Using regression and Multifactorial Analysis of Variance to assess the effect of ascorbic, citric, and malic acids on spores and activated spores of Alicyclobacillusacidoterrestris.

The type strain of the species Alicyclobacillus acidoterrestris (DSM 3922) and the strain CB1 (accession number: KP144333) were studied in this research to assess the effects of three weak acids (malic, citric, and ascorbic acids), pH (3 or 4), and spore status (spores, and activated spores). Acids were used to prepare 7 different blends, and the blends used to reduce the pH of Malt Extract broth to 3 and 4; then, media were inoculated with spores or activated spores, stored at 45 °C (optimal temperature for A. acidoterrestris growth), and analyzed immediately and after 2 and 7 days. Data were preliminary standardized as increase/reduction of microbial population, compared to the initial concentration, and modelled through two different statistical approaches (multifactorial ANOVA, and multiple regression). Finally, a binary code (0-no growth or reduction of viable count; 1-growth) was used to perform a multiple regression analysis on the growth probability of A. acidoterrestris. Generally, ascorbic acid was the most effective compound, but other acids (e.g., malic acid) could contribute to increase the inactivation ratio; concerning spore status, the highest sensitivity of activated spores suggests that acids probably act during the outgrowing phase. Finally, the two strains showed different trends at pH 3.0, being the type strain the most resistant one.

Wine Microbiology and Predictive Microbiology: A Short Overview on Application, and Perspectives.

Predictive microbiology (PM) is an essential element in food microbiology; its aims are the determination of the responses of a given microorganism combining mathematical models with experimental data under certain environmental conditions, and the simulation a priori of the growth/inactivation of a population based on the known traits of a food matrix. Today, a great variety of models exist to describe the behaviour of several pathogenic and spoilage microorganisms in foods. In winemaking, many mathematical models have been used for monitoring yeast growth in alcoholic fermentation as well as to predict the risk of contamination of grapes and grape products by mycotoxin producing fungi over the last years, but the potentialities of PM in wine microbiology are underestimated. Thus, the goals of this review are to show some applications and perspectives in the following fields: (1) kinetics of alcoholic and malolactic fermentation; (2) models and approaches for yeasts and bacteria growth/inactivation; (3) toxin production and removal.

Healthy and pro-inflammatory gut ecology plays a crucial role in the digestion and tolerance of a novel Gluten Friendly™ bread in celiac subjects: a randomized, double blind, placebo control in vivo study.

Gluten Friendly™ (GF) is a new gluten achieved through a physicochemical process applied to wheat kernels. The goal of this research was to assess the in vivo effects of Gluten Friendly™ bread on celiac gut mucosa and microbiota. In a double-blind placebo-controlled intervention study, 48 celiac disease (CD) patients were randomized into 3 groups to eat 100 g of bread daily, containing different doses (0; 3 g; 6 g) of GF for 12 weeks. The small-bowel morphology (VH/CrD), intraepithelial densities of CD3+, celiac serology, MUC2, CB1, gut permeability, proinflammatory cytokines, gluten in stools, symptoms, and gut microbial composition were assessed. All 48 CD subjects experienced no symptoms. K-means analysis evidenced celiac subjects clustering around unknown parameters independent of GF dosage: K1 35%; K2 30%; K3 35%. VH/CrD significantly decreased in K1 and K2. VH/CrD did not correlate with IEL increase in K2. 33-mer was not detected in 47% and 73% of patients in both K1 and K2, respectively. VH/CrD and IEL did not change significantly and strongly correlated with the absence of 33-mer in K3. Inflammation and VH/CrD decrease are strongly related with the presence of proinflammatory species at the baseline. A boost in probiotic, butyrate-producing genera, is strongly related with GF tolerance at the end of the trial. Our research suggests that a healthy and proinflammatory ecology could play a crucial role in the digestion and tolerance of the new gluten molecule in celiac subjects. However, GF can be completely digested by gut microbiota of CD subjects and shapes it toward gut homeostasis by boosting healthy butyrate-producing populations. The clinical trial registry number is NCT03137862 (https://clinicaltrials.gov).

Viability, Sublethal Injury, and Release of Cellular Components From Alicyclobacillus acidoterrestris Spores and Cells After the Application of Physical Treatments, Natural Extracts, or Their Components.

