A method for isolating potential probiotic lactobacilli directly fromtraditional milk-based foods was developed. The novel digestion/enrichment protocol was set up taking care to minimize the protective effect of milk proteins and fats and was validated testing three commercial fermented milks containing well-known probiotic Lactobacillus strains. Only probiotic bacteria claimed in the label were isolated from two out of three commercial fermented milks. The application of the new protocol to 15 raw milk samples and 6 traditional fermented milk samples made it feasible to isolate 11 potential probiotic Lactobacillus strains belonging to Lactobacillus brevis, Lactobacillus fermentum, Lactobacillus gasseri, Lactobacillus johnsonii, Lactobacillus plantarum, Lactobacillus reuteri, and Lactobacillus vaginalis species. Even though further analyses need to ascertain functional properties of these lactobacilli, the novel protocol set-up makes it feasible to isolate quickly potential probiotic strains from traditional milk-based foods reducing the amount of time required by traditional procedures that, in addition, do not allow to isolate microorganisms occurring as subdominant populations.

The aim of this study was to test the antibacterial activity of Bacillus subtilis TR50 isolated from a traditional cured sausage against ten food-borne pathogenic bacteria. Most of the pathogens were sensitive to cell-free concentrated supernatant from TR50. The inhibitory activity was retained at pH values between 4 and 8, up to80 _C for 30 min and after treatment with pepsin and trypsin, whereas it disappeared after chymotrypsin hydrolysis; lipase treatment abolished inhibitory activity only against one target strain. The ammonium sulphate precipitation of TR50 concentrated cell-free culture retained the most residual antibacterial activity unlike the ethyl acetate and acid fractions. All these results suggest that the antimicrobial compound(s) produced by TR50 could be of proteinaceous nature and require a lipid moiety for activity against the pathogens tested. This is the first report on the antimicrobial activity against food-borne pathogens of a B. subtilis strain occurring in a traditional sausage potentially useful for food safety improvement.

The aim of this work was to verify the antimicrobial activity of propolis and green tea extracts against foodborne Staphylococcus aureus and Escherichia coli. Viability of the strains, assayed in broth against eight concentration of glycerol extracts ranging from 0 to 25%, was clearly affected only at concentration higher than 6%. Then, the 24h microbial growth kinetic for both pathogens was drawn for both extracts only at 25 and 12.5%. The values of inhibition index turbidity ratio [1] were found always higher for green tea than for propolis; in addition, the inhibition index turbidity ratio produced by green tea extract was found always higher than 90% for S. aureus strains whereas it was close to 70% for the E. coli strains. Then, both extracts were incorporated into inert absorbent polysaccharide hydrogels at 10 and 70% and assayed, following the ASTM 2180, only against the most resistant E. coli O157:H7 strain. After 24 hours of contact, the concentration of viable cells showed a slight reduction for both extracts without any statistical (P<0.05) significant difference. In conclusion, antimicrobial compounds occurring in natural extracts showed a loss of antimicrobial activity when included into hydrogels. The causes of this phenomenon, still under examination, could be ascribed to a reduction in the specific antimicrobial activity, or in the mobility, of different natural compounds. AcknowledgementThis work was carried out by MAIN spa and OR.MA. srl (Pisticci, Mt, Italy) within a research project funded by Italian Ministry of Economic Development within the project (DM 04/09/2013).References1. Baruzzi F, Pinto L, Quintieri L, Carito A, Calabrese N, Caputo L (2015) Efficacy of lactoferricin B in controlling ready-to-eat vegetable spoilage caused by Pseudomonas spp. International Journal of Food Microbiology 215: 179-186.

Several spices are used to process 'Nduja, a traditional Calabrian sausage, well knownfor its typical flavor and piquancy. Despite wide use of spices for meat curing andcontrolling undesired microorganisms, no studies reported antimicrobial effectivenessof the spices in the 'Nduja processing. The aim of this work was to evaluate theantimicrobial activities against 10 foodborne pathogens of processed spices used inthe 'Nduja industrial manufacturing. Spice extracts (using ethanol, methanol andwater as solvents) were investigated by disk diffusion assay for their antimicrobialactivity. Methanol extracts showed the best results, mainly against Bacillus cereusstrains. Thus, they were further evaluated, through microdilution automated spectrofluorimetricabsorbance readings. Many spice methanol extracts showed low antimicrobialactivities, although B. cereus strains were still more sensitive. In addition, twoout of the three spice mixtures showed medium and high inhibition activity againstEscherichia coli ATCC 35401 and Yersinia enterocolitica DSM 4780, respectively.

The aim of this work was to check the efficacy of bovine lactoferrin (BLF) and its pepsin-digested hydrolysate (LFH) to control spoilage bacteria contaminating the governing liquid of high moisture (HM) Mozzarella cheese during cold storage. These natural substances resulted effective when tested in vitro against five potential spoilage bacteria contaminating cold-stored HM Mozzarella cheese. Among six LFH fractions, only the fraction containing lactoferricins, mainly represented by LfcinB?????, resulted effective against Escherichia coli K12 at the same extent of the whole pepsin-digested hydrolysate. LFH tested throughout seven days for its antimicrobial activity against the main bacterial groups growing in cold-stored commercial HM Mozzarella cheese samples delayed significantly the growth of pseudomonads and coliforms in comparison with the un-treated samples. This is the first report providing a direct evidence of the ability of LFH to inhibit the growth of cheese spoilage bacteria.

Psychrotrophic Pseudomonas spp. were proved to cause dramatic quality losses in fresh foods like leafy greens and dairy products resulting in the shortening of their shelf-life. These spoilage microorganisms are resistant to most disinfection treatments. This work was addressed to assay the antimicrobial efficacy of the food-grade bovine lactoferrin (BLF) and its hydrolysates (LFH), obtained after digestion with pepsin or the enzymatic cardoon extract. The inhibition assays were carried out in vitro on selected Pseudomonas spp. strains involved in off-color development in ready-to-eat vegetables (RTE) and in mozzarella cheese. A higher antimicrobial activity against most of the tested strains was found for pepsin digested BLF in comparison with BLF digested with the enzymatic cardoon extract. A solution of the antimicrobial peptide lactoferricin B (LFcinB), purified from the pepsin-LFH, was applied to both cold stored RTE lettuce leaves and mozzarella cheeses, inoculated with the spoilage strains. A significant reduced tissue browning of RTE lettuce leaves, as well as absence of mozzarella cheese discoloration applying LFcinB. The results obtained show the possibility to use milk-derived antimicrobial peptides in the control of food spoilage caused by Pseudomonas strains.

Red chicory is a leafy vegetable, currently used for the preparation of ready-to-eat salads, that undergoes heavy losses during cold storage mainly due to microbial soft rot development. In this work, low doses of gaseous ozone (0.1-1 ppm) were applied during 21 days of cold storage (1°C) under passive cooling in order to reduce postharvest losses of this vegetable. Conventional refrigeration in absence of gaseous ozone was used as the reference storage method. In addition, in order to evaluate the antimicrobial effect, a challenge test was performed inoculating sound red chicory heads with a specific spoilage microbiota, accounting approximately 8 log cfu/mL bacteria and 7 log cfu/mL yeasts and moulds, responsible for soft rot development under cold storage. The results showed that soft rot developed from day 8, especially in the inoculated samples, even though the leaf appearance was unchanged. At the end of cold storage, non-inoculated red chicory heads stored under conventional conditions loosed all characteristics related to vegetable fresh appearance; on the contrary, color brightness and tissue turgidity were retained in samples stored under passive cooling. Oxidative discoloration of leaf margins was recorded in red chicory heads stored under 1 ppm ozone. At the day 21 microbial counts of spoiled vegetable, regardless the different cold storage conditions, did not show any significant difference. Conversely, as expected, evident difference in soft rot severity was recorded between inoculated and non-inoculated samples. Passive cooling determined a lower amount of food waste in comparison with conventional refrigeration. Control samples, inoculated or not, showed 100% of losses whereas those under passive cooling produced 40.09 ± 4.24 and 90.69 ± 12.03 % of food waste for inoculated and non-inoculated samples, respectively. A further reduction in the amount of vegetable waste (ranging from 10 to 30%) was registered when passive refrigeration was associated with gaseous ozone treatment at 0.1 ppm. Higher ozone doses caused no significant additional improvements. In conclusions, the combined application of gaseous ozone and passive cooling resulted a valid approach to counteract soft rot development on red chicory heads and to reduce the amount of food waste. However, ozone exposure need to be carefully calibrated to preserve vegetable tissues from oxidative damages caused by this gas.

