Microbial dynamics and quality characteristics of spontaneously fermented salamis produced by replacing pork fat with avocado pulp.

The aim of this study was to develop a novel and healthier fermented meat product by replacing pork fat with avocado pulp (AVP) during salami production. Experimental salamis were produced under laboratory conditions by substituting pork fat with AVP partially (10-AVP) and totally (20-AVP), while control salamis (CTR) remained AVP-free. The microbial composition of control and experimental salamis was assessed using a combined culture-dependent and -independent approach. Over a 20-days ripening period, lactic acid bacteria, coagulase-negative staphylococci, and yeasts dominated the microbial community, with approximate levels of 9.0, 7.0 and 6.0 log CFU/g, respectively. Illumina technology identified 26 taxonomic groups, with leuconostocs being the predominant group across all trials [constituting 31.26-59.12 % of relative abundance (RA)]. Gas Chromatography-Mass Spectrometry (GC-MS) analysis revealed changes in fatty acid composition and volatile organic compounds due to the substitution of pork fat with AVP. Specifically, monounsaturated fatty acids and terpene compounds increased, while saturated fatty acids and lipid oxidation products decreased. Although AVP influenced the sensory characteristics of the salamis, the highest overall satisfaction ratings were observed for the 10-AVP salamis. Consequently, substituting pork fat with AVP emerges as a viable strategy for producing healthier salamis and diversifying the meat product portfolio.

Enhancing the quality and safety of Nocellara del Belice green table olives produced using the Castelvetrano method.

The Castelvetrano method is the most widely used among the various table olive processing styles in Sicily. After debittering, the product is stored at low temperatures to prevent the growth of undesirable microorganisms. In an effort to enhance the production process, yeast isolates underwent genotypic characterization and technological screening. The screening process identified two yeast strains Candida norvegica OC10 and Candida boidinii LC1, which can grow at low temperatures and tolerate high pH values (up to 10) and salinity [10% (w/v)]. During the monitoring period, the inoculated trials showed limited presence of spoilage/pathogenic microorganisms. Additionally, the yeasts limited oxidative phenomena and softening of the drupes. The organic compounds detected were higher in the inoculated trials than in the control, and cold storage induced aromatic decay, which was less pronounced in the trial inoculated with C. norvegica. Sensory analysis revealed that the inoculated trials scored higher in sweetness, hardness and crispness.

Description of Ewiss cheese, a new ewe milk cheese processed by Swiss cheese manufacturing techniques: Microbiological, physicochemical, and sensory aspects.

Typically, Swiss-type cheese is made from cow milk. However, in the present work an attempt to expand the sheep supply chain and product offering in this field was made by developing a new type of cheese using Swiss-type cheese technology. The cheese was manufactured under industrial conditions, and fermentations were carried out using freeze-dried commercial starters that are traditionally used in the production of Swiss cheese. Two experimental "Ewiss cheese" (EC) products were produced using raw milk (RM) and pasteurized milk (PM), respectively. Fourteen microbial groups were investigated by plate counts from curd until ripened cheeses. According to microbiological analyses, no statistically significant differences were found between the 2 productions with respect to the group of lactic acid bacteria (LAB). The curds were mainly characterized by mesophilic LAB cocci (7.45 log10 cfu/g in RM-EC and 7.33 log10 cfu/g in PM-EC). However, at the end of the ripening period (9 mo), the cheeses exhibited a higher presence of mesophilic LAB rods. Undesired microbiological groups were found only in the curd of raw milk cheese in the range of 104 to 105 cfu/g, but they were reaching undetectable levels by plate count in the cheese at the end of ripening. The RM-EC and PM-EC were characterized by 76% and 68% of DM, respectively. These cheeses contained 29.30% and 34.36% of protein, and 51.31% and 50.38% of fat, respectively. Textural analysis showed differences in terms of hardness, chewiness, and gumminess between the experimental cheeses and Swiss cheese sold on the market. These differences could be attributed to the higher protein content of ewe milk. The main fatty acids in the cheeses were palmitic acid, myristic acid, oleic acid, and capric acid. Among the organic acids, RM-EC had higher concentrations of lactic acid, whereas PM-EC was higher in propionic acid. The ewe cheeses emitted 46 volatile compounds, including acids, aldehydes, ketones, esters, alcohols, and other compounds. The PM-EC was characterized by the main compounds of Swiss-type cheese: acetic acid, butyric acid, ethyl butyrate, ethyl caproate, propanoic acid, and tetramethylpyrazine. Sensory evaluation showed that the new dairy products were generally appreciated, and PM-EC was the most preferred by the judges. This research has enabled the development of new ewe milk products, which could stimulate the valorization of a sector that has been long neglected and still has a large margin of improvement.

