Traditional and innovative approaches to evaluate microbial contribution in long ripened fermented foods: the case of Parmigiano Reggiano cheese

La crescita economica e lo sviluppo delle industrie casearie si basano ampiamente sugli starter microbici utilizzati per produrre la gran parte dei prodotti fermentati industriali. D’altro canto, le colture naturali impiegate nella produzione di formaggi tradizionali sono responsabili di diversi tratti distintivi di qualità, che rendono questi prodotti apprezzabili in tutto il mondo. Nonostante siano stati fatti molti progressi nella comprensione della fisiologia dei batteri lattici e nei processi che indirizzano lo sviluppo di prodotti fermentati di qualità, molto rimane ancora da comprendere. Per questo motivo, è stato scelto come oggetto di questo studio, un formaggio italiano, a Denominazione di Origine Protetta, apprezzato e di rilevanza economica come il Parmigiano Reggiano, allo scopo di ottenere una miglior comprensione di come e quando i batteri lattici siano coinvolti in questi processi. A tale fine, sono stati utilizzati sia approcci tradizionali che innovativi. Nel corso di questa tesi di dottorato, un accurato e laborioso campionamento ha permesso di analizzare un formaggio Parmigiano Reggiano in tutti gli stadi della produzione e nei due anni di stagionatura. Sono state effettuate conte microbiche tradizionali poi confrontate con i dati ottenuti da terreni di conta innovativi e attraverso conte dirette in grado di stimare anche la vitalità cellulare. E’ stato anche valutato il grado di autolisi cellulare tramite conta microbica diretta e grazie alla quantificazione sia del DNA che degli enzimi rilasciati in seguito a lisi cellulare. Inoltre, è stata utilizzata la tecnica Length heterogeneity-PCR per monitorare l’evoluzione delle specie microbiche nel corso di un completo processo di caseificazione. Infine, è stato sviluppato un modello matematico in grado di stabilire l’effetto di NaCl, così come di pH e temperatura sull’attività aminopeptidasica.Future development and economic growth of dairy industry largely depends on microbial starter used to produce the great majority of industrial fermented products. Moreover, natural cultures implied in traditional cheeses manufacturing, are responsible for many good recognizable traits which make them appreciable all over the word. Even if, great progress has been made toward understanding lactic acid bacteria physiology, and processes that drive quality fermented food development, much remains to be learned. For this reason, a Protected Designation of Origin, appreciated and economically important Italian cheese like Parmigiano Reggiano, has been chosen as subject for this study, to try a better understanding of how and when lactic acid bacteria are involved in these processes. To reach this issue, both traditional and innovative approaches have been used. Troughout this Ph.D. thesis, an accurate and laborious sampling allowed to analyse Parmigiano Reggiano cheese in all stages of manufacturing and through two years of ripening. Traditional microbial counts have been attended and compared with data obtained by means of innovative counting media and direct enumeration considering even cell viability. Cell autolysis has been also evaluated by direct microbial count and by the quantification of both cell lysis-released DNA and cell lysis-released enzymes. Moreover, length heterogeneity-PCR has been held to monitor microbial species evolution during a complete cheesemaking process. Furthermore, a mathematical model able to assess the effect of NaCl concentration, as well as pH and temperature on aminopeptidases activities has been performed

Izdvajanje i identifikacija mikroflore iz kravljeg kolostruma tradicionalnim i molekularnim metodama

