Insect frass in stored cereal products as a potential source of Lactobacillus sanfranciscensis for sourdough ecosystem.

AIM: Insect frass samples were collected from Drosophila melanogaster, Plodia interpunctella, Rhyzopertha dominica, Sitophilus granarius, Sitophilus oryzae, Sitophilus zeamais, Tribolium confusum and Tribolium castaneum to elucidate if they can be the origin of Type I sourdough micro-organisms (Lactobacillus sanfranciscensis and Candida milleri). METHODS AND RESULTS: Selective enrichments were carried out to isolate lactic acid bacteria (LBA) and yeast. A metagenetic analysis, targeted on bacterial 16S rRNA gene and fungal ITS region, was performed by using Illumina MiSeq protocol. In cultivation conditions, Lactococcus garvieae and Rhodotorula mucilaginosa were the most frequently species among LAB and yeasts respectively. The Next Generation Sequencing approach revealed that Enterobacteriaceae, Pseudomonadacae and Bacillaceae were the dominating taxa, accounting for 61% of the bacterial community. Lactobacillus genus showed a relative abundance of only 0·36%, but L. sanfranciscensis proved to be the species most frequent between lactobacilli and predominant in faecal samples of T. castaneum and T. confusum larvae. The core fungal microbiota was constituted by Saccharomycetales, Pleosporaceae and Nectriaceae that attained the 51% of recognized OTUs. While the most abundant yeast genus was Candida (17·1%), sequences belonging to C. milleri were not found. CONCLUSIONS: Frass released by the insects of stored cereal products can be the natural reservoir of L. sanfranciscensis. SIGNIFICANCE AND IMPACT OF THE STUDY: Insect dejections are potential sources of novel strains for controlled bakery productions.

Characterization of malolactic bacteria isolated from Aosta Valley wines and evidence of psychrotrophy in some strains.

AIM: A survey on indigenous malolactic bacteria populations isolated from wines produced in 13 different wineries of Aosta Valley, the highest vine-growing area in Europe, was carried out in order to characterize the dominant strains in spontaneous malolactic fermentation (MLF) and to reveal the appearance of psychrotrophic ones. METHODS AND RESULTS: Fifty-four isolates were identify by ITS rDNA region analysis and partial sequencing of 16S rDNA gene: 80% were ascribed to Oenococcus oeni while the remaining 20% were attributed to Pediococcus damnosus species. The genetic diversity of 43 O. oeni isolates was investigated through PFGE analysis after ApaI and SfiI restriction: 28 different pulso-types were discriminated at a level of similarity of 90%. In general, the MLF was led by more than one strain simultaneously. No connection between genotype and grape variety or vine-training system or geographical site of isolation was observed. The histidine decarboxylase gene was not found in any isolate. Pediococci proved to be more resistant than oenococci to lysozyme. Three O. oeni strains (2A1, 6A2 and 11A4) were able to develop at 10°C in Petit Rouge wine. CONCLUSIONS: Psychrotrophy is a phenotypic trait present in O. oeni species and it may be possible to select strains for the management of MLF in cold climate territories where this biologic transformation is very difficult to control. SIGNIFICANCE AND IMPACT OF THE STUDY: The natural emergence of strains able to perform the MLF at 10°C in wine is a new finding, interesting because it confirms the ecological ability of O. oeni species to adapt itself to environmental conditions by strain phenotype variations. It can be also a starting point for more sustainable oenological practices, since it would be alternative to the conditioning systems of the tanks or of the wineries where they are costly in terms of investment and energy consumption.

Development of a Type I gluten-free sourdough.

UNLABELLED: The aim of this study was the setting up of a gluten-free sourdough from selected lactobacilli and yeasts isolated from a traditional wheat-based Type I sourdough. A gluten-free matrix was inoculated with Lactobacillus sanfranciscensis and Candida humilis, fermented to pH 4·0, and constantly propagated for ten times. A stable association between micro-organisms was observed from the second refreshment with mean values of 9·08 ± 0·25 log CFU g(-1) for lactobacilli and 7·81 ± 0·07 log CFU g(-1) for yeasts. In order to have a good workability of the dough, a 230 BU consistency was considered. Rheofermentographic indices remained constant over the ten refreshments, showing an average value of 23·2 mm dough height in about 7·5 h. The CO2 production and retention volumes reached average values of 1430 and 1238 ml respectively. The microbiological and technological data obtained highlighted that a GF sourdough was effectively developed. SIGNIFICANCE AND IMPACT OF THE STUDY: Type I sourdough has a long tradition as a leavening agent of baked goods as its use results in an improved texture, flavour, taste and extended shelf-life of the final products. In this study a Type I gluten-free sourdough was obtained. After few refreshments in controlled conditions, the sourdough presented a stable association between Lactobacillus sanfranciscensis and Candida humilis, constant fermentation times and technological properties (in terms of dough consistency, dough maximum height, CO2 production and retention). The results showed that the gluten-free sourdough developed in this study can improve the overall quality of gluten-free baked products.

