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Summary We present a global atlas of 4,728 metagenomic samples from mass-transit systems in 60 cities over 3 years, representing the first systematic, worldwide catalog of the urban microbial ecosystem. This atlas provides an annotated, geospatial profile of microbial strains, functional characteristics, antimicrobial resistance (AMR) markers, and genetic elements, including 10,928 viruses, 1,302 bacteria, 2 archaea, and 838,532 CRISPR arrays not found in reference databases. We identified 4,246 known species of urban microorganisms and a consistent set of 31 species found in 97% of samples that were distinct from human commensal organisms. Profiles of AMR genes varied widely in type and density across cities. Cities showed distinct microbial taxonomic signatures that were driven by climate and geographic differences. These results constitute a high-resolution global metagenomic atlas that enables discovery of organisms and genes, highlights potential public health and forensic applications, and provides a culture-independent view of AMR burden in cities., Graphical abstract, Highlights • Cities possess a consistent “core” set of non-human microbes • Urban microbiomes echo important features of cities and city-life • Antimicrobial resistance genes are widespread in cities • Cities contain many novel bacterial and viral species, This systematic, worldwide catalog of urban microbiomes represents a metagenomic atlas important for understanding the ecology, virulence, and antibiotic resistance of city-specific microbial communities.

Background Since its invention, Atomic Force Microscopy has demonstrated to be one of the most interesting and useful techniques in many fields such as biology, optics and electronics to investigate nanoscale phenomena. Not only can it provide high resolution three-dimensional imaging of surfaces, but it also allows quantitative characterization of topographies, forces, mechanical and viscoelastic properties of interfaces at nanometer level. Scope and approach Here we review current literature in the food packaging field where AFM has been proposed for the quantitative characterization of surfaces with functional layers allowing for exploitation of preservative properties as a result for example of higher permeability or antimicrobial activity. In fact, probing microscopes allow analysis of physical or mechanical properties of interfaces providing relevant information at the nanoscale with regard to different parameters such as dimensions, shapes, evolution and adhesion. Furthermore, recent developments in AFM have shown how fast imaging techniques can be implemented to allow time evolution description even at relatively high temperatures. What we aim is to establish how AFM is effectively promising in the research and development of innovative food packaging. Key findings and conclusions AFM is largely used to characterize (bio)plastic materials for food packaging but also the comprehension of materials modification due to their activation is approached by using AFM, frequently combined to others instrumental analysis. In particular, even though AFM is basically used for topographical analysis of activated plastic materials, new and advanced AFM analysis are carried out for the characterization of different chemical and physical properties.

Cigarette butts (CBs) are the most common litter item on Earth but no long-term studies evaluate their fate and ecological effects. Here, the role of nitrogen (N) availability and microbiome composition on CBs decomposition were investigated by a 5-years experiment carried out without soil, in park grassland and sand dune. During decomposition, CBs chemical changes was assessed by both 13C CPMAS NMR and LC-MS, physical structure by scanning electron microscope and ecotoxicity by Aliivibrio fischeri and Raphidocelis subcapitata. Microbiota was investigated by high-throughput sequencing of bacterial and eukaryotic rRNA gene markers. CBs followed a three-step decomposition process: at the early stage (∼30 days) CBs lost ∼15.2% of their mass. During the subsequent two years CBs decomposed very slowly, taking thereafter different trajectories depending on N availability and microbiome composition. Without soil CBs showed minor chemical and morphological changes. Over grassland soil a consistent N transfer occurs that, after de-acetylation, promote CBs transformation into an amorphous material rich in aliphatic compounds. In sand dune we found a rich fungal microbiota able to decompose CBs, even before the occurrence of de-acetylation. CBs ecotoxicity was highest immediately after smoking. However, for R. subcapitata toxicity remained high after two and five years of decomposition.

Long ripened cheeses, such as Grana Padano (GP), a Protected Designation of Origin (PDO) Italian cheese, harbor a viable microbiota mainly composed of non-starter lactic acid bacteria (NSLAB), which contribute to the final characteristics of cheese. The NSLAB species Lactobacillus rhamnosus, Lb. casei and Lb. paracasei are frequently found in GP, and form a closely related taxonomic group (Lb. casei group), making it difficult to distinguish the three species through 16S rRNA sequencing. SpxB, a metabolic gene coding for pyruvate oxidase in Lb. casei group, was recently used to distinguish the species within this bacterial group, both in pure cultures and in cheese, where it could provide an alternative energy source through the conversion of pyruvate to acetate. The aim of this work was to study the evolution of the metabolically active microbiota during different stages of GP ripening, targeting 16S rRNA to describe the whole microbiota composition, and spxB gene to monitor the biodiversity within the Lb. casei group. Furthermore, activation of pyruvate oxidase pathway was measured directly in cheese by reverse transcription real time PCR (RT-qPCR). The results showed that Lb. casei group dominates throughout the ripening and high-throughput sequencing of spxB allowed to identify four clusters inside the Lb. casei group. The dynamics of the sequence types forming the clusters were followed during ripening. Pyruvate oxidase pathway was expressed in cheese, showing a decreasing trend over ripening time. This work highlights how the composition of the microbiota in the early manufacturing stages influences the microbial dynamics throughout ripening, and how targeting of a metabolic gene can provide an insight into the activity of strains relevant for dairy products.

Food Research International Volume 89, 1 November 2016, Pages 790-796 Microbial diversity in pitted sweet cherries (Prunus avium L.) as affected by High-Hydrostatic Pressure treatment (Article) Toledo del Árbol, J.ab, Pérez Pulido, R.a, La Storia, A.b, Grande Burgos, M.J.a, Lucas, R.a, Ercolini, D.b, Gálvez, A.a a Área de Microbiología, Departamento de Ciencias de la Salud, Facultad de Ciencias Experimentales, Universidad de Jaén, Jaén, Spain b Department of Agricultural Sciences, Division of Microbiology, University of Naples Federico II, Via Università 100, Portici, Italy View references (42) Abstract Sweet cherries are a highly appreciated seasonal fruit rich in anthocyanins. The purpose of the present study was to determine the effect of High-Hydrostatic Pressure (HHP) processing on the microbiological quality and bacterial biodiversity of sweet cherries. Pitted cherries inoculated with their own epiphyte microbiota to simulate a worst-case scenario of contamination during preparation and processing were treated or not by HHP (600 MPa, 8 min) and stored at 4 °C for 60 days. HHP treatment reduced total viable counts by 4.65 log cycles. The surviving bacterial fraction did not increase significantly (p < 0.05) for the first 15 days of storage. Concentrations of yeasts and molds were reduced below detectable levels. Upon prolonged storage (60 days), microbial growth was observed. Bacterial biodiversity studied by high-throughput sequencing of the 16S rRNA gene revealed that Proteobacteria had highest relative abundance (88.70%) in the spiked cherries followed by Firmicutes (11.04%). Gluconobacter and Enterobacteriaceae together with Leuconostoc were the most abundant Operational Taxonomic Units (OTUs). Upon application of HHP treatment, 97.62% of OTUs from the surviving fraction belonged to Proteobacteria. The relative abundance of Enterobacteriaceae also decreased markedly while Acetobacteraceae (represented mainly by Gluconobacter) increased to 89.18%. Gluconobacter dominated during storage. Results from the present study provide insights on the microbiota of sweet cherries and the dynamics of the bacterial populations surviving HHP treatments that may be useful to improve the non-thermal preservation of cherries.

