Editorial: Microbiota of Grapes: Positive and Negative Role on Wine Quality

During spontaneous food/beverage fermentations, the microbiota associated with the raw material has a considerable importance: this microbial consortium evolves in reason of the nutrient content and of the physical, chemical, and biological determinants present in the food matrix, shaping fermentation dynamics with significant impacts on the ‘qualities’ of final productions. The selection from the indigenous micro-biodiversity of ‘virtuous’ ecotypes that coupled pro-technological and biotechnological aptitudes provide the basis for the formulation of ‘tailored’ starter cultures. In the fermenting food and beverage arena, the wine sector is generally characterized by the generation of a high added value. Together with a pronounced seasonality, this feature strongly contributes to the selection of a large group of starter cultures. In the last years, several studies contributed to describe the complexity of grapevine-associated microbiota using both culture-dependent and culture-independent approaches. The grape-associated microbial communities continuously change during the wine-making process, with different dominances that correspond to the main biotechnological steps that take place in wine. In order to simplify, following a time trend, four major dominances can be mainly considered: non-Saccharomyces, Saccharomyces, lactic acid bacteria (LAB), and spoilage microbes. The first two dominances come in succession during the alcoholic fermentation: the impact of Saccharomyces (that are responsible of key enological step of ethanol production) can be complemented/integrated by the contributions of compatible non-Saccharomyces strains. Lactic acid bacteria constitute the malolactic consortium responsible of malolactic fermentation, a microbial bioconversion often desired in wine (especially in red wine production). Finally, the fourth dominance, the undesired microbiota, represents a panel of microorganisms that, coupling spoilage potential to the resistance to the harsh conditions typical of wine environment, can cause important economic losses. In each of these four dominances a complex microbial biodiversity has been described. The studies on the enological significance of the micro-biodiversity connected with each of the four dominances highlighted the presence of a dichotomy: in each consortia there are species/strains that, in reason of their metabolisms, are able to improve wine ‘qualities’ (resource of interest in starter cultures design), and species/strains that with their metabolism are responsible of depreciation of wine. Articles describing new oenological impacts of yeasts and bacteria belonging to the four main categories above mentioned (non-Saccharomyces, Saccharomycetes, lactic acid bacteria, and spoilage microbes) are welcome. Moreover, in this Research Topic, we encourage mini-review submissions on topics of immediate interest in wine microbiology that link microbial biodiversity with positive/negative effects in wine

Potential of Lactobacillus casei , Culture Permeate, and Lacti Acid To Control Microorganisms in Ready-To-Use Vegetables.

The effects of various treatments (i.e., the addition of a strain of Lactobacillus that produces antimicrobial agents, Lactobacillus casei IMPC LC34, its sterile permeate, and 0.5 or 1% lactic acid) on the growth of microorganisms associated with ready-to-use mixed salad vegetables were compared during refrigerated (8°C) storage. The addition of 3% culture permeate to mixed salads reduced the total mesophilic bacteria counts from 6 to 1 log CFU/g, and suppressed coliforms, enterococci, and Aeromonas hydrophila after 6 days of storage at 8°C. A similar effect was shown when the L. casei culture was inoculated in the vegetables. One percent lactic acid had a bacteriostatic effect on the bacterial groups examined, except for total and fecal coliforms, which were reduced by about 2 and 1 log unit, respectively, while 0.5% lactic acid did not affect the indigenous microflora of the vegetables. The potential of these new hurdles to prevent the growth of spoilage and pathogenic bacteria in ready-to-use salad vegetables is suggested

Editorial: Biogenic amines in foods

Various issues related to the qualitative and quantitative presence of biogenic amines in cheese, dry sausages, wine, and fish were addressed. The possible scavenging of these compounds by technological processes and amineoxidase activity of some microorganisms is also reported

Exploring Antibiotic Resistance Diversity in Leuconostoc spp. by a Genome-Based Approach: Focus on the {lsaA} Gene