Alicyclobacillus acidoterrestris is a spoiling microorganism regarded as one of the most important causes of spoilage of fruit juices and acidic products. In this paper, four strains of A. acidoterrestris (type strain-DSM 3922; two wild strains isolated from soil-C8 and C24; wild strain isolated from a spoiled pear juice CB1) were treated through natural extracts/active compounds from essential oils (EOs), and physical treatments were used to assess their susceptibility and the presence of sublethal injury. The characterization of damage was also performed. The results suggest that it is possible to control A. acidoterrestris through alternative approaches, although the effect relied upon the age of spores. In addition to the mere antimicrobial effect, some treatments could cause a sublethal injury on spores. Lemon extract was the most effective treatment for both the antimicrobial effect and the sublethal injury, as evidenced by the release of proteins, and calcium dipicolinate [dipicolinic acid (DPA)] by fresh spores and only DPA (with an exception for C8) by old spores. A sublethal injury with protein release was also found for physical treatments [US (ultrasound) or heating]. For the first time, this paper reports on the existence of a sublethal injury for A. acidoterrestris, and this evidence could also be a challenge, because injured microorganisms could restore their metabolism, or an opportunity to design new preserving treatments.

Effect of Physical and Chemical Treatments on Viability, Sub-Lethal Injury, and Release of Cellular Components from Bacillus clausii and Bacillus coagulans Spores and Cells.

Bacterial spores are of concern to the food industry due to their ability to survive processing and their potential to subsequently germinate and grow in food. In this paper, two strains belonging to the genus Bacillus (B. clausii DSM 8716 and B. coagulans DSM 1) were studied under in vitro conditions after the application of essential oils, and physical treatments; cells and spores' susceptibility, the extent of sub-lethal injury and the release of cellular components as a function of treatment and targets (cells, spores, old or activated spores) were studied. The highest antimicrobial effect was found for cells treated through citrus extract, while both essential oils and physical treatments could cause a sub-lethal injury on the surviving cells and spores; in addition, the spores of B. coagulans released dipicolinic acid (DPA) and proteins. Sub-lethal injury should be considered when designing a food processing treatment, because injured microorganisms could either repair the damage or be inactivated with a different effect on microbial stability of foods.

Antifungal and Antibacterial Effect of Propolis: A Comparative Hit for Food-Borne Pseudomonas, Enterobacteriaceae and Fungi.

Propolis is a natural brownish resinous substance collected by honeybees (Apis mellifera), with a documented bioactivity against many microorganisms. In this study, the activity of propolis was investigated using some strains of Pseudomonas spp., Enterobacteriaceae, Lactobacillus plantarum, yeasts (Saccharomyces cerevisiae and Debaryomyces hansenii) and Fusarium oxysporum. Two approaches were used (a modified microdilution protocol and viable count), and the microorganisms were inoculated at two levels (low or high inoculum). The antimicrobial effect of propolis relies upon several factors, like the kind of microorganisms (for example S. cerevisiae was more resistant than D. hansenii, while Lactobacillus plantarum was never affected), the cell concentration (at high inoculum higher amounts of propolis were required for an antimicrobial action), and the mode of action (a delay of growth rather than a complete inhibition). The results of this paper point out, for the first time, the antimicrobial activity of propolis against some spoilers, with a focus on the possible effect; thus, they could be the background to designing an effective tool to prolong the shelf life of foods.

Nonthermal Technologies for Fruit and Vegetable Juices and Beverages: Overview and Advances.

In recent years, there has been a growing interest in the design of novel nonthermal processing systems that minimally modify sensory, nutritional, and functional properties of fruit and vegetable juices and beverages. The benefits of nonthermal treatments are strongly dependent on the food matrix. Thus, an understanding of the effects that these technologies exert on the properties of juices and beverages is important to design and optimize technological parameters to produce value-added products. This review covers research on nonthermal electrical treatments, high pressure processing, ultrasound, radiation processing, inert gas treatments, cold plasma, and membrane processing. Advances towards optimization of processing conditions, and combined technologies approaches have been also extensively reviewed. This information could be useful to: (1) manage processing systems and optimize resources; (2) preserve nutritional value and organoleptic properties, and (3) provide processing conditions for validation of these technologies at the industrial scale.

An In Vitro Fermentation Study on the Effects of Gluten FriendlyTM Bread on Microbiota and Short Chain Fatty Acids of Fecal Samples from Healthy and Celiac Subjects.