Microbial succession over decomposing litter is controlled by biotic interactions, dispersal limitation, grazing pressure, and substrate chemical changes. Recent evidence suggests that the changes in litter chemistry and microbiome during decomposition are interdependent. However, most previous studies separately addressed the microbial successional dynamics or the molecular changes of decomposing litter. Here, we combined litter chemical characterization by 13C NMR spectroscopy with next generation sequencing to compare leaf litter chemistry and microbiome dynamics using 30 litter types, either fresh or decomposed for 30 and 180 days. We observed a decrease of cellulose and C/N ratio during decomposition, while lignin content and lignin/N ratio showed the opposite pattern. 13C NMR revealed significant chemical changes as microbial decomposition was proceeding, with a decrease in O-alkyl C and an increase in alkyl C and methoxyl C relative abundances. Overall, bacterial and eukaryotic taxonomical richness increased with litter age. Among Bacteria, Proteobacteria dominated all undecomposed litters but this group was progressively replaced by members of Actinobacteria, Bacteroidetes, and Firmicutes. Nitrogen-fixing genera such as Beijerinckia and Rhizobium occurred both in undecomposed as well as in aged litters. Among Eukarya, fungi belonging to the Ascomycota phylum were dominant in undecomposed litter with the typical phyllospheric genus Aureobasidium. In aged litters, phyllospheric species were replaced by zygomycetes and other ascomycetous and basidiomycetous fungi. Our analysis of decomposing litter highlighted an unprecedented, widespread occurrence of protists belonging to the Amebozoa and Cercozoa. Correlation network analysis showed that microbial communities are non-randomly structured, showing strikingly distinct composition in relation to litter chemistry. Our data demonstrate that the importance of litter chemistry in shaping microbial community structure increased during the decomposition process, being of little importance for freshly fallen leaves.

Essential oils (EOs) are one of the most important groups of plant metabolites responsible for their biological activities. This study was carried out to study the chemical composition and the antimicrobial effects of Artemisia herba-alba and Origanum majorana essential oils against some Gram-positive and Gram-negative bacteria, and a fungal strain isolated from spoiled butter. The plants were collected in the region Azzemour of South West Morocco and the EOs, extracted by hydrodistillation, were analyzed by GC-MS. The antimicrobial activity was determined using the agar paper disc method. The main components of A. herba-alba EO were cis-thujone, trans-thujone and vanillyl alcohol, in O. majorana EO terpinen-4-ol, isopulegol and &beta, phellandrene predominated. Both essential oils exhibited growth inhibiting activities in a concentration-dependent manner on several microorganism species. Our results demonstrated that O. majorana and A. herba-alba EOs could be effective natural antibacterial agents in foods.

Microbial spoilage of raw meat causes huge economic losses every year. An understanding of the microbial ecology associated with the spoilage and its dynamics during the refrigerated storage of meat can help in preventing and delaying the spoilage-related activities. The raw meat microbiota is usually complex, but only a few members will develop during storage and cause spoilage upon the pressure from several external factors, such as temperature and oxygen availability. We characterized the metagenome of beef packed aerobically or under vacuum during refrigerated storage to explore how different packaging conditions may influence the microbial composition and potential spoilage-associated activities. Different population dynamics and spoilage-associated genomic repertoires occurred in beef stored aerobically or in vacuum packaging. Moreover, the pangenomes of Pseudomonas fragi strains were extracted from metagenomes. We demonstrated the presence of specific, storage-driven strain-level profiles of Pseudomonas fragi, characterized by different gene repertoires and thus potentially able to act differently during meat spoilage. The results provide new knowledge on strain-level microbial ecology associated with meat spoilage and may be of value for future strategies of spoilage prevention and food waste reduction. IMPORTANCE This work provides insights on the mechanisms involved in raw beef spoilage during refrigerated storage and on the selective pressure exerted by the packaging conditions. We highlighted the presence of different microbial metagenomes during the spoilage of beef packaged aerobically or under vacuum. The packaging condition was able to select specific Pseudomonas fragi strains with distinctive genomic repertoires. This study may help in deciphering the behavior of different biomes directly in situ in food and in understanding the specific contribution of different strains to food spoilage.

Bacillus thuringiensis is a widely used bacterial entomopathogen producing insecticidal toxins, some of which are expressed in insect-resistant transgenic crops. Surprisingly, the killing mechanism of B. thuringiensis remains controversial. In particular, the importance of the septicemia induced by the host midgut microbiota is still debated as a result of the lack of experimental evidence obtained without drastic manipulation of the midgut and its content. Here this key issue is addressed by RNAi-mediated silencing of an immune gene in a lepidopteran host Spodoptera littoralis, leaving the midgut microbiota unaltered. The resulting cellular immunosuppression was characterized by a reduced nodulation response, which was associated with a significant enhancement of host larvae mortality triggered by B. thuringiensis and a Cry toxin. This was determined by an uncontrolled proliferation of midgut bacteria, after entering the body cavity through toxin-induced epithelial lesions. Consequently, the hemolymphatic microbiota dramatically changed upon treatment with Cry1Ca toxin, showing a remarkable predominance of Serratia and Clostridium species, which switched from asymptomatic gut symbionts to hemocoelic pathogens. These experimental results demonstrate the important contribution of host enteric flora in B. thuringiensis-killing activity and provide a sound foundation for developing new insect control strategies aimed at enhancing the impact of biocontrol agents by reducing the immunocompetence of the host.

