Currently, there is very little information available regarding the microbiome associated with the wine production chain. Here, we used an amplicon sequencing approach based on high-throughput sequencing (HTS) to obtain a comprehensive assessment of the bacterial community associated with the production of three Apulian red wines, from grape to final product. The relationships among grape variety, the microbial community, and fermentation was investigated. Moreover, the winery microbiota was evaluated compared to the autochthonous species in vineyards that persist until the end of the winemaking process. The analysis highlighted the remarkable dynamics within the microbial communities during fermentation. A common microbial core shared among the examined wine varieties was observed, and the unique taxonomic signature of each wine appellation was revealed. New species belonging to the genus Halomonas were also reported. This study demonstrates the potential of this metagenomic approach, supported by optimized protocols, for identifying the biodiversity of the wine supply chain. The developed experimental pipeline offers new prospects for other research fields in which a comprehensive view of microbial community complexity and dynamics is desirable.

Fumonisin B1 (FB1) is among the most common mycotoxins found in maize kernels and maize productsworldwide. The microbiological process of detoxification and transformation of toxic organic pollutants is a promisingmethod for foodstuffs decontamination. Some basidiomycetes, such as the Pleurotus eryngii species complex,include several important commercial edible varieties that can detoxify polycyclic organic compounds and a rangeof wastes and pollutants. We investigated the potential role of P. eryngii, one of the most consumed mushrooms, inthe decontamination of FB1 in maize. In addition, selected antioxidant enzymes, (soluble peroxidase (POD), catalase(CAT) and ascorbate peroxidase), primarily involved in control of cell hydrogen peroxide levels, and lignindegradation, were analyzed, to evaluate their contributions to the molecular mechanisms of FB1 by P. eryngii. FB1decontamination by P. eryngii and involvement of CAT and POD enzymes in the control of toxic decontaminationlevels of H2O2 were demonstrated. A consistent reduction of FB1 was observed at different incubation times. Theaverage decrease levels of FB1, with respect to the control cultures, ranged from 45 to 61% (RSD < 15%). This studyis a possible eco-friendly approach to reducing this mycotoxin in the feed supply chains.

At the three-leaf stage, sterile seedlings of resistant (CO433) and susceptible (CO389) maize lines was spiked with fumonisin B1 (FB1) (dissolved in PBS, pH =7.4, at the final concentration of 1mg/mL) in the part of the stem between the collar and the insertion of the first leaf. At various times, the FB1 content was determined by the use of HPLC/FLD previously derivatized with an o-phthaldeyde (OPA). The stem was ground with liquid nitrogen and extracted with a solution methanol/water (70:30, v/v). Recovery was 106% (RSD < 8%). The concentration of FB1 after 3 hours (88%, RSD 12%) and 48 hours (92%, RSD 7%) after spiking showed that no translocation in seedling maize occurred. To evaluate if the defence systems were alerted, the leaves were used to monitor the ascorbate-glutathione cycle, phenolics and enzymes protective from oxidative stress (catalase, superoxide dismutase and cytosolic peroxidases) at 3 and 48 hours post-spiking. The study was also extended to the analysis of total antioxidants, hydrogen peroxide and malondialdehyde contents to evaluate the oxidation level after FB1 treatment. Defense response promptly appeared activated in leaves of resistant line; particularly, after 3h, ascorbate, ascorbate peroxidase, SOD were augmented, underlining a higher fitness in the counteracting the phytotoxic action of FB1. In contrast, in the susceptible line, catalase, phenolics and ascorbate increased at longer time, conferring a lower readiness to the FB1 treatment. Same trend in total antioxidants, cytosolic peroxidases and other components analyzed was observed in both CO433 and CO389 after spiking. FB1, although did not cause seedlings suffering, induced metabolic perturbations. These data are useful for further investigation on molecular mechanisms that are to the basis of the FB1 and others mycotoxins contamination in maize, in order to improve resistance to fungal pathogens.