Recently, an innovative gluten detoxification method called Gluten FriendlyTM (GF) has been developed. It induces structural modifications, which abolish the antigenic capacity of gluten and reduce the in vitro immunogenicity of the most common epitopes involved in celiac disease, without compromising the nutritional and technological properties. This study investigated the in vitro effects of GF bread (GFB) on the fecal microbiota from healthy and celiac individuals by a three-stage continuous fermentative system, which simulates the colon (vessel 1, proximal colon; vessel 2, transverse colon; and vessel 3, distal colon), as well as on the production of short chain fatty acids (SCFA, acetate, propionate, butyrate). The system was fed with GFB and the changes in microbiota through fluorescence in situ hybridization and in SCFA content were assessed. GFB exerted beneficial modulations such as bifidogenic effects in each compartment of the model both with healthy- and celiac-derived samples, as well as growth in Clostridium clusters XIVa+b in celiac-derived samples. Furthermore, increased levels of acetic acid were found in vessel 1 inoculated with the fecal microbiota of healthy individuals, as well as acetic and propionic in vessel 1 and 2 with celiac-derived samples. In addition, the use of multivariate approaches showed that the supplementation of GFB could result in a different modulation of the fecal microbiota and SCFA, as a function of initial equilibrium.

Microbial Resources and Enological Significance: Opportunities and Benefits.

Among the innovative trends in the wine sector, the continuous exploration of enological properties associated with wine microbial resources represents a cornerstone driver of quality improvement. Since the advent of starter cultures technology, the attention has been focused on intraspecific biodiversity within the primary species responsible for alcoholic fermentation (Saccharomyces cerevisiae) and, subsequently, for the so-called 'malolactic fermentation' (Oenococcus oeni). However, in the last decade, a relevant number of studies proposed the enological exploitation of an increasing number of species (e.g., non-Saccharomyces yeasts) associated with spontaneous fermentation in wine. These new species/strains may provide technological solutions to specific problems and/or improve sensory characteristics, such as complexity, mouth-feel and flavors. This review offers an overview of the available information on the enological/protechnological significance of microbial resources associated with winemaking, summarizing the opportunities and the benefits associated with the enological exploitation of this microbial potential. We discuss proposed solutions to improve quality and safety of wines (e.g., alternative starter cultures, multistrains starter cultures) and future perspectives.

Encapsulation of Active Compounds in Fruit and Vegetable Juice Processing: Current State and Perspectives.

The production of value-added and/or functional juices has increased significantly in recent years, following an increased consumer demand to promote health and/or prevent disease through diet and nutrition. Micro and nano-encapsulation are promising technologies to protect and deliver sensitive compounds, allowing a controlled release in the target sites. This paper offers an overview of current applications, limits and challenges of encapsulation technologies in the production of fruit and vegetable juices, with a particular emphasis on products derived from different botanical sources.

Thermal Treatments for Fruit and Vegetable Juices and Beverages: A Literature Overview.

Fruit and vegetable juices and beverages are generally preserved by thermal processing, currently being the most cost-effective means ensuring microbial safety and enzyme deactivation. However, thermal treatments may induce several chemical and physical changes that impair the organoleptic properties and may reduce the content or bioavailability of some nutrients; in most cases, these effects are strongly dependent on the food matrix. Moreover, the efficacy of treatments can also be affected by the complexity of the product and microorganisms. This review covers researches on this topic, with a particular emphasis on products derived from different botanical sources. Technologies presented include conventional and alternative thermal treatments. Advances toward hurdle-based technology approaches have been also reviewed.

A Focus on Quality and Safety Traits of Saccharomyces cerevisiae Isolated from Uva di Troia Grape Variety.

The aim of this work was to study Saccharomyces cerevisiae strains isolated from vineyards of the autochthonous grape variety "Uva di Troia" located in different geographical areas of Apulian region (Southern Italy). Four hundred isolates were studied in relation to H2 S production, ß-glucosidase activity, and pigments adsorption from grape skin. Thus, 81 isolates were selected, identified through the amplification of the interdelta region, and grouped in 19 biotypes (from I to XIX). The enological performances were assessed to determine the content of residual sugars, ethanol, glycerol, and volatile acidity, after a microfermentation in Uva di Troia must for each isolate. The ability to remove ochratoxin A (OTA) was studied as an additional tool to select promising strains. A geographical-dependent technological variability was found for glycerol and volatile acidity, suggesting that the different indigenous yeasts can have a peculiar impact on the final characteristics of the corresponding wine ("Nero di Troia"). Only 2 biotypes (VI and XVII) were able to remove OTA throughout fermentation, with the highest reduction achieved by the biotype XVII (ca. 30%).