Le pigmentazioni anomale di mozzarelle sono state associate a ceppi psicrotrofi di Pseudomonas fluorescens presenti a basse concentrazioni nel liquido di governo già nelle prime fasi di frigoconservazione di tale formaggio fresco. Il mancato controllo di questi batteri alterativi ne causa la crescita determinando il ritiro dal mercato dei prodotti alterati. Scopo di questo lavoro è stato valutare l'efficacia antimicrobica in vitro ed in mozzarella dell'idrolizzato peptico di lattoferrina bovina (HLF) contro P. fluorescens 84095 isolato da mozzarelle blu. I risultati hanno mostrato che la crescita in vitro a 4?C del ceppo è stata inibita da 50 mg/mL di HLF. La stessa concentrazione, applicata a mozzarelle inoculate, ha determinato una significativa riduzione delle conte di 84095 rispetto a quelle registrate in campioni senza HLF fino al 14 giorno di frigoconservazione. Il trattamento ha inoltre bloccato lo sviluppo della colorazione blu. Le analisi LC-MS hanno confermato la presenza della leucoindigoidina, forma ridotta ed incolore del pigmento blu indigoidina solo nei campioni di mozzarella inoculati non trattati con HLF. Questo lavoro, pertanto, suggerisce l'applicazione di un idrolizzato antimicrobico di una proteina del latte, efficace per prevenire la colorazione blu, senza modificare la composizione della mozzarella. Parole chiave: peptidi antimicrobici, shelf-life, controllo alterazioni

In questo lavoro, mais triturato combinato alternativamente con frumento, frumento parboiled,riso e patata è stato bollito in eccesso di acqua e sottoposto a fermentazione lattica per circa16 ore al fine di realizzare 4 tipologie di bevande (B1, B2, B3 e B4) ad elevato contenuto di amidiresistenti (RS), una frazione dell'amido che sfugge alla digestione del primo tratto intestinaleed è fermentata nel colon. Le bevande, dal contenuto medio di acqua (in peso) del 76,8%,hanno mostrato valori medi di pH di 4,06 e di acidità totale titolabile di 0,53% (w/w) dopofrigoconservazione e viscosità apparente variabile in funzione della loro composizione (da 19,4a 37,9 P.s). Larga parte della conta batterica totale, prossima in tutti i casi ai 9 log ufc/g, risultavacostituita da batteri lattici. A seguito di determinazioni enzimatico-spettrofotometriche lebevande hanno mostrato percentuali di RS, sulla sostanza secca, superiori al 5,5%. Quantitàcrescenti di amilosio, registrate nelle miscele di mais e frumento, hanno determinato incrementisignificativi di RS nelle rispettive bevande fino a raggiungere in media il 48% del contenutototale di amido. Il presente lavoro pone le basi per lo sviluppo di un nuovo tipo di bevandapotenzialmente utile per il controllo della risposta glicemica post-prandiale, come disciplinatodal Regolamento Europeo n. 432/2012 in materia di indicazioni nutrizionali-salutistiche.

La qualità e la shelf-life della mozzarella è strettamente correlata ai cambiamenti della composizione chimico-fisica e microbiologica del liquido di governo, nel quale si riversano zuccheri, peptidi e sali minerali provenienti dalla mozzarella che favoriscono la crescita microbica anche in condizioni di frigo conservazione. Nel presente lavoro, il liquido di governo di bocconcini di mozzarella è stato sostituito con biopolimeri ottenuti da soluzioni di siero neutralizzati, pastorizzati e arricchiti con differenti concentrazioni di alginato di sodio (E401). Le mozzarelle sono state, quindi, conservate a 4°C per 30 giorni, monitorando i parametri reologici, chimici e microbiologici.A seguito di valutazioni preliminari, solo sul prodotto conservato con il biopolimero al 2% è proseguito il monitoraggio, valutando che le mozzarelle conservate in alginato mostravano percentuali maggiori di deformazioni, rispetto a quelle riscontrate sulle mozzarelle di riferimento. Parallelamente, la capacità di recupero elastico di queste ultime risultava significativamente maggiore di quella dei campioni inclusi in alginato, suggerendo un possibile indebolimento della struttura del formaggio trattato.A differenza del normale liquido di governo che tende ad acidificarsi significativamente a partire dal settimo giorno di frigoconservazione, il siero alginato mostrava maggiore stabililtà raggiungendo i medesimi valori di pH del liquido di governo con un ritardo di circa 14 giorni. Il trend di acidificazione è stato registrato anche in mozzarella, sebbene le differenze tra i due metodi di conservazione risultino inferiori. Come atteso, in condizioni di stress fisico controllato, è stato osservato che le mozzarelle in liquido di governo rilasciavano molto più siero interno, soprattutto entro sette giorni di frigoconservazione, rispetto alle mozzarelle conservate con il biopolimero di siero alginato. I dati microbiologici non rivelano significative differenze tra i campioni in liquido di governo e quelli in siero alginato, sia nelle cinetiche che nei valori della carica microbica delle popolazioni lattiche e non. I risultati ottenuti in questo lavoro, sebbene preliminari, dimostrano che il siero alginato può essere utilizzato come metodo di conservazione alternativo al liquido di governo per mantenere i parametri strutturali e chimici della mozzarella nel corso della conservazione. Tale metodo inoltre può essere ulteriormente migliorato al fine di favorire il parziale recupero del siero, attualmente considerato un sottoprodotto delle lavorazioni lattiero casearie dagli onerosi costi di smaltimento.

Bovine lactoferrin and lactoferricin B, well-known for their antimicrobial properties, were individually immobilized on two different coatings functionalized with - COOH groups deposited in the inner part of polyethylene micro tubes by means of a plasma deposition (PE-CVD) process fed with ethylene and acrylic acid vapors. The resulting functionalized tubes were tested for antimicrobial activity against three Pseudomonas strains responsible for casein hydrolysis and cheese pigmentation. The cell counts of these spoilage bacteria, incubated for 30 h under their optimal growth conditions, were found to be significantly reduced after 24 h in micro tubes functionalized with lactoferricin B, whereas a very low antimicrobial activity against the same strains, often undistinguishable from that of control samples, was observed in tubes functionalized with lactoferrin.This is the first work in which a plasma coating functionalized by lactoferricin B was studied to make an active packaging useful to control cheese spoilage by Pseudomonas.Industrial relevance: The current study describes a new method to immobilize two food grade proteinaceous natural compounds. The resulting plasma-functionalized lactoferricin B-immobilized coating is a promising tool for the control of spoilage microorganisms and shelf-life extension of cheeses.

Microbial spoilage is one of the main factors affecting the quality of fresh fruits and vegetables, leading to off-flavor, fermented aroma, and tissue decay. The knowledge of microbial growth kinetics is essential for estimating a correct risk assessment associated with consuming raw vegetables and better managing the development of spoilage microorganisms. This study shows, for the first time, that only a part of total microbial community, originally present on fresh harvested female zucchini flowers, was able to adapt itself to refrigerated conditions. Through the study of microbial growth kinetics it was possibleto isolate forty-four strains belonging to twenty-two species of the genera Acinetobacter, Arthrobacter, Bacillus, Enterobacter, Erwinia, Klebsiella, Pantoea, Pseudoclavibacter, Pseudomonas, Serratia, Staphylococcus, andWeissella, suggesting Enterobacteriaceae as potentially responsible for pistil spoilage.

TR50 CFCS showed antimicrobial activities against several food-borne pathogens and seven strains belonging to B. pumilus, B. licheniformis and B. thuringensis. The EA fraction retained inhibitory activity against six food-borne pathogenic strains. An inhibition halo against B. cereus DSM 4312 was produced by one peptide band with an apparent molecular weight of about 2.5 kDa, resolved on Tricine SDS-PAGE. The loss of inhibitory activities of TR50 CFCS against some tested strains and the comparison between CFCS and EA RP-HPLC profiles sustains the hypothesis that TR50 produces various active compounds responsible for its antimicrobial activity. Inhibition halos produced by bands scraped from TLC indicates the presence of lipopeptides. These preliminary results highlight that the antibacterial potential of B. subtilis TR50 could be exploited in several food sectors and in the sausage curing process. Further analyses are in progress to identify the active compounds.

Mozzarella is one of the most known Italian traditional fresh cheeses, usually stored into a cold governing liquid in order to maintain its typical organoleptic characteristics. These storage conditions favour the growth of psychrotrophic bacteria among which some are responsible for cheese spoilage. In this work, bovine lactoferrin (BLF) and its pepsin-digested hydrolysate (LFH) were tested in vitro to evaluate their efficacy as potential control agents of Mozzarella cheese spoilage bacteria growing during refrigeration period. At first, five bacterial strains (Pseudomonas fragi 23A, P. gessardi 2A, Serratia proteamaculans 19A, Aeromonas salmonicida 22DB and Rahnella aquatilis 19B), previously isolated from different cold-stored Mozzarella cheese samples, were tested for their ability to grow at 4 °C in the governing liquid-based medium, in which Mozzarella releases debris, sugars and salts, and to promote proteolytic activities against casein fractions. In order to check the efficacy of BLF or LFH to control spoilage bacteria, target strains were inoculated in the governing liquid-based medium at two levels of initial inoculum (3 and 6 log cfu/mL), and incubated at 4 °C or 30 °C for two weeks. At the initial inoculum of 3 log cfu/mL all strains grew well in the governing liquid at 4 °C, reaching 7.1 log cfu/mL, on average, after four days of cold storage. The most proteolytic strains were P. gessardi 2A and P fragi 23A that were able to digest completely all casein fractions, flowed into the governing liquid, after five and eight days, respectively. The remaining strains caused the disappearance of some casein bands on SDS-PAGE patterns, after twelve days of incubation. Viable cells of P. gessardi 2A and R. aquatilis 19B were undetectable when 3 log cfu/ml of each microorganism were inoculated in the governing liquid supplemented with 10 mg/mL of LFH. S. proteomaculans 19A, A. salmonicida 22DB and P. fragi 23A resulted partially inhibited as their bacterial loads resulted 2.3 log cfu/mL, on average, lower than those of the un-supplemented control up to day 5 of cold storage. After additional two days at 4 °C, this difference decreased by 1 log cfu/mL. The inhibitory activity of LFH resulted higher than BLF against all strains grown in the governing liquid-based medium. No antimicrobial activity was observed when the governing liquid, amended with BLF or LFH, was inoculated with target strains at 6 log cfu/mL. Also the increase in temperature of incubation advantaged microbial cells, reducing the inhibitory efficacy of BLF and LFH: after one day of incubation at 30 °C, a decrease of bacterial growth was registered only for P. fragi 23A and R. aquatilis 19B and only at 3 log cfu/ml of initial inoculum level. As expected, casein band intensities in SDS-PAGE patterns were comparable with those of the uninoculated control sample only in experiments showing the lowest microbial viable cell concentration. In conclusion, we demonstrat