Effect of grape pomace from red cultivar 'Nero d'Avola' on the microbiological, physicochemical, phenolic profile and sensory aspects of ovine Vastedda-like stretched cheese.

AIMS: The purpose of this study was to functionalize an ovine stretched cheese belonging to 'Vastedda' typology with red grape pomace powder (GPP) of Nero d'Avola cultivar and to characterize the microbiological, physicochemical, phenolic profile and sensory characteristics of the final cheeses. METHODS AND RESULTS: Before cheeses production, GPP was characterized for its microbiological profile, antibacterial activity and polyphenolic content. No colonies of bacteria and yeasts were detected in the GPP. GPP showed a large inhibition spectrum against spoilage and pathogenic bacteria. Three classes of polyphenolic compounds belonging to flavan-3-ols, flavonol and phenolic acids were identified. Two cheeses [0 and 1% (w w-1 ) of GPP] were produced with pasteurized ewe's milk and commercial starter cultures. Plate counts and randomly amplified polymorphic DNA analysis demonstrated the ability of the starter strains to drive the fermentation process in the presence of GPP. GPP enrichment resulted in an increase of protein, phenolic compounds, sensory traits and reduced fat. CONCLUSIONS: GPP addition to cheese represents an optimal strategy for the valorization of winemaking by-products and to obtain polyphenol-enriched cheese. SIGNIFICANCE AND IMPACT OF THE STUDY: This study allowed to achieve an ovine cheese with specific physicochemical, nutraceutical and sensorial characteristics able to enlarge the functional dairy product portfolio.

Use of sequentially inoculation of Saccharomyces cerevisiae and Hanseniaspora uvarum strains isolated from honey by-products to improve and stabilize the quality of mead produced in Sicily.

Mead is a beverage produced by alcoholic fermentation of honey-must. The starter yeasts that are commonly used for the alcoholic fermentation of honey-must are oenological Saccharomyces cerevisiae strains. The objective of the present work was, for the first time, to apply yeasts of honey by-products origin to evaluate the influences the taste-olfactory attributes of mead. For this purpose, three experimental productions were set up, which included: (i) single inoculation of S. cerevisiae; (ii) single inoculation of Hanseniaspora uvarum; (iii) sequential inoculation of H. uvarum/S. cerevisiae. Two control trials were performed, using a commercial strain of S. cerevisiae of oenological origin and a spontaneous fermentation. The results of the chemical parameters showed differences between the trials in terms of residual sugars, acetic acid, glycerol, ethanol and volatile organic compounds. Sensorial analysis also showed a high heterogeneity among trials. The attributes of sweetness, honey and floral were found in mead fermented with H. uvarum, whereas all meads obtained with S. cerevisiae were dry, balanced and without off-odors and off-flavours. The results obtained showed that the controlled application of conventional and non-conventional yeast strains isolated from honey by-products origin could be a promising approach to improve the quality of meads.

Technological screening and application of Saccharomyces cerevisiae strains isolated from fermented honey by-products for the sensory improvement of Spiritu re fascitrari, a typical Sicilian distilled beverage.

