The effect of seed germination and Bacillus spp. on the ripening of plant cheese analogs.

Consumer demand for plant cheeses is increasing, but challenges of improving both flavor and quality remain. This study investigated the microbiological and physicochemical impact of seed germination and fermentation with Bacillus velezensis and Bacillus amyloliquefaciens on the ripening of plant cheese analogs. Chlorine treatment or addition of Lactiplantibacillus plantarum and Lactococcus lactis controlled microbial growth during seed germination. Lp. plantarum and Lc. lactis also served as starter cultures for the acidification of soy and lupine milk and were subsequently present in the unripened plant cheese as dominant microbes. Acidification also inhibited the growth and metabolic activity of bacilli but Bacillus spores remained viable throughout ripening. During plant cheese ripening, Lc. lactis was inactivated before Lp. plantarum and the presence of bacilli during seed germination delayed Lc. lactis inactivation. Metagenomic sequencing of full-length 16S rRNA gene amplicons confirmed that the relative abundance of the inoculated strains in each ripened cheese sample exceeded 99%. Oligosaccharides including raffinose, stachyose, and verbascose were rapidly depleted in the initial stage of ripening. Both germination and the presence of bacilli during seed germination had impact on polysaccharide hydrolysis during ripening. Bacilli but not seed germination enhanced proteolysis of plant cheese during ripening. In conclusion, the use of germination with lactic acid bacteria in combination with Bacillus spp. exhibited the potential to improve the quality of ripened plant cheeses with a positive effect on the reduction of hygienic risks. IMPORTANCE: The development of novel plant-based fermented food products for which no traditional templates exist requires the development of starter cultures. Although the principles of microbial flavor formation in plant-based analogs partially overlap with dairy fermentations, the composition of the raw materials and thus likely the selective pressure on the activity of starter cultures differs. Experiments that are described in this study explored the use of seed germination, the use of lactic acid bacteria, and the use of bacilli to reduce hygienic risks, to acidify plant milk, and to generate taste-active compounds through proteolysis and fermentative conversion of carbohydrates. The characterization of fermentation microbiota by culture-dependent and culture-independent methods also confirmed that the starter cultures used were able to control microbial communities throughout 90 d of ripening. Taken together, the results provide novel tools for the development of plant-based analogs of fermented dairy products.

Factors influencing conjugated linoleic acid content of dairy products: challenges and strategies.

Conjugated linoleic acid (CLA), a bioactive fatty acid that provides various physiological benefits, has gained increasing attention in the food industry, and various studies have focused on enhancing its content in dairy products. The factors influencing CLA content in dairy products vary significantly, including lactation stage, breed type, seasonality, feed, management methods of the animals, the manufacturing processes, storage, and ripening periods of the product. Additionally, the incorporation of CLA-producing probiotic bacteria, such as Lactobacillus, Lactococcus, Bifidobacterium, and Propionibacterium, is an emerging study in this field. Studies have revealed that factors affecting the CLA content in milk affect that in dairy products as well. Furthermore, the species and strains of CLA-producing bacteria, fermentation conditions, ripening period, and type of dairy product are also contributing factors. However, production of CLA-enhanced dairy products using CLA-producing bacteria while maintaining their optimal viability and maximizing exposure to free linoleic acid remains limited. The current review emphasized the factors affecting the CLA content and related mechanisms, challenges in the application of CLA-producing probiotic bacteria, and strategies to address these challenges and enhance CLA production in dairy products. Therefore, the development of functional dairy products with enhanced CLA levels is expected to be possible.

Bacterial proteome adaptation during fermentation in dairy environments.