Contamination of vineyards from black Aspergilli is a well-known condition that cause the accumulation of ochratoxin A (OTA) in grapes and derived products. This contamination is strongly related to climatic conditions, geographical regions (South Mediterranean climate is highly conducive), grape varieties, damage by insects, although, great variations may occur from one year to another. Among the black Aspergilli commonly found in infected grapes, Aspergillus carbonarius is considered the main responsible of OTA contamination, with A. niger at secondary extent. To minimise the black Aspergilli infection and limit OTA concentrations in grapes, several strategies are commonly adopted, including the implementations of good agricultural practices and the use of pesticides and fungicides. These strategies are essential to manage the problem, but since they are insufficient when extremely favourable condition occurs in the vineyard, new strategies, aimed to reduce OTA risk in vineyards, are necessary. In this respect, implementation of electrolysed oxidising water (EOW) in agriculture has arising during the last decade as an interesting alternative to replace or limit the use of chemicals. The efficacy of EOW was also demonstrated in post-harvest for reduction of gray mold and brown rot on surfaces of peaches and grapes. In this study, we screened for the first time the efficacy of EOW generated by EVA System® 100 apparatus (Industrie De Nora S.p.A., Milan, Italy) at different concentrations of free chlorine (ranging from 0.0125 to 0.4 g/L) on conidial germination and growth of A. carbonarius and A. niger. A good fungicidal activity was achieved after 2-10 minutes treatment with EOW containing 0.4-0.2 g/L of free chlorine, although A. carbonarius conidia were more resistant to EOW than A. niger conidia. Then EOW at 0.4 g/L free chlorine was tested on detached berries of Primitivo and Aglianico wine grape varieties that were singularly infected with A. niger and A. carbonarius. Treatments with Switch (cyprodinil and fludioxonil) at 0.8 g/L, a fungicide regularly used in vineyard against black Aspergilli, were used as positive controls. Percentage of infected berries, McKinney index, and OTA concentrations were used to evaluate the efficacy of the EOW treatment. On Aglianico grape berries EOW and Switch produced an almost complete reduction on percentage of infection, McKinney index and OTA concentration compared to the control. On Primitivo grape berries EOW treatment reduced more than half the percentage of infections and McKinney index for both A. carbonarius and A. niger, although Switch showed a better performance. A significant reduction of OTA concentration was observed for EOW and Switch treatments. These results evidence for the first time that EOW is effective to reduce black Aspergilli inoculum and OTA contamination on grape berries.

Fumonisins are mycotoxins with cancerpromotingactivity and are associated with a number ofanimal and human diseases. The potential risk of contaminationby fumonisin B2 (FB2), although at low levels, hasbeen demonstrated in must and wine. Black aspergilli ingeneral and Aspergillus niger in particular are considered tobe the major responsible agents of FB2 contamination ingrape and its by-products. Contamination by FB2 thereforeis yet another safety concern of grape and wine producers,as ochratoxin A, produced mainly by A. carbonarius, mayprove to be a major mycotoxicological problem in thegrape-wine chain.

This study reports the fungal and bacterial metabolites associated with natural contamination of 38 composite samples of locally processed rice from five Agro-ecological Zones of Nigeria (AEZs). The samples were evaluated for the presence of microbial metabolites with the Liquid Tandem Mass Spectrometry (LC-MS/MS) technique. Among the identified metabolites, 63 fungal and 5 bacterial metabolites were measured at varying concentrations and occurrence levels. Fusarium toxins had the highest incidence of 79% but occurred in low amounts with fumonisin B1 (FB1) having the highest percentage incidence of 39.5% and a mean of 18.52 µg/kg. Among the Aspergillus toxins, aflatoxins (AFs) occurred in 36.9% of the rice samples with aflatoxin B1 (AFB1) having the highest occurrence level of 18.4% and a mean value of 5 µg/kg. About 12 metabolites had incidence levels >50%, including beauvericin (BEA) and tryptophol which had occurrence levels of 100%. Among the emerging toxins under evaluation by international organizations such as the European Food Safety Authority (EFSA) and the Food and Agriculture Organization of the United Nations (FAO), citrinin, sterigmatocystin (STER) and beauvericin were detected with maximum values of 207, 125 and 131 ?g/kg respectively. This paper also reports the first documented evidence of the contamination of Nigerian rice by bacterial and Alternaria metabolites, nivalenol, kojic acid, STER, moniliformin, fusaric acid, fumonisin B3, citrinin, 3-nitropropionic acid, andrastin A, cytochalasins, emodin and physicon.