Ochratoxin A Removal by Yeasts after Exposure to Simulated Human Gastrointestinal Conditions.

Yeasts can remove ochratoxin A (OTA) in foods; however, we do not know if they can experience this ability in the gut. Thus, the aim of this paper was to study OTA binding by 3 wine strains of Saccharomyces cerevisiae (W13, W28, and the commercial isolate BM45) and S. cerevisiae var. boulardii ATCC MYA-796 (a probiotic yeast) after exposure to conditions simulating the passage through human gastrointestinal tract. Yeasts removed OTA by up to 44%, with a better ability found after gastrointestinal and sequential assays. However, the phenomenon was reversible, because ca. 18% to 28% of toxin was not stably bound. After the evaluation of the net amount of OTA bound by yeasts (that is, after toxin release), S. cerevisiae W13 and S. cerevisiae var. boulardii ATCC MYA-796 were selected as the most promising strains for further studies (for example, clinical trials) due to their ability to remove 21% of OTA.

Using physical approaches for the attenuation of lactic acid bacteria in an organic rice beverage.

A wild strain of Lactobacillus plantarum, isolated from an Italian sourdough, was inoculated in an organic rice drink; however, it caused a strong acidification. Thus, it was preliminary processed through homogenization (single or multiple passes) or sonication (US) and then inoculated in the beverage. The samples were stored at 4 °C and analyzed to assess pH, production of lactic acid, viable count and sensory scores. A US-2-step process (power, 80%) could control acidification; viability and sensory traits were never affected by sonication. This result was confirmed on two commercial probiotics (Lactobacillus casei LC01 and Bifidobacterium animalis subsp. lactis Bb12). In the 2nd step samples inoculated with attenuated strains were also stored under thermal abuse conditions (25 or 37 °C for 4 or 24 h, then at 4 °C) and the results showed that US could control acidification for a short thermal abuse. Finally, US-attenuated starter cultures were inoculated in the rice drink containing ß-glucans as healthy compounds; the targets did not cause any significant change of prebiotic.

Differential Adsorption of Ochratoxin A and Anthocyanins by Inactivated Yeasts and Yeast Cell Walls during Simulation of Wine Aging.

The adsorption of ochratoxin A (OTA) by yeasts is a promising approach for the decontamination of musts and wines, but some potential competitive or interactive phenomena between mycotoxin, yeast cells, and anthocyanins might modify the intensity of the phenomenon. The aim of this study was to examine OTA adsorption by two strains of Saccharomyces cerevisiae (the wild strain W13, and the commercial isolate BM45), previously inactivated by heat, and a yeast cell wall preparation. Experiments were conducted using Nero di Troia red wine contaminated with 2 µg/L OTA and supplemented with yeast biomass (20 g/L). The samples were analyzed periodically to assess mycotoxin concentration, chromatic characteristics, and total anthocyanins over 84 days of aging. Yeast cell walls revealed the highest OTA-adsorption in comparison to thermally-inactivated cells (50% vs. 43% toxin reduction), whilst no significant differences were found for the amount of adsorbed anthocyanins in OTA-contaminated and control wines. OTA and anthocyanins adsorption were not competitive phenomena. Unfortunately, the addition of yeast cells to wine could cause color loss; therefore, yeast selection should also focus on this trait to select the best strain.

Selection of autochthonous Saccharomyces cerevisiae strains as wine starters using a polyphasic approach and ochratoxin A removal.

Over the last few years, the selection of autochthonous strains of Saccharomyces cerevisiae as wine starters has been studied; however, researchers have not focused on the ability to remove ochratoxin A (OTA) as a possible trait to use in oenological characterization. In this article, a polyphasic approach, including yeast genotyping, evaluation of phenotypic traits, and fermentative performance in a model system (temperature, 25 and 30°C; sugar level, 200 and 250 g liter(-1)), was proposed as a suitable approach to select wine starters of S. cerevisiae from 30 autochthonous isolates from Uva di Troia cv., a red wine grape variety grown in the Apulian region (Southern Italy). The ability to remove OTA, a desirable trait to improve the safety of wine, was also assessed using enzyme-linked immunosorbent assay. The isolates, identified by PCR-restriction fragment length polymorphism analysis of the internal transcribed spacer region and DNA sequencing, were differentiated at strain level through the amplification of the interdelta region; 11 biotypes (I to XI) were identified and further studied. Four biotypes (II, III, V, VIII) were able to reduce OTA, with the rate of toxin removal from the medium (0.6 to 42.8%, wt/vol) dependent upon the strain and the temperature, and biotypes II and VIII were promising in terms of ethanol, glycerol, and volatile acidity production, as well as for their enzymatic and stress resistance characteristics. For the first time, the ability of S. cerevisiae to remove OTA during alcoholic fermentation was used as an additional trait in the yeast-selection program; the results could have application for evaluating the potential of autochthonous S. cerevisiae strains as starter cultures for the production of typical wines with improved quality and safety.

