. The flow cytometry application with changes in monitoring membrane fatty acid composition and in surface charging rates allowed the characterization of sorted subpopulations that may contribute to further understanding of diversity and heterogeneity in physiology of bacterial populations