The enzymatic repertoire of starter cultures belonging to the Lactococcus genus determines various important characteristics of fermented dairy products but might change in response to the substantial environmental changes in the manufacturing process. Assessing bacterial proteome adaptation in dairy and other food environments is challenging due to the high matrix-protein concentration and is even further complicated in particularly cheese by the high fat concentrations, the semi-solid state of that matrix, and the non-growing state of the bacteria. Here, we present bacterial harvesting and processing procedures that enable reproducible, high-resolution proteome determination in lactococcal cultures harvested from laboratory media, milk, and miniature Gouda cheese. Comparative proteome analysis of Lactococcus cremoris NCDO712 grown in laboratory medium and milk revealed proteome adaptations that predominantly reflect the differential (micro-)nutrient availability in these two environments. Additionally, the drastic environmental changes during cheese manufacturing only elicited subtle changes in the L. cremoris NCDO712 proteome, including modified expression levels of enzymes involved in flavour formation. The technical advances we describe offer novel opportunities to evaluate bacterial proteomes in relation to their performance in complex, protein- and/or fat-rich food matrices and highlight the potential of steering starter culture performance by preculture condition adjustments.

Selection of adjunct cultures for the ripening of plant cheese analogues.

Fermentation contributes to the taste and odor of plant cheeses. The selection of functional cultures for the fermentation of plant cheeses, however, is in its infancy. This study aimed to select lactic acid bacteria for ripening of soy and lupin cheese analogues. Bacillus velezensis and B. amyloliquefaciens were used for germination of seeds to produce proteolytic enzymes; Lactococcus lactis and Lactiplantibacillus plantarum served as primary acidifying cultures. Levilactobacillus hammesii, Furfurilactobacillus milii, or Lentilactobacillus buchneri were assessed as adjunct cultures for the ripening of plant cheese. Growth of bacilli was inhibited at low pH. Both Lc. lactis and Lp. plantarum were inactived during plant cheese ripening. Cell counts of Lv. hammesii remained stable over 45 d of ripening while Ff. milii and Lt. buchneri grew slowly. Sequencing of full length 16S rRNA genes confirmed that the inocula the plant cheeses accounted for more than 98% of the bacterial communities. HPLC analysis revealed that Lt. buchneri metabolized lactate to acetate and 1,2-propanediol during ripening. Bacilli enhanced proteolysis as measured by quantification of free amino nitrogen, and the release of glutamate. LC-MS/MS analysis quantified kokumi-active dipeptides. The concentrations of γ-Glu-Leu, γ-Glu-Ile, and γ-Glu-Ala, γ-Glu-Cys in unripened cheeses were increased by seed germination but γ-Glu-Phe was degraded. Lt. buchneri but not Lv. hammesii or Ff. milii accumulated γ-Glu-Val, γ-Glu-Ile or γ-Glu-Leu during ripening, indicating strain-specific differences. In conclusion, a consortium of bacilli, acidification cultures and adjunct cultures accumulates taste- and kokumi-active compounds during ripening of plant cheeses.

Interrelationships among physical and chemical traits of cheese: Explanatory latent factors and clustering of 37 categories of cheeses.

Cheese presents extensive variability in physical, chemical, and sensory characteristics according to the variety of processing methods and conditions used to create it. Relationships between the many characteristics of cheeses are known for single cheese types or by comparing a few of them, but not for a large number of cheese types. This case study used the properties recorded on 1,050 different cheeses from 107 producers grouped into 37 categories to analyze and quantify the interrelationships among the chemical and physical properties of many cheese types. The 15 cheese traits considered were ripening length, weight, firmness, adhesiveness, 6 different chemical characteristics, and 5 different color traits. As the 105 correlations between the 15 cheese traits were highly variable, a multivariate analysis was carried out. Four latent explanatory factors were extracted, representing 86% of the covariance matrix: the first factor (38% of covariance) was named Solids because it is mainly linked positively to fat, protein, water-soluble nitrogen, ash, firmness, adhesiveness, and ripening length, and negatively to moisture and lightness; the second factor (24%) was named Hue because it is linked positively to redness/blueness, yellowness/greenness, and chroma, and negatively to hue; the third factor (17%) was named Size because it is linked positively to weight, ripening length, firmness, and protein; and the fourth factor (7%) was named Basicity because it is linked positively to pH. The 37 cheese categories were grouped into 8 clusters and described using the latent factors: the Grana Padano cluster (characterized mainly by high Size scores); hard mountain cheeses (mainly high Solids scores); very soft cheeses (low Solids scores); blue cheeses (high Basicity scores), yellowish cheeses (high Hue scores), and 3 other clusters (soft cheeses, pasta filata and treated rind, and firm mountain cheeses) according to specific combinations of intermediate latent factors and cheese traits. In this case study, the high variability and interdependence of 15 major cheese traits can be substantially explained by only 4 latent factors, allowing us to identify and characterize 8 cheese type clusters.