Heat inactivation of wine spoilage yeast Dekkera bruxellensis by hot water treatment.

UNLABELLED: Cell suspensions of four Dekkera bruxellensis strains (CBS 2499, CBS 2797, CBS 4459 and CBS 4601) were subjected to heat treatment in deionized water at four different temperatures (55·0, 57·5, 60·0 and 62·5°C) to investigate their thermal resistance. The decimal reduction times at a specific temperature were calculated from the resulting inactivation curves: the D-values at 55·0°C ranged from 63 to 79·4 s, at 57·5°C from 39·6 to 46·1 s, at 60·0°C from 19·5 to 20·7 s, at 62·5°C from 10·2 to 13·7 s. The z-values were between 9·2 and 10·2°C, confirming that heat resistance is a strain-dependent character. A protocol for the sanitization of 225 l casks by immersion in hot water was set up and applied to contaminated 3-year-old barrels. The heat penetration through the staves was evaluated for each investigated temperature by positioning a thermal probe at 8 mm deep. A treatment at 60°C for an exposure time of 19 min allowed to eliminate the yeast populations up to a log count reduction of 8. SIGNIFICANCE AND IMPACT OF THE STUDY: Brettanomyces/Dekkera bruxellensis is the main yeast involved in red wine spoilage that occurs during ageing in barrel, generating considerable economic losses. Current sanitization protocols, performed using different chemicals, are ineffective due to the porous nature of the wood. The thermal inactivation of D. bruxellensis cells by hot water treatment proves to be efficacious and easy to perform, provided that the holding time at the killing temperature takes into account the filling time of the vessel and the time for the heat penetration into the wood structure.

Short communication: Characterization of Shiga toxin 2-carrying bacteriophages induced from Shiga-toxigenic Escherichia coli isolated from Italian dairy products.

Forty samples of raw milk cheese and 25 samples of raw milk itself were subjected to enrichment culture for Shiga-toxigenic Escherichia coli (STEC), and a single Shiga toxin 2- (Stx(2)) positive strain was obtained from one of the cheese samples. Thus, aged cheeses in which the curd is subsequently heat treated (48°C) cannot be presumed to be STEC free. Infective Stx(2) bacteriophages were induced from 3 STEC strains isolated elsewhere from raw milk and 1 STEC strain from aged cheese sourced in Italy. Data on E. coli host range, morphology, genome size, and genetic variation determined by restriction fragment length polymorphism and multi-locus genotyping are presented. Although all 4 bacteriophages were found to be short-tailed Podoviridae, they exhibited considerable variation in both genome size and content. This extended to the Stx(2) genes themselves, whose sequences contained several point mutations, but these did not translate to amino acid substitutions.

Inactivation of wine spoilage yeasts Dekkera bruxellensis using low electric current treatment (LEC).

AIMS: The objective of this study was to investigate the inactivation of a selected yeast Dekkera bruxellensis strain 4481 in red wine by application of low electric current treatment (LEC). METHODS AND RESULTS: LEC (200 mA) was applied for 60 days to a red wine, Montepulciano d'Abruzzo, in an alternative strategy to the SO(2) addition during wine storage. The LEC effect on both cell activity and microflora viability was assessed. LEC decreased significantly the survival viable cells and increased the death rate of D. bruxellensis strain 4481 yeast. A final comparison was made of the main physico-chemical parameters of the wine after the different treatments. The study suggests the importance of an appropriate LEC treatment which limits wine deterioration in terms of off-flavours synthesis. CONCLUSIONS: The results demonstrate that the growth of undesirable Dekkera can be inhibited by low voltage treatment; LEC was shown to be useful to prevent wine spoilage and has the potential of being a concrete alternative method for controlling wine spoilage. SIGNIFICANCE AND IMPACT OF THE STUDY: Wine spoilage can be avoided by preventing the growth of undesirable Dekkera yeasts, through the effective use of LEC in the winemaking process.

Genetic diversity and physiological traits of Brettanomyces bruxellensis strains isolated from Tuscan Sangiovese wines.

Eighty four isolates of Brettanomyces bruxellensis, were collected during fermentation of Sangiovese grapes in several Tuscan wineries and characterized by restriction analysis of 5.8S-ITS and species-specific PCR. The isolates were subsequently analysed, at strain level, by the combined use of the RAPD-PCR assay with primer OPA-02 and the mtDNA restriction analysis with the HinfI endonuclease. This approach showed a high degree of polymorphism and allowed to identify seven haplotypes, one of them being the most represented and widely distributed (72 isolates, 85.7%). Physiological traits of the yeasts were investigated under a wine model condition. Haplotypes clustered into two groups according to their growth rates and kinetics of production of 4-ethylphenol and 4-ethylguaiacol. Hexylamine was the biogenic amine most produced (up to 3.92 mg l(-1)), followed by putrescine and phenylethylamine. Formation of octapamine was detected by some haplotypes, for the first time.