Nisin is an antimicrobial peptide widely used as natural food preservative. Unfortunately, some factors lead to a reduction in its antimicrobial activity in food matrices. In this work nisin was microencapsulated in alginate matrix by vibrating technology with a high efficiency (75%) and microcapsules were characterized by having homogeneity in shape and surface morphology. Antimicrobial activity of microcapsules was evaluated against Brochothrix thermosphacta 7R1 immediately after microencapsulation process and during storage (24, 48, 72, 120, 144 and 168 h) under different conditions (i.e. 4 and 20 °C, pH 2.5, 4.5 and 6.0). We proposed a new method to evaluate the residual activity of encapsulated antimicrobial substances measured in Active Microcapsule Units per ml (AMU/ml). The test revealed that during a storage at 4 °C and pH 6.0, microcapsules retained their antimicrobial activity better than under other tested storage conditions. Furthermore, microcapsules efficiently protected nisin from the activity of protease. Resting cell experiments for the determination of antimicrobial activity of microcapsules during 120 h of storage under several conditions indicated greater activity at pH 6.0 with stirring. Nisin release from microcapsules determined by HPLC analysis showed that no trace of nisin was detected in the storage solution.

Beef burgers were stored at 4°C in a vacuum in nisin-activated antimicrobial packaging. Microbial ecology analyses were performed on samples collected between days 0 and 21 of storage to discover the population diversity. Two batches were analyzed using RNA-based denaturing gradient gel electrophoresis (DGGE) and pyrosequencing. The active packaging retarded the growth of the total viable bacteria and lactic acid bacteria. Culture-independent analysis by pyrosequencing of RNA extracted directly from meat showed that Photobacterium phosphoreum , Lactococcus piscium , Lactobacillus sakei , and Leuconostoc carnosum were the major operational taxonomic units (OTUs) shared between control and treated samples. Beta diversity analysis of the 16S rRNA sequence data and RNA-DGGE showed a clear separation between two batches based on the microbiota. Control samples from batch B showed a significant high abundance of some taxa sensitive to nisin, such as Kocuria rhizophila , Staphylococcus xylosus , Leuconostoc carnosum , and Carnobacterium divergens , compared to control samples from batch A. However, only from batch B was it possible to find a significant difference between controls and treated samples during storage due to the active packaging. Predicted metagenomes confirmed differences between the two batches and indicated that the use of nisin-based antimicrobial packaging can determine a reduction in the abundance of specific metabolic pathways related to spoilage. The present study aimed to assess the viable bacterial communities in beef burgers stored in nisin-based antimicrobial packaging, and it highlights the efficacy of this strategy to prolong beef burger shelf life.

The application of high hydrostatic pressure (HHP, 600 MPa, 8 min) on brined green asparagus and the changes in bacterial diversity after treatments and during storage at 4 °C (30 days) or 22 °C (10 days) were studied. HHP treatments reduced viable cell counts by 3.6 log cycles. The residual surviving population did not increase during storage at 4 °C. However, bacterial counts significantly increased at 22 °C by day 3, leading to rapid spoilage. The microbiota of green asparagus was composed mainly by Proteobacteria (mainly Pantoea and Pseudomonas), followed by Firmicutes (mainly Lactococcus and Enterococcus) and to a less extent Bacteroidetes and Actinobacteria. During chill storage of untreated asparagus, the relative abundance of Proteobacteria as well as Enterococcus and Lactococcus decreased while Lactobacillus increased. During storage of untreated asparagus at 22 °C, the abundance of Bacteroidetes decreased while Proteobacteria increased during late storage. The HHP treatment determined a reduction of the Proteobacteria both early after treatment and during chill storage. In the HHP treated samples stored at 22 °C, the relative abundance of Pseudomonas rapidly decreased at day 1, with an increase of Bacteroidetes. This was followed by a marked increase in Enterobacteriaceae (Escherichia) simultaneously with increase in viable counts and spoilage. Results from the study indicate that the effect of HHP treatments on the viability of microbial populations in foods also has an impact on the dynamics of microbial populations during the storage of the treated foods.

In Italy, fermented sausages (called “salami”) are consumed in large quantities. Salami samples from a local meat factory in the area of Torino were analyzed at 0, 3, 7, 30 and 45 days of ripening. Swab samples from the production environment were also collected at the beginning of the experiment. The diversity of metabolically active microbiota occurring during the natural fermentation of salami was evaluated by using RT-PCR-DGGE coupled with RNA-based pyrosequencing of the 16S rRNA gene. A culture-dependent approach was also applied to identify and characterize isolated Staphylococcaceae and LAB populations. Staphylococcus succinus , Staphylococcus xylosus and Lactobacillus sakei were the species most frequently isolated during the maturation time. Rep-PCR analysis showed that S . succinus and S . xylosus isolated from swabs and salami samples clustered together, suggesting possible contamination during the production process. RT-PCR-DGGE and rRNA-based pyrosequencing showed that the metabolically active populations were dominated by S . succinus , Lb . sakei and Leuconostoc carnosum . In this specific case study, only a few species belonging to Staphylococcaceae, Lactobacillaceae and Leuconostocaceae may be metabolically active and contribute to determine the final characteristics of the products.

BackgroundCystic fibrosis (CF) is a disorder affecting the respiratory, digestive, reproductive systems and sweat glands. This lethal hereditary disease has known or suspected links to the dysbiosis gut microbiota. High-throughput meta-omics-based approaches may assist in unveiling this complex network of symbiosis modifications.ObjectivesThe aim of this study was to provide a predictive and functional model of the gut microbiota enterophenotype of pediatric patients affected by CF under clinical stability.MethodsThirty-one fecal samples were collected from CF patients and healthy children (HC) (age range, 1-6 years) and analysed using targeted-metagenomics and metabolomics to characterize the ecology and metabolism of CF-linked gut microbiota. The multidimensional data were low fused and processed by chemometric classification analysis.ResultsThe fused metagenomics and metabolomics based gut microbiota profile was characterized by a high abundance of Propionibacterium, Staphylococcus and Clostridiaceae, including Clostridium difficile, and a low abundance of Eggerthella, Eubacterium, Ruminococcus, Dorea, Faecalibacterium prausnitzii, and Lachnospiraceae, associated with overexpression of 4-aminobutyrate (GABA), choline, ethanol, propylbutyrate, and pyridine and low levels of sarcosine, 4-methylphenol, uracil, glucose, acetate, phenol, benzaldehyde, and methylacetate. The CF gut microbiota pattern revealed an enterophenotype intrinsically linked to disease, regardless of age, and with dysbiosis uninduced by reduced pancreatic function and only partially related to oral antibiotic administration or lung colonization/infection.ConclusionsAll together, the results obtained suggest that the gut microbiota enterophenotypes of CF, together with endogenous and bacterial CF biomarkers, are direct expression of functional alterations at the intestinal level. Hence, it's possible to infer that CFTR impairment causes the gut ecosystem imbalance.This new understanding of CF host-gut microbiota interactions may be helpful to rationalize novel clinical interventions to improve the affected children's nutritional status and intestinal function.

