Potential of histamine-degrading microorganisms and diamine oxidase (DAO) for the reduction of histamine accumulation along the cheese ripening process.

Lentilactobacillus parabuchneri is the main bacteria responsible for the accumulation of histamine in cheese. The goal of this study was to assess the efficiency of potential histamine-degrading microbial strains or, alternatively, the action of the diamine oxidase (DAO) enzyme in the reduction of histamine accumulation along the ripening process in cheese. A total of 8 cheese variants of cow milk cheese were manufactured, all of them containing L. parabuchneri Deutsche Sammlung von Mikroorganismen 5987 (except for the negative control cheese variant) along with histamine-degrading strains (Lacticaseibacillus casei 4a and 18b; Lactobacillus delbrueckiisubsp. bulgaricus Colección Española de Cultivos Tipo (CECT) 4005 and Streptococcus salivariussubsp.thermophilus CECT 7207; two commercial yogurt starter cultures; or Debaryomyces hansenii), or DAO enzyme, tested in each cheese variant. Histamine was quantified along 100 days of cheese ripening. All the degrading measures tested significantly reduced the concentration of histamine. The highest degree of degradation was observed in the cheese variant containing D. hansenii, where the histamine content decreased up to 45.32 %. Cheese variants with L. casei, or L. bulgaricus and S. thermophilus strains, also decreased in terms of histamine content by 43.05 % and 42.31 %, respectively. No significant physicochemical changes (weight, pH, water activity, color, or texture) were observed as a consequence of the addition of potential histamine-degrading adjunct cultures or DAO in cheeses. However, the addition of histamine-degrading microorganisms was associated with a particular, not unpleasant aroma. Altogether, these results suggest that the use of certain histamine-degrading microorganisms could be proposed as a suitable measure in order to decrease the amount of histamine accumulated in cheeses.

Revealing the microbial heritage of traditional Brazilian cheeses through metagenomics.

Brazilian artisanal cheeses date from the first Portuguese settlers and evolved via local factors, resulting in unique products that are now part of the patrimony and identity of different Brazilian regions. In this study, we combined several culture-independent approaches, including 16S/ITS metagenetics, assembly- and deep profiling-metagenomics to characterize the originality of the microbiota of five varieties of Brazilian artisanal cheeses from the South and Southeast regions of Brazil. Their core microbiota contained mainly lactic acid bacteria (LAB), of which Lactococcus lactis subsp. lactis was the most frequent, followed by Streptococcus thermophilus in the South region. Moreover, several samples from the Southeast region contained, as dominant LAB, two other food Streptococci belonging to a new species of the salivarius group and S. infantarius. Rinds of samples from the Southeast region were dominated by the halotolerant bacterium Corynebacterium variabile, and the yeasts Diutina catenulata, followed by Debaryomyces hansenii and Kodamaea ohmeri. Rinds from the South region contained mainly LAB due to their short ripening time, and the predominant yeast was D. hansenii. Phylogenomic analysis based on L. lactis metagenome-assembled genomes (MAGs) showed that most Brazilian strains are closely related and form a different clade from those whose genomes are available at this time, indicating that they belong to a specific group. Lastly, functional analysis showed that S. infantarius acquired a âˆ¼ 26 kb DNA fragment from S. thermophilus starter strains that carry the LacSZ system, allowing fast lactose assimilation, an adaptation advantage for growth in milk. Finally, our study identified several areas of concern, such as the presence of somatic cell DNA and high levels of antibiotic resistance genes in several cheese microbiota, suggesting that milk from diseased animals may still be used occasionally. Overall, the data from this study highlight the potential value of the traditional and artisanal cheese production network in Brazil, and provide a metagenomic-based scheme to help manage this resource safely.

Novel Variants of Streptococcus thermophilus Bacteriophages Are Indicative of Genetic Recombination among Phages from Different Bacterial Species.