"Spiritu re fascitrari" is a Sicilian alcoholic beverage obtained through distillation of a decoction of spontaneously fermented honey by-products (FHP). The production process often leads to sensorial defects due to the unstable alcoholic fermentation. The objective of this work was to select Saccharomyces cerevisiae strains from FHP to be used as starter in decoction fermentation. Based on chemical, microbiological and technological data, from a total of 91 strains three S. cerevisae were selected for further testing to produce FHP at laboratory scale level. After FHP distillation, the analysis of volatile organic compounds showed a complex mixture of sensory active molecules, mainly alcohols and aldehydes. Among the alcohols, 3-methyl-1-butanol, 2-methyl-1-butanol, phenylethyl alcohol, hexadecanol and octadecanol were found at the highest concentrations. Among the carboxylic acids, acetic acid was mainly detected in the spontaneously fermented samples. FHP fermented with the three selected strains were not characterized by the presence of off-odors or off-flavours. The results obtained in this work demonstrate that the selected S. cerevisiae strains are promising starters to stabilize the production of distilled alcoholic beverages produced from honey by-products.

In-Depth Investigation of the Safety of Wooden Shelves Used for Traditional Cheese Ripening.

The main goal of this research was to characterize the bacterial diversity of the wooden boards used for aging traditional Sicilian cheeses and to evaluate whether pathogenic bacteria are associated with these surfaces. Eighteen cheese dairy factories producing three traditional cheese typologies (PDO Pecorino Siciliano, PDO Piacentinu Ennese, and Caciocavallo Palermitano) were selected within the region of Sicily. The wooden shelf surfaces were sampled by a destructive method to detach wood splinters as well as by a nondestructive brushing to collect microbial cells. Scanning electron microscopy showed the presence of almost continuous bacterial formations on the majority of the shelves analyzed. Yeasts and fungal hyphae were also visualized, indicating the complexity of the plank communities. The amplicon library of the 16S rRNA gene V3-V4 region was paired-end sequenced using the Illumina MiSeq system, allowing the identification of 14 phyla, 32 classes, 52 orders, 93 families, and 137 genera. Staphylococcus equorum was identified from all wooden surfaces, with a maximum abundance of 64.75%. Among cheese-surface-ripening bacteria, Brevibacterium and Corynebacterium were detected in almost all samples. Several halophilic (Halomonas, Tetragenococcus halophilus, Chromohalobacter, Salimicrobium, Marinococcus, Salegentibacter, Haererehalobacter, Marinobacter, and Idiomarinaceae) and moderately halophilic (Salinicoccus, Psychrobacter, and Salinisphaera) bacteria were frequently identified. Lactic acid bacteria (LAB) were present at low percentages in the genera Leuconostoc, Lactococcus, Lactobacillus, Pediococcus, and Streptococcus. The levels of viable microorganisms on the wooden shelves ranged between 2.4 and 7.8 log CFU/cm2. In some cases, LAB were counted at very high levels (8.2 log CFU/cm2). Members of the Enterobacteriaceae family were detected in a viable state for only six samples. Coagulase-positive staphylococci, Salmonella spp., and Listeria monocytogenes were not detected. Seventy-five strains belonged to the genera Leuconostoc, Lactococcus, Pediococcus, Enterococcus, Lactobacillus, and Weissella. IMPORTANCE This study provides evidence for the lack of pathogenic bacteria on the wooden shelves used to ripen internal bacterially ripened semihard and hard cheeses produced in Sicily. These three cheeses are not inoculated on their surfaces, and surface ripening is not considered to occur or, at least, does not occur at the same extent as surface-inoculated smear cheeses. Several bacterial groups identified from the wooden shelves are typically associated with smear cheeses, strongly suggesting that PDO Pecorino Siciliano, PDO Piacentinu Ennese, and Caciocavallo Palermitano cheese rind contributes to their final organoleptic profiles.

Non-conventional yeasts from fermented honey by-products: Focus on Hanseniaspora uvarum strains for craft beer production.