Lactobacillus curvatus 54M16 produced bacteriocins sak X, sak Tα, sak Tβ and sak P. The aim of this study was to investigate the anti-listerial activity of the bacteriocins-producing strain against Listeria monocytogenes in vitro co-culture experiments and during the manufacture of fermented sausages. In MRS broth, Lb. curvatus 54M16 was able to inhibit L. monocytogenes to undetectable levels after 48 h at 20 °C or 5 days at 15 °C. Anti-listerial activity was lower during the production of fermented sausages with pathogen inoculation at levels of approximately 4 Log CFU g-1. However, total inhibition of L. monocytogenes native to the raw ingredients was achieved over the course of the fermentation. Moreover, 16S rRNA-based analysis revealed the ability of Lb. curvatus 54M16 to dominate and affect the bacterial ecosystem, whereas spoilage-associated bacterial genera, such as Brochothrix, Psychrobacter, Pseudomonas and some Enterobacteriaceae, were found until the end of ripening in sausages without Lb. curvatus 54M16. The use of the bacteriocins-producing Lb. curvatus 54M16 in fermented sausages could be an important contribution to product safety, provided that eco-physiological factors and other preservation methods are maintained at levels required for the inhibition of pathogens in controlled conditions.

Microbial contamination in foodservice environments plays a fundamental role in food quality and safety. In such environments the composition of the microbiota is influenced by the characteristics of the specific surfaces and by food handling and processing and a resident microbiota may be present in each site. In this study, the bacterial biogeographical patterns in a hospital cooking center was studied by 16S rRNA-based culture-independent high-throughput amplicon sequencing in order to provide a comprehensive mapping of the surfaces and tools that come in contact with foods during preparation. Across all area, surface swab-samples from work surfaces of different zones were taken: food pre-processing rooms (dedicated to fish, vegetables, and red and white meat), storage room and kitchen. The microbiota of environmental swabs was very complex, including more than 500 operational taxonomic units (OTUs) with extremely variable relative abundances (0.02–99%) depending on the species. A core microbiota was found that was common to more than 70% of the samples analyzed and that included microbial species that were common across all areas such as Acinetobacter , Chryseobacterium , Moraxellaceae, and Alicyclobacillus , although their abundances were below 10% of the microbiota. Some surfaces were contaminated by high levels of either Pseudomonas , Psychrobacter , Paracoccus , or Kocuria . However, beta diversity analysis showed that, based on the composition of the microbiota, the environmental samples grouped according to the sampling time but not according to the specific area of sampling except for the case of samples from the vegetable pre-processing room that showed a higher level of similarity. The cleaning procedures can have a very strong impact on the spatial distribution of the microbial communities, as the use of the same cleaning tools can be even a possible vector of bacterial diffusion. Most of the microbial taxa found are not those commonly found in food as spoilers or hazardous bacteria, which indicates that food and storage conditions can be very selective in the growth of possible contaminants.

Flower pollen is collected by honeybee foragers, adhered on their rear legs and transported into the hives in the form of pellets. Once in the hives, bee pollen is moisturised with nectar and bee mouth secretions and due to enzymatically modifications it becomes the so-called bee-bread, the protein reservoir of young bees. Bee pollen can be artificially removed from bee legs and collected by using specific systems, the bee pollen traps. Bee pollen is commercialized for human consumption as fresh product and after freezing or drying. Although bee pollen is nowadays largely consumed in developed countries, as food or food supplement according to local legislation, little is known on its safety related to microbiological hazards. In this work, we aimed to characterize for the first time the microbiological profile of Italian bee pollen in fresh, frozen and dried form collected along an entire harvesting season. Moreover, monthly microbiological analyses were performed on frozen (storage at -18°C) and dried (storage at room temperature) bee pollen over a 4 months period. Further aim of this work was the evaluation of the possible impact on production level of three different traps used for pollen collection. Our results on microbial contamination of fresh and frozen bee pollen show that a more comprehensive microbiological risk assessment of bee pollen is required. On the other side, dried pollen showed very low microbial contamination and no pathogen survived after the drying process and during storage.

Vegetable cultivation under plastic tunnels provides high-quality crop yields but negatively affects soil fertility, especially when conventional agricultural system (e.g., use of synthetic mineral fertilizer and agrochemicals) was adopted. In this regard, the use of organic amendments has been proposed as a reliable and effective approach for soil fertility recovery. The aim of this study was to assess the effects of different organic amendments and application frequency on crop yield, soil fertility and soil microbial communities. A 1-year long mesocosm experiment was performed by conditioning a soil with different organic amendment types and frequency of application, compared with the ordinary soil management based on mineral fertilizers and fumigation. Soil fertility was assessed by determining physical and chemical soil parameters, whereas microbial community functioning and structure were assessed by high-throughput sequencing of bacterial and eukaryotic rRNA gene markers and BIOLOG EcoPlates™. Compared to the organic amendment, the use of synthetic fertilizer had a higher crop production but negatively affected pH, soil organic carbon content and soil aggregation. Diversity and richness of bacteria and eukaryotic were lower in the synthetic than in the organic amendments. The addition of organic amendments promoted the growth of Acidobacteria and Gemmatimonadetes bacteria. On the contrary, members of Actinobacteria and Proteobacteria were more abundant in the soil treated with synthetic fertilizer. This study increases our current knowledge on the effect of the synthetic and organic amendment applications on crop yield, soil fertility and soil microbial community functionality.

Fontina Protected Denomination of Origin (PDO) cheese is a full-fat semi-cooked cheese traditionally made in Northwest Italy (Aosta Valley) and manufactured from raw cow's milk. The management of cattle farms in Aosta Valley calls for seasonal migration to high pastures during the summer and the concentration of calving during the autumn and the beginning of the winter. Based on cattle physiology and given to calving seasonality, three cow lactation phases i.e. post-partum , oestrus and early gestation, can be identified and an effect could be hypothesized on average milk composition and on cheese quality. The aim of the present paper was to investigate the bacterial dynamics during Fontina PDO cheese manufacturing and ripening, in relation to the different lactation stages, in order to evaluate a possible correlation between microbiota and phase of lactation. For this purpose, microbial RNA analysis was carried out by RT-PCR coupled with DGGE and high-throughput sequencing. A good performance of the starter cultures was highlighted throughout Fontina PDO manufacturing and ripening; in fact, the starter prevailed against the autochthonous microbiota. Thus, the microbial activity, which was supposed to affect the final quality of Fontina PDO cheese, appeared to be strictly associated to the presence of the starter, which did not show any difference in its performance according to the different stages of cow lactation. Therefore, the results of this research highlighted a negligible correlation between the microbiota of raw milk and the organoleptic quality and typicity of Fontina cheese in relation to lactation seasonality.