Bacteriophages are the main cause of fermentation failures in dairy plants. The majority of Streptococcus thermophilus phages can be divided into either cos- or pac-type phages and are additionally characterized by examining the V2 region of their antireceptors. We screened a large number of S. thermophilus phages from the Chr. Hansen A/S collection, using PCR specific for the cos- or pac-type phages, as well as for the V2 antireceptor region. Three phages did not produce positive results with the assays. Analysis of phage morphologies indicated that two of these phages, CHPC577 and CHPC926, had shorter tails than the traditional S. thermophilus phages. The third phage, CHPC1151, had a tail size similar to those of the cos- or pac-type phages, but it displayed a different baseplate structure. Sequencing analysis revealed the genetic similarity of CHPC577 and CHPC926 with a subgroup of Lactococcus lactis P335 phages. Phage CHPC1151 was closely related to the atypical S. thermophilus phage 5093, homologous with a nondairy streptococcal prophage. By testing adsorption of the related streptococcal and lactococcal phages to the surface of S. thermophilus and L. lactis strains, we revealed the possibility of cross-interactions. Our data indicated that the use of S. thermophilus together with L. lactis, extensively applied for dairy fermentations, triggered the recombination between phages infecting different bacterial species. A notable diversity among S. thermophilus phage populations requires that a new classification of the group be proposed.IMPORTANCEStreptococcus thermophilus is a component of thermophilic starter cultures commonly used for cheese and yogurt production. Characterizing streptococcal phages, understanding their genetic relationships, and studying their interactions with various hosts are the necessary steps for preventing and controlling phage attacks that occur during dairy fermentations.

Exopolysaccharides modify functional properties of whey protein concentrate.

The objective of this research was to produce whey protein concentrate (WPC) with modified functionality using exopolysaccharide- (EPS) producing cultures. Two different EPS-producing cultures, Lactococcus lactis ssp. cremoris JFR and Streptococcus thermophilus, producing EPS1 and EPS2 respectively, were used in this study. One EPS-nonproducing commercial cheese culture (DVS 850; Chr. Hansen, Milwaukee, WI) was used as the control. Reconstituted sweet whey powder was used in this study to eliminate variations from fresh whey. Cultures grown overnight in reconstituted WPC (10% wt/vol) were added, directly or after overnight cooling (cooled EPS), at 2% (wt/vol) to 6% (wt/wt) solution of reconstituted whey. Whey was then high-temperature, short-time pasteurized at 75 °C for 35s and ultrafiltered to a volume reduction factor of 5. Ultrafiltered whey (retentate) was spray dried at inlet and outlet air temperatures of 200 and 90 °C, respectively, to obtain WPC. In general, the solubility of WPC was higher at pH 7 than at pH 3. Whey protein concentrate containing EPS2 exhibited higher protein solubility than did WPC containing no EPS. Also, the presence of EPS in WPC decreased protein denaturation. The emulsifying ability of WPC containing EPS was higher than that in control. Addition of EPS to WPC significantly enhanced its gelling ability. Foam overrun and hydrophobicity of WPC were not affected by addition of EPS. In conclusion, data obtained from this study show that EPS modify WPC functionality. The extent of modification depends on the type of EPS. Cooling of culture containing EPS before its addition to whey further reduced WPC protein denaturation and increased its solubility at pH 7 and gel hardness.

Culture-independent detection of microorganisms in traditional Slovakian bryndza cheese.

In order to investigate the microflora of Slovakian bryndza cheese (a cheese containing unpasteurized or pasteurized ewes' milk component) by a culture-independent method, DNA was extracted directly from 7 bryndza samples and analysed by an innovative method. Using the universal prokaryotic and fungal primers, ribosomal DNA internal transcribed spacer (ITS) regions with variable length were amplified. The standard universal reverse primer L1 aligning to bacterial 23s rDNA was found unsuitable for some lactic acid bacteria and other species based on in silico analysis. Therefore, L1 primer was replaced by a combination of novel primers GplusR and GminusR aligning to the adjacent, more conserved DNA region. The amplification profiles were visualised by both standard electrophoresis and by fluorescent capillary gel electrophoresis. From representative samples, major amplicons were excised from the gel, cloned and sequenced. Sequencing revealed that the samples contained Lactobacillus delbrueckii, Lactobacillus brevis, Streptococcus thermophilus, Lactococcus lactis, Lactococcus raffinolactis, Streptococcus macedonicus, Leuconostoc pseudomesenteroides, Debaromyces hansenii, Mucor fragilis, Yarrowia lipolytica and Galactomyces geotrichum. These results represent an extension of the knowledge on the microflora of Slovakian bryndza cheese. The introduced automated ribosomal DNA intergenic spacer analysis of the bacterial and fungal genomes proved to be very effective in the application of studying microflora of cheese.