In the present study, we evaluated the antimicrobial activity of neutral electrolyzed water (NEW) against 14 strains of spoilage Pseudomonas of fresh cut vegetables under cold storage. The NEW, produced from solutions of potassium and sodium chloride, and sodium bicarbonate developed up to 4000 mg/L of free chlorine, depending on the salt and relative concentration used. The antimicrobial effect of the NEW was evaluated against different bacterial strains at 105 cells/ml, with different combinations of free chlorine concentration/contact time; all concentrations above 100 mg/L, regardless of the salt used, were found to be bactericidal already after 2 min. When catalogna chicory and lettuce leaves were dipped for 5 min in diluted NEW, microbial loads of mesophilic bacteria and Enterobacteriaceae were reduced on average of 1.7 log cfu/g. In addition, when lettuce leaves were dipped in a cellular suspension of the spoiler Pseudomonas chicorii I3C strain, diluted NEW was able to reduce Pseudomonas population of about 1.0 log cfu/g. Thanks to its high antimicrobial activity against spoilage microorganisms, and low cost of operation, the application of cycles of electrolysis to the washing water looks as an effective tool in controlling fresh cut vegetable microbial spoilage contamination occurring during washing steps. (C) 2015 Elsevier Ltd. All rights reserved.

In this work the development of table grape sour rot, a disease sustained by a consortium of non-Saccharomyces yeasts (NSY) and acetic acid bacteria (AAB), was observed under postharvest conditions inoculating wounded berries with 8 NSY strains, or 7 AAB, or their combinations (56). Candida zemplinina CBS 9494 (Cz) and Acetobacter syzygii LMG 21419 (As) produced the highest percentage of rotten berries (100%) after 6 days at 25 °C. Subsequently, the efficacy of low doses of gaseous ozone (2.14 mg m-3) was evaluated in wounded berries inoculated with this simplified As-Cz microbial consortium after ten days at 4 °C. At the end of cold storage, no sample showed sour rot symptoms, although ozonation mainly affected the As viability. In order to simulate sale conditions, all samples were additionally stored at 25°C for 12 days. After this period, the sour rot index of inoculated As-Cz berries previously cold-stored under ozone or air accounted for 22.6 ± 3.7% and 66.7 ± 4.5%, respectively. Inoculated rotten berries showed a slight reduction in monosaccharide content and the different increases in concentration of acetic acid, gluconic acid, ethanol and glycerol. Microbiological analyses of sound and rotten berries revealed, in addition to As and Cz, the occurrence of 15 new strains potentially responsible for sour rot development and mainly belonging to Gluconobacter albidus and Hanseniaspora uvarum. In this work we demonstrated that cold ozonation was useful to extend the shelf-life of cold stored table grapes even when, at the end of treatment, they were further stored at room temperature.

The objective of this study was to develop a new synbiotic beverage evaluating the ability of some bifidobacteria strains to grow in this beverage which was fortified with whey proteins up to 20 g L-1, and enriched with 10 g L-1 of prebiotic inulin or resistant starch. The ability of Bifidobacterium strains to survive for 30 days at 4°C was evaluated in two synbiotic whey protein fortified beverages formulated with 2% of whey proteins and 1% of inulin or resistant starch. Microbial growth was significantly affected by the whey protein amount as well as by the kind of prebiotic fibre. Resistant starch promoted the growth of the Bifidobacterium pseudocatenulatum strain and its viability under cold storage, also conferring higher sensory scores. The development of this new functional beverage will allow to carry out in vivo trials in order to validate its pre- and probiotic effects.

Probiotics species appear to differentially regulate the intestinal immune response. Moreover, we have shown that different immune-modulatory abilities can be found among probiotic strains belonging to the same species. In this study, we further addressed this issue while studying L. gasseri, a species that induces relevant immune activities in human patients. Results: We determined the ability of two strains of L. gasseri, OLL2809 and L13-Ia, to alter cell surface antigen expression, cytokine production and nuclear erythroid 2-related factor 2 (Nrf2)-mediated cytoprotection in murine bone marrow-derived dendritic cells (DCs) and MODE-K cells, which represent an enterocyte model. Differential effects of L. gasseri strains were observed on the expression of surface markers in mature DCs; nevertheless, both strains dramatically induced production of IL-12, TNF- and IL-10. Distinctive responses to OLL2809 and L13-Ia were also shown in MODE-K cells by analyzing the expression of MHC II molecules and the secretion of IL-6; however, both L. gasseri strains raised intracellular glutathione. Treatment of immature DCs with culture medium from MODE-K monolayers improved cytoprotection and modified the process of DC maturation by down-regulating the expression of co-stimulatory markers and by altering the cytokine profile. Notably, bacteria-conditioned MODE-K cell medium suppressed the expression of the examined cytokines, whereas cytoprotective defenses were significantly enhanced only in DCs exposed to OLL2809-conditioned medium. These effects were essentially mediated by secreted bacterial metabolites. Conclusions: We have demonstrated that L. gasseri strains possess distinctive abilities to modulate in vitro DCs and enterocytes. In particular, our results highlight the potential of metabolites secreted by L. gasseri to influence enterocyte-DC crosstalk. Regulation of cellular mechanisms of innate immunity by selected probiotic strains may contribute to the beneficial effects of these bacteria in gut homeostasis. © 2013 Luongo et al.; licensee BioMed Central Ltd.

Bio-composite coatings, consisting of an organic matrix embedding a bioactive molecule, have been deposited by means of atomizer-assisted atmospheric pressure plasma. Ethylene was chosen as the precursor of the matrix, while the atomizer was fed with a water solution of lysozyme. Coatings chemical composition was investigated by XPS, FTIR and MALDI-TOF spectroscopies, and it has been proved that the one-step inclusion of protein domains in the composite coatings is successful and lysozyme chemical structure is only slightly altered. The amount of embedded lysozyme is as high as 14?g/cm² as evaluated from water release test. Finally, the activity of the plasma-embedded protein is close to that of pure lysozyme as verified against Micrococcus lysodeikticus ATCC 4698 through an agar plate diffusion test.

Pseudomonas fluorescens is a genetically and phenotypically heterogeneous species that is often reported as a spoiler of fresh foods, but it has recently been implicated in clinical infection. In this study, we sequenced the genome of P. fluorescens strain ITEM 17298, isolated from mozzarella cheese and able to cause several alterations under cold storage.

The aim of this work was to check the efficacy of an annular ultraviolet reactor for stabilizing the growth of lactic acid bacteria (LAB), the main components of the microbiota of three raw fermented beverages, Boza, Ayran and Matsoni during cold storage. The ultraviolet (UV) dose of 260 mJ cm-2 caused a reduction of 2.6 and 1 log CFU mL-1 in the LAB loads of Boza and Ayran samples, respectively, whereas multiple UV treatments were required for Matsoni; they caused the decrease of 3 log CFU mL-1 in LAB counts. Twenty-two bacterial strains, belonging to 11 species, were identified in the natural microflora of the three beverages before and after the UV treatments. Only a minimal post-acidification process was registered up to day 30 in all the three beverages. This ultraviolet device is at disposal of beverage manufacturers, who could apply it for producing microbially-stable drinks with an extended shelf-life

The aim of this work was to check the efficacy of an annular ultraviolet reactor for stabilizing the growth of lactic acid bacteria (LAB), the main components of the microbiota of three raw fermented beverages, Boza, Ayran and Matsoni during cold storage. The ultraviolet (UV) dose of 260 mJ cm-2 caused a reduction of 2.6 and 1 log CFU mL-1 in the LAB loads of Boza and Ayran samples, respectively, whereas multiple UV treatments were required for Matsoni; they caused the decrease of 3 log CFU mL-1 in LAB counts. Twenty-two bacterial strains, belonging to 11 species, were identified in the natural microflora of the three beverages before and after the UV treatments. Only a minimal post-acidification process was registered up to day 30 in all the three beverages. This ultraviolet device is at disposal of beverage manufacturers, who could apply it for producing microbially-stable drinks with an extended shelf-life.

