Unexplored microbial diversity from 2,500 food metagenomes and links with the human microbiome.

Complex microbiomes are part of the food we eat and influence our own microbiome, but their diversity remains largely unexplored. Here, we generated the open access curatedFoodMetagenomicData (cFMD) resource by integrating 1,950 newly sequenced and 583 public food metagenomes. We produced 10,899 metagenome-assembled genomes spanning 1,036 prokaryotic and 108 eukaryotic species-level genome bins (SGBs), including 320 previously undescribed taxa. Food SGBs displayed significant microbial diversity within and between food categories. Extension to >20,000 human metagenomes revealed that food SGBs accounted on average for 3% of the adult gut microbiome. Strain-level analysis highlighted potential instances of food-to-gut transmission and intestinal colonization (e.g., Lacticaseibacillus paracasei) as well as SGBs with divergent genomic structures in food and humans (e.g., Streptococcus gallolyticus and Limosilactobabillus mucosae). The cFMD expands our knowledge on food microbiomes, their role in shaping the human microbiome, and supports future uses of metagenomics for food quality, safety, and authentication.

Bifidobacterium fermentum sp. nov. and Bifidobacterium aquikefiricola sp. nov., isolated from water kefir.

Four strains, representing two novel Bifidobacterium species, were isolated from water kefir, a fermented beverage. 16S rRNA gene analysis suggested that the novel species share high identities (98.82-98.89%) with Bifidobacterium aquikefiri LMG 28769T. Complete genomes were assembled with a short- and long-read hybrid sequencing approach. In agreement with the 16S rRNA gene analysis, phylogenetics with 117 marker genes places the novel species closest to B. aquikefiri LMG 28769T as well. The isolates have average nucleotide identity (ANI) scores ranging from 81.46 to 84.84% and digital DNA-DNA hybridization (dDDH) scores from 23.9 to 38.5% with the closest related species, as well as ANI scores between the proposed new species of 80.50%, indicating that the isolates represent two novel species. Matrix-assisted laser desorption/ionization-time of flight chemotaxonomic analysis supported the gene-based taxonomic placement. We propose the names Bifidobacterium fermentum sp. nov. and Bifidobacterium aquikefiricola sp. nov. for these novel species within the Bifidobacterium genus. The proposed type strain B. fermentum WK012_4_13T (= LMG 33104T = DSM 116073T; GenBank accession number GCF_041080835.1) has a genome size of 2.43 Mbp, with a G+C content of 56.00 mol%. The proposed type strain for B. aquikefiricola WK041_4_12T (= LMG 33105T = DSM 116074T; GenBank accession number GCF_041080795.1) has a genome size of 2.36 Mbp and a G+C content of 53.94 mol%. B. fermentum cells are Gram-positive staining, non-motile, non-spore-forming, fructose-6-phosphate phosphoketolase (F6PPK)-positive, catalase- and oxidase-negative and bacillary club shaped. B. aquikefiricola cells are Gram-positive staining, non-motile, non-spore-forming, F6PPK-positive, catalase- and oxidase-negative and square rod shaped.

In-depth characterization of food and environmental microbiomes across different meat processing plants.

BACKGROUND: Processing environments can be an important source of pathogenic and spoilage microorganisms that cross contaminate meat and meat products. The aim of this study was to characterize the microbiome of raw materials, processing environments and end products from 19 facilities producing different meat products. RESULTS: The taxonomic profiles of the microbial communities evolved along processing, from raw materials to end products, suggesting that food contact (FC) surfaces play an important role in modulating the microbiome of final products. Some species persisted with the highest relative abundance in raw materials, food processing environments and/or in the final product, including species from the genera Pseudomonas, Staphylococcus, Brochothrix, Acinetobacter and Psychrobacter. Processing environments showed a very diverse core microbiota, partially shared with the products. Pseudomonas fragi and Pseudomonas sp. Lz4W (in all sample and facility types) and Brochothrix thermosphacta, Psychrobacter sp. and Psychrobacter sp. P11F6 (in raw materials, FC surfaces and end products) were prominent members of the core microbiota for all facilities, while Latilactobacillus sakei was found as a dominant species exclusively in end products from the facilities producing fermented sausages. Processing environments showed a higher amount of antimicrobial resistance genes and virulence factors than raw materials and end products. One thousand four hundred twenty-one medium/high-quality metagenome-assembled genomes (MAGs) were reconstructed. Of these, 274 high-quality MAGs (completeness > 90%) corresponded to 210 putative new species, mostly found in processing environments. For two relevant taxa in meat curing and fermentation processes (S. equorum and L. sakei, respectively), phylogenetic variation was observed associated with the specific processing facility under study, which suggests that specific strains of these taxa may be selected in different meat processing plants, likely contributing to the peculiar sensorial traits of the end products produced in them. CONCLUSIONS: Overall, our findings provide the most detailed metagenomics-based perspective up to now of the microbes that thrive in meat, meat products and associated environments and open avenues for future research activities to better understand the microbiome functionality and potential contribution to meat quality and safety. Video Abstract.