This study reports the distribution of fungal isolates and fungal metabolites that naturallycontaminate locally processed rice from five agro-ecological zones of Nigeria. The fungal specieswere isolated by the dilution plate technique and identified by appropriate diagnostics, whilemetabolites were determined by a liquid chromatographic tandem mass spectrometric method.Aspergillus and Penicillium species were the predominant isolates found in the rice samples whileFusarium spp. were not isolated. The mean fungal count differed significantly (p < 0.05) across thezones and ranged from 9.98 × 102 cfu g-1 in the Southern Guinea Savannah to 96.97 × 102 cfu g-1in the Derived Savannah. For 16 fungal metabolites, selected from 63 positively identified fungalmetabolites based on their concentration and spread across the zones, an occurrence map wasconstructed. The Northern Guinea Savannah recorded the highest contamination of fungalmetabolites while the Sudan Savannah zone recorded the least.

Powdery mildew (PM), caused by the fungus Erysiphe necator, is one of the most widespread fungal disease of grape and maycause extensive openings on the berry surface during the infection. We evaluated the effect of damage caused by PM in grape berrieson the growth of and mycotoxin production by Aspergillus and on the oxidative stress in infected berries. Berries of Vitis vinifera L.cv. Negroamaro with sound skin (SS) and those naturally infected by PM were surface sterilized and inoculated with eitherfumonisin B2 (FB2)-producing strains of Aspergillus niger or ochratoxin A (OTA)-producing strains of Aspergillus carbonariusand incubated at 20 and 30uC. The PM berries were significantly more susceptible to both Aspergillus colonization (5 to 15 timesmore susceptible) and OTA and FB2 contamination (2 to 9 times more susceptible) than were SS berries. The highest toxinconcentration was detected in inoculated PM berries both for OTA (9 ng/g) at 20uC and for FB2 (687 ng/g) at 30uC. In inoculated SSand PM berries, although malondialdehyde and hydrogen peroxide concentrations did not increase, the two black Aspergillusspecies caused a significant decrease in ascorbate content, thus inducing a pro-oxidant effect. These results indicate that grape berriesaffected by PM are more susceptible to black Aspergillus growth and to production and/or accumulation of FB2 and OTA. Thus,preventive control of E. necator on grape berries could reduce the mycotoxicological risk from black Aspergillus infection.

In this study, the effects of pure olive phenolic compounds and olive mill wastewater (OMWW) (after membrane filtration treatments) on Aspergillus flavus growth and aflatoxin B1 (AFB1) production, were investigated. Five OMWWs coming from Greek (Lianolia, Koroneiki and Asprolia) and Italian (Cellina di Nardò and Coratina) olive oil cultivars, opportunely filtered using a membrane system, were added at two concentrations (5 and 15%) to growth medium, in order to evaluate their effect on A. flavus growth and AFB1 production. The OMWW fractions treatment, after 6 days of incubation, did not inhibit the fungal growth rate, but at 15% concentration significantly reduced the AFB1 production (ranging from 88 to 100%). A similar approach was used for caffeic acid, hydroxytyrosol, tyrosol and verbascoside, the major pure phenolic compounds identified in OMWW fractions. They were evaluated at increasing doses (10, 50 and 100 ?g/ml) following both AFB1 production and fungal growth. At the highest concentration (100 ?g/ml) all pure compounds showed a reduction of about 99% of AFB1 production without any influence on fungal growth. This is the first time in which OMWWs and their main phenolics were used in the treatments against AFB1 production. The results obtained could provide possible new strategies for preventing AFB1 food contamination using olive polyphenols and OMWW fractions with anti-aflatoxigenic effect, and permitting to harness in a sustainable way an olive oil by-product.

