Graviera Cheese Research Articles

A Study of Greek Graviera Cheese by NMR-Based Metabolomics.

Graviera is a very popular yellow hard cheese produced in mainland Greece and the Aegean islands, and in three PDO (protected denomination of origin) locations. Apart from geographic location, type of milk and production practices are also factors that affect cheese composition, and make this dairy product unique in taste and aroma. In this work, 1H nuclear magnetic resonance (NMR) spectroscopy in combination with chemometrics has been used to determine the metabolite profile (40 compounds) of graviera cheese produced in different geographic locations, with emphasis on cheeses produced on the island of Crete. Organic acids and amino acids were the main components quantified in the polar cheese fraction, while the fatty acid (FA) composition of the lipid fraction was also obtained. Analysis of variance (Anova) of the dataset showed that γ-aminobutyric acid (GABA), conjugated linoleic acids (CLA) and linoleic acid differentiate gravieras produced in different areas of Crete, and that the total amino acid content was higher in cheeses produced in eastern Crete. Targeted discriminant analysis models classified gravieras produced in mainland Greece, Cyclades and Crete based on differences in 1,2-diglycerides, sterols, GABA and FA composition. Targeted and untargeted orthogonal partial least squares discriminant analysis (OPLS-DA) models were capable of differentiating gravieras produced in the island of Crete and hold promise as the basis for the authentication of PDO graviera products.

Indigenous Lactic Acid Bacteria Isolated from Raw Graviera Cheese and Evaluation of Their Most Important Technological Properties.

The aim of the present study was to characterize LAB isolates from raw-milk cheeses, to evaluate some of their technological properties and to select a few 'wild' LAB strains that could potentially be used as starter cultures. LAB strains were isolated and identified from raw milk, curd, and cheese at 30, 60, and 90 days of ripening. A total of 100 strains were isolated, 20 from each phase of ripening. All isolates were tested for acidification ability, curd formation, and aroma production at 32 °C and 42 °C after 24 and 48 h. Following the acidification test, 42 strains were selected for identification and characterization of their technological properties. A high proportion of lactic acid bacteria and Gram + cocci were found throughout the cheese-making process. Enterococci reached their maximum proportion on the 7th day of ripening while Lactobacilli increased significantly during the first month of ripening. Forty-two strains were identified by phenotypic, biochemical, and molecular techniques. Lactococci were predominant in raw milk and curd while Lactobacilli in the ripening of the cheese. Four LAB strains including one Leuconostoc pseudomenteroides, two Lacticaseibacillus paracasei subsp. paracasei and one Enterococcus hirae, were proposed for their potential use as starters or secondary cultures.

Physicochemical Characteristics, Fatty Acids Profile and Lipid Oxidation during Ripening of Graviera Cheese Produced with Raw and Pasteurized Milk.

The production of cheese can be made from either pasteurized or non-pasteurized milk, depending on the country or dietary habits. In this work, the effect of pasteurization of milk on the progress of the physicochemical properties, fatty acids profile and lipid oxidation of cheese throughout a maturation period of 90 days is presented. This research was carried out on two types of Graviera cheese produced in Greece, one made from raw milk and the other from pasteurized milk. The proximal composition of each sample was evaluated, the fatty acids profile was analyzed by Gas Chromatography, whereas lipid oxidation was determined on the basis of the formation of malondialdehyde (MDA). Significant differences (p < 0.05) in the values of pH, fat and density between raw and pasteurized milk were observed. The physicochemical parameters during the ripening of the cheeses showed significant differences according to the type and the stage of maturation. Specifically, the two types of cheese differed significantly (p < 0.05) in terms of pH, protein, fat in dry matter (FDM), and water-soluble nitrogen/total nitrogen (WSN/TN). Although the fatty acids profile was similar for the two types of cheese, differences were observed during the ripening stages as well as between the milk and the final product. The lipid oxidation levels increased during maturation, whereas they seemed to be lower in the pasteurized cheeses. Therefore, it can be concluded that the use of raw or pasteurized milk affects the physicochemical characteristics, fatty acids profile and lipid oxidation of Graviera cheese during ripening.

Dietary Intake Assessment of Pre-Packed Graviera Cheese in Greece and Nutritional Characterization Using the Nutri-Score Front of Pack Label Scheme.