Microbiome mapping in dairy industry reveals new species and genes for probiotic and bioprotective activities.

The resident microbiome in food industries may impact on food quality and safety. In particular, microbes residing on surfaces in dairy industries may actively participate in cheese fermentation and ripening and contribute to the typical flavor and texture. In this work, we carried out an extensive microbiome mapping in 73 cheese-making industries producing different types of cheeses (fresh, medium and long ripened) and located in 4 European countries. We sequenced and analyzed metagenomes from cheese samples, raw materials and environmental swabs collected from both food contact and non-food contact surfaces, as well as operators' hands and aprons. Dairy plants were shown to harbor a very complex microbiome, characterized by high prevalence of genes potentially involved in flavor development, probiotic activities, and resistance to gastro-intestinal transit, suggesting that these microbes may potentially be transferred to the human gut microbiome. More than 6100 high-quality Metagenome Assembled Genomes (MAGs) were reconstructed, including MAGs from several Lactic Acid Bacteria species and putative new species. Although microbial pathogens were not prevalent, we found several MAGs harboring genes related to antibiotic resistance, highlighting that dairy industry surfaces represent a potential hotspot for antimicrobial resistance (AR) spreading along the food chain. Finally, we identified facility-specific strains that can represent clear microbial signatures of different cheesemaking facilities, suggesting an interesting potential of microbiome tracking for the traceability of cheese origin.

Improved sampling and DNA extraction procedures for microbiome analysis in food-processing environments.

Deep investigation of the microbiome of food-production and food-processing environments through whole-metagenome sequencing (WMS) can provide detailed information on the taxonomic composition and functional potential of the microbial communities that inhabit them, with huge potential benefits for environmental monitoring programs. However, certain technical challenges jeopardize the application of WMS technologies with this aim, with the most relevant one being the recovery of a sufficient amount of DNA from the frequently low-biomass samples collected from the equipment, tools and surfaces of food-processing plants. Here, we present the first complete workflow, with optimized DNA-purification methodology, to obtain high-quality WMS sequencing results from samples taken from food-production and food-processing environments and reconstruct metagenome assembled genomes (MAGs). The protocol can yield DNA loads >10 ng in >98% of samples and >500 ng in 57.1% of samples and allows the collection of, on average, 12.2 MAGs per sample (with up to 62 MAGs in a single sample) in ~1 week, including both laboratory and computational work. This markedly improves on results previously obtained in studies performing WMS of processing environments and using other protocols not specifically developed to sequence these types of sample, in which <2 MAGs per sample were obtained. The full protocol has been developed and applied in the framework of the European Union project MASTER (Microbiome applications for sustainable food systems through technologies and enterprise) in 114 food-processing facilities from different production sectors.

High pressure processing at the early stages of ripening enhances the safety and quality of dry fermented sausages elaborated with or without starter culture.

To study the quality of chorizo de León dry fermented sausages (DFS), high pressure processing (HPP) applied at the early stages of ripening and the use of a functional starter culture were evaluated as additional safety measures. Furthermore, the ability to control the populations of artificially inoculated Listeria monocytogenes and Salmonella Typhimurium was investigated and the evolution of microbial communities was assessed by amplicon 16S rRNA metataxonomics. The use of HPP and the starter culture, independently or combined, induced a reduction of Listeria monocytogenes of 1.5, 4.3 and > 4.8 log CFU/g respectively, as compared to control. Salmonella Typhimurium counts were under the detection limit (<1 log) in all treated end-product samples. Both additional measures reduced the activity of undesirable microbiota, such as Serratia and Brochothrix, during the production of DFS. Moreover, the starter culture highly influencedthe taxonomic profile of samples.No adverse sensory effects were observed, and panelists showed preference for HPP treated DFS. In conclusion, this new approach of applying HPP at the early stages of ripening of DFS in combination with the use of a defined starter culture improved the safety and quality of the meat product.

