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A special issue of Applied Sciences (ISSN 2076-3417). This special issue belongs to the section "Applied Microbiology".

Deadline for manuscript submissions: closed (20 May 2023) | Viewed by 31898

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Prof. Athina S. Tzora

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Dean of the School of Agriculture, Department of Agriculture, University of Ioannina, 47100 Arta, Greece
Interests: microbiology; immunology; food born diseases; food safety; fermented foods; Lactic Acid Bacteria; microbial resistance; selective breeding programs
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Special Issue Information

Dear Colleagues,

Fermented foods have played an important role in the human diet and constitute a special feature of some dietary patterns. Fermented dairy products have been found in human nutrition from ancient times, and nowadays they continue to be fundamental components of a balanced diet. Fermented dairy products such as different varieties of cheeses, which are differentiated according to their organoleptic characteristics, are now available for consumers, and although a small proportion of these products are homemade, most of them are industrially produced. Dairy products constitute an important microbial reservoir that imparts quality characteristics and impacts human health. Food safety is a topic of continued concern, inextricably linked to the production process and quality of these products, making it important to understand their microbiota. Using state-of-the-art techniques, we will focus on the most abundant microbes, which will be correlated to the microbial community as well as their changes during different ripening times. Emphasis will be put on microbe composition and on how microbes can be affected by specific factors (quality of milk production, pH, water activity, manufacturing and processing methodologies).

This Special Issue aims to highlight the role of the microorganisms present in fermented dairy products. As they are nutritionally rich media, all microbial groups (spoilage microorganisms, pathogens or carriers of antimicrobial resistance, and beneficial microorganisms such as lactic acid bacteria and probiotics) may affect human health and disease.

Prof. Athina S. Tzora
Guest Editor

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Keywords

Dairy products
Microbiota
Food safety
Ripening time
Antimicrobial resistance
Pathogens
Spoilage microorganisms
Lactic acid bacteria
Probiotics
Human health

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Published Papers (12 papers)

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Editorial

Jump to: Research

3 pages, 188 KiB

Open AccessEditorial

Food Microbiology: Dairy Products’ Microbiota

by Athina Tzora

Appl. Sci. 2023, 13(22), 12111; https://doi.org/10.3390/app132212111 - 7 Nov 2023

Viewed by 1523

Abstract

The examination of dairy products involves the application of omics approaches to screen varied collections of microorganisms [...] Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

Research

Jump to: Editorial

22 pages, 1187 KiB

Open AccessArticle

Effect of Immobilized Pediococcus acidilactici ORE5 Cells on Pistachio Nuts on the Functional Regulation of the Novel Katiki Domokou-Type Cheese Microbiome

by Chrysoula Pavlatou, Anastasios Nikolaou, Ioanna Prapa, Konstantinos Tegopoulos, Stavros Plesssas, Maria E. Grigoriou, Eugenia Bezirtzoglou and Yiannis Kourkoutas

Appl. Sci. 2023, 13(14), 8047; https://doi.org/10.3390/app13148047 - 10 Jul 2023

Cited by 2 | Viewed by 1222

Abstract

Nowadays, functional foods supplemented with health-promoting microorganisms have attracted consumer attention due to their health benefits. However, maintaining high cell loads, which consist of an essential requirement for conferring the health effect, is a real bottleneck for the food industry due to viability declines during food processing and storage. Hence, freeze-drying and cell immobilization have been suggested to enhance cell viability. The aim of our study was to assess the effect of freeze-dried immobilized P. acidilactici ORE5 on pistachio nuts on the functional regulation of the Katiki Domokou-type cheese microbiome. Supplementation of Katiki Domokou-type cheese with free or immobilized P. acidilactici ORE5 culture resulted in cell loads > 8.5 logcfu/g up to 7 days of storage. Both free and immobilized P. acidilactici ORE5 cells suppressed the growth of L. monocytogenes after deliberate inoculation, acting as a protecting shield. HS-SPME GC/MS analysis showed that the incorporation of P. acidilactici ORE5 culture in cheese resulted in an improved volatile compounds profile, as verified by the preliminary sensory evaluation. According to Next-Generation Sequencing analysis, a wide range of bacterial diversity was revealed among samples. The most abundant genus was Lactococcus in all samples, while the results showed an increased presence of Pediococcus spp. in cheese fortified with P. acidilactici ORE5 culture, highlighting the ability of the strain to survive in the final product. Furthermore, the incorporation of P. acidilactici ORE5 culture in cheese had a significant impact on cheese microbiome composition, as the presence of spoilage bacteria, such as Chryseobacterium, Acinetobacter and Pseudomonas, was significantly less compared to the control cheese, indicating quality improvement and prolongation of the product’s shelf-life. Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