The impact of seasonal calving systems with varying pasture allowance on Cheddar cheese composition, nutritional quality and ripening characteristics.

The objective of this study was to examine the impact of stage of lactation (early, mid and late) and proportion of pasture in the cows diet (high: GRS, medium: PMR and no: TMR) on the composition and quality of Cheddar cheese. Triplicate trials were carried out in each stage of lactation, and milk protein and fat contents were standardized for Cheddar cheese manufacture at pilot scale. As cheese milks were standardized for milk fat and protein contents, gross composition did not differ as a result of diet. Fatty acid profiles of GRS cheese were significantly different from TMR, while PMR profiles were less distinct and more similar to both GRS and TMR profiles, as illustrated by partial least squares discriminatory analysis. Fatty acids including CLA C18:2 cis-9, trans-11, C22:1 n-9 and C18:3 n-3 were most influential in this separation of profiles. Fatty acid profiling revealed that GRS derived cheese contained higher proportions of nutrients considered beneficial for human health including higher proportions of unsaturated fatty acids and omega-3 fatty acids. A biomarker model utilizing the proportions of 5 fatty acids was constructed and was effective at distinguishing between cheese of GRS, TMR and PMR feeding systems. Proportions of ρ-κ-casein, αs2-casein and αs1-casein in cheese also differed between diets while proportions of ρ-κ-casein, αs1-casein and ß-casein were lowest in late lactation cheese. The impact of diet was less influential compared with that of stage of lactation on the ripening characteristics of cheese. An index of primary proteolysis was highest in late lactation cheese. The peptides derived from the proteolysis of κ-casein and ß-casein and levels of secondary proteolysis, in particular, the proportions of 12 free amino acids were most influenced by stage of lactation. Overall this study demonstrated the effects of increasing pasture allowance and stage of lactation on the nutritional quality and ripening properties of Cheddar cheese.

Microbiological, physicochemical, and sensory changes throughout ripening of an experimental soft smear-ripened cheese in relation to salt concentrations.

To evaluate the effect of NaCl content on microbiological, biochemical, physicochemical, and sensorial characteristics, Munster cheeses were prepared from pasteurized milk seeded with 3 yeasts (Kluyveromyces marxianus, Debaryomyces hansenii, and Geotrichum candidum) and 5 ripening bacteria (Arthrobacter arilaitensis, Brevibacterium aurantiacum, Corynebacterium casei, Hafnia alvei, and Staphylococcus equorum). Experiments were performed in triplicate under 1.0%, 1.7%, and 2.4% NaCl levels in cheese. Ripening (d 2-27) was carried out at 12°C and 96% relative humidity. These kinetics were both reproducible and repeatable at a 99% confidence level. For each microbial, biochemical, and physicochemical parameter, 2 kinetic descriptors (the maximal or minimal rate and its occurrence time) were defined. On d 2, the physicochemical variables (water activity, dry matter, and water content) were strongly dependent on the salting level. From d 2 to d 27, K. lactis was insensitive to salt, whereas D. hansenii was stimulated. Geotrichum candidum growth appeared very sensitive to salt in cheese: at 1.0% NaCl, G. candidum exhibited overgrowth, negatively affecting rind appearance, underrind consistency and thickness, and off-flavor flaws. A salt concentration of 2.4% induced death of G. candidum. A total of 4 bacteria (A. arilaitensis, B. aurantiacum, C. casei, and H. alvei) were moderately sensitive to salt, but S. equorum was insensitive to it. Salt level in cheese had a significant effect on carbon substrate consumption rates. The lactate consumption rate in 1.0% salted cheeses was approximately twice higher than under 2.4% NaCl. Data analysis of microorganism, biochemical, and physicochemical kinetics, as well as sensory analysis, showed that 1.7% NaCl was the best salt level in Munster-type cheeses to achieve an optimum balance between cheese characteristics, sensory qualities, and marketability.