In recent years a rising common concern is looking at biodiversity concept with a new sight, attempting to evaluate its economical value, as ground step for supporting measures proposed by national governments and international committees. Although this utilitarian view applied to a complex concept could cause an underestimation of the true potential of biological resources, nowadays a wide spectrum of direct and indirect quantifiable values has been recognized as tightly correlated to biodiversity. Fifty percent of the living biomass on the planet is microbial and microorganisms provide an important source of genetic information for molecular biology and biotechnology. At this respect, the direct-use values is easily perceived and continuously growing thanks to the relevant contribution of biotechnologies, and the possibility to preserve biological resources through long-term conservation of genetic resources. Fungi play a major role in bio-regulatory systems in natural ecosystems and could represent an extraordinary source of new compounds, with a large range of secondary metabolites having biological activities of great ecological relevance, from crop protection to negative impact on humans and domesticated animals.The Agro-Food Microbial Culture Collection "ITEM" (http://server.ispa.cnr.it/ITEM/Collection/), joined to the work for years of researchers in the Institute of Sciences of Food Productions, allows to produce, purify, and characterize novel bioactive metabolites obtained by growing fungal pathogens belonging to several genera. Thousands strains belonging to toxigenic genera of Fusarium, Aspergillus, Alternaria, and Penicillium, represented a great biodiversity in the ITEM collection to deepen the knowledge on fungal biology and strategies development for reducing mycotoxin contamination. Yeast and lactic bacteria strains with peculiar properties has been also preserved and characterized for autochthonous industrial fermentation of typical Apulian wines, table olive and dairy products. Probiotic bacteria are applied for functional foods. A new species of Penicillium from dryed-meat has been isolated and characterized, with possible application for safe seasoning. In general, microorganisms of agro-food interest are preserved and may represent a new frontier of discovery of novel metabolites to be used as safe and environmentally friendly agrochemicals. ITEM take part of the Italian Network of Genetic Resource - BioGenRes (www.biogenres.cnr.it/); and of the European Project on Microbial Resource Research Infrastructure - MIRRI (www.mirri.org/).

Dried vine fruits may be heavily colonized by Aspergillus species. The molecular biodiversity of an Aspergillus population (234 strains) isolated from dried vine fruit samples of worldwide origin were analyzed by investigating four housekeeping gene loci (calmodulin, beta-tubulin, elongation factor 1-alpha, RPB2). Aspergillus Sect. Nigri was dominant and the strains were identified as A. tubingensis (138), A. awamori (38), A. carbonarius (27),A. uvarum (16) and A. niger (11). Four Aspergillus flavus strains were also identified from Chilean raisins. Two clusters closely related to the A. tubingensis species with a significant bootstrap (60% and 99%) were identified as distinct populations. Among the four loci, RPB2 showed the highest genetic variability. This is the first complete study on the worldwide distribution of black Aspergilli occurring on dried vine fruits identified by a molecular approach.

Grape berries attacked by Lobesia botrana larvae are more easily infected by Aspergillus section Nigri (black aspergilli) ochratoxigenic species. Two-year field trials were carried out in Apulia (Italy) to evaluate a bioinsecticide control strategy against L. botrana and the indirect effect on reducing ochratoxin A (OTA) contamination in vineyards. A commercial Bacillus thuringiensis formulate and an experimental Beauveria bassiana (ITEM-1559) formulate were tested in two vineyards cultivated with the same grape variety, Negroamaro, but with two different training systems (espalier and little-arbor techniques). In both years and training systems the treatments by B. bassiana ITEM-1559 significantly controlled L. botrana larvae attacks with effectiveness similar to B. thuringensis (more than 20%). A significant reduction of OTA concentrations (up to 80% compared to untreated controls) was observed only in the first year in both training systems, when the metereological parameters prior to harvest were more favorable to the insect attack. Results of field trials showed that B. bassiana ITEM-1559 is a valid bioinsecticide against L. botrana and that grape moth biocontrol is a strategy to reduce OTA contamination in vineyard in seasons with heavy natural infestation.

The healthy consumers make a strong pressure to natural products that can prevent the chronic diseases and improve the general health status, and therefore an important aspect that have to be considered is the safe level of the nutraceuticals. This study reports the occurrence of Ochratoxin A (OTA) and associated fungal contamination in 35 samples of dried vine fruits imported in the European community potentially used for the development of new nutraceutical supplements. High pressure liquid chromatography analysis identified 18 samples as contaminated by OTA with an average level of 2.6 ?g/kg. OTA was measured in 4 samples of currants (mean value of 6.6 ?g/kg) and 13 samples of raisins (mean value of 1.4 ?g/kg). In one sample of currants and one of raisins from Turkey OTA exceeded the limits set by European Commission of 10 ?g/kg, being contaminated with 12.61 and 15.99 ?g/kg, respectively. All the positive samples were confirmed by Orbitrap Q Exactive through their molecular weight and the corresponding fragmentation. The worldwide consumption of dried vine fruits contributed to OTA exposure in several group of consumers. In particular, considering the potential nutraceutical approach, this consumption may be represent a severe risk for healthy consumers that consider these products like healthy and salutistic for their contents in antioxidants, flavonoids, and polyphenols. Data reported in this study confirmed the need to regularly monitor mycotoxin levels in these food products and optimize the process of fruits drying in order to reduce the development of toxigenic molds.