Spontaneous alcoholic fermentation of grape must is a complex process, carried out by indigenous yeast populations arising from the vineyard or the winery environment and therefore representing an autochthonous microbial terroir of the production area. Microbial diversity at species and biotype level is extremely important in order to develop the composite and typical flavour profile of DOCG (Appellation of Controlled and Guaranteed Origin) wines. In this study, we monitored fungal populations involved in spontaneous fermentations of Aglianico and Greco di Tufo grape must by high-throughput sequencing (HTS) of 18S rRNA gene amplicons. We firstly proposed an alternative/addition to ITS as target gene in HTS studies and highlighted consistency between the culture-dependent and –independent approaches. A complex mycobiota was found at the beginning of the fermentation, mainly characterized by non-Saccharomyces yeasts and several moulds, with differences between the two types of grapes. Moreover, Interdelta patterns revealed a succession of several Saccharomyces cerevisiae biotypes and a high genetic diversity within this species.

Mozzarella (M), Grana Padano (GP) and Parmigiano Reggiano (PR) are three of the most important traditional Italian cheeses. In the three cheese manufactures the initial fermentation is carried out by adding natural whey cultures (NWCs) according to a back-slopping procedure. In this study, NWCs and the corresponding curds from M, GP and PR manufactures were analyzed by culture-independent pyrosequencing of the amplified V1–V3 regions of the 16S rRNA gene, in order to provide insights into the microbiota involved in the curd acidification. Moreover, culture-independent high-throughput sequencing of lacS gene amplicons was carried out to evaluate the biodiversity occurring within the S. thermophilus species. Beta diversity analysis showed a species-based differentiation between GP-PR and M manufactures indicating differences between the preparations. Nevertheless, all the samples shared a naturally-selected core microbiome, that is involved in the curd acidification. Type-level variability within S. thermophilus species was also found and twenty-eight lacS gene sequence types were identified. Although lacS gene did not prove variable enough within S. thermophilus species to be used for quantitative biotype monitoring, the possibility of using non rRNA targets for quantitative biotype identification in food was highlighted.

The salivary microbiota has been linked to both oral and non-oral diseases. Scant knowledge is available on the effect of environmental factors such as long-term dietary choices on the salivary microbiota and metabolome. This study analyzed the microbial diversity and metabolomic profiles of the saliva of 161 healthy individuals who followed an omnivore or ovo-lacto-vegetarian or vegan diet. A large core microbiota was identified, including 12 bacterial genera, found in >98% of the individuals. The subjects could be stratified into three "salivary types" that differed on the basis of the relative abundance of the core genera Prevotella, Streptococcus/Gemella and Fusobacterium/Neisseria. Statistical analysis indicated no effect of dietary habit on the salivary microbiota. Phylogenetic beta-diversity analysis consistently showed no differences between omnivore, ovo-lacto-vegetarian and vegan individuals. Metabolomic profiling of saliva using (1)H-NMR and GC-MS/SPME identified diet-related biomarkers that enabled a significant discrimination between the 3 groups of individuals on the basis of their diet. Formate, urea, uridine and 5-methyl-3-hexanone could discriminate samples from omnivores, whereas 1-propanol, hexanoic acid and proline were characteristic of non-omnivore diets. Although the salivary metabolome can be discriminating for diet, the microbiota has a remarkable inter-individual stability and did not vary with dietary habits. Microbial homeostasis might be perturbed with sub-standard oral hygiene or other environmental factors, but there is no current indication that a choice of an omnivore, ovo-lacto-vegetarian or vegan diet can lead to a specific composition of the oral microbiota with consequences on the oral homeostasis.

Traditional Caciocavallo of Castelfranco is a semi-hard "pasta-filata" cheese produced from raw cows' milk in Campania region. The aim of the present research is mainly focused on the study, by 16S rRNA gene pyrosequencing and viable counts, of the dynamics of bacterial communities during manufacture and ripening of traditional Caciocavallo cheese. Moreover, the possible correlation between cheese microbiota and cows' feeding based on silage or hay was also evaluated. In general, except for enterococci, the technological process significantly affected all the microbial groups. According to 16S rRNA, raw cows' milk was dominated by Streptococcus thermophilus, L. lactis and Pseudomonas sp. in hay cheese production, whereas Lactococcus lactis and Acinetobacter sp. dominated silage production. Differences in the taxonomic structure of the milk's microbiota within diet groups were not related to silage and hay cows' feeding. Moreover, S. thermophilus was the unique species that dominate from raw milks to fermented intermediates and cheese in both hay and silage cheese productions. Feeding and ripening time influenced significantly sensory characteristics of the cheeses.

Microbial contamination in food processing plants can play a fundamental role in food quality and safety. The aims of this study were to learn more about the possible influence of the meat processing environment on initial fresh meat contamination and to investigate the differences between small-scale retail distribution (SD) and large-scale retail distribution (LD) facilities. Samples were collected from butcheries ( n = 20), including LD ( n = 10) and SD ( n = 10) facilities, over two sampling campaigns. Samples included fresh beef and pork cuts and swab samples from the knife, the chopping board, and the butcher's hand. The microbiota of both meat samples and environmental swabs were very complex, including more than 800 operational taxonomic units (OTUs) collapsed at the species level. The 16S rRNA sequencing analysis showed that core microbiota were shared by 80% of the samples and included Pseudomonas spp., Streptococcus spp., Brochothrix spp., Psychrobacter spp., and Acinetobacter spp. Hierarchical clustering of the samples based on the microbiota showed a certain separation between meat and environmental samples, with higher levels of Proteobacteria in meat. In particular, levels of Pseudomonas and several Enterobacteriaceae members were significantly higher in meat samples, while Brochothrix , Staphylococcus , lactic acid bacteria, and Psychrobacter prevailed in environmental swab samples. Consistent clustering was also observed when metabolic activities were considered by predictive metagenomic analysis of the samples. An increase in carbohydrate metabolism was predicted for the environmental swabs and was consistently linked to Firmicutes , while increases in pathways related to amino acid and lipid metabolism were predicted for the meat samples and were positively correlated with Proteobacteria . Our results highlighted the importance of the processing environment in contributing to the initial microbial levels of meat and clearly showed that the type of retail facility (LD or SD) did not apparently affect the contamination. IMPORTANCE The study provides an in-depth description of the microbiota of meat and meat processing environments. It highlights the importance of the environment as a contamination source of spoilage bacteria, and it shows that the size of the retail facility does not affect the level and type of contamination.