Leuconostoc spp. are environmental microorganisms commonly associated with fermented foods. Absence of antibiotic resistance (AR) in bacteria is a critical issue for global food safety. Herein, we updated the occurrence of AR genes in the Leuconostoc genus through in silico analyses of the genomes of 17 type strains. A total of 131 putative AR traits associated with the main clinically relevant antibiotics were detected. We found, for the first time, the lsaA gene in L. fallax ATCC 700006T and L. pseudomesenteroides NCDO 768T. Their amino acid sequences displayed high similarities (59.07% and 52.21%) with LsaA of Enterococcusfaecalis V583, involved in clindamycin (CLI) and quinupristin-dalfopristin (QUD) resistance. This trait has different distribution patterns in Leuconostoc nontype strains-i.e., L. pseudomesenteroides, L. lactis and L. falkenbergense isolates from fermented vegetables, cheeses, and starters. To better explore the role of lsaA, MIC for CLI and QUD were assessed in ATCC 700006T and NCDO 768T; both strains were resistant towards CLI, potentially linking lsaA to their resistant phenotype. Contrarily, NCDO 768T was sensitive towards QUD; however, expression of lsaA increased in presence of this antibiotic, indicating an active involvement of this trait and thus suggesting a revision of the QUD thresholds for this species

Suitability of the Nisin Z-producer Lactococcus lactis subsp. lactis CBM 21 to be Used as an Adjunct Culture for Squacquerone Cheese Production

This research investigated the technological and safety effects of the nisin Z producer Lactococcus lactis subsp. lactis CBM 21, tested as an adjunct culture for the making of Squacquerone cheese in a pilot-scale plant. The biocontrol agent remained at a high level throughout the cheese refrigerated storage, without having a negative influence on the viability of the conventional Streptococcus thermophilus starter. The inclusion of CBM 21 in Squacquerone cheesemaking proved to be more effective compared to the traditional one, to reduce total coliforms and Pseudomonas spp. Moreover, the novel/innovative adjunct culture tested did not negatively modify the proteolytic patterns of Squacquerone cheese, but it gave rise to products with specific volatile and texture profiles. The cheese produced with CBM 21 was more appreciated by the panelists with respect to the traditional one

A survey of Saccharomyces populations associated with wine fermentations from the Apulia region (South Italy)

The aim of this paper was to investigate the genetic and phenotypic characteristics of yeasts isolated from samples of grape musts collected from four different areas of Apulia region. The 68 yeast isolates were identified asSaccharomyces cerevisiae by PCR-RFLP of 5.8S-ITS region of the rRNA gene. Individual isolates were differentiated by RAPD-PCR and AFLP. The following oenological traits were studied: fermentation power, resistance to cycloheximide, alcohol and SO2, formation of SO2 and H2S, β-glucosidase activity, and production of biogenic amines and secondary compounds. Many phenotypes were common to several yeasts isolated from the four different areas, such as high SO2 resistance and fermentation power. In addition, someS. cerevisiae isolates showed a β-glucosidase activity and others had a high resistance to cycloheximide. All the strains formed biogenic amines. Solid Phase Microextraction was used to determine secondary compounds produced in wine by the single yeast cultures

Transcriptional and Metabolic Response of Wine-Related Lactiplantibacillus plantarum to Different Conditions of Aeration and Nitrogen Availability

Lactic acid bacteria (LAB) perform the process of malolactic fermentation (MLF) in wine. Availability of oxygen and nitrogen nutrients could influence LAB growth, malolactic activity, and other metabolic pathways, impacting the subsequent wine quality. The impact of these two factors has received limited investigation within LAB, especially on a transcriptome level. The aim of this study was to evaluate metabolic changes in the strain Lactiplantibacillus plantarum IWBT B063, growing in synthetic grape juice medium (GJM) under different oxygen exposure conditions, and with low availability of nitrogen-based nutrients. Next-generation sequencing was used to analyze expression across the transcriptome (RNA-seq), in combination with conventional microbiological and chemical analysis. L. plantarum consumed the malic acid present in all the conditions evaluated, with a slight delay and impaired growth for nitrogen limitation and for anaerobiosis. Comparison of L. plantarum transcriptome during growth in GJM with and without O-2 revealed differential expression of 148 functionally annotated genes, which were mostly involved in carbohydrate metabolism, genetic information processing, and signaling and cellular processes. In particular, genes with a protective role against oxidative stress and genes related to amino acid metabolism were differentially expressed. This study confirms the suitability of L. plantarum IWBT B063 to carry out MLF in different environmental conditions due to its potential adaption to the stress conditions tested and provides a better understanding of the genetic background of an industrially relevant strain