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13 pages, 2900 KiB

Open AccessArticle

Inhibition of Listeria monocytogenes by Broth Cultures of Surface Microbiota of Wooden Boards Used in Cheese Ripening

by Kirty Wadhawan, Andrew Steinberger, Scott Rankin, Garret Suen and Charles Czuprynski

Appl. Sci. 2023, 13(10), 5872; https://doi.org/10.3390/app13105872 - 10 May 2023

Cited by 2 | Viewed by 1934

Abstract

Listeria monocytogenes is a significant concern in cheese production. It has been assumed by some that wood surfaces pose a greater risk for pathogen contamination during cheese production. The goal of this study was to evaluate the effect of the microbiota obtained from wooden boards used in cheese ripening on the growth of Listeria monocytogenes. Bacterial communities from the surface of wooden boards obtained from cheese-ripening facilities were inoculated into tryptic soy broth (TSB) and incubated at 11 °C for 48 h. These communities (10⁸ CFU/mL stationary phase cells) were co-incubated with 10⁴–10⁵ CFU/mL L. monocytogenes 2203 at 11 °C for up to eight days. At various times, samples were removed, diluted in sterile saline and plated on modified Oxford agar. Bacterial communities from each of the five boards from three different facilities significantly inhibited growth of L. monocytogenes in vitro, compared to growth of L. monocytogenes 2203 alone. Using 16S rRNA analysis, we identified sequences belonging to the genera Carnobacterium, Leuconostoc and Staphylococcus as the most abundant in the communities grown in TSB. Leuconostoc mesenteroides and Staphylococcus equorum isolated from the most inhibitory community significantly inhibited growth of L. monocytogenes in TSB at 11 °C, compared to growth of L. monocytogenes 2203 alone. These findings suggest that some members of the complex microbial communities on wooden boards in cheese aging facilities might inhibit the growth of L. monocytogenes. Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

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10 pages, 2039 KiB

Open AccessArticle

A Sustainable Method: Production of the Fermented Rice Milk Yogurt by Using Three Efficient Lactic Acid Bacteria

by Wael N. Hozzein, Sameh M. Hisham and Dalal Hussien M. Alkhalifah

Appl. Sci. 2023, 13(2), 907; https://doi.org/10.3390/app13020907 - 9 Jan 2023

Cited by 5 | Viewed by 4425

Abstract

Lactic acid bacteria (LAB) have long been used as starters in non-dairy cereal fermentation, as they aid in the production of products such as yoghurt and cheese. Broken rice milk is a plant-based milk alternative that is high in carbs and low in fat, providing excellent nutritional value to human users. The current study intends to ferment broken rice milk supplemented with 6% skim milk using three Lactobacillus strains for the development of yoghurt products, as well as to evaluate the growth, changes in physio-chemical properties, and sensory qualities of the yoghurt produced. Lactobacillus bulgaricus, Lactobacillus casei, Lactobacillus acidophilus, and a commercial yoghurt culture consortium fermented broken rice milk after 8 h. Rather than employing L. acidophilus or a commercial yogurt culture consortia, L. bulgarics was the most efficient starter for yoghurt manufacturing, followed by L. casei. L. bulgaricus had the highest viability counts of 8.5 Log CFU/mL, 0.18 specific growth rate, and 3.78 doubling time. Furthermore, it produces a significant reduction in pH to 4.3 and increases total titratable acidity to 0.09 percent with high overall acidity values of 1.4 mg/L of acetic and lactic acid contents. The maximum acidification rate (Vmax) was 0.2125, the maximum acidification time (Tmax) was 4 h, and the time to reach pH 4.6 (Te) was 5 to 8 h. As a result, L. bulgaricus was chosen as the most efficient isolate for the production of fermented rice milk yoghurt. More research is needed, however, to investigate the new rice-based yoghurt product’s sensory qualities as well as its toxicological effects on normal and malignant human cells. Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

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11 pages, 299 KiB

Open AccessArticle

Staphylococcus aureus from Minas Artisanal Cheeses: Biocide Tolerance, Antibiotic Resistance and Enterotoxin Genes

by Josisleine Recalde Allaion, Karina Ghougassian Barrionuevo, Maria Jose Grande Burgos, Antonio Gálvez and Bernadette Dora Gombossy de Melo Franco

