Ozone modified hypothalamic signaling enhancing thermogenesis in the TDP-43A315T transgenic model of Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS), a devastating progressive neurodegenerative disease, has no effective treatment. Recent evidence supports a strong metabolic component in ALS pathogenesis. Indeed, metabolic abnormalities in ALS correlate to disease susceptibility and progression, raising additional therapeutic targets against ALS. Ozone (O3), a natural bioactive molecule, has been shown to elicit beneficial effects to reduce metabolic disturbances and improved motor behavior in TDP-43A315T mice. However, it is fundamental to determine the mechanism through which O3 acts in ALS. To characterize the association between O3 exposure and disease-associated weight loss in ALS, we assessed the mRNA and protein expression profile of molecular pathways with a main role in the regulation of the metabolic homeostasis on the hypothalamus and the brown adipose tissue (BAT) at the disease end-stage, in TDP-43A315T mice compared to age-matched WT littermates. In addition, the impact of O3 exposure on the faecal bacterial community diversity, by Illumina sequencing, and on the neuromuscular junctions (NMJs), by confocal imaging, were analysed. Our findings suggest the effectiveness of O3 exposure to induce metabolic effects in the hypothalamus and BAT of TDP-43A315T mice and could be a new complementary non-pharmacological approach for ALS therapy.

Enterococcus populations in artisanal Manchego cheese: biodiversity, technological and safety aspects.

Enterococci represent a considerable proportion of the microbiota in Manchego cheeses. In this study, a total of 132 enterococci isolated from good quality Manchego cheeses from two dairies at different ripening times were genotypically characterized and identified using molecular techniques. Representative isolates from the clusters obtained after genotyping were assayed for some enzymatic activities considered to have a potential role in cheese ripening, and for 2,3-butanedione and acetoin production, evaluation of odor intensity and appearance in milk and safety evaluation. Enterococcus faecalis was the predominant specie, accounting for 81.8% of the total isolates, while Enterococcus faecium, Enterococcus hirae and Enterococcus avium were present in low proportions. The number of genotypes involved at each ripening time varied both between dairies and with the ripening times; genotype E. faecalis Q1 being present in almost all the samples from both dairies. Eight isolates showed a higher proteolytic activity and 3 isolates produced high quantities of acetoin-diacetyl, for which reason they are interesting from a technological standpoint. A low antibiotic resistance was found and almost all the strains were susceptible to clinically important antibiotics. On the contrary, only four isolates (E. faecalis C4W1 and N0W5, and E. faecium N32W1 and C16W2) did not harbor some of the virulence genes assayed.

Genotypic and technological characterization of Leuconostoc isolates to be used as adjunct starters in Manchego cheese manufacture.

Twenty-seven Leuconostoc (Ln.) isolates from Manchego cheese were characterized by phenotypic and genotypic methods, and their technological abilities studied in order to test their potential use as dairy starter components. While phenotypic diversity was evaluated by studying the biochemical characteristics of technological interest (i.e. acidifying and aminopeptidase activities), genotypic diversity was evidenced by using Randomly Amplified Polymorphic DNA-Polymerase Chain Reaction (RAPD-PCR). Additional technological abilities such as lipolytic, proteolytic and autolytic activities, salt and pH tolerance and production of dextran, flavour compounds and biogenic amines, were investigated. The marked differences among strains reflected the existing biodiversity in naturally fermented products. After statistically evaluating their performance, strains C0W2, belonging to Ln. lactis, and C16W5 and N2W5, belonging to Ln. mesenteroides subsp. dextranicum, revealed the best properties to be used in mixed dairy starter cultures. This study evidences the fact that natural environments can be considered as a proper source of useful strains, for the dairy industry.

Genetic diversity, dynamics, and activity of Lactobacillus community involved in traditional processing of artisanal Manchego cheese.

A total of 248 strains of predominant lactobacilli isolated during the manufacture and ripening of artisanal Manchego cheeses obtained from two dairies were obtained and the genetic diversity of 197 investigated using random amplified polymorphic DNA (RAPD-PCR). 51 isolates could not be lysed and were therefore not genotyped. Forty-two distinct RAPD patterns, grouped in six major clusters at a similarity level of 54%, were obtained. Phenotypic characterization of isolates enabled their assignment to the species L. plantarum, L. brevis, L. paracasei subsp. paracasei, L. fermentum, L. pentosus, L. acidophilus and L. curvatus. In samples from both dairies, the species L. plantarum, L. brevis and L. paracasei subsp. paracasei dominated during ripening. Three genotypes showed excellent physiological characteristics and were therefore proposed as adjunct cultures for Manchego cheese manufacture.