Gravieras are ‘gruyere’ type hard cheeses with a variety of different products and the second highest consumption in Greece. In this study, we present a dietary intake assessment and a nutritional characterization of pre-packed graviera products sold in the Greek market using Nutri-Score Front of Pack Label (FoPL). The nutrient contents of 92 pre-packed graviera products were combined with daily individual consumption data extracted from the Hellenic National Nutrition Health Survey (n = 93), attempting to evaluate the contribution of graviera’s consumption to the Greek diet. The analysis of nutrients’ intake as a Reference Intake (RI) percentage ranked saturated fat first on the nutrients’ intake list, with RI percentage ranging from 36.1 to 109.2% for the 95th percentile of consumption. The respective % RI for energy, total fat, carbohydrates, sugars, proteins and salt ranged from 12.7–20.7%, 21.6–50.4%, 0–3.1%, 0–6.1%, 37–57.1% and 6.3–42%. Nutri-Score classified 1% of the products to C—light orange class, 62% to D—orange and 37% to E—dark orange, while no products were classified to A—dark green or B—green classes. The comparison between the Nutri-Score classification and the nutrients’ intake assessment, also separately conducted within the classes, showed a higher salt intake after the consumption of products classified as D—orange and E—dark orange.

Physicochemical, Spectroscopic, and Chromatographic Analyses in Combination with Chemometrics for the Discrimination of the Geographical Origin of Greek Graviera Cheeses.

Seventy-eight graviera cheese samples produced in five different regions of Greece were characterized and discriminated according to geographical origin. For the above purpose, pH, titratable acidity (TA), NaCl, proteins, fat on a dry weight basis, ash, fatty acid composition, volatile compounds, and minerals were determined. Both multivariate analysis of variance (MANOVA) and linear discriminant analysis (LDA) were applied to experimental data to achieve sample geographical discrimination. The results showed that the combination of fatty acid composition plus minerals provided a correct classification rate of 89.7%. The value for the combination of fatty acid compositions plus conventional quality parameters was 94.9% and for the combination of minerals plus conventional quality parameters was 97.4%. When cheeses of the above five geographical origins were combined with previously studied graviera cheeses from six other geographical origins collected during the same seasons in Greece, the respective values for the discrimination of geographical origin of all eleven origins were 89.3% for conventional quality parameters plus minerals; 94.0% for conventional quality parameters plus fatty acids; 94.1% for minerals plus fatty acids; and 95.2% for conventional quality parameters plus minerals plus fatty acids. Such high correct classification rates demonstrate the robustness of the developed statistical model.

Characterisation and differentiation of geographical origin of Graviera cheeses produced in Greece based on physico-chemical, chromatographic and spectroscopic analyses, in combination with chemometrics

A total of 90 Graviera cheese samples, produced by selected local dairies in six different regions of Greece, were analysed for pH, titratable acidity, NaCl, proteins, fat on dry weight basis, ash, fatty acid composition by gas chromatography, volatile compounds by gas chromatography/mass spectrometry, and minerals by inductively coupled plasma - optical emission spectrometry, in an effort to differentiate Graviera cheeses according to geographical origin. Statistical treatment of the data included multivariate analysis of variance and linear discriminant analysis. The results showed that individual analytical parameters gave a correct classification of 46.7% for volatile compounds (VCs), 86.7% for physico-chemical parameters (PCPs), 91.1% for fatty acids (FAs) and 91.2% for minerals. Respective values for combinations of analytical parameters were 93.3% for PCPs plus FAs, 93.4% for PCPs plus FAs plus minerals, and 97.8% for FAs plus minerals.

Greek Graviera Cheese Assessment through Elemental Metabolomics-Implications for Authentication, Safety and Nutrition.

This study presents the comprehensive elemental profile of Greek Graviera (Gruyère) cheeses. In total, 105 samples from nine different geographic regions produced from sheep, goat and cow milk and their mixtures were assessed. Elemental signatures of 61 elements were investigated for determination of geographic origin and milk type. Regional and milk type classification through Linear Discriminant Analysis was successful for almost all cases, while a less optimistic cross validation exercise presented lower classification rates. That points to further research using a much larger sample set, increasing confidence for cheese authentication utilizing also bioinformatics tools under development. This is the first study reporting signatures of 61 elements in dairy products including all sixteen rare earth elements and all seven precious metals. Safety and quality were assessed regarding toxic and nutritive elements. According to both EU and USA regulations and directives, Graviera is a nutritional source for trace and macro elements with low levels of toxic elements.

