Sourdough is one of the oldest starters traditionally used for making baked goods, offering several advantages to the sensory, rheology, and shelf life of final products. The present study investigated, for the first time, the microbiota of spontaneously fermented Maiorca dough samples collected from bakeries located in Sicily (Italy). Four sourdough samples (M1, M2, M3, and M4), were produced using Triticum vulgare Host. var. albidum Koern (Maiorca grain) were subjected to LAB and yeasts isolation and identification at the species level. The in-depth characterization of the lactobacilli population revealed that Lactiplantibacillus plantarum and Levilactobacillus brevis unquestionably dominated the Maiorca sourdough ecosystem. Concerning the yeasts community, high species diversity was found. Saccharomyces cerevisiae and Wickerhamomyces anomalus were the most frequently isolated species. In addition, Torulaspora delbrueckii, Pichia kluyveri, Candida boidinii, and Candida diddensiae were also detected. Investigations on both pro-technological and functional traits of the isolated strains could lead to the selection of starters for the production of baked goods.

Mucins are large glycoproteins whose degradation requires the expression of several glycosil hydrolases to catalyze the cleavage of the oligosaccharide chains and release monosaccharides that can be assimilated. In this study, we present a characterization on the strains Clostridium celatum WC0700, Clostridium tertium WC0709, and Paraclostridium bifermentans WC0705. These three strains were previously isolated from enrichment cultures on mucin of fecal samples from healthy subjects and can use mucin as sole carbon and nitrogen source. Genome analysis and in vitro functional analysis of these strains elucidated their physiological and biochemical features. C. celatum WC0700 harbored the highest number of glycosyl hydrolases specific for mucin degradation, while P. bifermentans WC0705 had the least. These predicted differences were confirmed growing the strains on 5 mucin-decorating monosaccharides (L-fucose, N-Acetylneuraminic acid, galactose, N-acetylgalactosamine, and N-acetylglucosamine) as only source of carbon. Fermenting mucin, they all produced formic, acetic, propionic, butyric, isovaleric, and lactic acids, and ethanol; acetic acid was the main primary metabolite. Further catabolic capabilities were investigated, as well as antibiotic susceptibility, biofilm formation, tolerance to oxygen and temperature. The potential pathogenicity of the strains was evaluated through in silico research of virulence factors. The merge between comparative and functional genomics and biochemical/physiological characterization provided a comprehensive view of these mucin degraders, reassuring on the safety of these species and leaving ample scope for deeper investigations on the relationship with the host and for assessing if some relevant health-promoting effect could be ascribed to these SCFA producing species.

Aim:Clostridia are relevant commensals of the human gut due to their major presence and correlations to the host. In this study, we investigated intestinal Clostridia of 51 healthy subjects and reconstructed their taxonomy and phylogeny. The relatively small number of intestinal Clostridia allowed a systematic whole genome approach based on average amino acid identity (AAI) and core genome with the aim of revising the current classification into genera and determining evolutionary relationships. Methods: 51 healthy subjects’ metagenomes were retrieved from public databases. After the dataset’s validation through comparison with Human Microbiome Project (HMP) samples, the metagenomes were profiled using MetaPhlAn3 to identify the population ascribed to the class Clostridia. Intestinal Clostridia genomes were retrieved and subjected to AAI analysis and core genome identification. Phylogeny investigation was conducted with RAxML and Unweighted Pair Group Method with Arithmetic Mean (UPGMA) algorithms, and SplitsTree for split decomposition. Results: 225 out of 406 bacterial taxonomic units were ascribed to Bacillota [Firmicutes], among which 124 were assigned to the class Clostridia. 77 out of the 124 taxonomic units were referred to a species, altogether covering 87.7% of Clostridia abundance. According to the lowest AAI genus boundary set at 55%, 15 putative genera encompassing more than one species (G1 to G15) were identified, while 19 species did not cluster with any other one and each appeared to belong to a diverse genus. Phylogenetic investigations highlighted that most of the species clustered into three main evolutive clades. Conclusion: This study shed light on the species of Clostridia colonizing the gut of healthy adults and pinpointed several gaps in knowledge regarding the taxonomy and the phylogeny of Clostridia.

Archaea are an understudied component of the human microbiome. In this study, the gut archaeome and bacteriome of 60 healthy adults from different region were analyzed by whole-genome shotgun sequencing. Archaea were ubiquitously found in a wide range of abundances, reaching up to 7.2 %. The dominant archaeal phylum was Methanobacteriota, specifically the family Methanobacteriaceae, encompassing more than 50 % of Archaea in 50 samples. The previously underestimated Thermoplasmatota, mostly composed of Methanomassiliicoccaceae, dominated in 10 subjects (>50 %) and was present in all others except one. Halobacteriota, the sole other archaeal phylum, occurred in negligible concentration, except for two samples (4.6–4.8 %). This finding confirmed that the human gut archaeome is primarily composed of methanogenic organisms and among the known methanogenic pathway: i) hydrogenotrophic reduction of CO₂ is the predominant, being the genus Methanobrevibacter and the species Methanobrevibacter smithii the most abundant in the majority of the samples; ii) the second pathway, that involved Methanomassiliicoccales, was the hydrogenotrophic reduction of methyl-compounds; iii) dismutation of acetate or methyl-compounds seemed to be absent. Co-occurrence analysis allowed to unravel correlations between Archaea and Bacteria that shapes the overall structure of the microbial community, allowing to depict a clearer picture of the human gut archaeome.

In the present work, two cell-envelope proteinases (CEPs) from Lacticaseibacillus casei strains PRA205 and 2006 were characterized at both the biochemical and genetic levels. The genomes of both L. casei strains included two putative CEPs genes prtP2 and prtR1, but only prtR1 was transcribed. The extracted PrtR1 proteinases were serine proteinases with optimal activity at 40 °C and pH 7.5, and were activated by Ca²⁺ ions. Interestingly, PrtR1 from L. casei PRA205 exhibited high residual activity at pH 4 and at 5 °C, suggesting its possible exploitation for fermented food production. The caseinolytic activity against αS1- and β-casein indicated that both PrtR1s belonged to the PI/PIII type. These PrtR1s cleaved β-casein peptide bonds preferentially when amino acid M or N was present at the P1 subsite and amino acids A and D were at the P1′ subsite. Several bioactive peptides were found to be released from PrtR1 after αs1- and β-casein hydrolysis.