Cultivation of vegetables under plastic tunnels is a steadily growing farming system in the agricultural sector but raises concerns about its environmental sustainability. The aim of the present work was to assess the impact of organic farming, compared to conventional cultivation, on agro-ecosystem functions and soil microbial communities. Two farms that practiced organic cultivation for 10 and 20 years were compared with one conventional farm, all with cultivations under plastic tunnel. Soil functions were assessed with multi-species bioassays on plant growth and organic matter decomposition, and microbial communities were characterized by high-throughput sequencing of bacterial and eukaryotic rRNA gene markers. Plant growth and organic matter decomposition were higher in organic compared to conventionally management soils. Agronomical practices showed a significant effect on microbial diversity and composition. Soil bacterial diversity was lower in both organic farms than in the conventional farm. Soil eukaryotic diversity was slightly higher in the 10-year organic farm but lower in the 20-year organic farm compared to the conventional farm. At phylum level, Acidobacteria, Firmicutes, and TM7 were higher, while Planctomycetes, Verrucomicrobia, and Actinobacteria were lower in organic-farmed soils compared to conventional farm. Noteworthy, Metazoa raised from 0.1% of eukarya relative abundance in conventional-farmed to 20.9% in 20 year organic-farmed soil. Correlation analysis between microbial OTUs relative abundance and ecosystem functions suggest that eukarya play a major role, compared to bacteria, in controlling plant growth and organic carbon cycling. Integration of microbial diversity data with plant growth and decomposition of organic matter decomposition allowed us to provide a linkage between agricultural management, microbial community composition, and soil functionality.

Objectives Habitual diet plays a major role in shaping the composition of the gut microbiota, and also determines the repertoire of microbial metabolites that can influence the host. The typical Western diet corresponds to that of an omnivore; however, the Mediterranean diet (MD), common in the Western Mediterranean culture, is to date a nutritionally recommended dietary pattern that includes high-level consumption of cereals, fruit, vegetables and legumes. To investigate the potential benefits of the MD in this cross-sectional survey, we assessed the gut microbiota and metabolome in a cohort of Italian individuals in relation to their habitual diets. Design and results We retrieved daily dietary information and assessed gut microbiota and metabolome in 153 individuals habitually following omnivore, vegetarian or vegan diets. The majority of vegan and vegetarian subjects and 30% of omnivore subjects had a high adherence to the MD. We were able to stratify individuals according to both diet type and adherence to the MD on the basis of their dietary patterns and associated microbiota. We detected significant associations between consumption of vegetable-based diets and increased levels of faecal short-chain fatty acids, Prevotella and some fibre-degrading Firmicutes, whose role in human gut warrants further research. Conversely, we detected higher urinary trimethylamine oxide levels in individuals with lower adherence to the MD. Conclusions High-level consumption of plant foodstuffs consistent with an MD is associated with beneficial microbiome-related metabolomic profiles in subjects ostensibly consuming a Western diet. Trial registration number This study was registered at clinical trials.gov as NCT02118857.

An antimicrobial polyethylene (PE) film was obtained by coating a nisin-based antimicrobial solution. PE sheets were coated on both sides and were used for the packaging of beefsteaks to be stored in air or modified atmosphere packaging (MAP, 60% O₂-40% CO₂). Microbial populations, species diversity, headspace volatile organic compounds, colour and sensory properties were monitored after 0, 1, 7 and 12 days of storage at 4 °C. The viable counts showed that there was an effect of MAP and antimicrobial film on the development of all the spoilage associated microbial populations. Carnobacterium spp., Brochothrix thermosphacta, Pseudomonas fragi and Rhanella aquatilis were found in most of the samples. C. maltaromaticum was identified in MRS bulk cells from samples stored in air as well as MAP. Quantitative data of headspace-SPME-GC/MS analysis showed that during storage the production of volatile organic compounds (VOCs) was affected by the use of the treated film and the MAP storage. Compounds such as phenylethylalcohol, nonanal, decanal and ethylbutanoate were produced only from 7 to 12 day of storage and only in the samples stored in air. In agreement with the microbiological and VOCs data, the meat stored in active packaging scored the best rankings in the sensory evaluation. Principal component analysis of microbial, sensory and instrumental data showed that beefsteaks stored with the combination of MAP and active packaging for 12 days at 4 °C differed from the other samples that were more associated to high microbial loads, VOCs concentration and meat off odour perception. In conclusion, the antimicrobial sheets in combination with MAP storage at 4 °C were effective for the storage of beefsteaks by retarding the growth of spoilage bacteria, determining lower concentration of VOCs and keeping acceptable levels of colour and other sensory parameters for more than 10 days.

Antimicrobial packaging can be considered an extremely challenging technology that could have a significant impact on shelf-life extension and food safety of fresh meat and meat products. In this study, different commercial polyethylene films differing in vinyl acetate ethylene, erucamide contents, and oxygen permeability were used for the coating treatment with a nisin-based antimicrobial solution (NS). Detection and measurement of the activity of the NS was determined against different food spoilage bacteria. NS was then spread manually on food contact layer of different plastic films using coating rods providing thickness of 6, 40, 60, and 100 μm. The polyethylene films before and after treatment were analysed by atomic force microscopy (AFM). NS was active against Gram-positive bacteria and the best activity was obtained against Brochothrix thermosphacta. Viable staining and epifluorescence microscopy analysis of indicator strains in contact with activated plastic films showed that the effect of the film on the various indicator strains changed very much on the basis of both type of film and indicator strain. The highest numbers of lysed cells were shown by two polyethylene films that, according to the AFM and roughness parameters analyses, were characterized by significant increase or decrease of roughness after the coating treatment. AFM analysis showed that the homogeneity of the coating was much influenced by the type of plastic films used. In order to test the efficacy in food, portions of beef chuck tender slices were prepared and covered with the antimicrobial plastic films on both sides. After 1 h and 1, 7, and 12 days of storage at 4 °C the meat samples were analyzed by standard plate counting targeting spoilage associated microbial populations. The antimicrobial plastic films after 1 h of contact with the meat caused a significant reduction of lactic acid bacteria and B. thermosphacta. The most effective antimicrobial activity of films was shown against the same populations after 24 h of storage.