Appl. Sci. 2022, 12(3), 1019; https://doi.org/10.3390/app12031019 - 19 Jan 2022

Cited by 3 | Viewed by 2029

Abstract

Staphylococcus aureus is a common contaminant in artisanal raw-milk cheeses. Tolerance of S. aureus to biocides is a threat to disinfection in the cheese production environment, while antibiotic resistance and enterotoxin production are additional health concerns. This study aimed to evaluate the tolerance of S. aureus isolated from Minas artisanal cheeses to the biocides benzalkonium chloride, hexadecylpyridinium chloride, cetrimide, triclosan, hexachlorophene, and chlorhexidine, and the simultaneous occurrence of genes coding for antibiotic resistance (mecA, aacA-aphD, and tetK), efflux pumps [qacA/B and smr (qacC/D)], and enterotoxins (sea, seb, sec, sed, see, seg, seh, sei, and sej). Among the tested isolates, 38.2% were resistant to at least one biocide, and 73.1% were positive for one or more antibiotic resistance gene. Most of the biocide-tolerant and antibiotic-resistant isolates harbored efflux pump genes, and were positive for at least one staphylococcal enterotoxin gene. The study highlights the need for correct hygiene monitoring programs to ensure the safety of these products. Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

9 pages, 282 KiB

Open AccessArticle

Assessing the Microbiological Safety Parameters of Minas Artisanal Cheese Samples in Retail Environments in São Paulo, Brazil

by Josisleine Recalde Allaion, Karina Ghougassian Barrionuevo and Bernadette Dora Gombossy de Melo Franco

Appl. Sci. 2021, 11(19), 9331; https://doi.org/10.3390/app11199331 - 8 Oct 2021

Cited by 6 | Viewed by 2455

Abstract

Minas artisanal cheese is the best known and most consumed type of cheese in Brazil. Prepared with raw bovine milk and starter cultures, these cheeses face many opportunities for post-processing contamination during their transport to commercial establishments as well as in how they are stored, displayed, and maintained for cutting/slicing at retail establishments. It is a common practice for retailers to purchase entire cheeses from the producers, cut them into smaller pieces for retail sale, and store them at room temperature instead of properly refrigerating them. This study evaluated the microbiological safety parameters of samples of Minas artisanal cheeses collected at retail establishments in the city of São Paulo, Brazil, to more realistically assess consumer exposure to the most common pathogens. Samples were submitted for investigation of Salmonella spp., Listeria monocytogenes, and the counts of total coliforms, Escherichia coli, and coagulase-positive enterotoxigenic staphylococci using culture and real-time PCR methods. A worrisome number of samples failed to comply with the current Brazilian legislation for foods in retail environments and presented more than one non-compliance issue. Results highlighted that quality and safety management tools, such as good hygiene practices and HACCP, in retail environments deserve more attention to reduce the possible risks to consumer health. Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

15 pages, 2052 KiB

Open AccessArticle

Polyphasic Characterization of Microbiota of “Mastredda”, a Traditional Wooden Tool Used during the Production of PDO Provola dei Nebrodi Cheese

by Gabriele Busetta, Giuliana Garofalo, Guido Mangione, Luigi Botta, Elena Franciosi, Rosalia Di Gerlando, Massimo Todaro, Giuseppe Licitra, Maria Luisa Scatassa, Raimondo Gaglio and Luca Settanni

Appl. Sci. 2021, 11(18), 8647; https://doi.org/10.3390/app11188647 - 17 Sep 2021

Cited by 8 | Viewed by 2083

Abstract

The biofilms of the wooden tables used for the acidification of the curd were investigated for PDO Provola dei Nebrodi cheese, a traditional stretched cheese made in eastern Sicily (southern Italy) from raw cows’ milk. To this purpose the wooden tables of four dairy facilities were analysed for their microbiota by scanning electron microscopy (SEM) analysis and a combined culture-independent and -dependent microbiological approach. SEM inspection showed an almost continuous biofilm formation. MiSeq Illumina analysis identified 8 phyla, 16 classes, 25 orders, 47 families and 50 genera. Corynebacterium, Bifidobacterium and lactic acid bacteria (LAB) were detected in all samples. In particular, the LAB genera detected on all wooden tables were Lactobacillus, Streptococcus and Lactococcus. LAB dominated the surfaces of all wooden tables with levels higher than 7.0 Log CFU/cm². In particular, the LAB found at the highest levels were mesophilic cocci. Coagulase positive staphylococci, Salmonella spp., Listeria monocytogenes and Shiga-toxigenic Escherichia coli were never detected. Twenty-seven dominating LAB strains were identified within the genera Enterococcus, Lactobacillus, Lacticaseibacillus, Lactiplantibacillus, Levilactobacillus, Lactococcus, Leuconostoc, Pediococcus and Streptococcus. This work showed that the wooden table used during the production of PDO Provola dei Nebrodi cheese is a safe system and a microbiologically active tool. Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