New Genetic Determinants for qPCR Identification and the Enumeration of Selected Lactic Acid Bacteria in Raw-Milk Cheese.

Lactic acid bacteria (LAB) play an important role in the ripening of cheeses and contribute to the development of the desired profile of aroma and flavor compounds. Therefore, it is very important to monitor the dynamics of bacterial proliferation in order to obtain an accurate and reliable number of their cells at each stage of cheese ripening. This work aimed to identify and conduct a quantitative assessment of the selected species of autochthonous lactic acid bacteria from raw cow's milk cheese by the development of primers and probe pairs based on the uniqueness of the genetic determinants with which the target microorganisms can be identified. For that purpose, we applied real-time quantitative PCR (qPCR) protocols to quantify Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus, and Lactococcus lactis subsp. cremoris cells in cheese directly after production and over three-month and six-month ripening periods. While L. lactis subsp. cremoris shows good acidification ability and the ability to produce antimicrobial compounds, L. delbrueckii subsp. bulgaricus has good proteolytic ability and produces exo-polysaccharides, and S. thermophilus takes part in the formation of the diacetyl flavor compound by metabolizing citrate to develop aroma, they all play an important role in the cheese ripening. The proposed qPCR protocols are very sensitive and reliable methods for a precise enumeration of L. delbrueckii subsp. bulgaricus, S. thermophilus, and L. lactis subsp. cremoris in cheese samples.

Monitoring cheese ripening by single-sided nuclear magnetic resonance.

The noninvasive, longitudinal study of products and food processing is of interest for the dairy industry. Here, we demonstrated that single-sided nuclear magnetic resonance (NMR) can be used for noninvasive monitoring of the cheese ripening process. The maturation of soft-ripened Camembert-like molded cheese samples was monitored for 20 d measuring 1-dimensional and 2-dimensional NMR relaxation and diffusion data at various depths, ranging from the hard surface layer to the soft center. Gelation and gel shrinkage were observed throughout ripening, and a complete loss of free water signal was observed at the cheese rind. Transversal (T2) relaxation distributions include 3 components that evolve with ripening time and position, corresponding to water inside the casein gel network, water trapped in casein, and fat. Two-dimensional T1-T2 relaxation experiments provided enhanced resolution of the 3 components, allowing quantification of the relative proportions of each phase. Furthermore, diffusion (D)-T2 relaxation correlation experiments revealed the bimodal size distribution of fat globules. The study demonstrated that single-sided NMR can provide spatially resolved signal intensity, relaxation, and diffusion parameters that reflect structural changes during the ripening process and can be exploited to understand and monitor the ripening of cheeses.

Identifying Chemical Differences in Cheddar Cheese Based on Maturity Level and Manufacturer Using Vibrational Spectroscopy and Chemometrics.

Cheese is a nutritious dairy product and a valuable commodity. Internationally, cheddar cheese is produced and consumed in large quantities, and it is the main cheese variety that is exported from Australia. Despite its importance, the analytical methods to that are used to determine cheese quality rely on traditional approaches that require time, are invasive, and which involve potentially hazardous chemicals. In contrast, spectroscopic techniques can rapidly provide molecular information and are non-destructive, fast, and chemical-free methods. Combined with partner recognition methods (chemometrics), they can identify small changes in the composition or condition of cheeses. In this work, we combined FTIR and Raman spectroscopies with principal component analysis (PCA) to investigate the effects of aging in commercial cheddar cheeses. Changes in the amide I and II bands were the main spectral characteristics responsible for classifying commercial cheddar cheeses based on the ripening time and manufacturer using FTIR, and bands from lipids, including ß'-polymorph of fat crystals, were more clearly determined through changes in the Raman spectra.

Characterization of autochthonal yeasts isolated from Spanish soft raw ewe milk protected designation of origin cheeses for technological application.