There is a variety of different food processing methods, which can be used to prepare ready-to-eat foods. However, the need to preserve the freshness and nutritional qualities leads to the application of mild technologies which may be insufficient to inactivate microbial pathogens. In this work, fresh chicory stems were packed under a vacuum in films, which were transparent to microwaves. These were then exposed to microwaves for different periods of time. The application of sous vide microwave cooking (SV-MW, 900W, 2450 MHz), controlled naturally occurring mesophilic aerobic bacteria, yeasts and molds for up to 30 d when vacuum-packed vegetables were stored at 4 degrees C. In addition, the process lethality of the SV-MW 90 s cooking was experimentally validated. This treatment led to 6.07 +/- 0.7 and 4.92 +/- 0.65 log cfu/g reduction of Escherichia coli and Listeria monocytogenes inoculated over the chicory stems (100 g), respectively. With an initial load of 9 log cfu/g for both pathogens, less than 10 cfu/g of surviving cells were found after 90 s cooking. This shows that short-time microwave cooking can be used to effectively pasteurize vacuum-packed chicory stems, achieving > 5 log cfu/g reduction of E. coli and L. monocytogenes.

The microbial content of plant tissues has been reported to cause the spoilage of ca. 30% of chlorine-disinfectedfresh vegetables during cold storage. The aim of this work was to evaluate the efficacy of antimicrobial peptidesin controlling microbial vegetable spoilage under cold storage conditions. A total of 48 bacterial isolateswere collectedfrom ready-to-eat (RTE) vegetables and identified as belonging to Acinetobacter calcoaceticus, Aeromonasmedia, Pseudomonas cichorii, Pseudomonas fluorescens, Pseudomonas jessenii, Pseudomonas koreensis, Pseudomonasputida, Pseudomonas simiae and Pseudomonas viridiflava species. Reddish or brownish pigmentation was foundwhen Pseudomonas strains were inoculated in wounds on leaves of Iceberg and Trocadero lettuce and escarolechicory throughout cold storage. Bovine lactoferrin (BLF) and its hydrolysates (LFHs) produced by pepsin, papainand rennin, were assayed in vitro against four Pseudomonas spp. strains selected for their heavy spoiling ability.As the pepsin-LFH showed the strongest antimicrobial effect, subsequent experiments were carried out using thepeptide lactoferricin B (LfcinB), well known to be responsible for its antimicrobial activity. LfcinB significantly reduced(P <= 0.05) spoilage by a mean of 36% caused by three out of four inoculated spoiler pseudomonads on RTElettuce leaves after six days of cold storage. The reduction in the extent of spoilage was unrelated to viable celldensity in the inoculated wounds. This is the first paper providing direct evidence regarding the application ofan antimicrobial peptide to control microbial spoilage affecting RTE leafy vegetables during cold storage.

This work aims at studying the efficacy of low doses of gaseous ozone in postharvest control of the table grape sour rot, a disease generally attributed to a consortium of non-Saccharomyces yeasts (NSY) and acetic acid bacteria (MB). Sour rot incidence of wounded berries, inoculated with 8 NSYstrains, or 7 AAB, or 56 yeast-bacterium associations, was monitored at 25 degrees C up to six days. Sour rot incidence in wounded berries inoculated with yeast-bacterium associations resulted higher than in berries inoculated with one single NSY or AAB strain. Among all NSY-AAB associations, the yeast-bacterium association composed of Candida zemplinina CBS 9494 (Cz) and Acetobacter syzygii LMG 21419 (As) showed the highest prevalence of sour rot; thus, after preliminary in vitro assays, this simplified As-Cz microbial consortium was inoculated in wounded berries that were stored at 4 degrees C for ten days under ozone (2.14 mg m(-3)) or in air. At the end of cold storage, no berries showed sour-rot symptoms although ozonation mainly affected As viable cell count. After additional 12 days at 25 degrees C, the sour rot index of inoculated As-Cz berries previously cold-stored under ozone or in air accounted for 22.6 +/- 3.7% and 66.7 +/- 4.5%, respectively. Molecular analyses of dominant AAB and NSY populations of both sound and rotten berries during post refrigeration period revealed the appearance of new strains mainly belonging to Gluconobacter albidus and Hanseniaspora uvarum species, respectively. Cold ozonation resulted an effective approach to extend the shelf-life of table grapes also after cold storage. (C) 2017 Elsevier Ltd. All rights reserved.

In the present study, we evaluated the antimicrobial activity of neutral electrolyzed water (NEW) against 14 strains of Pseudomonas responsible for spoilage of the leaves of lettuce 'iceberg', 'trocadero' chicory and endive under cold storage. Active chlorine concentration was found stable for 14 minutes also when it was in contact with mixed vegetables; in addition, NEW resulted to be bactericide, within 2 min, for all concentrations of free chlorine above 100 ppm. When catalogna chicory and lettuce leaves were dipped for 5 min in a NEW solution microbial loads of mesophilic bacteria counts and Enterobacteriaceae were reduced on average of 1.7 log cfu/g. This work demonstrates that in situ production of active chlorine, via electrolysis of a diluted sals solution, can be efficiently used against spoilage bacteria contaminating fresh cut vegetable.

This study demonstrates the efficacy of cold gaseous ozone treatments at low concentrations in the eradication of high Listeria monocytogenes viable cell loads from glass, polypropylene, stainless steel, and expanded polystyrene food-contact surfaces. Using a step by step approach, involving the selection of the most resistant strain-surface combinations, 11 Listeria sp. strains resulted inactivated by a continuous ozone flow at 1.07 mg m(-3) after 24 or 48 h of cold incubation, depending on both strain and surface evaluated. Increasing the inoculum level to 9 log CFU coupon(-1), the best inactivation rate was obtained after 48 h of treatment at 3.21 mg m(-3) ozone concentration when cells were deposited onto stainless steel and expanded polystyrene coupons, resulted the most resistant food-contact surfaces in the previous assays. The addition of naturally contaminated meat extract to a high load of L. monocytogenes LMG 23775 cells, the most resistant strain out of the 11 assayed Listeria sp. strains, led to its complete inactivation after 4 days of treatment. To the best of our knowledge, this is the first report describing the survival of L. monocytogenes and the effect of ozone treatment under cold storage conditions on expanded polystyrene, a commonly used material in food packaging. The results of this study could be useful for reducing pathogen cross-contamination phenomena during cold food storage.

La collaborazione planetaria tra centri di ricerca ha permesso di conoscere velocemente tutti i fattori di virulenza del “batterio killer”, un ceppo di Escherichia coli, responsabile della recente tossinfezione alimentare da germogli di semi. Ma l’emergenza non può considerarsi conclusa.

Nduja is a traditional Calabrian (Italy) sausage made from pork trimmings (underbelly, shoulder and head) and manufactured with high amounts of different spices (ca. 5%), mostly represented by the red hot chili pepper that confers its typical flavour and piquancy. In addition, spices are well known for their antimicrobial activity against several microorganisms; in fact, no yeast and mould growth was found on this sausage throughout ripening and storage. In this study, methanolic, ethanolic and aqueous extracts of processed sweet, hot chili pepper powder (SP and HCP), and three spice mixtures (SM1, SM2, SM3), used in industrial manufacturing of 'Nduja, were assayed for their antimicrobial activity against ten food-borne bacteria (Figura 1) by using agar disc diffusion assay.

Allo scopo di incrementare la resa casearia della caciotta sono state condotte prove di caseificazioni con latte vaccino arricchito con differenti quantità (2,5 - 10%) di siero concentrato in polvere (WPC) proveniente da reflui caseari.I risultati preliminari hanno evidenziato che ad elevate concentrazioni (dal 5 al 10%) l'aggiunta di WPC aumentava significativamente i tempi di coagulazione e diminuiva la consistenza del formaggio rispetto a quella del campione controllo. Al contrario l'aggiunta di WPC al 2,5% ha prodotto un aumento della resa casearia, nonostante la maggior parte delle sieroproteine (SP) aggiunte fossero perse durante il processo di coagulazione. Durante la stagionatura la riduzione dell'umidità dei formaggi, inferiore nelle caciotte con WPC rispetto a quelle controllo, ha consentito un sensibile aumento del livello di SP unicamente nei formaggi arricchiti con WPC. E' stato possibile infatti integrare nella caciotta stagionata oltre il 10% della quantità di SP aggiunta nel latte, ottenendo rese casearie superiori per circa l'1,5%. La produzione di ricotta proveniente dal siero delle caciotte arricchite in WPC ha inoltre permesso un recupero di SP pari a circa il 50% in più di quanto ottenuto dalle ricotte controllo.A termine della stagionatura (1 mese), le caciotte con e senza WPC sono state confezionate sottovuoto e conservate per tre mesi a 4°C per la valutazione della loro shelf life. Le popolazioni batteriche mesofile (CBT), i batteri lattici e i coliformi hanno mostrato una modesta riduzione delle cariche vitali senza particolari differenze tra le due tipologie di formaggio. Le prove di assaggio hanno evidenziato una maggiore cremosità della caciotta arricchita in WPC rispetto a quella controllo. Tuttavia a fine del periodo di shelf-life i formaggi con WPC presentavano un lieve retrogusto amaro che era assente nelle caciotte controllo.Le valutazioni organolettiche delle ricotte realizzate tra le 24 e 48 ore dalla loro produzione non hanno mostrato sostanziali differenze.I risultati ottenuti mostrano per la prima volta che, applicando un processo di lavorazione integrato, è possibile recuperare le SP presenti nel siero per ottenere formaggi con aumentata resa casearia ed un migliorato profilo proteico. Il controllo dei fenomeni di proteolisi legati alla produzione di peptidi amari necessita di ulteriore sperimentazione.