The increasing interest in novel beer productions focused on non-Saccharomyces yeasts in order to pursue their potential in generating groundbreaking sensory profiles. Traditional fermented beverages represent an important source of yeast strains which could express interesting features during brewing. A total of 404 yeasts were isolated from fermented honey by-products and identified as Saccharomyces cerevisiae, Wickerhamomyces anomalus, Zygosaccharomyces bailii, Zygosaccharomyces rouxii and Hanseniaspora uvarum. Five H. uvarum strains were screened for their brewing capability. Interestingly, Hanseniaspora uvarum strains showed growth in presence of ethanol and hop and a more rapid growth than the control strain S. cerevisiae US-05. Even though all strains showed a very low fermentation power, their concentrations ranged between 7 and 8 Log cycles during fermentation. The statistical analyses showed significant differences among the strains and underlined the ability of YGA2 and YGA34 to grow rapidly in presence of ethanol and hop. The strain YGA34 showed the best technological properties and was selected for beer production. Its presence in mixed- and sequential-culture fermentations with US-05 did not influence attenuation and ethanol concentration but had a significant impact on glycerol and acetic acid concentrations, with a higher sensory complexity and intensity, representing promising co-starters during craft beer production.

Molecular analysis of the dominant lactic acid bacteria of chickpea liquid starters and doughs and propagation of chickpea sourdoughs with selected Weissella confusa.

Fermented chickpea liquid is used as a leavening agent in chickpea bread production. In the present study, traditional chickpea liquid starter and dough samples were collected from bakeries in Turkey and microbiologically investigated. Culture-independent analysis for microbiota diversity, performed by MiSeq Illumina, identified Clostridium perfringens as major group in all samples, while Weissella spp. Dominated LAB community. A culture-dependent methodology was applied and 141 isolates were confirmed to be members of the LAB group based on 16s rRNA gene sequence analysis. In particular, 11 different LAB species were identified confirming the high frequency of isolation of weissellas, since Weissella confusa and Weissella cibaria constituted 47.8 and 12.4%, respectively, of total LAB isolated. The other species were Enterococcus faecium, Enterococcus lactis, Lactobacillus brevis, Lactobacillus plantarum, Leuconostoc mesenteroides, Leuconostoc mesenteroides subsp. Dextranium, Pediococcus acidilactici, Pediococcus pentosaceus and Streptococcus lutetiensis. Due to high frequency of isolation, W. confusa strains were investigated at technological level and W. confusa RL1139 was used as mono-culture starter in the experimental chickpea sourdough production. Chemical and microbiological properties, as well as volatile organic compounds (VOCs) of the chickpea liquid starters and doughs were subjected to a multivariate analysis. Control and W. confusa inoculated chickpea liquid starter and dough samples were close to each other in terms of some characteristics related to chemical, microbiological and VOCs profile, but the inoculated sourdough showed a higher generation of certain VOCs, like butanoic acid (81.52%) and ethyl acetate (8.15%) than control sourdough. This is important in order to maintain typical characteristics of the traditional chickpea dough, but at the same time improving the aroma profile. This work demonstrated that W. confusa RL1139 can be applied at large scale production level without compromising the typical characteristics of the final product.

Persistence of a mixed lactic acid bacterial starter culture during lysine fortification of sourdough breads by addition of pistachio powder.

Pistachio powder was added to flour or semolina to evaluate its contribution to increase the amount of lysine in bread. Bread production was carried out by sourdough technology using a selected 3-species (Lactobacillus sanfranciscensis/Leuconostoc citreum/Weissella cibaria) lactic acid bacterial (LAB) starter culture. All sourdoughs were subjected to a long-time fermentation (21 h) and showed levels of LAB around 109 CFU/g, indicating the suitability of pistachio powder for lactic fermentation. Yeasts were also detected, in particular in semolina trials. MiSeq Illumina technology was applied to investigate the bacterial composition of sourdoughs evidencing a different distribution of LAB species among the trials with Lactobacillus as major LAB group in almost all sourdoughs. Physicochemical parameters were comparable among the trials. After baking, pistachio powder was found not to influence the height of the breads, but pistachio breads were more firm than control breads. Color of the breads, void fraction and cell density, were influenced by pistachio powder. The amount of lysine increased consistently thanks to pistachio supplementation which also determined a higher presence of o-xylene, p-cymene and limonene and the appearance of α-pinene and 1-octen-3-ol in breads. Sensory tests showed the best appreciation scores for the breads produced with flour and pistachio powder.