The yeasts involved in the ripening process of artisanal soft raw ewe milk Protected Designation of Origin (PDO) Torta del Casar and Queso de la Serena cheeses produced in Extremadura, Spain, were isolated throughout their ripening process, strain typed, and characterized for some important technological properties. A total of 508 yeast isolates were obtained and identified by inter-single sequence repeat anchored PCR amplification analysis and subsequent sequencing of the internal transcribed spacer ITS1/ITS2 5.8S rRNA. A total of 19 yeast species representing 8 genera were identified. Debaryomyces hansenii, Pichia kudriavzevii, Kluyveromyces lactis, and Yarrowia lipolytica were the predominant species. We selected 157 isolates, by genotyping and origin, for technological characterization. The evaluation of yeast isolates' growth under stress conditions of cheese ripening showed that 87 presented better performance. Among them, 71 isolates were not able to catabolize tyrosine to produce a brown pigment. Principal component analysis of the biochemical features of these isolates showed that 9 strains stood out, 3 K. lactis strains (2287, 2725, and 1507), 2 Pichia jadinii (1731 and 433), 2 Yarrowia alimentaria (1204 and 2150), Y. lipolytica 2495 and P. kudriavzevii 373. These strains displayed strong extracellular proteolytic activity on skim milk agar as well as an adequate enzymatic profile (strong aminopeptidase and weak protease activity), suggesting their great potential for cheese proteolysis. Extracellular lipolytic activity was mainly restricted to Yarrowia spp. isolates and weakly present in P. kudriavzevii 373 and K. lactis 2725, although enzymatic characterization by API-ZYM (bioMérieux SA) evidenced that all may contribute, at least in part, to the lipolysis process. Moreover, these strains were able to assimilate lactose, galactose, and glucose at NaCl concentrations higher than that usually found in cheese. However, lactate and citrate assimilation were limited to Y. lipolytica 2495, P. kudriavzevii 373, and P. jadinii 433, and may contribute to the alkalinizing process relevant to biochemical processes that take place in the last stages of ripening. By contrast, K. lactis strains showed acidifying capacity and ß-galactosidase activity and may take part in the initial stages of ripening, together with lactic acid bacteria. Thus, considering the technological characteristics studied, the 9 selected strains presented biochemical features well suited to their potential use as adjunct cultures, alone or in combination with autochthonous starter bacteria in the cheesemaking process, to overcome the heterogeneity of these PDO cheeses, preserving their unique sensory characteristics.

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.

Feeding concentrates with different protein sources to high-yielding, mid-lactation Norwegian Red cows: Effect on cheese ripening.

Soybean meal is one of the most important protein sources in concentrate feeds for dairy cows. The objective of the present study was to provide knowledge on the effects of using a novel yeast microbial protein source (Candida utilis) in concentrate feed for dairy cows on the production and quality of a Gouda-type cheese. Forty-eight Norwegian Red dairy cows in early to mid lactation were fed a basal diet of grass silage, which was supplemented with 3 different concentrate feeds. The protein source of the concentrates was based on conventional soybean meal (SBM), novel yeast (C. utilis; YEA), or barley (BAR; used as negative control because barley has a lower protein content). The experiment was carried out for a period of 10 wk, with the first 2 wk as an adaptation period where all dairy cows were fed grass silage and the SBM concentrate. The cows were then randomly allocated to 1 of the 3 different compound feeds: SBM, yeast, or barley. Cheeses were made during wk 8 and 9 of the experiment, with 4 batches of cheese made from milk from each of the 3 groups. The cheeses made from milk from cows fed SBM concentrate (SBM cheese) had a higher content of dl-pyroglutamic acid and free amino acids than the other cheeses, indicating a faster ripening in the SBM cheeses. Despite these differences, the sensory properties, the microbiota, and the Lactococcus population at 15 wk of ripening were not significantly different between the cheeses. This experiment showed that although the raw materials used in the concentrate feed clearly influenced the ripening of the cheeses, this did not affect cheese quality. Yeast (C. utilis) as a protein source in concentrate feed for dairy cows can be used as a replacement for soybean meal without compromising the quality of Norwegian Gouda-type cheeses.