A gallium-modified chitosan/poly(acrylic acid) bilayer was obtained by electrochemical techniques ontitanium to reduce orthopaedic and/or dental implants failure. The bilayer in vitro antibacterial propertiesand biocompatibility were evaluated against Escherichia coli and Pseudomonas aeruginosa and on MG63osteoblast-like cells, respectively. Gallium loading into the bilayer was carefully tuned by the electro-chemical deposition time to ensure the best balance between antibacterial activity and cytocompatibility.The 30 min deposition time was able to reduce in vitro the viable cell counts of E. coli and P. aeruginosa of2 and 3 log cfu/sheet, respectively. Our results evidenced that the developed antibacterial coating did notconsiderably alter the mechanical flexural properties of titanium substrates and, in addition, influencedpositively MG63 adhesion and proliferation. Therefore, the gallium-modified chitosan/poly(acrylic acid)bilayer can be exploited as a promising titanium coating to limit bacterial adhesion and proliferation,while maintaining osseointegrative potential.

Le malattie cardiovascolari, cancro, malattie non infettive dell'apparato respiratorio e diabete sono malattie croniche, anche dette "noncommunicable diseases" (NCDs) legate a disordini metabolici come l'incremento della pressione sanguigna, sovrappeso ed obesità, iperglicemia e iperlipidemia. I dati raccolti dalla Word Health Organization indicano come nei Paesi altamente sviluppati l'87% della mortalità è attribuita alle NCDs il cui più importante fattore di rischio è rappresentato dall'elevata pressione sanguigna. E' stato ampiamente dimostrato che gli alti livelli di sodio nella dieta sono legati all'incremento della pressione sanguigna e, per tale motivo, la riduzione della quantità di sodio assunta con la dieta è un obiettivo portato avanti a livello nazionale e comunitario. Dal momento che circa il 70-75% del sodio assunto proviene da alimenti processati, ridurre l'uso del sale nei vari processi di trasformazione alimentare potrebbe portare ad un ridotto consumo giornaliero di sodio. Dal momento che l'aggiunta in etichetta di indicazioni nutrizionali come "a basso (o bassissimo) contenuto di sodio/sale" è favorevolmente accolta dai consumatori, numerose ricerche sono in atto per superare gli ostacoli tecnici legati alle modifiche di processo necessarie per l'ottenimento di formaggi sicuri e di buona qualità a basso contenuto di sodio. Il Cacioricotta è un formaggio a pasta semidura che può raggiungere un contenuto di sale superiore al 6% a fine stagionatura. La salatura è cruciale per l'ottenimento delle caratteristiche chimiche fisiche, sensoriali e microbiologiche ottimali di questo formaggio, per cui, la riduzione del sale potrebbe determinare la riduzione dei livelli di sicurezza e la shelf-life di tale formaggio. In questo studio la salatura del Cacioricotta è stata realizzata sostituendo il sale con quantità crescenti di un concentrato minerale contenente circa il 10% di sodio. Le caratteristiche chimico-fisiche, microbiologiche, le concentrazioni di sodio e di altri metalli e i profili di consistenza strumentale (TPA) sono stati confrontate con quelli dei campioni di riferimento (0 e 100% cloruro di sodio) e monitorate durante 90 giorni di frigoconservazione sotto vuoto.Durante i 90 giorni di frigoconservazione i valori di pH e acidità totale sono risultati simili in tutti i campioni analizzati mentre il valore dell'attività dell'acqua è risultato inversamente proporzionale alla concentrazione di sodio aggiunta con valori medi di 0,901 ± 0,03 al termine della frigoconservazione. Le differenti concentrazioni di sodio, monitorate mediante ICP-OES, non hanno influenzato le cariche vitali di enterococchi, stafilococchi, funghi e lieviti e batteri lattici, mentre le popolazioni di coliformi ed Escherichia coli risultavano controllate in maniera più efficace all'aumentare della quantità di cloruro di sodio usato. I formaggi con parziale e totale sostituzione del sale mostravano valori di durezza in media più elevati (86,26 N) di qu

Ayran is a traditional Turkish milk drink which is fermented and salted. Inadequate production andstorage conditions contribute to its variable organoleptic quality and stability during shelf-life. A thoroughphysico-chemical, nutritional and microbial characterization of artisanal Ayran was carried out inorder to standardize its overall quality without altering its original traits. Ayran microbial ecosystem waslargely dominated by Streptococcus thermophilus (ST) and Lactobacillus delbrueckii subsp. bulgaricus(LDB). High counts of other lactic acid bacteria species, including Lactobacillus helveticus (LH), Lactobacillusfermentum (LF), and Lactobacillus paracasei (LP), were also found. Selected LDB, LP and LH strainsgrew well in milk displaying fast acidification and high proteolysis, differently from ST and LF strains thatdid not cause noticeable changes. A selected autochthonous three-strain culture (TSC), composed of onestrain of LDB, LP and ST, was applied for the pilot-scale production of traditional Ayran. The Ayranproduced with this TSC resulted in the most extensive shelf-life (one month) and in the best terms of itsnutritional and sensory quality nevertheless altering its typical pleasant yogurt and cottage cheese notes.This TSC is at disposal of SMEs who need to standardize the overall quality of this traditional fermentedmilk, preserving its typical traits

The quality and shelf-life of ready-to-use lettuces depend on genotypic traits of raw material and on several aspects from preharvest to postharvest processing. In general, preharvest factors should be aimed to optimize their impact on postharvest quality, in order to limit the use of chemical preservative during processing. In this study the influence of soil and soilless growing systems on quality and microbial traits of three multi-leaf lettuce cultivars (two green Eztoril and Ezabel, and one red Ezra) was evaluated at harvest and after 7 and 13 days of storage at 8 °C. At harvest, Ezra showed a respiration activity and a total phenol content respectively 2-fold and 25% significantly higher than the green cultivars. Soil lettuces resulted more stressed than those grown in soilless, as indicated by their initial content in antioxidants. As for nitrate content, soilless grown lettuces at harvest showed an average concentration higher than soil-grown ones (838 vs 432 mg/kg fw), although values are generally lower than limits imposed by the EU Regulation (No.1258/2011). The initial microbial load was significantly higher in soil lettuce compared with lettuce cultivated in soilless condition. Moreover the latter resulted cleaner than soil grown samples, with evident beneficial effect regarding the potential application of mild washing during processing in view of extended cold storage. During storage, soilless lettuces showed no ammonium accumulation, differently from those cultivated in soil. In addition, lettuce cultivars grown in soilless condition showed unchanged content in the antioxidant activity and total phenols, and lower microbial counts than soil lettuces. Results of the present study showed that soilless growing system can positively affect qualitative and microbiological parameter of lettuces studied, and it can be a valid agricultural system in order to obtain high quality multi-leaf lettuces for ready-to-use industry.

In this study 28 bacterial strains, isolated fromgreenwaters-polluted-soil, were investigated for their abilityto grow in presence of phenols added to Mineral BasalMedium (MBM) in aerobic conditions. In particular, threeof them were found to be able to use as sole carbon sourcephenol, cathecol, caffeic acid and ferulic acid with efficiencyranging from 76% (phenol in 5 days, millimolarconcentration from 3.7 10-2 to 9 10-3) to 95% (ferulic acidin 2 days millimolar concentration from 6.8 10-1 to 310-2). For these strains the taxonomic position was studiedby amplification and sequencing of 16S rRNA genes. Theisolated strains were classified belonging to Arthrobactersulfureus, Pseudomonas synxantha and Pseudomonasoryzihabitans. Noteworthy, for the first time such Pseudomonasstrains have been shown to be able to use polyphenolsas the only carbon source in vitro. In fact, to thebest of our knowledge, this kind of study were not done onPs. Synxantha, while it was recently shown the ability ofP. oryzihabitans to degrade catechol. These findings mayopen to new biotechnological applications for the degradationof polyphenols.