Selection of lactic acid bacteria as biopreservation agents and optimization of their mode of application for the control of Listeria monocytogenes in ready-to-eat cooked meat products.

In order to meet consumers´ demands for more natural foods and to find new methods to control foodborne pathogens in them, research is currently being focused on alternative preservation approaches, such as biopreservation with lactic acid bacteria (LAB). Here, a collection of lactic acid bacteria (LAB) isolates was characterized to identify potential biopreservative agents. Six isolates (one Lactococcus lactis, one Lacticaseibacillus paracasei and four Lactiplantibacillus plantarum) were selected based on their antimicrobial activity in in vitro assays. Whole genome sequencing showed that none of the six LAB isolates carried known virulence factors or acquired antimicrobial resistance genes, and that the L. lactis isolate was potentially a nisin Z producer. Growth of L. monocytogenes was successfully limited by L. lactis ULE383, L. paracasei ULE721 and L. plantarum ULE1599 throughout the shelf-life of cooked ham, meatloaf and roasted pork shoulder. These LAB isolates were also applied individually or as a cocktail at different inoculum concentrations (4, 6 and 8 log10 CFU/g) in challenge test studies involving cooked ham, showing a stronger anti-Listerial activity when a cocktail was used at 8 log10 CFU/g. Thus, a reduction of up to ~5.0 log10 CFU/g in L. monocytogenes growth potential was attained in cooked ham packaged under vacuum, modified atmosphere packaging or vacuum followed by high pressure processing (HPP). Only minor changes in color and texture were induced, although there was a significant acidification of the product when the LAB cultures were applied. Remarkably, this acidification was delayed when HPP was applied to the LAB inoculated batches. Metataxonomic analyses showed that the LAB cocktail was able to grow in the cooked ham and outcompete the indigenous microbiota, including spoilage microorganisms such as Brochothrix. Moreover, none of the batches were considered unacceptable in a sensory evaluation. Overall, this study shows the favourable antilisterial activity of the cocktail of LAB employed, with the combination of HPP and LAB achieving a complete inhibition of the pathogen with no detrimental effects in physico-chemical or sensorial evaluations, highlighting the usefulness of biopreservation approaches involving LAB for enhancing the safety of cooked meat products.

Microbiological Safety and Shelf-Life of Low-Salt Meat Products-A Review.

Salt is widely employed in different foods, especially in meat products, due to its very diverse and extended functionality. However, the high intake of sodium chloride in human diet has been under consideration for the last years, because it is related to serious health problems. The meat-processing industry and research institutions are evaluating different strategies to overcome the elevated salt concentrations in products without a quality reduction. Several properties could be directly or indirectly affected by a sodium chloride decrease. Among them, microbial stability could be shifted towards pathogen growth, posing a serious public health threat. Nonetheless, the majority of the literature available focuses attention on the sensorial and technological challenges that salt reduction implies. Thereafter, the need to discuss the consequences for shelf-life and microbial safety should be considered. Hence, this review aims to merge all the available knowledge regarding salt reduction in meat products, providing an assessment on how to obtain low salt products that are sensorily accepted by the consumer, technologically feasible from the perspective of the industry, and, in particular, safe with respect to microbial stability.

Application of lactic acid bacteria for the biopreservation of meat products: A systematic review.

The increasing concern of consumers about food quality and safety and their rejection of chemical additives has promoted the breakthrough of the biopreservation field and the development of studies on the use of beneficial bacteria and their metabolites as potential natural antimicrobials for shelf life extension and enhanced food safety. Control of foodborne pathogens in meat and meat products represents a serious challenge for the food industry which can be addressed through the intelligent use of bio-compounds or biopreservatives. This article aims to systematically review the available knowledge about biological strategies based on the use of lactic acid bacteria to control the proliferation of undesirable microorganisms in different meat products. The outcome of the literature search evidenced the potential of several strains of lactic acid bacteria and their purified or semi-purified antimicrobial metabolites as biopreservatives in meat products for achieving longer shelf life or inhibiting spoilage and pathogenic bacteria, especially when combined with other technologies to achieve a synergistic effect.