Phenotypic and genotypic characterization of lactobacilli isolated from Spanish goat cheeses.

Lactic acid bacteria from 18 Spanish goat cheeses produced by seven dairies were isolated to evaluate the genetic diversity of this bacterial community. 136 representative isolates were characterized by phenotyping and Randomly Amplified Polymorphic DNA (RAPD-PCR) analysis. Ten species were identified with predominance of Lactobacillus paracasei subsp. paracasei. The presence of L. curvatus, L. pentosus, L. cellobiosus and L. rhamnosus has not hitherto been reported in Spanish goat cheeses. A high degree of genetic diversity was found for L. paracasei subsp. paracasei, L. curvatus and L. plantarum. Some of the identified strains displayed strong acidifying and proteolytic capacities.

Characterization of Lactobacillus strains and monitoring by RAPD-PCR in controlled fermentations of "Almagro" eggplants.

The characterization of 23 Lactobacillus strains was performed. The strains were assayed for biogenic amine-forming capacity, hydrogen peroxide production, pectin esterase, cellulase and polygalacturonase production, growth rate, acidifying capacity and salt tolerance. Three strains were selected which belonged to the species, Lactobacillus brevis, Lactobacillus plantarum and Lactobacillus fermentum. Different starter cultures prepared as combinations of these three strains were assayed in pilot scale fermentations and Randomly Amplified Polymorphic DNA (RAPD) analysis, using a previously selected random primer, was applied for monitoring the inoculated strains. The course of fermentations was similar in all batches but sensorial analysis of eggplants fermented using a mixed culture of the three strains displayed the best results, and no differences were obtained when compared with commercial eggplants.

Genetic diversity (RAPD-PCR) of lactobacilli isolated from "Almagro" eggplant fermentations from two seasons.

Genetic diversity of 323 strains of lactobacilli isolated from an Almagro eggplant manufacturing plant was analyzed by using random amplified polymorphic DNA (RAPD). Thirty-four distinct RAPD patterns were obtained with 95% of isolates grouped into 18 main clusters. Genetic diversity was higher in brines from season II, Lactobacillus plantarum and Lactobacillus fermentum/cellobiosus being the species with the greatest number of genotypes. A single L. fermentum/cellobiosus genotype comprised isolates from both seasons and could be considered endemic to that factory.

Identification of lactic acid bacteria from spontaneous fermentation of 'Almagro' eggplants by SDS-PAGE whole cell protein fingerprinting.

In order to complete the previously performed phenotypic identification [Sánchez et al., 2000. Int. J. Food Microbiol. 59 (2000) 9], whole cell protein patterns obtained using Sodium Dodecyl Sulphate Polyacrylamide Gel Electrophoresis (SDS-PAGE) of 149 lactic acid bacteria (LAB) strains, isolated from 'Almagro' eggplants spontaneous fermentation, were analysed. Phenotypic identification of those strains had revealed the presence of the species Lactobacillus (Lb) plantarum, Lb. pentosus, Lb. brevis biotype 2, Lb. brevis biotype 3, Lb. fermentum, Lactobacillus spp. and Aerococcus viridans. The SDS-PAGE technique generated complex and stable patterns that were easy to interpret and compare with those of the 17 reference strains used in this study. After numerical analysis of the resulting electrophoretic protein patterns, six well-delineated clusters were discriminated that, with some exceptions, correlated well with the different groups phenotypically found. Only two strains showed unique protein patterns without meaningful homology to any of the reference strains used for this study and remained unidentified.

Effect of freezing on the spontaneous fermentation and sensory attributes of Almagro eggplants.

A study of the physico-chemical and microbiological processes taking place during the spontaneous fermentation of eggplant fruits that had been frozen after blanching revealed a delay in the onset of the active fermentation stage, which has been ascribed to the adverse effect of the freezing temperature employed on the lactic acid bacteria involved in fermentation. The species most affected by freezing were L. fermentum and L. brevis biotype 2. Despite this finding, the pattern of fermentation was similar to the fermentation process in unfrozen eggplants. Freezing of the fruits did not significantly affect the sensory acceptability of the product and, in fact, in some cases, Almagro eggplants made from frozen fruits were preferred to commercially manufactured products.