The bovine gastrointestinal tract (GIT) is the main reservoir for enterohaemorrhagic Escherichia coli (EHEC) responsible for food-borne infections. Therefore, it is crucial to develop strategies, such as EHEC suppression by antagonistic microorganisms, to reduce EHEC survival in the GIT of cattle and to limit shedding and food contamination. Most human-derived Lactobacillus reuteri strains produce hydroxypropionaldehyde (HPA), an antimicrobial compound, during anaerobic reduction of glycerol. The capacity of L. reuteri LB1-7, a strain isolated from raw bovine milk, to produce HPA and its antimicrobial activity against an O157:H7 EHEC strain (FCH6) were evaluated in bovine rumen fluid (RF) under strict anaerobiosis. EHEC was totally suppressed when incubated in RF inoculated with L. reuteri LB1-7 and supplemented with 80 mM glycerol (RF-Glyc80). The addition of LB1-7 or glycerol alone did not modify EHEC survival in RF. Glycerol was converted to HPA (up to 14 mM) by LB1-7 during incubation in RF-Glyc80, and HPA production appeared to be responsible for EHEC suppression. The bactericidal activity of L. reuteri LB1-7, the concentration of glycerol required and the level of HPA produced depended on physiological and ecological environments. In vitro experiments also showed that EHEC inoculated in rumen fluid and exposed to L. reuteri and glycerol had a very limited growth in rectal contents. However, L. reuteri exerted an antimicrobial activity against the rumen endogenous microbiota and perturbed feedstuff degradation in the presence of glycerol. The potential administration of L. reuteri and glycerol in view of application to finishing beef cattle at the time of slaughter is discussed. Further in vivo studies will be important to confirm the efficiency of L. reuteri and glycerol supplementation against EHEC shedding in ruminants.

Cheesemaking procedures applied for the production of traditional dairy products, including Protected Designation Origin (PDO) cheeses, usually include wild bacteria from raw milk and natural starters. These sources of microbial diversity have been largely exploited for the improvement of dairy products quality and safety. However, recent studies demonstrated these traditional foods are also a good source of probiotic bacteria. The application of a validated novel digestion/enrichment protocol to 15 raw milk samples allowed the isolation of 11 lactic acid bacteria strains belonging to Enterococcus faecium, Lactobacillus brevis, L. gasseri, , L. johnsonii, L. reuteri L. vaginalis and Lactococcus lactis species. This work reports the isolation of potentially probiotic lactobacilli from raw milk samples paying particular attention to antimicrobial activities played by the natural isolate Lactobacillus reuteri LB1-7 against shiga toxin-producing enterohaemorrhagic Escherichia coli (EHEC) strains. L. reuteriLB1-7 was assayed for the production of hydroxypropionaldehyde (HPA) verifying its antimicrobial activity against several EHEC strains. Antimicrobial assays were carried out in bovine rumen fluid as well as in samples collected from the bovine rectum. Under both conditions the EHEC was totally suppressed when incubated together with L. reuteri LB1-7 and supplemented with 80 mM glycerol. The addition of LB1-7 or glycerol alone did not modify EHEC survival in any condition. This work underlines that microbial diversity still occurring in raw milk could be an useful source of strains to be employed in the improving of food quality and safety. In addition, results related to the suppression of EHEC strains open the possibility to counteract the EHEC shedding by cattle thanks to a direct-fed microbial supplementation.

Cactus pear (Opuntia ficus-indica (L.) Mill., 'Gialla') is a delicious and nourishing fresh fruit having a number of documented positive health effects. One of the reasons this fruit is not widely commercialized in international and domestic markets is because of the presence of residual glochids (thorny hairs) on the fruit surface. Marketing cactus pear as a ready to eat product may significantly improve its consumption, preserving its quality, safety and marketability. In this work green-yellow cactus pear fruits were peeled and stored for 13 days at 4 and 8°C, packaged in passive and in active modified atmosphere. Samples stored in air were used as control. The passive modified atmosphere improved the marketability of cactus pears stored at 8°C for 6 days. The rapid decrease in O2 and the increase in CO2 concentration in packages stored at 8°C caused the loss of marketability of fruits stored at high temperature. Samples stored at 4°C in passive and active modified atmosphere resulted in a marketablility of 9 days, since each one in air was just below the salable limit at 6 days. A very low initial microbial growth was detected, while an increase in mesophilic and psychrophilic bacteria during storage at both temperatures was found in the air samples. In conclusion, it was found that it is possible to store fresh-cut cactus pear, obtaining a marketability of 9 days at 4°C; while, when the temperature is higher than 4°C, it is not possible to preserve the fruit quality for more than 6 days.

Fifty-one yeasts were randomly isolated from four different preparations of Boza, a Turkish cereal-based beverage in order to throw light upon their biotechnological potential. Sequence analysis of 26S rDNA D1/D2 domain revealed that the isolated yeasts belonged to Candida, Clavispora, Cryptococcus, Cystofilobasidium, Geotrichum, Coniochaeta, Pichia, Rhodotorula, Saccharomyces, Torulaspora and Trichosporon genera. Interestingly, Pichia fermentans resulted the dominant species with 21 isolates; a high polymorphism among these yeasts resulted by RAPD-PCR analysis which allowed to distinguish 15 strains that were characterized for their biochemical and technological features. In addition, all P. fermentans strains did not assimilate d-maltose and d-sucrose and expressed a very low invertase activity, suggesting their possible coexistence with maltose- and sucrose-fermenting lactic acid bacteria in Boza microbial ecosystem. Three yeasts produced a high amount of the rose-like fragrance 2-phenylethanol that might be involved in flavor development of this traditional beverage. Moreover, these yeasts also displayed an antagonistic activity against Trichosporon cutaneum, a pathogen occurring in Boza. These selected yeasts are at disposal of Boza manufacturers who could apply them, in combination with LAB, for producing a high quality beverage in accordance with the Turkish quality standards.(IF 3.150)

The aim of this study was to analyse non-lactic acid bacteria populations (NLABPs) and evaluate their role in proteolysis of cold-stored high moisture (HM) Mozzarella cheese. NLABPs reached values close to 8log cfumL -1 after seven days of cold storage. Sequencing of 16 rDNA and rpoB genes and molecular biotyping allowed to identify 66 bacterial strains belonging to 25 species from 15 genera, mainly represented by Pseudomonas, Acinetobacter, and Rahnella. Fifteen strains showed proteolytic activity values higher than 1000.00?g GlymL -1 after 24h of growth in skimmed milk. Moreover, as shown by Urea-PAGE, 11 proteolytic strains caused partial or total disappearance of at least one of the caseins. Their proteolytic behaviour was assessed even when they grew inside the governing liquid together with HM Mozzarella cheese at 4°C for 12 days. This is the first report that throws light on the complexity of NLABPs in HM Mozzarella cheese, demonstrating that some strains caused the partial hydrolysis of ?, ?, and ? caseins on its outer surface where a concomitant wrinkling and successive exfoliation became visible without significant changes in texture characteristics. (IF 3.283)

Bacillus genus is a widespread heterogeneous group of Gram-positive, facultative anaerobic, endospore-forming bacteria occurring in many types of foods. Bacillus has been reported to produce more than 45 antimicrobial molecules: some of them are of clinical value, whereas others have been assayed in vitro against food-borne microorganisms. In this work, the antibacterial activity of the cell-free concentrate supernatant of Bacillus subtilis TR50, a strain isolated from the natural microflora of an Italian traditional cured sausage, was tested by the disc diffusion assay against B. cereus DSM4312, B. cereus DSM4313, Enterococcus faecalis V583, Listeria monocytogenes DSM20600, Staphylococcus aureus NCTC8325, Aeromonas hydrophila DSM30187, Escherichia coli ATCC35401, Helicobacter pylori ATCC43504, Salmonella enterica ATCC13311, Yersinia enterocolitica DSM4780 and 17 Bacillus spp. strains isolated from traditional and industrial cured sausages. All pathogens, except E. faecalis, resulted inhibited by the cell-free concentrated TR50 supernatant. The average size of the inhibition halos on the bacterial lawns of the Bacillus species assayed decreased in the following order: B. pumilus > B. licheniformis > B. thuringensis, whereas no antimicrobial activity was detected against B. amyloliquefaciens and the other B. subtilis strains. The cell-free concentrate TR50 supernatant was extracted with ethyl acetate and the resultant organic fraction proved to retain antimicrobial activity. The same fraction was both spotted on silica thin layer chromatography (TLC) plates and loaded on Tricine SDS-PAGE gel; both TLC plates and the Tricine SDS-PAGE gel were overlaid with the Isosensitest soft-agar inoculated with Bacillus cereus DSM 4312 or B. cereus 4313. The halos on the bacterial lawn agar resulted correlated with the retention factor (Rf) of the spots, that were subsequently scraped from TLC and extracted again with methanol; among the spots (Rf 0.50-0.88), only the compounds with Rf ranging from 0.49 to 0.58 and from 0.69 to 0.77 retained antimicrobial activity. Among bands revolved by Tricine SDS-PAGE, one with an apparent molecular weight of about 2.5 kDa, displayed inhibitory activity against the same indicator strains. Finally, the organic fraction further purified by reverse phase (RP)-HPLC showed that only 20% of the TR50 specific peaks were retained; some of them fell in the chromatographic region common to commercial iturins. These preliminary results highlight the antibacterial potential of B. subtilis TR50 against widespread meat-borne pathogens that could be exploited in the sausage curing process. Further analyses are in progress to identify active compounds even though results of analytical techniques suggest the presence of lipopeptides responsible antimicrobial activity.

