This work presents the first study on the bacterial communities in Pico cheese, a traditional cheese of the Azores (Portugal), made from raw cow's milk. Pyrosequencing of tagged amplicons of the V3-V4 regions of the 16S rDNA and Operational Taxonomic Unit-based (OTU-based) analysis were applied to obtain an overall idea of the microbiota in Pico cheese and to elucidate possible differences between cheese-makers (A, B and C) and maturation times.Pyrosequencing revealed a high bacterial diversity in Pico cheese. Four phyla (Firmicutes, Proteobacteria, Actinobacteria and Bacteroidetes) and 54 genera were identified. The predominant genus was Lactococcus (77% of the sequences). Sequences belonging to major cheese-borne pathogens were not found. Staphylococcus accounted for 0.5% of the sequences. Significant differences in bacterial community composition were observed between cheese-maker B and the other two units that participated in the study. However, OTU analysis identified a set of taxa (Lactococcus, Streptococcus, Acinetobacter, Enterococcus, Lactobacillus, Staphylococcus, Rothia, Pantoea and unclassified genera belonging to the Enterobacteriaceae family) that would represent the core components of artisanal Pico cheese microbiota. A diverse bacterial community was present at early maturation, with an increase in the number of phylotypes up to 2 weeks, followed by a decrease at the end of ripening. The most remarkable trend in abundance patterns throughout ripening was an increase in the number of sequences belonging to the Lactobacillus genus, with a concomitant decrease in Acinetobacter, and Stenotrophomonas. Microbial rank abundance curves showed that Pico cheese's bacterial communities are characterized by a few dominant taxa and many low-abundance, highly diverse taxa that integrate the socalled "rare biosphere".2

As a genus that has evolved for resistance against adverse environmental factors and that readily exchanges genetic elements, enterococci are well adapted to the cheese environment and may reach high numbers in artisanal cheeses. Their metabolites impact cheese flavor, texture, and rheological properties, thus contributing to the development of its typical sensorial properties. Due to their antimicrobial activity, enterococci modulate the cheese microbiota, stimulate autolysis of other lactic acid bacteria (LAB), control pathogens and deterioration microorganisms, and may offer beneficial effects to the health of their hosts. They could in principle be employed as adjunct/protective/probiotic cultures; however, due to their propensity to acquire genetic determinants of virulence and antibiotic resistance, together with the opportunistic character of some of its members, this genus does not possess Qualified Presumption of Safety (QPS) status. It is, however, noteworthy that some putative virulence factors described in foodborne enterococci may simply reflect adaptation to the food environment and to the human host as commensal. Further research is needed to help distinguish friend from foe among enterococci, eventually enabling exploitation of the beneficial aspects of specific cheese-associated strains. This review aims at discussing both beneficial and deleterious roles played by enterococci in artisanal cheeses, while highlighting the need for further research on such a remarkably hardy genus.

Autochthonous lactic acid bacteria may provide a means of promoting the quality and safety of traditional fermented food products, in particular, artisanal cheeses. Pico cheese is an artisanal, dairy specialty of the Azores in risk of disappearing. Efforts to maintain its quality to the requirements of the modern markets are, thus, necessary. Lactic acid bacteria were isolated from artisanal Pico cheese, identified by sequencing of the 16S rRNA gene, and their potential as starter cultures was evaluated by studying their acidification ability, enzymatic activities (caseinolysis, lipolysis and API-ZYM profile), diacetyl and expolysaccharide production, autolysis, antimicrobial activity against Listeria monocytogenes ATCC 7466, Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 29523, Pseudomonas aeruginosa ATCC 27853 and Clostridium perfringens ATCC 8357, sensory evaluation of odour formation in milk, syneresis and firmness of the curd. Several of the studied lactic acid bacteria isolates showed interesting properties for practical application as starters in artisanal cheese production. The isolates with the highest number of positive traits and, therefore, the most promising for starter development were Lactococcus lactis ssp. lactis L1C21M1, Lactobacillus paracasei L1B1E3, Leuconostoc pseudomesenteroides L1C1E6, Lactobacillus casei L1A1E5 and L1C1E8.

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Animal products, in particular dairy and fermented products, are major natural sources of lactic acid bacteria (LAB). These are known for their antimicrobial properties, as well as for their roles in organoleptic changes, antioxidant activity, nutrient digestibility, the release of peptides and polysaccharides, amino acid decarboxylation, and biogenic amine production and degradation. Due to their antimicrobial properties, LAB are used in humans and in animals, with beneficial effects, as probiotics or in the treatment of a variety of diseases. In livestock production, LAB contribute to animal performance, health, and productivity. In the food industry, LAB are applied as bioprotective and biopreservation agents, contributing to improve food safety and quality. However, some studies have described resistance to relevant antibiotics in LAB, with the concomitant risks associated with the transfer of antibiotic resistance genes to foodborne pathogens and their potential dissemination throughout the food chain and the environment. Here, we summarize the application of LAB in livestock and animal products, as well as the health impact of LAB in animal food products. In general, the beneficial effects of LAB on the human food chain seem to outweigh the potential risks associated with their consumption as part of animal and human diets. However, further studies and continuous monitorization efforts are needed to ensure their safe application in animal products and in the control of pathogenic microorganisms, preventing the possible risks associated with antibiotic resistance and, thus, protecting public health.

Pico cheese is a traditional variety manufactured on the eponymous Azorean Island (Portugal). In total, 153 cheeses were analysed. The average total solids (TS) content by the end of maturation was 63.3 ± 0.3 g/100 g, Aw 0.943–0.966, salt‐in‐moisture 2.12–4.17 g/100 g and pH 4.89–5.11 ± 0.02. Proteolysis was similar to that of other Iberian raw milk cheeses, but the free fatty acid content was generally higher. Lactococci were the predominant bacteria. Coagulase‐positive staphylococci constituted the most severe hygiene problem. Positive correlations were found between maturation time and TS, titratable acidity, Aw, proteolysis indices and Enterobacteriaceae and coagulase‐positive staphylococcus viable numbers.

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Animal products, in particular dairy and fermented products, are natural, major sources of lactic acid bacteria (LAB). Due to their antimicrobial properties, LAB are used in humans and in animals, with beneficial effects, as probiotics or in the treatment of a variety of diseases. In livestock production, LAB contribute to animal performance, health, and productivity. In the food industry, LAB are applied as bioprotective and biopreservation agents, contributing to improve food safety and quality. However, some studies have described resistance to relevant antibiotics in LAB, with the concomitant risks associated to the transfer of antibiotic resistance genes to foodborne pathogens, their potential dissemination throughout the food chain, and the environment. Here, we summarize the application of LAB in livestock and animal products, as well as the health impact of LAB in animal food products. In general, the beneficial effects of LAB on the human food chain seem to outweigh the potential risks associated with their consumption as part of animal and human diets. However, further studies and continuous monitorization efforts are needed to ensure their safe application in animal products and in the control of pathogenic microorganisms, preventing the possible risks associated with antibiotic resistance and, thus, protecting public health.