Cheesomics: the future pathway to understanding cheese flavour and quality.

Cheese is a fermented dairy product, harboring diverse microbial communities (microbiota) that change over time and vary depending on the type of cheese and their respective starter and adjunct cultures. These microorganisms play a crucial role in determining the flavor, quality and safety of the final product. Exploring the composition of cheese microbiota and the underlying molecular mechanisms involved in cheese ripening has been the subject of many studies. Recent advances in next generation sequencing (NGS) methods and the development of sophisticated bioinformatics tools have provided deeper insights into the composition and potential functionality of cheese microbiota far beyond the information provided by culture-dependent methods. These advances, which include rRNA gene amplicon sequencing and metagenomics, have been complemented and expanded in recent years by the application of metatranscriptomics, metaproteomics and metabolomics. This paper reviews studies in which application of these meta-omics technologies has led to a better understanding of the microbial composition and functionality of cheese and highlights opportunities by which the integration of outputs from diverse multi-omics analytical platforms (cheesomics) could be used in the future to advance our knowledge of the cheese ripening process and identify biomarkers for predicting cheese flavor, quality, texture and safety, and bioactive metabolites with potential to influence human health.

Effects of the depletion of whey proteins from unconcentrated milk using microfiltration on the yield, functionality, and nutritional profile of Cheddar cheese.

Some European dairies use low concentration factor microfiltration (MF) in their cheese plants. Removal of whey protein (WP) from milk before cheesemaking using microfiltration without concentration provides the opportunity to produce a value-added by-product, milk-derived whey. However, few studies have focused on the effects on cheese properties caused by the depletion of WP from cheese milk. Most studies have concentrated cheese milk using MF in addition to depletion of WP. In our approach, cheese milk was not concentrated during WP depletion using MF. We wanted to quantify residual WP levels in cheese made from MF milk and to explore whether WP depletion from milk would influence functionality, nutritional profile, and cheese quality during ripening. Casein (CN) contents for all milks were kept at ∼2.5%, to eliminate the confounding factor of concentration of CN, which was observed in some previous MF studies. Cheese milks had similar ratios of CN to fat. Three standardized milks were produced with various CN:true protein (TP) ratios: (a) control with a CN:TP ratio of 83:100, (b) 35% WP depletion, 89:100 CN:TP, and (c) 70% WP depletion, 95:100 CN:TP. Cheddar cheeses were made from MF milk with various WP depletion levels and aged for 9 mo, and their functionality was evaluated during ripening. We found no major differences in cheese composition or pH values between samples. Cheese yield, solids recovery, and nitrogen recovery were slightly higher in the 95:100 CN:TP cheeses compared with the control. These enhanced recoveries reflect that MF-treated milk started with a higher fraction of CN-based protein solids, rather than WP solids. The standardized milk from the 95:100 CN:TP treatment also had a slightly higher fat content compared with the control, likely helping to increase cheese yield. Rheological properties of cheeses during heating were similar between treatments. Hardness initially decreased with age for all cheeses due to proteolysis or solubilization, or both, of calcium phosphate. Maximum loss tangent (LT), an index of cheese meltability, was slightly lower for the control cheese until 30 d of ripening, but after 30 d, all treatments exhibited similar maximum LT values. The temperature where LT = 1 (crossover temperature), an index of softening point during heating, was slightly lower for MF cheese compared with the control cheeses during ripening. Microfiltration treatment had no significant influence on proteolysis. Sensory properties were similar between the cheeses, except for bitterness. Bitterness intensity was slightly lower in the MF cheeses than in the control cheeses and increased in all cheeses during ripening. We detected no major differences in the concentrations of key nutrients or vitamins between the various cheeses. Depletion of WP in cheese milk by MF did not negatively affect cheese quality, or its nutritional profile, and resulted in similar cheesemaking yields.

SPME-GC-MS and Multivariate Analysis of Sensory Properties of Cheese in a Sack Matured with Probiotic Starter Cultures.