Evolution of indigenous starter microorganisms and physicochemical parameters in spontaneously fermented beef, horse, wild boar and pork salamis produced under controlled conditions.

The present work was carried out to evaluate the microbiological and physicochemical composition of salamis produced with the meat of beef, horse, wild boar and pork. Salami productions occurred under controlled laboratory conditions to exclude butchery environmental contaminations, without the addition of nitrate and nitrite. All trials were monitored during the ripening (13 °C and 90% relative humidity) extended until 45 d. The evolution of physicochemical parameters showed that beef and pork salamis were characterized by a higher content of branched chain fatty acids (FA) and rumenic acid than horse and wild boar salamis, whereas the last two productions showed higher values of secondary lipid oxidation. Plate counts showed that lactic acid bacteria (LAB), yeasts and coagulase-negative staphylococci (CNS) populations dominated the microbial community of all productions with Lactobacillus and Staphylococcus as most frequently isolated bacteria. The microbial diversity evaluated by MiSeq Illumina showed the presence of members of Gammaproteobacteria phylum, Moraxellaceae family, Acinetobacter, Pseudomonas, Carnobacterium and Enterococcus in all salamis. This study showed the natural evolution of indigenous fermented meat starter cultures and confirmed a higher suitability of horse and beef meat for nitrate/nitrite free salami production due to their hygienic quality at 30 d.

Formation and Characterization of Early Bacterial Biofilms on Different Wood Typologies Applied in Dairy Production.

The main hypothesis of this work was that Sicilian forestry resources are suitable for the production of equipment to be used in cheese making and indigenous milk lactic acid bacteria (LAB) are able to develop stable biofilms providing starter and nonstarter cultures necessary for curd fermentation and cheese ripening, respectively. Hence, the present work was carried out with deproteinized whey to evaluate LAB biofilm formation on different woods derived from tree species grown in Sicily. Microbiological and scanning electron microscopy analyses showed minimal differences in microbial levels and compositions for the neoformed biofilms. The specific investigation of Salmonella spp., Listeria monocytogenes, Escherichia coli, coagulase-positive staphylococci (CPS), and sulfite-reducing anaerobes did not generate any colony for all vats before and after bacterial adhesion. LAB populations dominated all vat surfaces. The highest levels (7.63 log CFU/cm2) were registered for thermophilic cocci. Different colonies were characterized physiologically, biochemically, and genetically (at strain and species levels). Six species within the genera Enterococcus, Lactobacillus, Lactococcus, and Streptococcus were identified. The species most frequently present were Lactobacillus fermentum and Lactococcus lactis LAB found on the surfaces of the wooden vats in this study showed interesting characteristics important for dairy manufacture. To thoroughly investigate the safety of the wooden vat, a test of artificial contamination on new Calabrian chestnut (control wood) vats was carried out. The results showed that LAB represent efficient barriers to the adhesion of the main dairy pathogens, probably due to their acidity and bacteriocin generation.IMPORTANCE This study highlights the importance of using wooden vats for traditional cheese production and provides evidence for the valorization of the Sicilian forest wood resources via the production of dairy equipment.

Shelf life evaluation of fresh-cut red chicory subjected to different minimal processes.

Microbiological, chemical and physical parameters of minimally processed red chicory (Cichorium intybus L.) subjected to two different transformation processes were investigated. A classic ready-to-eat (RTE) process (P1) and a production without cutting (P2) were monitored during refrigerated (4 °C) storage (15 d). Total mesophilic microorganisms, total psychrotrophic microorganisms and pseudomonads were detected at the highest cell densities in all samples. Presumptive Pseudomonas population dominated the cultivable microbial community of RTE red chicory and were characterized genetically. Twenty-two randomly amplified polymorphic DNA (RAPD) types were investigated by 16S rRNA gene sequencing, resulting in members of Rahnella and Pseudomonas. The identification of Pseudomonas species was further determined by sequencing of gyrB, rpoB and rpoD genes resulting in 16 species. A highest visual quality and a lower weight loss and colour variation were registered for P2, while soluble solid, nitrate and ascorbic acid contents were not affected by processing and storage. The integrated microbiological, chemical and physical approach applied in this study demonstrated the longer shelf-life of P2 red chicory.