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17 pages, 755 KiB

Open AccessArticle

Characterization of Lactic Acid Bacteria in Pecorino di Farindola Cheese and Manufacturing with a Lacticaseibacillus paracasei Autochthonous Culture

by Giuseppe Aprea, Alessandra Alessiani, Franca Rossi, Lorena Sacchini, Arianna Boni, Daniela D’Angelantonio, Silvia Scattolini, Anna Franca Sperandii, Gabriella Centorotola, Diana Neri, Francesco Pomilio, Elisabetta Di Giannatale, Ilaria Del Matto, Patrizia Tucci and Giacomo Migliorati

Appl. Sci. 2021, 11(17), 7897; https://doi.org/10.3390/app11177897 - 27 Aug 2021

Cited by 5 | Viewed by 2408

Abstract

This study focused on the characterization of lactic acid bacteria (LAB) in a renowned traditional Italian cheese, Pecorino di Farindola, in order to select an autochthonous culture and investigate its potential for the improvement of safety and functional properties. Two hundred and six LAB isolated throughout production and maturation from nine cheese lots of three farms were identified by 16S rRNA gene sequencing and tested for the presence of genes encoding virulence factors, vancomycin resistance (for enterococci), biogenic amines (BAs) and bacteriocin production for antimicrobial activity; and for the capacity to survive in the gastrointestinal tract (GIT) based on tolerance to low pH and bile salts and adhesion to CaCo-2 cells. A Lacticaseibacillus paracasei isolate was used in cheese making and determined a decline of spiked Listeria monocytogenes and Escherichia coli O157 faster than in the control cheese. The autochthonous bacterial groups were numerically unaffected, apart from lactobacilli that were recovered in higher numbers in cheese with the addition of the L. paracasei strain. Based on repetitive extragenic palyndrome (Rep) polymerase chain reaction (PCR) profiles, the added strain possibly dominated until day 88 in the cheese. Results encourage further trials with autochthonous cultures in order to efficiently inhibit hazardous bacteria and to enrich a functional microbiota in Pecorino di Farindola and similar cheeses. Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

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13 pages, 1238 KiB

Open AccessFeature PaperArticle

Association of Breed of Sheep or Goats with Somatic Cell Counts and Total Bacterial Counts of Bulk-Tank Milk

by Daphne T. Lianou, Charalambia K. Michael, Natalia G. C. Vasileiou, Dimitra V. Liagka, Vasia S. Mavrogianni, Mariangela Caroprese and George C. Fthenakis

Appl. Sci. 2021, 11(16), 7356; https://doi.org/10.3390/app11167356 - 10 Aug 2021

Cited by 12 | Viewed by 2075

Abstract

The objective was to describe potential associations of somatic cell counts (SCC) and total bacterial counts (TBC) in bulk-tank milk from sheep and goat farms with breeds of these animals in Greece. In total, 325 dairy sheep flocks and 119 dairy goat herds were visited for the collection of milk; the breed of animals in farms was evaluated for a potential association with SCC or TBC. The most frequently seen sheep breeds were the Lacaune (95 flocks) and the Chios (44 flocks). The most frequently seen goat breeds were the indigenous Greek (Capra prisca) (50 herds) and the Murciano-Granadina (13 herds). In a multivariable analysis, the breed and the application of machine-milking in sheep flocks, and the breed and the management system in goat herds emerged as significant factors for increased SCC (>0.75 × 10⁶ cells mL⁻¹) in bulk-tank milk. Further, the month of lactation at sampling in sheep flocks emerged a significant factor for increased TBC (>1500 × 10³ cfu mL⁻¹) in bulk-tank milk. Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

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11 pages, 287 KiB

Open AccessArticle

White Brined Cheese Production by Incorporation of a Traditional Milk-Cereal Prebiotic Matrix with a Candidate Probiotic Bacterial Strain