RESEARCH BACKGROUND: Cheese in a sack is a traditional cheese produced in Croatia. Types of cheese with similar production technology are made in other countries but chemical and microbiological composition varies between regions. Traditionally, cheese in a sack is produced without the addition of starter cultures. Addition of beneficial probiotic cultures to numerous dairy products has documented advantages. Effects that the addition of probiotic bacteria to traditional cheese have on aroma compounds and sensory properties have not been fully investigated. The aim of this study is to determine the sensory properties and differences in the aromatic profiles between cheese samples ripened in a lambskin sack, produced traditionally without the addition of any starter culture, or with the addition of probiotic bacteria. EXPERIMENTAL APPROACH: In this study, cheese in a sack was produced with the addition of probiotic cultures Lactobacillus plantarum B and L. lactis ssp. lactis S1. During ripening volatile aroma compounds were analysed with a solid-phase microextraction gas chromatography-mass spectrometry. Sensory properties were evaluated by trained tasters who are familiar with the traditional taste of the cheese from a sack. The results of aroma composition and taste scores were then compared using factorial and principal component analyses. RESULTS AND CONCLUSIONS: Chromatography showed differences in the composition of aroma compounds and the sensory properties between the cheese produced with Lactobacillus starter cultures and the control cheese, traditionally produced without a starter culture. The addition of probiotic cultures L. plantarum B and L. lactis ssp. lactis S1 resulted in products with better sensory properties and chemical profile of volatile aromatic compounds. NOVELTY AND SCIENTIFIC CONTRIBUTION: This study investigates the usage of naturally present probiotic cultures as starter cultures in cheese in a sack production. Their effects on aroma profiles and sensory characteristics have been compared for the first time using factorial and principal component analyses.

Characterisation of fungal contamination sources for use in quality management of cheese production farms in Korea.

OBJECTIVE: This study was conducted to determine the composition and diversity of the fungal flora at various control points in cheese ripening rooms of 10 dairy farms from six different provinces in the Republic of Korea. METHODS: Floor, wall, cheese board, room air, cheese rind and core were sampled from cheese ripening rooms of ten different dairy farms. The molds were enumerated using YM petrifilm, while isolation was done on yeast extract glucose chloramphenicol agar plates. Morphologically distinct isolates were identified using sequencing of internal transcribed spacer region. RESULTS: The fungal counts in 8 out of 10 dairy farms were out of acceptable range, as per hazard analysis critical control point regulation. A total of 986 fungal isolates identified and assigned to the phyla Ascomycota (14 genera) and Basidiomycota (3 genera). Of these Penicillium, Aspergillus, and Cladosporium were the most diverse and predominant. The cheese ripening rooms was overrepresented in 9 farms by Penicillium (76%), while Aspergillusin a single farm. Among 39 species, the prominent members were Penicillium commune, P. oxalicum, P. echinulatum, and Aspergillus versicolor. Most of the mold species detected on surfaces were the same found in the indoor air of cheese ripening rooms. CONCLUSION: The environment of cheese ripening rooms persuades a favourable niche for mold growth. The fungal diversity in the dairy farms were greatly influenced by several factors (exterior atmosphere, working personnel etc.,) and their proportion varied from one to another. Proper management of hygienic and production practices and air filtration system would be effective to eradicate contamination in cheese processing industries.

Characteristics of ripened Tronchón cheese from raw goat milk containing legally admissible amounts of antibiotics.