Background: Epidemiology associates whole-grain (WG) consumption with several health benefits. Mounting evidence suggests that WG wheat polyphenols play a role in mechanisms underlying health benefits. Objective: The objective was to assess circulating concentration, excretion, and the physiologic role of WG wheat polyphenols in subjects with suboptimal dietary and lifestyle behaviors. Design: A placebo-controlled, parallel-group randomized trial with 80 healthy overweight/obese subjects with low intake of fruits and vegetables and sedentary lifestyle was performed. Participants replaced precise portions of refined wheat (RW) with a fixed amount of selected WG wheat or RW products for 8 wk. At baseline and every 4 wk, blood, urine, feces, and anthropometric and body composition measures were collected. Profiles of phenolic acids in biological samples, plasma markers of metabolic disease and inflammation, and fecal microbiota composition were assessed. Results: WG consumption for 4‐8 wk determined a 4-fold increase of serum dihydroferulic acid (DHFA) and a 2-fold increase of fecal ferulic acid (FA) compared with RW consumption (no changes). Similarly, urinary FA at 8 wk doubled the baseline concentration only in WG subjects. Concomitant reduction of plasma tumor necrosis factor-a (TNF-a) after 8 wk and increased interleukin (IL)-10 only after 4 wk with WG compared with RW (P = 0.04) were observed. No significant change in plasma metabolic disease markers over the study period was observed, but a trend toward lower plasma plasminogen activator inhibitor 1 with higher excretion of FA and DHFA in the WG group was found. Fecal FA was associated with baseline low Bifidobacteriales and Bacteroidetes abundances, whereas after WG consumption, it correlated with increased Bacteroidetes and Firmicutes but reduced Clostridium. TNF-a reduction correlated with increased Bacteroides and Lactobacillus. No effect of dietary interventions on anthropometry and body composition was found. Conclusions: WG wheat consumption significantly increased excreted FA and circulating DHFA. Bacterial communities influenced fecal FA and were modified by WG wheat consumption. This trial was registered at clinicaltrials.gov as NCT01293175. Am J Clin Nutr doi: 10.3945/ajcn.114.088120.

Aims: To evaluate the effect of a bacteriocin-activated polythene film on resting and growing populations of Listeria monocytogenes. Methods and Results: The active polythene films were industrially obtained by coating a solution of bacteriocin 32Y from Lactobacillus curvatus upon the surface of the film to be in contact with the packaged material. The behaviour of live Listeria populations was examined in liquid suspensions directly in contact with the bacteriocin-activated film, packed in antimicrobial film, and in a challenge test of storage of frankfurters superficially contaminated by L. monocytogenes and packed in antimicrobial film. In all the experiments, live and dead cells of L. monocytogenes were counted in epifluorescence microscopy after viable staining, which proved to be a suitable method to evaluate the action of bacteriocins on populations of L. monocytogenes. The results showed that the direct contact between active film surface and L. monocytogenes cells is effective for a fast and irreversible inactivation of the population by determining a direct cell disruption. This was confirmed by the results of the challenge test indicating that the antimicrobial package was effective in inhibiting the growth and survival of the pathogen on the surface of frankfurters during storage. Conclusions: The use of the antimicrobial film is encouraged especially for solid food products where the superficial contaminants come immediately in contact with the antimicrobial film. Significance and Impact of the Study: A fast inactivation of the bacterial population, coupled with appropriate conditions of storage, can improve the quality and safety and prolong the shelf-life of the food products packed in antimicrobial films.

The microbial composition of artisan and industrial animal rennet pastes was studied by using both culture-dependent and -independent approaches. Pyrosequencing targeting the 16S rRNA gene allowed to identify 361 operational taxonomic units (OTUs) to the genus/species level. Among lactic acid bacteria (LAB), Streptococcus thermophilus and some lactobacilli, mainly Lactobacillus crispatus and Lactobacillus reuteri , were the most abundant species, with differences among the samples. Twelve groups of microorganisms were targeted by viable plate counts revealing a dominance of mesophilic cocci. All rennets were able to acidify ultrahigh-temperature-processed (UHT) milk as shown by pH and total titratable acidity (TTA). Presumptive LAB isolated at the highest dilutions of acidified milks were phenotypically characterized, grouped, differentiated at the strain level by randomly amplified polymorphic DNA (RAPD)-PCR analysis, and subjected to 16S rRNA gene sequencing. Only 18 strains were clearly identified at the species level, as Enterococcus casseliflavus , Enterococcus faecium , Enterococcus faecalis , Enterococcus lactis , Lactobacillus delbrueckii , and Streptococcus thermophilus , while the other strains, all belonging to the genus Enterococcus , could not be allotted into any previously described species. The phylogenetic analysis showed that these strains might represent different unknown species. All strains were evaluated for their dairy technological performances. All isolates produced diacetyl, and 10 of them produced a rapid pH drop in milk, but only 3 isolates were also autolytic. This work showed that animal rennet pastes can be sources of LAB, mainly enterococci, that might contribute to the microbial diversity associated with dairy productions.

Pyrosequencing of the 16S rRNA gene, community-level physiological profiles determined by the use of Biolog EcoPlates, and proteolysis analyses were used to characterize Canestrato Pugliese Protected Designation of Origin (PDO) cheese. The number of presumptive mesophilic lactococci in raw ewes' milk was higher than that of presumptive mesophilic lactobacilli. The numbers of these microbial groups increased during ripening, showing temporal and numerical differences. Urea-PAGE showed limited primary proteolysis, whereas the analysis of the pH 4.6-soluble fraction of the cheese revealed that secondary proteolysis increased mainly from 45 to 75 days of ripening. This agreed with the concentration of free amino acids. Raw ewes' milk was contaminated by several bacterial phyla: Proteobacteria (68%; mainly Pseudomonas ), Firmicutes (30%; mainly Carnobacterium and Lactococcus ), Bacteroidetes (0.05%), and Actinobacteria (0.02%). Almost the same microbial composition persisted in the curd after molding. From day 1 of ripening onwards, the phylum Firmicutes dominated. Lactococcus dominated throughout ripening, and most of the Lactobacillus species appeared only at 7 or 15 days. At 90 days, Lactococcus (87.2%), Lactobacillus (4.8%; mainly Lactobacillus plantarum and Lactobacillus sakei ), and Leuconostoc (3.9%) dominated. The relative utilization of carbon sources by the bacterial community reflected the succession. This study identified strategic phases that characterized the manufacture and ripening of Canestrato Pugliese cheese and established a causal relationship between mesophilic lactobacilli and proteolysis.