Enteric bacteria of food ice and their survival in alcoholic beverages and soft drinks.

This study aimed to evaluate the levels of enteric bacteria in ice cubes produced in different environments (home-made, prepared in bars and pubs with ice machines and produced in industrial plants) and to determine their survival in different alcoholic beverages and soft drinks. Members of the Enterobacteriaceae family were found in almost all samples analysed. All industrial and the majority of home-made samples did not contain coliforms. Enterococci were not identified in domestic samples while they were detected in two industrial and three bar/pub samples. The samples collected from bars and pubs were characterized by the highest levels of enteric bacteria. Fourteen strains representing 11 species of eight bacterial genera were identified, some of which are known agents of human infections. The most numerous groups included Enterococcus and Stenotrophomonas. The survival of Enterococcus faecium ICE41, Pantoea conspicua ICE80 and Stenotrophomonas maltophilia ICE272, that were detected at the highest levels (100-400 CFU/100 mL thawed ice) in the ice cubes, was tested in six drinks and beverages characterized by different levels of alcohol, CO2, pH and the presence of antibacterial ingredients. The results showed a species-specific behaviour and, in general, a reduction of the microbiological risks associated with ice after its transfer to alcoholic or carbonated beverages.

Evaluation of different conditions to enhance the performances of Lactobacillus pentosus OM13 during industrial production of Spanish-style table olives.

The main objective was to set up a methodology to improve the high volume production of green table olives, cv. Nocellara del Belice. Lactobacillus pentosus OM13 was applied during three different industrial processes of table olives as follows: trial one (IOP1) was subjected to an addition of lactic acid until a brine level of pH 7.0 was reached; trial two (IOP2) subjected to same addition of lactic acid as in trial one plus nutrient adjuvant and trial three (IOP3) subjected to same addition as trial two, but with the strain L. pentosus OM13 acclimatized in brine for 12 h before inoculation. These trials were compared against two untreated controls (spontaneously fermented and addition of L. pentosus OM13 only). Within the third day of fermentation, the pH of the brines decreased significantly, reaching pH 4.85 for trial three, pH 5.15 for trial two, and pH 5.92 for trial one. The pH of both controls decreased more slowly, and had values below pH 5.0 only after the fifteenth day of fermentation (control one) and the sixty-fifth day of fermentation (control two). Trial three reached the highest lactic acid bacteria (LAB) concentration on the third day of fermentation. After six days of fermentation, all trials showed similar values of LAB counts that were significantly higher compared to control number one. The result from genotypic identification showed that L. pentosus OM13 was the most frequently isolated in the inoculated trials. Lactobacillus plantarum, Lactobacillus coryniformis and Pediococcus pentosaceous were also detected at very low concentrations. Homoguaiacol, 2-butanol, 4-ethylphenol, phenylethyl alcohol and 4-ethylphenol were the volatile organic compounds detected at the highest levels in all experimental trials. Trial three showed a higher concentration of squalene that was not detected in other trials. The highest sensory scores of green olive aroma and overall satisfaction were found for all experimental olives, especially for those of trial one and trial two, that differed significantly from the untreated controls. This study provides evidence that the addition of lactic acid, nutrient adjuvants and, most importantly, the acclimatization of LAB cells significantly shortens the acidification process of olive brine, and improves safety and sensory quality. Shorter acidification processes result in a more rapid transformation of table olives, with reduced commodity loss and lower costs of production compared to conventional manufacturing protocols.

Inhibitory Activity and Chemical Characterization of Daucus carota subsp. maximus Essential Oils.