Colostrum is the first milk produced by the mammary gland during the early postpartum period. In addition to nutrients, antimicrobial and growth factors, bovine colostrum contains valuable microflora, including members of the genera Lactobacillus and Bifidobacterium, which have been used widely in probiotic food. The aim of this work is to identify and quantify the microflora of colostrum samples from two Holstein dairy cows on the first three postpartum days through culture-dependent 16S rRNA gene sequencing and length heterogeneity polymerase chain reaction (LH-PCR) techniques, using an adapted method to extract the total DNA. Twenty-nine strains isolated in synthetic media were identified by their 16S rRNA gene sequences, revealing two potential probiotic strains (Lactobacillus casei and Bifidobacterium pseudolongum). Colostrum samples were subjected to LH-PCR analysis to obtain fingerprint profiles of the microflora. Lactobacillus casei was the main species present in the samples. The probiotic potential of these bacteria for use in fermented and functional foods remains to be evaluated.Kolostrum je prvo mlijeko, koje proizvodi mliječna žlijezda tijekom ranog postpartalnog razdoblja. Osim što sadrži hranjiva, antimikrobne tvari i hormone rasta, kravlji kolostrum sadržava dragocjenu mikrofloru, uključujući sojeve rodova Lactobacillus i Bifidobacterium što se koriste u proizvodnji probiotičke hrane. Svrha je ovoga rada bila identificirati i kvantificirati mikrofloru u uzorcima kolostruma dvaju Holstein mlječnih krava, uzetih tijekom prva tri dana postpartalnog razdoblja. Mikroorganizmi su određeni sekvenciranjem 16S rRNK gena i LH-PCR metodom kojom se ekstrahira ukupna DNK. U sintetičkoj je podlozi izolirano dvadeset i devet sojeva, identificiranih pomoću sekvencija 16S rRNK gena, te su izdvojena dva potencijalno probiotička soja (Lactobacillus casei i Bifidobacterium pseudolongum). Uzorci su kolostruma analizirani LH-PCR metodom kako bi se utvrdio DNK profil mikroflore, te je utvrđeno da u uzorcima prevladava vrsta Lactobacillus casei. Probiotička svojstva ove bakterije za proizvodnju fermentiranih i probiotičkih proizvoda tek treba istražiti

Izdvajanje i identifikacija mikroflore iz kravljeg kolostruma tradicionalnim i molekularnim metodama

Colostrum is the first milk produced by the mammary gland during the early postpartum period. In addition to nutrients, antimicrobial and growth factors, bovine colostrum contains valuable microflora, including members of the genera Lactobacillus and Bifidobacterium, which have been used widely in probiotic food. The aim of this work is to identify and quantify the microflora of colostrum samples from two Holstein dairy cows on the first three postpartum days through culture-dependent 16S rRNA gene sequencing and length heterogeneity polymerase chain reaction (LH-PCR) techniques, using an adapted method to extract the total DNA. Twenty-nine strains isolated in synthetic media were identified by their 16S rRNA gene sequences, revealing two potential probiotic strains (Lactobacillus casei and Bifidobacterium pseudolongum). Colostrum samples were subjected to LH-PCR analysis to obtain fingerprint profiles of the microflora. Lactobacillus casei was the main species present in the samples. The probiotic potential of these bacteria for use in fermented and functional foods remains to be evaluated.Kolostrum je prvo mlijeko, koje proizvodi mliječna žlijezda tijekom ranog postpartalnog razdoblja. Osim što sadrži hranjiva, antimikrobne tvari i hormone rasta, kravlji kolostrum sadržava dragocjenu mikrofloru, uključujući sojeve rodova Lactobacillus i Bifidobacterium što se koriste u proizvodnji probiotičke hrane. Svrha je ovoga rada bila identificirati i kvantificirati mikrofloru u uzorcima kolostruma dvaju Holstein mlječnih krava, uzetih tijekom prva tri dana postpartalnog razdoblja. Mikroorganizmi su određeni sekvenciranjem 16S rRNK gena i LH-PCR metodom kojom se ekstrahira ukupna DNK. U sintetičkoj je podlozi izolirano dvadeset i devet sojeva, identificiranih pomoću sekvencija 16S rRNK gena, te su izdvojena dva potencijalno probiotička soja (Lactobacillus casei i Bifidobacterium pseudolongum). Uzorci su kolostruma analizirani LH-PCR metodom kako bi se utvrdio DNK profil mikroflore, te je utvrđeno da u uzorcima prevladava vrsta Lactobacillus casei. Probiotička svojstva ove bakterije za proizvodnju fermentiranih i probiotičkih proizvoda tek treba istražiti

Development of an Innovative Nutraceutical Fermented Beverage from Herbal Mate (Ilex paraguariensis A.St.-Hil.) Extract