The bacterial ecology during rye and wheat sourdough preparation was described by 16S rRNA gene pyrosequencing. Viable plate counts of presumptive lactic acid bacteria, the ratio between lactic acid bacteria and yeasts, the rate of acidification, a permutation analysis based on biochemical and microbial features, the number of operational taxonomic units (OTUs), and diversity indices all together demonstrated the maturity of the sourdoughs during 5 to 7 days of propagation. Flours were mainly contaminated by metabolically active genera ( Acinetobacter , Pantoea , Pseudomonas , Comamonas , Enterobacter , Erwinia , and Sphingomonas ) belonging to the phylum Proteobacteria or Bacteroidetes (genus Chryseobacterium ). Their relative abundances varied with the flour. Soon after 1 day of propagation, this population was almost completely inhibited except for the Enterobacteriaceae . Although members of the phylum Firmicutes were present at very low or intermediate relative abundances in the flours, they became dominant soon after 1 day of propagation. Lactic acid bacteria were almost exclusively representative of the Firmicutes by this time. Weissella spp. were already dominant in rye flour and stably persisted, though they were later flanked by the Lactobacillus sakei group. There was a succession of species during 10 days of propagation of wheat sourdoughs. The fluctuation between dominating and subdominating populations of L. sakei group, Leuconostoc spp., Weissella spp., and Lactococcus lactis was demonstrated. Other subdominant species such as Lactobacillus plantarum were detectable throughout propagation. As shown by PCR-denaturing gradient gel electrophoresis (PCR-DGGE) analysis, Saccharomyces cerevisiae dominated throughout the sourdough propagation. Notwithstanding variations due to environmental and technology determinants, the results of this study represent a clear example of how the microbial ecology evolves during sourdough preparation.

A nisin-EDTA solution was used for activation of the internal surface of plastic bags that were used to store beef chops at 1°C after vacuum packaging. The aim of the work was to evaluate the effect of the antimicrobial packaging on beef during storage. Volatile compounds and microbial populations were monitored after 0, 9, 20, 36, and 46 days of storage. The active packaging retarded the growth of lactic acid bacteria. Brochothrix thermosphacta was unable to grow for the whole storage time in treated samples, while the levels of Carnobacterium spp. in treated samples were below the detection limit for the first 9 days and reached loads below 5 Log CFU/cm(2) after 46 days. On the other hand, Enterobacteriaceae and Pseudomonas spp. were not affected by the use of the antimicrobial packaging and grew in all of the samples, with final populations of about 4 Log CFU/cm(2). Carnobacterium divergens was identified by PCR-denaturing gradient gel electrophoresis analysis of DNA extracted from beef after 36 days of storage. During beef storage, alcohols, aldehydes, ketones, and carboxylic acids were detected in the headspace of beef samples by solid-phase microextraction-gas chromatography-mass spectrometry analysis. The microbial metabolic activity was affected by the use of the antimicrobial film from the beginning up to 36 days with a maximum in the differences of volatile metabolites in samples analyzed at 20 days. The volatiles were also determined by electronic nose, allowing differentiation based on the time of storage and not on the type of packaging. The active packaging reduces the loads of spoilage microbial populations and the release of metabolites in the headspace of beef with a probable positive impact on meat quality.

Intermediates of production of two batches of traditional mozzarella cheese were analyzed by culture-independent pyrosequencing. The quantitative distribution of taxa within the samples suggested that thermophilic lactic acid bacteria from the natural starter were mainly responsible for the fermentation, while microorganisms found in raw milk did not develop during fermentation.

Beef chops were stored at 4°C under different conditions: in air (A), modified-atmosphere packaging (MAP), vacuum packaging (V), or bacteriocin-activated antimicrobial packaging (AV). After 0 to 45 days of storage, analyses were performed to determine loads of spoilage microorganisms, microbial metabolites (by solid-phase microextraction [SPME]-gas chromatography [GC]-mass spectrometry [MS] and proton nuclear magnetic resonance [ 1 H NMR]), and microbial diversity (by PCR–denaturing gradient gel electrophoresis [DGGE] and pyrosequencing). The microbiological shelf life of meat increased with increasing selectivity of storage conditions. Culture-independent analysis by pyrosequencing of DNA extracted directly from meat showed that Brochothrix thermosphacta dominated during the early stages of storage in A and MAP, while Pseudomonas spp. took over during further storage in A. Many different bacteria, several of which are usually associated with soil rather than meat, were identified in V and AV; however, lactic acid bacteria (LAB) dominated during the late phases of storage, and Carnobacterium divergens was the most frequent microorganism in AV. Among the volatile metabolites, butanoic acid was associated with the growth of LAB under V and AV storage conditions, while acetoin was related to the other spoilage microbial groups and storage conditions. 1 H NMR analysis showed that storage in air was associated with decreases in lactate, glycogen, IMP, and ADP levels and with selective increases in levels of 3-methylindole, betaine, creatine, and other amino acids. The meat microbiota is significantly affected by storage conditions, and its changes during storage determine complex shifts in the metabolites produced, with a potential impact on meat quality.

The aim of this study was to assess the microbial populations causing the spoilage of chilled beef during storage and to evaluate the effect of the use of an antimicrobial packaging for the meat storage. A nisin activated antimicrobial packaging was developed by using a nisin, HCL and EDTA solution and used for the storage of beef cuts at 1 degrees C. The common spoilage related microbial groups were monitored during the storage of beef in activated and non activated plastic bags by using selective media. The use of the antimicrobial packaging caused an overall significant reduction of viable counts of Gram positive bacteria such as carnobacteria, lactic acid bacteria and Brochotrix thermosphacta whose development was inhibited for at least 11 days of storage compared to the control. Moreover, a 1-3 log cycles reduction of enterobacteria was also registered between 22 and 32 days of storage. The microbiota was assessed at species level by using Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analysis of 16S rRNA gene of DNA extracted directly from meat and from bulk cells from selective media plates and showed that the species occurring within the targeted microbial groups did not change according to storage conditions. In conclusion, the use of the nisin activated packaging reduced the number of spoilage populations but did not affect the species diversity. Improved antimicrobial packaging is needed, possibly coupled with vacuum storage, to possibly achieve a simultaneous inhibition of more spoilage microbial groups and to preserve the microbiological quality of beef during chilled storage.

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 (P

The objective of the work was to study the viability and antimicrobial activity of bacteriocin-producing lactic acid bacteria (LAB) incorporated into whey protein/inulin/gelatine (WP) edible films in presence or absence of nutrient (modified MRS broth). Moreover, the role of the cell on the film structure and properties has been investigated. The results of the work showed that WP-based films were able to ensure a high viability of the bacteriocin-producing strain L. curvatus 54M16 during 28 days of storage at 4 °C. The addition of nutrient in the film matrix slightly affected the viability of the cells, but it was critical for the antimicrobial activity of the films. Films in presence of nutrient showed a good antimicrobial activity against L. innocua C6 as in vitro system as on cooked ham. The presence of LAB has a significant effect on the structure of the film: it reduced the viscosity of the film forming solution and improved the elasticity and the percentage of elongation. Whereas, no effect was observed for water vapour transmission rate and solubility. Thus, WP-based films in presence of modified MRS broth can be used as effective delivery and carrier systems for lactic acid bacteria to develop bioactive edible film or coating with antimicrobial properties.