The essential oils (EOs) of green seeds from Daucus carota subsp. maximus growing wild in Pantelleria Island (Sicily, Italy) were characterized. EOs were extracted by steam distillation, examined for their inhibitory properties against food-borne Gram-positive and Gram-negative bacteria and analyzed for the chemical composition by gas chromatography (GC) and mass spectrometry (MS). Undiluted EOs showed a large inhibition spectrum against Gram-positive strains and also vs. Acinetobacter spp. and Stenotrophomonas maltophilia. The minimum inhibition concentration (MIC) was in the range 1.25 - 2.50 µl/ml for the most sensitive strains. The chemical analysis indicated that D. carota subsp. maximus EOs included 34 compounds (five monoterpene hydrocarbons, six oxygenated monoterpenes, 14 sesquiterpene hydrocarbons, four oxygenated sesquiterpenes, camphorene and four other compounds), accounting for 95.48% of the total oil, and that the major chemicals were carotol, ß-bisabolene, and isoelemicin.

Selection of Amine-Oxidizing Dairy Lactic Acid Bacteria and Identification of the Enzyme and Gene Involved in the Decrease of Biogenic Amines.

Accumulation of biogenic amines (BAs) in cheese and other foods is a matter of public health concern. The aim of this study was to identify the enzyme activities responsible for BA degradation in lactic acid bacteria which were previously isolated from traditional Sicilian and Apulian cheeses. The selected strains would control the concentration of BAs during cheese manufacture. First, 431 isolates not showing genes encoding the decarboxylases responsible for BA formation were selected using PCR-based methods. Ninety-four out of the 431 isolates degraded BAs (2-phenylethylamine, cadaverine, histamine, putrescine, spermine, spermidine, tyramine, or tryptamine) during cultivation on chemically defined medium. As shown by random amplification of polymorphic DNA-PCR and partial sequencing of the 16S rRNA gene, 78 of the 94 strains were Lactobacillus species (Lactobacillus casei, Lb. fermentum, Lb. parabuchneri, Lb. paracasei, Lb. paraplantarum, and Lb. rhamnosus), Leuconostoc species (Leuconostoc lactis and Ln. mesenteroides), Pediococcus pentosaceus, Lactococcus lactis, Streptococcus species (Streptococcus gallolyticus and S. thermophilus), Enterococcus lactis, and Weissella paramesenteroides A multicopper oxidase-hydrolyzing BA was purified from the most active strain, Lb. paracasei subsp. paracasei CB9CT. The gene encoding the multicopper oxidase was sequenced and was also detected in other amine-degrading strains of Lb. fermentum, Lb. paraplantarum, and P. pentosaceus Lb. paracasei subsp. paracasei CB9CT and another strain (CACIO6CT) of the same species that was able to degrade all the BAs were singly used as adjunct starters for decreasing the concentration of histamine and tyramine in industrial Caciocavallo cheese. The results of this study disclose a feasible strategy for increasing the safety of traditional cheeses while maintaining their typical sensorial traits. IMPORTANCE: Because high concentrations of the potentially toxic biogenic amines may be found in traditional/typical cheeses, the safety of these food items should be improved. Lactic acid bacteria selected for the ability to degrade biogenic amines may be used during cheese making to control the concentrations of biogenic amines.

Microbial Activation of Wooden Vats Used for Traditional Cheese Production and Evolution of Neoformed Biofilms.

Three Lactococcus lactis subsp. cremoris strains were used to develop ad hoc biofilms on the surfaces of virgin wooden vats used for cheese production. Two vats (TZ) were tested under controlled conditions (pilot plant), and two vats (TA) were tested under uncontrolled conditions (industrial plant). In each plant, one vat (TA1 and TZ1) was used for the control, traditional production of PDO Vastedda della Valle del Belìce (Vastedda) cheese, and one (TA2 and TZ2) was used for experimental production performed after lactococcal biofilm activation and the daily addition of a natural whey starter culture (NWSC). Microbiological and scanning electron microscopy analyses showed differences in terms of microbial levels and composition of the neoformed biofilms. The levels of the microbial groups investigated during cheese production showed significant differences between the control trials and between the control and experimental trials, but the differences were not particularly marked between the TA2 and TZ2 productions, which showed the largest numbers of mesophilic lactic acid bacterium (LAB) cocci. LAB populations were characterized phenotypically and genotypically, and 44 dominant strains belonging to 10 species were identified. Direct comparison of the polymorphic profiles of the LAB collected during cheese making showed that the addition of the NWSC reduced their biodiversity. Sensory evaluation showed that the microbial activation of the wooden vats with the multistrain Lactococcus culture generated cheeses with sensory attributes comparable to those of commercial cheese. Thus, neoformed biofilms enable a reduction of microbial variability and stabilize the sensorial attributes of Vastedda cheese.