by Stavros Plessas, Vasillios Ganatsios, Ioanna Mantzourani and Loulouda Bosnea

Appl. Sci. 2021, 11(13), 6182; https://doi.org/10.3390/app11136182 - 3 Jul 2021

Cited by 19 | Viewed by 3026

Abstract

The aim of the present study is the evaluation of a novel potentially probiotic Lactobacillus paracasei SP5, previously isolated from dairy products, as a starter culture of white brined cheese production, either free or immobilized on a traditional food, “trahanas”, in order to provide protection to the starter culture and a prebiotic effect. All produced cheeses were compared with cheese manufactured by renin enzyme. Several parameters that affect the acceptability, quality, and shelf life of white brined cheese were investigated, including microbial populations, physicochemical characteristics, and cheese volatiles through 70 days of ripening and storage. White brined cheese production by free or immobilized L. paracasei SP5 resulted in significantly higher acidity (over 0.8 g of lactic acid/100 g of cheese at the 70th day of ripening) and significantly reduced counts (around 50%) of coliforms, yeasts, and fungi compared to cheese produced with no starter culture. The use of the freeze-dried novel starter culture, either free or immobilized, improved the aromatic profile of cheeses as was proven through a GC-MS analysis. In addition, it should be underlined that the application of the novel strain led to white brined cheese with improved overall quality and sensory characteristics. The results indicate the potential industrial use of freeze-dried L. paracasei SP5 as a starter culture for the production of good-quality functional white brined cheeses. Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

11 pages, 1091 KiB

Open AccessArticle

Effects of 2′-Fucosyllactose-Based Encapsulation on Probiotic Properties in Streptococcus thermophilus

by Shadi Pakroo, Gloria Ghion, Armin Tarrah, Alessio Giacomini and Viviana Corich

Appl. Sci. 2021, 11(13), 5761; https://doi.org/10.3390/app11135761 - 22 Jun 2021

Cited by 4 | Viewed by 2585

Abstract

Streptococcus thermophilus is widely used in dairy fermentation as a starter culture for yogurt and cheese production. Many strains are endowed with potential probiotic properties; however, since they might not survive in adequate amounts after transit through the human gastrointestinal tract, it is advisable to improve cell survivability during this passage. The present study evaluates the use of 2′-fucosyllactose, a prebiotic molecule from human milk, compared with other known molecules, such as gelatin and inulin, to form alginate-based microcapsules to fulfill these requirements. Such microcapsules, obtained by the extrusion technique, were evaluated in terms of encapsulation efficiency, storage stability, gastrointestinal condition resistance, and cell release kinetics. Results reveal that microcapsules made using 2′-fucosyllactose and those with inulin resulted in the most efficient structure to protect S. thermophilus strain TH982 under simulated gastrointestinal conditions (less than 0.45 log CFU/g decrease for both agents). In addition, a prompt and abundant release of encapsulated cells was detected after only 30 min from microcapsules made with sodium alginate plus 2′-fucosyllactose in simulated gastrointestinal fluid (more than 90% of the cells). These encouraging results represent the first report on the effects of 2′-fucosyllactose used as a co-encapsulating agent. Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

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14 pages, 3497 KiB

Open AccessArticle

Microbiota “Fingerprint” of Greek Feta Cheese through Ripening

by Athina Tzora, Aikaterini Nelli, Chrysoula Voidarou, George Fthenakis, Georgios Rozos, Georgios Theodorides, Eleftherios Bonos and Ioannis Skoufos

Appl. Sci. 2021, 11(12), 5631; https://doi.org/10.3390/app11125631 - 18 Jun 2021

Cited by 20 | Viewed by 3681

Abstract

Feta is a Greek protected designation of origin (PDO) brined curd white cheese made from small ruminants’ milk. In the present research, Greek Feta cheese bacterial diversity was evaluated via matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS). Analysis of 23 cheese samples, produced in different regions of the country, was performed in two ripening times (three or six months post-production). The identified microbiota were primarily constituted of lactic acid bacteria. A total of 13 different genera were obtained. The dominant species in both ripening times were Lactobacillus plantarum (100.0% and 87.0%, at three or six months post-production, respectively), Lactobacillus brevis (56.5% and 73.9%), Lactobacillus paracasei (56.5% and 39.1%), Lactobacillus rhamnosus (13.0% and 17.4%), Lactobacillus paraplantarum (4.3% and 26.1%), Lactobacillus curvatus (8.7% and 8.7%). Other species included Enterococcus faecalis (47.8% and 43.5%), Enterococcus faecium (34.8% and 17.4%), Enterococcus durans (13.0% and 17.4%), Enterococcus malodoratus (4.3% and 4.3%), and Streptococcus salivarius subsp. thermophilus (21.7% and 30.4%). The increased ripening time was found to be correlated to decreased total solids (r = 0.616; p = 0.002), protein (r = 0.683; p < 0.001), and PH (r = 0.780; p < 0.001). The results of this study contribute to a better understanding of the core microbiota of Feta cheese. Full article

(This article belongs to the Special Issue Food Microbiology: Dairy Products Microbiota)

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