Herbal mate (Ilex paraguariensis A.St.-Hil.) leaves are traditionally used for their stimulant, antioxidant, antimicrobial, and diuretic activity, presenting as principal components polyphenolic compounds. The aim of this work was to develop an innovative, non-dairy, functional, probiotic, fermented beverage using herbal mate extract as a natural ingredient which would also be hypocholesterolemic and hepatoprotective. Among different strains used, Lactobacillus acidophilus was selected as the best for fermentation. The addition of honey positively affected the development of L. acidophilus and the formulated beverage maintained microbial stability during shelf life. Key ingredients in the extract included xanthines, polyphenols and other antioxidants with potential health benefits for the consumer. Caffeine levels and antioxidant activity were also studied. Acceptable levels of caffeine and large antioxidant capacity were observed for the formulation when compared to other antioxidant beverages. An advantage of this product is the compliance to organic claims, while providing caffeine, other phyto-stimulants and antioxidant compounds without the addition of synthetic components or preservatives in the formulation. Sensorial analysis demonstrated that the beverage had good consumer acceptance in comparison to two other similar commercial beverages. Therefore, this beverage could be used as a new, non-dairy vehicle for probiotic consumption, especially by vegetarians and lactose intolerant consumers. It is expected that such a product will have good market potential in an era of functional foods

Mogućnosti primjene probiotika

Probiotics, live cells with different beneficiary characteristics, have been extensivelly studied and explored commercially in many different products in the world. Their benefits to human and animal health have been proven in hundreds of scientific research. Lactobacillus and Bifidobacterium are the main probiotic groups; however, there are reports on the probiotic potential of Pediococcus, Lactococcus, Bacillus and yeasts. Some of the identified probiotic strains exhibit powerful anti-inflammatory, antiallergic and other important properties. Apart from that, the consumption of dairy and non-dairy products stimulates the immunity in different ways. Various food matrices have been used with probiotics, which are briefly documented. In this review, the history of probiotics, their application in the health and food areas and new trends in probiotic products and processes are presented.Probiotici su žive stanice koje imaju različite pozitivne osobine, pa se intenzivno proučavaju i komercijalno primjenjuju u mnogim proizvodima diljem svijeta. U više stotina znanstvenih istraživanja dokazan je njihov pozitivan učinak na zdravlje ljudi i životinja. Glavne su skupine probiotika bakterije vrste Lactobacillus i Bifidobacterium, ali spominju se i probiotička svojstva bakterija vrsta Pediococcus, Lactococcus i Bacillus, te nekih kvasaca. Neki od identificiranih probiotičkih sojeva imaju izraženi protuupalni i antialergijski učinak, pa se konzumiranjem probiotičkih mliječnih i nemliječnih proizvoda povećava imunitet. U radu su ukratko spomenute razne namirnice što se koriste u kombinaciji s probioticima. Također je dan povijesni pregled razvoja probiotika, navedene su mogućnosti njihove primjene u prehrambenoj industriji, a i u zdravstvu, te prikazani novi pravci proizvodnje probiotičkih proizvoda

Biohydrogenation of Linoleic Acid by Lactic Acid Bacteria for the Production of Functional Cultured Dairy Products: A Review

Conjugated linoleic acid (CLA) isomers have attracted significant attention due to their important physiological properties, which have been observed in humans. Many lactic acid bacteria (LAB) demonstrate the ability to produce CLA isomers (C18:2 cis-9, trans-11 and C18:2 trans-10, cis-12) from the linoleic acid (LA) present in milk or in synthetic media. CLA isomers can be synthesized in vitro by LAB using vegetable oils rich in LA. The aim of this review is to present an update on the studies that have been conducted on the production of CLA isomers from LA mainly by LAB and of the factors that influence this conversion (source and concentration of LA and fermentation conditions). In addition, this review presents the relationship between the consumption of CLA isomers and their health benefits in humans such as anti-atherosclerosis and anti-carcinogenic effects. There is considerable variation between the studies concerning the beneficial effects of CLA in animal models, which have not been reflected in human studies. This can be attributed to the differences in the doses of CLA isomers used and to the different sources of CLA. Furthermore, the regulatory and scientific information classifying the physiological properties of CLA, which serve as support for the claims of its potential as a functional ingredient, are presented. More research is needed to determine whether CLA production by LAB can be enhanced and to determine the optimal requirements for these microbial cultures. Furthermore, safety and efficacy of CLA consumption have to be investigated in the future

New Insights into Food Fermentation

Food fermentation has been used for thousands of years for food preservation [...

