Characterization of the microbial community in ripened Pecorino Toscano cheese affected by pink discoloration.

Pink discoloration defect can cause economic losses for cheese producers due to the impossibility to sell the defected cheese, but few knowledge is currently available on the causes of this defect. To gain more insight on the causes that lead to the formation of pink discoloration in Pecorino Toscano cheese with the Protected Designation of Origin (PDO) status, the bacterial community in defected and not defected cheese was characterized by high-throughput sequencing of bacterial 16S rRNA gene. The bacterial community in the defected cheese significantly differed compared to the control. The relative abundance of the genera Acidipropionibacterium, Enterococcus, Escherichia/Shigella, Lactobacillus, Lentilactobacillus and Propionibacterium was higher in the cheese with pink discoloration defect. The concentration of short chain fatty acids and of lactic acid in cheese was measured and a shift towards the production of propionate in the cheese with pink discoloration defect was observed. Furthermore, the possible involvement of microbially produced vitamin B12 in the formation of pink discoloration was not supported by the data, since a tendency to a lower concentration of vitamin B12 was measured in the defected cheese compared to the control.

Rapid Detection of Brettanomyces bruxellensis in Wine by Polychromatic Flow Cytometry.

Brettanomyces bruxellensis is found in several fermented matrices and produces relevant alterations to the wine quality. The methods usually used to identify B. bruxellensis contamination are based on conventional microbiological techniques that require long procedures (15 days), causing the yeast to spread in the meantime. Recently, a flow cytometry kit for the rapid detection (1-2 h) of B. bruxellensis in wine has been developed. The feasibility of the method was assessed in a synthetic medium as well as in wine samples by detecting B. bruxellensis in the presence of other yeast species (Saccharomyces cerevisiae and Pichia spp.) and at the concentrations that produce natural contaminations (up to 105 cells/mL), as well as at lower concentrations (103-102 cells/mL). Wine samples naturally contaminated by B. bruxellensis or inoculated with four different strains of B. bruxellensis species together with Saccharomyces cerevisiae and Pichia spp., were analyzed by flow cytometry. Plate counts were carried out in parallel to flow cytometry. We provide evidence that flow cytometry allows the rapid detection of B. bruxellensis in simple and complex mixtures. Therefore, this technique has great potential for the detection of B. bruxellensis and could allow preventive actions to reduce wine spoilage.

Enumeration and rapid identification of yeasts during extraction processes of extra virgin olive oil in Tuscany.

The aim of this study was to evaluate the occurrence of yeast populations during different olive oil extraction processes, carried out in three consecutive years in Tuscany (Italy), by analysing crushed pastes, kneaded pastes, oil from decanter and pomaces. The results showed yeast concentrations ranging between 10(3) and 10(5) CFU/g or per mL. Seventeen dominant yeast species were identified by random amplified polymorphic DNA with primer M13 and their identification was confirmed by restriction fragments length polymorphism of ribosomal internal transcribed spacer and sequencing rRNA genes. The isolation frequencies of each species in the collected samples pointed out that the occurrence of the various yeast species in olive oil extraction process was dependent not only on the yeasts contaminating the olives but also on the yeasts colonizing the plant for oil extraction. In fact, eleven dominant yeast species were detected from the washed olives, but only three of them were also found in oil samples at significant isolation frequency. On the contrary, the most abundant species in oil samples, Yamadazyma terventina, did not occur in washed olive samples. These findings suggest a phenomenon of contamination of the plant for oil extraction that selects some yeast species that could affect the quality of olive oil.

Replacement of SO2 with an Unripe Grape Extract and Chitosan during Oak Aging: Case Study of a Sangiovese Wine.

The aim of this study was to evaluate the chemical, microbiological and sensory characteristics of a Sangiovese wine aged in barrique with the addition of an unripe grape extract (UGE) as an alternative to sulfur dioxide. Three samples were considered: control wine (TQ) with free SO2 of approximately 15 mg/L; sample A with chitosan (100 mg/L) and UGE (200 mg/L); and sample B with UGE (400 mg/L). The results achieved in this work demonstrated that the UGE, either alone or in combination with chitosan, was able to maintain the color characteristics of the Sangiovese wine and its sensory quality. Moreover, the addition of UGE contributed to an early and better stabilization of the color through the formation of polymeric pigments. The microbiological stabilization was comparable to SO2 when UGE was used at 200 mg/L in combination with chitosan. The market survey conducted in the present study confirmed how the use of UGE as an alternative to sulfitation was positively accepted by consumers, who are increasingly attentive not only to the quality of the wines they select but also to the sustainability of the production processes from which they derive and to the fact that they are not harmful to human health.

Trehalose Effect on the Aggregation of Model Proteins into Amyloid Fibrils.

Protein aggregation into amyloid fibrils is a phenomenon that attracts attention from a wide and composite part of the scientific community. Indeed, the presence of mature fibrils is associated with several neurodegenerative diseases, and in addition these supramolecular aggregates are considered promising self-assembling nanomaterials. In this framework, investigation on the effect of cosolutes on protein propensity to aggregate into fibrils is receiving growing interest, and new insights on this aspect might represent valuable steps towards comprehension of highly complex biological processes. In this work we studied the influence exerted by the osmolyte trehalose on fibrillation of two model proteins, that is, lysozyme and insulin, investigated during concomitant variation of the solution ionic strength due to NaCl. In order to monitor both secondary structures and the overall tridimensional conformations, we have performed UV spectroscopy measurements with Congo Red, Circular Dichroism, and synchrotron Small Angle X-ray Scattering. For both proteins we describe the effect of trehalose in changing the fibrillation pattern and, as main result, we observe that ionic strength in solution is a key factor in determining trehalose efficiency in slowing down or blocking protein fibrillation. Ionic strength reveals to be a competitive element with respect to trehalose, being able to counteract its inhibiting effects toward amyloidogenesis. Reported data highlight the importance of combining studies carried out on cosolutes with valuation of other physiological parameters that may affect the aggregation process. Also, the obtained experimental results allow to hypothesize a plausible mechanism adopted by the osmolyte to preserve protein surface and prevent protein fibrillation.

Extra Virgin Olive Oil Quality as Affected by Yeast Species Occurring in the Extraction Process.

In extra virgin olive oil (EVOO) extraction process, the occurrence of yeasts that could affect the quality of olive oil was demonstrated. Therefore, in this work, at first, the yeasts occurring during different extractive processes carried out in a Tuscany oil mill, at the beginning, in the middle, and the end of the harvesting in the same crop season, were quantified. Then, possible effects on quality of EVOO caused by the predominant yeast species, possessing specific enzymatic activities, were evaluated. Yeast concentrations were higher in extraction processes at the end of the harvesting. Twelve yeast species showing different isolation frequencies during olive oil extractive process and according to the harvesting date were identified by molecular methods. The yeast species dominating olive oil samples from decanter displayed enzymatic activities, potentially affecting EVOO quality according to zymogram analysis. HS-SPME-GC-MS analysis of the volatile compounds in commercial EVOO, inoculated with three yeast species (Nakazawaea molendini-olei, Nakazawaea wickerhamii, Yamadazyma terventina), pointed out significant differences depending on the strain inoculated. In conclusion, during the olive oil extractive processes, some yeast species colonize the extraction plant and may influence the chemical and sensory characteristics of EVOO depending on the cell concentrations and their enzymatic capabilities.