The aim of this work was to check the efficacy of bovine lactoferrin hydrolyzed by pepsin (LFH) to prevent blue discoloration of Mozzarella cheese delaying the growth of the related spoilage bacteria. Among 64 Pseudomonas fluorescens strains, isolated from 105 Mozzarella samples, only ten developed blue discoloration in cold-stored Mozzarella cheese slices. When Mozzarella cheese samples from dairy were treated with LFH and inoculated with a selected P.fluorescens strain, no pigmentation and changes in casein profiles were found up to 14 days of cold storage. In addition, starting from day 5, the count of P.fluorescens spoiling strain was steadily ca. one log cycle lower than that of LFH-free samples. ESI-Orbitrap-based mass spectrometry analyses allowed to reveal the pigment leucoindigoidine only in the blue LFH-free cheese samples indicating that this compound could be considered a chemical marker of this alteration. For the first time, an innovative mild approach, based on the antimicrobial activity of milk protein hydrolysates, for counteracting blue Mozzarella event and controlling psychrotrophic pigmenting pseudomonads, is here reported. © 2014 Elsevier Ltd.

Coatings consisting in gentamicin-containing nanocapsules have been synthetized by means of an aerosol-assisted atmospheric pressure plasma deposition process. The influence of different parameters affecting the process has been extensively investigated by means of a morphological and chemical characterization of the coatings. Scanning electron microscopy highlighted the presence of nanocapsules whose size and abundance depend on power input and deposition time. A detailed analysis carried out with matrix-assisted laser desorption ionization coupled to high-resolution mass spectrometry allowed to detect and identify the presence of gentamicin embedded in the coatings and its rearrangement, as a result of the interaction with the plasma. The release of gentamicin in water has been monitored by means of UV-vis fluorescence spectroscopy, and its biological activity has been evaluated as well by the disk diffusion assay against Staphylococcus aureus and Pseudomonas aeruginosa. It is confirmed that the antibacterial activity of gentamicin is preserved in the plasma-deposited coatings. Preliminary cytocompatibility investigations indicated that eukaryotic cells well tolerate the release of gentamicin from the coatings.

Fresh-cut Big Top' nectarines were dipped in 2% (w/v) ascorbic acid-1% (w/v) calcium lactate and stored at 4 degrees C for up to 12days in 10kPa O-2- and 10kPa CO2-modified atmosphere packaging (MAP). The used microperforated plastic film allowed O-2 and CO2 concentrations to reach steady values from the fifth day in storage onwards. Samples stored in MAP after chemical dipping showed the highest visual quality score, slight browning symptoms, increment in firmness and very low ethanol and acetaldehyde content. The chemical dipping also increased antioxidant capacity, probably due to the effect of ascorbic acid. The results suggested that the control of yeasts was mainly exerted by MAP, whereas only a slight effect was achieved by the chemical dipping. Therefore, MAP plus ascorbic acid/calcium lactate dipping was the best combination to preserve phytochemical content, antioxidant capacity and microbiological safety of fresh-cut nectarines during storage.

Changes in atmosphere composition and colour, tissue browning, decay,microbial viable cell count and sensorial attributes of fresh cut broccoli raab werestudied. Broccoli raab heads and young leaves were placed in polystyrene trays,covered with polyvinyl chloride (PVC) and polyamide/polyethylene (PA/PE) laminatedfilms and then stored under self controlled atmosphere (SCA) at 4°C for 16 d.Viable cell counts of packaged fresh-cut vegetables were lower than the legal limit forsafe consumption until the 12th day of storage in both PA/PE and PVC covered trays.However, after 8 and 12 d of storage, microbial counts of broccoli raab stored in PVCwere approx. 2 log cfu g-1 higher than those in PA/PE. Browning was only significantin samples packaged in PA/PE, probably due to high CO2 concentration inside thepackage. The results showed that PVC packaging was the best solution for freshbroccoli raab heads and young leaves stored at 4°C in SCA, leading to a shelf life of 12d compared to 7 d if vegetables were stored in PA/PE.

In questo lavoro, quattro tipologie di bevande (B1, B2, B3 e B4) a differentecomposizione (mais, frumento, riso e patata) e realizzate mediante cottura esuccessiva fermentazione lattica, sono state analizzate per il loro contenutodi amido resistenti (RS). Le bevande, conservate a 4°C, hanno mostrato elevateconcentrazioni di batteri lattici (in media 8,65 log ufc/g) e ridotti valori di pH(3,71 in media). Nonostante un contenuto in peso di acqua di circa il 76,8%, lebevande hanno presentato differenti valori di viscosità apparente (da 19,4 a37,9 Pa·s) e comportamento simile a quello di un fluido pseudoplastico. Inoltrele bevande hanno mostrato percentuali in peso di RS in media superiori al5,89%, in funzione di quantità crescenti di amilosio presenti negli ingredientiutilizzati. Le analisi hanno mostrato che, in media, il 48% del contenuto totaledi amido era costituito da RS. Il presente lavoro pone le basi per lo sviluppodi bevande a base di cereali potenzialmente utili al controllo della rispostaglicemica post-prandiale, come disciplinato dal Regolamento Europeo n.432/2012 in materia di indicazioni nutrizionali-salutistiche.

Bacillus subtilis TR50, a gram-positive endospore-forming bacteria isolated from a cured sausage, proved to be source of several antimicrobial compounds against food-borne pathogenic bacteria (Caputo et al., 2011). The wide chemical variability shown by these metabolites and their related antimicrobial efficacy depend on both the specific strain producer and on nutritional/environmental growth conditions (Baruzzi et al., 2011;Dhouha and Ellouze-Chaabouni, 2011).The difficulty for reliably identifying the antimicrobial compounds released by strains hampers their application in many fields such as controlling food microbial spoilage or fighting the emergence of antibiotic resistance to human pathogens. In general, identification of these antimicrobial compounds such as bacteriocins and lipopeptides in culture filtrates is accomplished by MALDI-TOF analysis or filtrate fractionation followed by HPLC separation and ESI -MS/MS analysis (Pabel et al., 2003; Kim et al., 2004; Caputo et al., 2011).In this work we describe a LC/MS method for the rapid screening of antimicrobial compounds produced by Bacillus strain TR50. By using HPLC separation and High Resolution MS analysis on a benchtop Orbitrap(TM)-based mass spectrometer equipped with a collision chamber we identified sixteen compounds belonging to three family of lipopeptides in the cell-free supernatant directly.This approach is herein described and proposed as a high throughput tool for the rapid identification of antimicrobial and antifungal compounds in microbial cultures.

Several Bacillus strains, typically isolated from different food sources, represent renowned producers of a multitude of low and high molecular weight compounds, including lipopeptides and macrolactones, with an importance for their antimicrobial activity. The high homology shared by many of these compounds also occurring as closely related isoforms poses a challenge in their prompt detection.Identification and structural elucidation is generally achieved by matrix-assisted laser desorption/ionization (MALDI) or liquid chromatography (LC) coupled to mass spectrometry (MS) after a pre-fractionation and/or purification step of the extract. In this paper we report the application of a method based on LC separation and high-resolution Orbitrap(TM)-based MS for the rapid screening of raw filtrate of the strain Bacillus subtilis TR50 endowed with antimicrobial activity, without requiring any sample pre-treatment.Upon direct analysis of the cell-free filtrate of Bacillus subtilis TR50 by high-resolution mass spectrometry (HRMS), different compounds families, that proved to exert a remarked antimicrobial activity against several foodborne pathogens, can be readily displayed along the chromatographic run. Among them, three different classes were identified and characterized belonging to the iturin, fengycin and surfactin groups. The high resolving power and accurate mass accuracy provided by the HRMS system in use ensured an enhanced selectivity compared to other mass spectrometers. In addition, after activation of the HCD cell, the HR-MS/MS spectra can provide insights in the structural elucidation of several compounds.The acquisition of HRMS spectra of raw filtrates of subtilis strains allows untargeted analysis of the major classes of compounds produced to be performed, thus facilitating identification of other unknown bioactive molecules after retrospective analysis. These features make this approach a fast tool applicable to the rapid screening and further identification of antimicrobial compounds released by Bacillus strains in raw filtrates

Washing water used for processing fruits and vegetables can convey spoilage fungi and bacteria. The common procedure to reduce microbial contamination involves the use of chlorine based compounds. Recently, electrolyzed water (EW) has been evaluated as an alternative measure in controlling microbial spoilage contamination occurring during washing steps. This work reviews results related to the application of EW for controlling microbial viability responsible for decay development during storage period. EW produced with sodium bicarbonate as electrolyte reduced Penicillium spp. population in water and, consequently, green mould decay in citrus fruits; the use of sodium chloride in EW production inactivated spores of Fusarium sp. in water and reduced pineapple decay during storage at 12 degrees C for 20 days as well as controlled yeast and mould population in date fruit up to six months of cold storage. EW was also found effective in controlling spoilage bacteria on ready-to-eat produce. Pseudomonas fluorescens, Pan toea agglomerans, and Rhanella aquatilis were undetectable in electrolyzed process water amended with sodium chloride although similar treatment slightly reduced Erwinia carotovora load inoculated onto lettuce. EW at low free chlorine concentration reduced viability of Pseudomonas spp. and psychrotrophic bacteria in both simulated and industrial washing water. EW treatment of fresh cut lettuce dipped in microbial contaminated water reduced Pseudomonas spp. of about 1 log cfu g(-1) delaying spoilage symptoms that occurred early in untreated vegetables. These results demonstrate that the use of EW can control spoilage microorganisms in washing water, reduce cross contamination phenomena and delay fruit and vegetable decay.