Sweeteners and sweet taste enhancers in the food industry

Abstract The search for new sweeteners technologies has increased substantially in the past decades as the number of diseases related to the excessive consumption of sugar became a public health concern. Low carbohydrates diets help to reduce ingested calories and to maintain a healthy weight. Most natural and synthetic high potency non-caloric sweeteners, known to date, show limitations in taste quality and are generally used in combination due to their complementary flavor characteristics and physicochemical properties in order to minimize undesirable features. The challenge of the food manufacturers is to develop low or calorie-free products without compromising the real taste of sugar expected by consumers. With the discovery of the genes coding for the sweet taste receptor in humans, entirely new flavor ingredients were identified, which are tasteless on their own, but potentially enhance the taste of sugar. These small molecules known as positive allosteric modulators (PAMs) could be more effective than other reported taste enhancers at reducing calories in consumer products. PAMs could represent a breakthrough in the field of flavor development after the increase in the knowledge of safety profile in combination with sucrose in humans

Editorial: Microbiological Safety and Quality Aspects of Fermented Dairy Products

Fermented dairy products include a wide range of foods that are highly appreciated worldwide. The production processes, the type of milk, practices adopted during feeding, milking, and beyond can affect the quality and safety characteristics of these products. A vast array of microorganisms can be found in milk, and microbial succession during fermentation of multiple products, as well as during cheese ripening contributes to the desired properties of these foods. In addition to sensory and safety aspects, microorganisms present in fermented dairy foods can positively affect the health of people due to their potential probiotic nature and the production of beneficial metabolites such. Despite recent innovations on microbial cultivation techniques, and the widespread use of a myriad of newly developed “omic” techniques, including genomic approaches through the use of High Throughput Sequencing in combination with advanced metabolomics, there are many dairy fermented products with limited information about their microbial composition (diversity) and dynamics (succession). This is particularly true for traditional regional products, which represent a rich niche for discoveries involving microorganisms with better conditions for recovery from disturbances and more reliable and robust for industrial production. The unknown microbiota can reveal new mechanisms of interactions and functions and new bioactive molecules with beneficial effects on the sensory aspect of food and on human health. The goal of this Research Topic was to deepen the current knowledge in the microbial safety and quality aspects of fermented dairy products, gathering studies on the different types of products, practices, and how the microbiota affected the quality and safety attributes of these much-appreciated foods. This Research Topic brings a series of 14 articles related to the microbiological safety and quality aspects of fermented dairy products.as vitamins, antioxidants, and antimicrobial compounds

Sweeteners and sweet taste enhancers in the food industry

<div><p>Abstract The search for new sweeteners technologies has increased substantially in the past decades as the number of diseases related to the excessive consumption of sugar became a public health concern. Low carbohydrates diets help to reduce ingested calories and to maintain a healthy weight. Most natural and synthetic high potency non-caloric sweeteners, known to date, show limitations in taste quality and are generally used in combination due to their complementary flavor characteristics and physicochemical properties in order to minimize undesirable features. The challenge of the food manufacturers is to develop low or calorie-free products without compromising the real taste of sugar expected by consumers. With the discovery of the genes coding for the sweet taste receptor in humans, entirely new flavor ingredients were identified, which are tasteless on their own, but potentially enhance the taste of sugar. These small molecules known as positive allosteric modulators (PAMs) could be more effective than other reported taste enhancers at reducing calories in consumer products. PAMs could represent a breakthrough in the field of flavor development after the increase in the knowledge of safety profile in combination with sucrose in humans.</p></div