Francesco Villani

Background: In recent years consumers, due to the growing interest for functional foods, have put particularly attention on products enriched with probiotic lactic acid bacteria for their ability to give human benefit by their assumption. Microorganisms to be considered as probiotic should be able to survive to gastrointestinal juices and colonize the intestinal tract to provide benefits to human health. The aim of the study consisted of the in vitro investigation of novel LAB strains, isolated during cheesemaking and ripening of Caciocavallo cheese, and on their probiotic potential for food application. Methods: In order to select potential probiotic bacteria, LAB isolates were preliminarily screened for their survival to simulated gastro-intestinal transitand assayed for other activities of probiotic and functional interest. Thanks to promising attitude as potential probiotics, four strains were tested to evaluate their adhesion ability on Caco-2 cell lines used as intestinal cell model. Microbiological data were analysed by one-way analysis of variance (ANOVA). Comparisons of means were carried out using post-hoc Tukey’s test. For vitro adhesion experiments, statistical significance was evaluated by Kruskal-Wallis test for equal medians. Results: Twenty-five LAB, presumptively identified as Lactobacillus spp., were isolated from cheese during 60 days of ripening and screened for their survival to simulated gastrointestinal transit. Sixteen isolates that showed a survival rate ≥85% were identifiedas 10 different profiles of Lb. paracasei subsp. paracasei. The 10 strains tolerated high concentration of bile salts by Minimal Inhibitory Concentration and growth rate assays and showed susceptibility or moderate susceptibility against antibiotics of human and veterinary importance, except for the resistanceto the class of aminoglycoside antibiotics. Eight out 10 strains showed in vitro cholesterol-lowering ability, whereas all strains showed antioxidant activity of their cell-free supernatants. Moreover, the four strains with highest survival to simulated gastrointestinal transit showed the ability to adhere to Caco-2 cells. Conclusion: The results suggest that some strains may be effective probiotics to be use as tool to design probiotic dairy products after confirmation probiotic activities in further in vivo studies. Findings of the present study suggest that four strains showed good or strong adherent ability on Caco-2 cell monolayer, that is one of prerequisite that probiotic bacteria must have to perform their functional properties. Keywords: adhesion, antioxidant activity, bile salts, cholesterol assimilation, dairy products, gastrointestinal resistance, probiotic lactobacilli.

Microbial growth on meat to unacceptable levels contributes significantly to change meat structure, color and flavor and to cause meat spoilage. The types of microorganisms initially present in meat depend on several factors and multiple sources of contamination can be identified. The aims of this study were to evaluate the microbial diversity in beefsteaks before and after aerobic storage at 4°C and to investigate the sources of microbial contamination by examining the microbiota of carcasses wherefrom the steaks originated and of the processing environment where the beef was handled. Carcass, environmental (processing plant) and meat samples were analyzed by culture-independent high-throughput sequencing of 16S rRNA gene amplicons. The microbiota of carcass swabs was very complex, including more than 600 operational taxonomic units (OTUs) belonging to 15 different phyla. A significant association was found between beef microbiota and specific beef cuts (PPseudomonas sp. and Brochothrix thermosphacta were the main contaminants, and Acinetobacter, Psychrobacter and Enterobacteriaceae were also found. Comparing the relative abundance of OTUs in the different samples it was shown that abundant OTUs in beefsteaks after storage occurred in the corresponding carcass. However, the abundance of these same OTUs clearly increased in environmental samples taken in the processing plant suggesting that spoilage-associated microbial species originate from carcasses, they are carried to the processing environment where the meat is handled and there they become a resident microbiota. Such microbiota is then further spread on meat when it is handled and it represents the starting microbial association wherefrom the most efficiently growing microbial species take over during storage and can cause spoilage.