Unravelling the Impact of Grape Washing, {SO}2, and Multi-Starter Inoculation in Lab-Scale Vinification Trials of Withered Black Grapes

Wine quality is strongly affected by chemical composition and microbial population of grape must, which, in turn, are influenced by several post-harvest treatments, including grape withering. Different strategies have been suggested to manage the fermenting must microbiota, as it plays a central role in the outcomes of both spontaneous and guided fermentations. This study aimed at evaluating the impact of grape washing, SO2 addition, and selected starter culture inoculation on population dynamics, fermentation kinetics, and main oenological parameters in lab-scale trials, focusing on withered grapes usually used for Amarone production. Although grape washing treatment was effective in removing heavy metals and undesirable microorganisms from grape berry surface, inoculation of multi-starter cultures impacted more fermentation rates. Further, both grape washing and starter inoculation procedures had a remarkable impact on wine chemical characteristics, while 30 mg/L SO2 addition did not significantly affect the fermentation process. In summary, the best strategy in terms of limiting off-flavors and potentially reducing the need for SO2 addition in wine from withered grapes was the use of yeast starters, particularly mixed cultures composed by selected strains of Metschnikowia spp. and Saccharomyces cerevisiae. Application of a washing step before winemaking showed a potential to improve organoleptic characteristics of win

Culture-independent analysis of diversity in microbial communities: pros and cons of the partial sequence-based profiling

Among the culture-independent techniques developed so far, DGGE/TGGE analysis of amplified 16S rRNA fragments is currently used to a great extent for the characterization of microbial consortia, due to its fast response and highly descriptive power. This approach relies on the separations of partial 16S rRNA gene amplicons targeting two specific hypervariable regions, namely V3 and V6-V8. Separation is followed by sequencing and identification of different bands based on BLAST comparisons. Some case studies are presented here concerning the identification of bacterial species and genera - belonging to different phyla such as Actinobacteria (Bifidobacterium, Rhodococcus), Firmicutes (Bacillus, Enterococcus, and Lactobacillus), Alphaproteobacteria (Sinorhizobium), Betaproteobacteria (Burkholderia), and Gammaproteobacteria (Pseudomonas) \u2013 all relevant to environmental and industrial biotechnology

Molecular Identification and Quantification of Tetracycline and Erythromycin Resistance Genes in Spanish and Italian Retail Cheeses

Large antibiotic resistance gene pools in the microbiota of foods may ultimately pose a risk for human health. This study reports the identification and quantification of tetracycline- and erythromycin-resistant populations, resistance genes, and gene diversity in traditional Spanish and Italian cheeses, via culturing, conventional PCR, real-time quantitative PCR (qPCR), and denaturing gradient gel electrophoresis (DGGE). The numbers of resistant bacteria varied widely among the antibiotics and the different cheese varieties; in some cheeses, all the bacterial populations seemed to be resistant. Up to eight antibiotic resistance genes were sought by gene-specific PCR, six with respect to tetracycline, that is, tet(K), tet(L), tet(M), tet(O), tet(S), and tet(W), and two with respect to erythromycin, that is, erm(B) and erm(F). The most common resistance genes in the analysed cheeses were tet(S), tet(W), tet(M), and erm(B). The copy numbers of these genes, as quantified by qPCR, ranged widely between cheeses (from 4.94 to 10.18 log10/g). DGGE analysis revealed distinct banding profiles and two polymorphic nucleotide positions for tet(W)-carrying cheeses, though the similarity of the sequences suggests this tet(W) to have a monophyletic origin. Traditional cheeses would therefore appear to act as reservoirs for large numbers of many types of antibiotic resistance determinants.The study was partially supported by a Spain-Italy bilateral collaboration program (Ref. IT2009-0080 and IT105MD12L). Financial support was further provided by projects from CICYT (Ref. AGL2011-24300-ALI) and INIA (Ref. RM2011-00005-00-00). A. B. Flórez and S. Delgado were supported by research contracts under Juan de la Cierva Program (Ref. JCI-2010-07457 and JCI-2008-02391, resp.). A. Alegría was awarded a scholarship of the Severo Ochoa program from FICYT (Ref. BP08-053).Peer Reviewe