Decontamination of ochratoxin A by yeasts: possible approaches and factors leading to toxin removal in wine.

Biological decontamination of mycotoxins using microorganisms is one of the well-known strategies for the management of mycotoxins in foods and feeds. Yeasts are an efficient biosorbant, used in winemaking to reduce the concentration of harmful substances from the must which affect alcoholic fermentation (medium-chain fatty acids) or which affect wine quality in a negative way (ethyl phenols and sulphur products). In recent years, several studies have demonstrated the ability of yeasts to remove ochratoxin A (OTA) by live cells, cell walls and cell wall extracts, yeast lees. In spite of the physical and chemical methods applied to remove the toxin, the biological removal is considered a promising solution, since it is possible to attain the decontamination without using harmful chemicals and without losses in nutrient value or palatability of decontaminated food. In addition, adsorption is recognized as economically viable, technically feasible and socially acceptable. This paper intends to review the current achievements of OTA removal mediated by yeasts, the recent updates in the selection of strains acting at the same time as starters and as biological tools to remove OTA and the factors affecting the removal process.

Ochratoxin A released back into the medium by Saccharomyces cerevisiae as a function of the strain, washing medium and fermentative conditions.

BACKGROUND: This study was aimed at investigating the removal of ochratoxin A (OTA) by two wild strains of Saccharomyces cerevisiae (W20 and W30) in a semi-synthetic medium under two temperatures (25, 30 °C) and sugar levels (200, 250 g L(-1) ), as well as the stability of OTA-yeast complex by evaluating the amount of bound toxin released back after some washing treatments with phosphate-buffered saline (PBS) or model wine (MW). In addition, the main products of fermentation were studied. RESULTS: Both W20 and W30 strains reduced OTA with removal percentages of 5.41-49.58%, and this process was affected by temperature and sugar concentration. Concerning the stability of the OTA-yeast complex, the amount of bound toxin decreased by 20-99% after five passes of washing, with a strong strain dependence and an effect of temperature and sugar concentration only for the W30 isolate. In addition, the two strains showed interesting technological properties in terms of fermentation products in a semi-synthetic medium (high ethanol yield, volatile acidity as acetic acid < 1.2 g L(-1) ; glycerol production exceeding 5.2 g L(-1) ). CONCLUSIONS: Apart from the removal of OTA, release of the toxin is a variable process and relies upon the strain effect; a significance of the other factors of the design (sugar concentration, temperature) was found only for a single isolate. Thus evaluation of the stability of the complex yeasts/OTA should be an additional trait to select promising functional yeasts.

Ochratoxin A removal by Saccharomyces cerevisiae strains: effect of wine-related physicochemical factors.

BACKGROUND: This study investigated the effect of some physicochemical parameters on the removal of ochratoxin A (OTA) by yeasts. RESULTS: Two wild strains of Saccharomyces cerevisiae (W47 and Y28) were used to assess OTA removal under various conditions of temperature, pH, ethanol content and incubation time. All samples were analysed for OTA concentration by enzyme-linked immunosorbent assay (ELISA). In addition, yeast oenological traits were investigated: qualitative and technological traits were assessed on appropriate laboratory media, while the main products of microfermentation (sugars, ethanol, glycerol, acetic acid) were evaluated by Fourier transform infrared spectroscopy (FTIR). The results showed OTA reduction by 36-42% in cultures containing 100 g L⁻¹ ethanol incubated at pH 3.5 and 37 °C. CONCLUSION: OTA removal was affected by contact time, pH and ethanol content, as it was increased at low pH and by 100 g L⁻¹ ethanol. Moreover, the phenomenon was reversible, as OTA was lowest after 4 days, then it was partially released in the medium.