The aim of this study was to evaluate the transfer of the most widely used antibiotics in dairy goats from milk to cheese as well as their effect on the cheese-making process and cheese characteristics during ripening. Antibiotic-free milk was spiked individually with 7 veterinary drugs (amoxicillin, benzylpenicillin, cloxacillin, erythromycin, ciprofloxacin, enrofloxacin, and oxytetracycline) at an equivalent concentration of the European Union maximum residue limit. Spiked goat milk was used to make mature Tronchón cheeses, which were analyzed at 0, 30, and 60 d of maturation to determine pH, chemical composition, proteolytic and lipolytic activities, and color and textural properties. A sensory evaluation of 60-d ripened cheeses was carried out. Cheeses from raw antibiotic-free goat milk were made simultaneously to be used as reference. The cheese-making process was unaffected by the presence of most antibiotics evaluated. Only erythromycin and oxytetracycline significantly increased the time required for cheese production (122 ± 29 and 108 ± 25 min, respectively). However, variable amounts of antibiotics, ranging from 7.4 to 68%, were transferred from milk to cheese, with oxytetracycline and quinolones showing the highest retention rates. In general, antibiotic residues present in the cheeses at the beginning of maturation decrease significantly along time. Thus, ß-lactams and erythromycin residues were not detectable after 30 d of ripening. However, relatively high concentrations of enrofloxacin (148 ± 12 µg/kg) and ciprofloxacin (253 ± 24 µg/kg) residues were found in the cheeses after 60 d of maturation. The quality characteristics of the Tronchón cheeses were only slightly affected by such substances, with few significant differences in the free fatty acid concentration and color and textural properties of the cheeses. Results herein indicate that the use of goat milk containing antibiotics, such as quinolones, at the European Union maximum residue limit for cheese production could adversely affect the safety of the final products because relatively high concentrations of these substances could be retained in soft and semi-mature cheeses, making it necessary to assess the risk for consumer health. Studies on the partition of the antibiotic substances during cheese-making, using specific technologies, would be convenient to guarantee the safety of cheese and related products.

Physicochemical and sensory evolutions of the lactic goat cheese Picodon in relation to temperature and relative humidity used throughout ripening.

To produce a wide variety of cheeses, it is necessary to control the ripening process. To do that, artisanal goat cheeses were ripened to evaluate the effects of temperature (10 and 14°C) and relative humidity (RH; 88 and 98%) on (1) 16 physicochemical characteristics throughout ripening and (2) 19 sensory characteristics at the end of ripening (d 12). Whatever the ripening time, the physicochemical characteristics were strongly dependent on the daily productions, which affected the sensory perception of the cheeses. Both physicochemical and sensory characteristics were strongly reliant on RH, whereas only a few of the characteristics were influenced by temperature changes. On d 12, whatever the ripening temperature, an RH increase from 88% to 98% modified many cheese characteristics (core pH, lactate consumption, underrind thickening, dry matter content, and hardness). As a result of these physicochemical properties, changes in perception were observed: the cheeses ripened under 88% RH were dry and hard compared with those ripened under 98% RH. An RH of 98% led to an acceleration of the ripening process, inducing a slightly ammonia and milky flavor and a sticky and creamy texture in the mouth. However, cheeses ripened under 14°C and 98% RH were also indicative of overripened cheeses: a temperature of 14°C induced an acceleration of the ripening process due to physicochemical modifications compared with a temperature of 10°C. Nevertheless, when the cheeses on d 0 were still very humid and soft, those ripened under 98% RH collapsed and were overripened with a liquid underrind. This study provides a means for achieving a better and more rational control of the ripening process in artisanal lactic goat cheeses.

Alcohol dehydrogenases from Proteus mirabilis contribute to alcoholic flavor.

BACKGROUND: Cheese ripening involves a complex series of metabolic reactions and numerous concomitant secondary transformations. Alcohol dehydrogenase (ADH) converts aldehydes into their corresponding alcohols, which enrich cheese aroma. RESULTS: In this study, we identified five ADH genes in Proteus mirabilis JN458, and these genes were overexpressed and characterized in Escherichia coli BL21 (DE3). The optimum pH was 7.0 for the purified recombinant ADH-1, ADH-2, and ADH-3 and 8.0 for ADH-4 and ADH-5. The optimum temperature was 40 °C for ADH-1, ADH-3, and ADH-5 and 45 °C for ADH-2 and ADH-4. The Km value of ADH-1, ADH-2, and ADH-3 was 34.45, 16.90, and 10.01 µmol L-1 for phenylacetaldehyde, respectively. The Km value of ADH-4 and ADH-5 was 14.81 and 24.62 µmol L-1 for 2-methylbutanal, respectively. CONCLUSION: Proteus species play important roles during cheese ripening. The results of our study are important for further research on cheese flavor and for quality control during cheese production. © 2019 Society of Chemical Industry.