The global interest in saving food resources is leading to recycle wasted-food materials to extract useful nutrients. In dairy industry, the recycling of whey proteins determines their utilization in the healthy-addressed foods, which, however, can cause immunological responses in allergic subjects. In this work, a whey protein concentrate (WPC) was alternatively hydrolyzed with pepsin, papain, trypsin and rennin in order to attenuate or abolish the ?-lactoglobulin (BLG) antigenicity. The electrophoretic profiles of both pepsin and papain WPC hydrolysates proved the disappearance of the BLG band, even though a slight antigenicity was still found by ELISA. Pepsin hydrolysates, filtered through a 10-kDa cut-off membrane, did not produce immunological response. A deeper investigation carried out on pepsin digested and ultrafiltered samples by LC-MS/MS showed the disappearance of the immunoreactive BLG-fragment IVTQMKGLDIQKVAGTW. The remaining peptides, partially overlapped to major IgE binding epitopes, were not able to give immunoreactivity response. The combined WPC pepsin digestion with ultrafiltration confirmed to be a user-friendly strategy to reduce markedly the WPC antigenicity. The improvement of this two-steps process could be used to produce novel hypoallergenic infant food formulas.

Several traditional dairy products are still fermented using unselected starter cultures largely represented by natural whey cultures (NWCs). Microbial biodiversity present in NWCs employed for the production of high moisture Mozzarella cheese areresponsible for differences in primary and secondary proteolysis as well as for the production of aroma compounds. In this work, Illumina (MiSeq)16S rDNA metagenomics analysis of raw milk samples and derived NWCs,was employed to investigate microbial changes occurring during natural fermentation.The microbiota composition, reported as average Operational Taxonomic Units (OUTs)per sample, showed significant differences immediately after the end of the first fermentation step (2h at 45°C) and after the firstandsecond overnight fermentations at 37°C (Pairwise-Wilconoxtest, p value 0.008, 0.008 and 0.014 respectively). More than 100 different OTUs were identified in raw milk samples, which were reduced to 32 at the end of the second overnight fermentation step. This switch in natural microbiota were mainly represented by Pseudomonadaceae (75%) and Lactococcusgarviae (7%) in raw milk to more than 80% of Streptococcusspp. and homofermentative Lactobacillus spp. after the second or third overnight fermentation steps. The application of metagenomic approach allowed us to identify 54 bacterial genus and 29 species and to monitor in detail changes occurring in microbiota when a NWC is produced from raw milk. Many of these species, accounting for less than 1% of total average OTUs per sample, were never reported isolated from and/or identified in milk or in NWC samples. In addition, the technological fermentation approach applied in this study made it feasible to enrich NWCs of useful lactic acid bacteria occurring in low amount in raw milk samples.

The present work focuses on the idea to prevent and/or inhibit the colonization of implant surfaces by microbial pathogens responsible for post-operative infections, adjusting antimicrobial properties of the implant surface prior to its insertion. An antibacterial coating based on chitosan and silver was developed by electrodeposition techniques on poly(acrylic acid)-coated titanium substrates. When a silver salt was added during the chitosan deposition step, a stable and scalable silver incorporation was achieved. The physico-chemical composition of the coating was studied by X-ray photoelectron spectroscopy (XPS), while atomic force microscopy in intermittent contact mode (ICAFM) was used to explore the coating morphology. The amount of silver released from the coating up to 21 days was evaluated by inductively coupled plasma mass spectrometry (ICP-MS). The capability of the proposed coating to interact in vitro with the biological environment in terms of compatibility and antibacterial properties was assessed using MG-63 osteoblast-like cell line and S. aureus and P. aeruginosa strains, respectively. These studies revealed that a coating showing a silver surface atomic percentage equal to 0.3% can be effectively used as antibacterial system, while providing good viability of osteoblast-like cells after 7 days. The antibacterial effectiveness of the prepared coating is mainly driven by a contact killing mechanism, although the low concentration of silver released (below 0.1 ppm up to 21 days) is enough to inhibit bacterial growth, advantaging MG-63 cells in the race for the surface.

La qualità e la shelf-life della mozzarella tradizionale è correlata ai cambiamentidella composizione chimico-fisica e microbiologica del liquido di governo, nel quale siriversano zuccheri, peptidi e sali minerali provenienti dalla mozzarella. Il liquido di governo,normalmente costituito da acqua e sale, si trasforma così in un substrato di crescita in cuiproliferano differenti popolazioni microbiche, incluse quelle alterative e patogene. Ne derivache il controllo microbiologico del liquido di governo rappresenta uno dei fattori principaliper la salubrità e la shelf-life della mozzarella durante la fase di conservazione e distribuzione.Una delle possibilità per contrastare l'alterazione della mozzarella è rappresentata dall'utilizzodi un siero a composizione definita e costante. Nel presente lavoro il liquido di governo dibocconcini di mozzarella è stato sostituito con soluzioni di siero neutralizzate, pastorizzatee gelificate differenti concentrazioni di alginato di sodio (siero-alginato). Le mozzarelle,conservate a 4°C per 25 giorni, sono state valutate per i parametri reologici, chimici emicrobiologici. I risultati non hanno evidenziato significative variazioni di pH del sieroalginatofino a 7 giorni di frigoconservazione. A differenza del liquido di governo che tendead acidificarsi raggiungendo valori di pH pari a 5,75 già al settimo giorno di frigoconservazione,il siero-alginato ha mostrato maggiore stabilità mostrando simili valori di pH dopo 17giorni (5,96); trend di acidificazione simili sono stati registrati anche nelle rispettivemozzarelle. Le mozzarelle in siero-alginato hanno registrato una minore deformazione epiù elevate percentuali di recupero quando sottoposte a sforzo meccanico, rispetto ai valoriregistrati per le mozzarelle controllo. Inoltre, le mozzarelle controllo rilasciavano circa il5% in più del liquido al settimo giorno di frigoconservazione, rispetto alle mozzarelleconservate in siero alginato probabilmente a causa dell'assorbimento di liquido di governoall'interno della matrice caseinica. I dati microbiologici non hanno rivelato significativedifferenze tra i campioni in liquido di governo e quelli in siero alginato, sia nelle cineticheche nei valori della popolazioni microbiche analizzate. I risultati ottenuti in questo lavoro,sebbene preliminari, dimostrano che il siero-alginato può essere utilizzato come metodo diconservazione alternativo al liquido di governo per stabilizzare almeno i parametri strutturalie chimici della mozzarella. Inoltre, l'implementazione in caseificio di tale metodo diconservazione, favorirebbe il parziale riutilizzo del siero riducendo gli onerosi costi del suosmaltimento.

The Institute of Sciences of Food Production (ISPA) is a centre of excellence acting in the fields of scientific research, innovation and technology transfer aimed to improve safety and quality of agro-food products. The Institute policy targets continuous innovation and socio-economic growth of territories by building and implementing strategic cooperative partnerships with world leader Institutions in the area of agro-food (FAO, EFSA, FSA, USDA, etc.) and with SMEs and large enterprises as well as local universities and Consortia of P.D.O. products within funding programs promoted by local, national and international bodies.

This work aimed to evaluate ozone effectiveness in reducing viable spoilage bacteria load throughout high moisture (HM) Mozzarella cheese-making process. At first, Mozzarella cheese samples were packaged with ozonated water (2 mg/L), stored at low temperature and monitored during shelf life. In a following phase cheese samples were put, before packaging, in direct contact with ozonated water at ozone concentrations of 2, 5 and 10 mg/L for 60 min. Then, gaseous ozone at concentrations of 10, 20 and 30 mg/m3 for different times were tested. In these experiments ozone was not effective in surface microbiological decontamination of cheeses. In all cases, there was no increase in the formation of primary and secondary lipid oxidation products.Finally, we treated only the cooling water, artificially contaminated with Pseudomonas spp. strains, obtaining a reduction of about four Log cfu/mL in the microbial load. HM Mozzarella cheeses cooled with pre-treated water was characterized by a low microbial load and an increased shelf life.We concluded that ozone can be successfully applied in Mozzarella cheese processing only to decontaminate water contaminated by potential spoilage bacteria before that it comes into contact with the product.Industrial relevance: HM Mozzarella cheese is a very popular fresh product. Microbiological contamination that occurs after the stretching step when HM Mozzarella cheese comes into contact with cooling water and preserving liquid heavily affects its shelf-life. For this reason, it was decided to investigate whether ozone applications can be extended to HM Mozzarella cheese-making process, as already occurs in other food industry sectors. This study demonstrates that water acting as carrier of spoilage microorganisms can be successfully treated with ozone in order to reduce microbial cheese contamination in each processing step after stretching and shaping.