A large factory-scale application of selected autochthonous lactic acid bacteria for PDO Pecorino Siciliano cheese production.

The main hypothesis of this study was that the autochthonous lactic acid bacteria (LAB) selected for their dairy traits are able to stabilize the production of PDO (Protected Denomination of Origin) Pecorino Siciliano cheese, preserving its typicality. The experimental plan included the application of a multi-strain lactic acid bacteria (LAB) culture, composed of starter (Lactococcus lactis subsp. lactis CAG4 and CAG37) and non starter (Enterococcus faecalis PSL71, Lactococcus garviae PSL67 and Streptococcus macedonicus PSL72) strains, during the traditional production of cheese at large scale level in six factories located in different areas of Sicily. The cheese making processes were followed from milk to ripened cheeses and the effects of the added LAB were evaluated on the microbiological, chemico-physical and sensorial characteristics of the final products. Results highlighted a high variability for all investigated parameters and the dominance of LAB cocci in bulk milk samples. The experimental curds showed a faster pH drop than control curds and the levels of LAB estimated in 5-month ripened experimental cheeses (7.59 and 7.27 Log CFU/g for rods and cocci, respectively) were higher than those of control cheeses (7.02 and 6.61 Log CFU/g for rods and cocci, respectively). The comparison of the bacterial isolates by randomly amplified polymorphic DNA (RAPD)-PCR evidenced the dominance of the added starter lactococci over native milk and vat LAB, while the added non starter LAB were found at almost the same levels of the indigenous strains. The sensory evaluation showed that the mixed LAB culture did not influence the majority of the sensory attributes of the cheeses and that each factory produced cheeses with unique characteristics. Finally, the multivariate statistical analysis based on all parameters evaluated on the ripened cheeses showed the dissimilarities and the relationships among cheeses. Thus, the main hypothesis of the work was accepted since the quality parameters of the final cheeses were stabilized, but all cheeses maintained their local typicality.

The influence of the wooden equipment employed for cheese manufacture on the characteristics of a traditional stretched cheese during ripening.

The influence of the wooden equipment used for the traditional cheese manufacturing from raw milk was evaluated on the variations of chemico-physical characteristics and microbial populations during the ripening of Caciocavallo Palermitano cheese. Milk from two farms (A, extensive; B, intensive) was processed in traditional and standard conditions. Chemical and physical traits of cheeses were affected by the farming system and the cheese making technology, and changed during ripening. Content in NaCl and N soluble was lower, and paste consistency higher in cheese from the extensive farm and traditional technology, whereas ripening increased the N soluble and the paste yellow and consistency. The ripening time decreased the number of all lactic acid bacteria (LAB) groups, except enterococci detected at approximately constant levels (10(4) and 10(5) cfu g(-1) for standard and traditional cheeses, respectively), till 120 d of ripening. In all productions, at each ripening time, the levels detected for enterococci were lower than those for the other LAB groups. The canonical discriminant analysis of chemical, physical and microbiological data was able to separate cheeses from different productions and ripening time. The dominant LAB were isolated, phenotypically characterised and grouped, genetically differentiated at strain level and identified. Ten species of LAB were found and the strains detected at the highest levels were Pediococcus acidilactici and Lactobacillus casei. Ten strains, mainly belonging to Lactobacillus rhamnosus and Lactobacillus fermentum showed an antibacterial activity. The comparison of the polymorphic profiles of the LAB strains isolated from the wooden vat with those of the strains collected during maturation, showed the persistence of three enterococci in traditional cheeses, with Enterococcus faecalis found at dominant levels over the Enterococcus population till 120 d; the absence of these strains in the standard productions evidenced the contribution of vat LAB during Caciocavallo Palermitano cheese ripening.