Major ecological shifts within the dominant nonstarter lactic acid bacteria in mature Greek Graviera cheese as affected by the starter culture type

Traditional Greek Graviera cheese is often produced from thermized milk to control undesirable bacterial contaminants. Since thermization also reduces the desirable lactic acid bacteria (LAB) microbiota of raw milk, natural undefined or commercially defined starters are used. This study evaluated effects of the type of starter added to bulk thermized milk on the microbiology of mature (day-90) Graviera cheese. Cheeses produced with a natural starter culture (NSC) in non-concentrated yogurt-like form or a commercial starter culture (CSC) containing Streptococcus thermophilus and various Lactococcus lactis strains in concentrated freeze-dried form, were analyzed microbiologically, and 200 LAB isolates (100 from each type of cheese) were identified. The LAB microbiota of the mature CSC-cheeses was dominated by nonstarter strains of Lactobacillus paracasei and Lb. plantarum whereas indigenous Enterococcus faecium and E. durans strains of high phenotypic and genotypic diversity predominated in the respective NSC-cheeses. Populations of enterococci in CSC-cheeses were subdominant by 10 to 100-fold compared with those in NSC-cheeses; E. faecium was the most frequently isolated Enterococcus species from the mature CSC-cheeses. Sporadic or no isolates of other LAB species, including the commercial S. thermophilus and Lc. lactis starter strains in the CSC-cheeses and the natural S. thermophilus and Lactobacillus delbrueckii subsp. bulgaricus starter strains plus indigenous Lactococcus, Leuconostoc and E. faecalis in the NSC-cheeses, were detected. In conclusion, the replacement of the NSC with the CSC controlled growth of dairy enterococci in favor of mesophilic nonstarter lactobacilli during ripening. While safety concerns associated with the inefficiency of NSCs to prevent outgrowth of indigenous enterococci suggest that CSCs should be preferred by traditional Greek Graviera cheese processors, panel sensory evaluations showed that the NSC-ripened cheeses were of slightly lower appearance but of occasionally higher flavor scores than the CSC-ripened cheeses.

Hurdle factors minimizing growth of Listeria monocytogenes while counteracting in situ antilisterial effects of a novel nisin A-producing Lactococcus lactis subsp. cremoris costarter in thermized cheese milks.

The capacity of growth, survival, and adaptive responses of an artificial contamination of a three-strain L. monocytogenes cocktail in factory-scale thermized (65 °C, 30 s) Graviera cheese milk (TGCM) was evaluated. Bulk TGCM samples for inoculation were sequentially taken from the cheese making vat before process initiation (CN-LM) and after addition of a commercial starter culture (CSC), the CSC plus the nisin A-producing (NisA+) costarter strain Lactococcus lactis subsp. cremoris M78 (CSC + M78), and all ingredients with the rennet last (CSC + M78-RT). Additional treatments included Listeria-inoculated TGCM samples coinoculated with the NisA+ costarter strain M78 in the absence of the CSC or with the CSC in previously sterilized TGCM to inactivate the background microbiota (CSC-SM). All cultures were incubated at 37 to 42 °C for 6 h, followed by additional 66 h at 22 °C, and 48 h at 12 °C after addition of 2% edible salt. L. monocytogenes failed to grow and declined in all CSC-inoculated treatments after 24 h. In contrast, the pathogen increased by 3.34 and 1.46 log units in the CN-LM and the CSC-SM treatments, respectively, indicating that the background microbiota or the CSC alone failed to suppress it, but they did so synergistically. Supplementation of the CSC with the NisA+ strain M78 did not deliver additional antilisterial effects, because the CSC Streptococcus thermophilus reduced the growth prevalence rates and counteracted the in situ NisA+ activity of the costarter. In the absence of the CSC, however, strain M78 predominated and caused the strongest in situ nisin-A mediated effects, which resulted in the highest listerial inactivation rates after 24 to 72 h at 22 °C. In all TGCM treatments, however, L. monocytogenes displayed a “tailing” survival (1.63 to 1.96 log CFU/mL), confirming that this pathogen is exceptionally tolerant to cheese-related stresses, and thus, can't be easily eliminated.

Growth, nisA Gene Expression, and In Situ Activity of Novel Lactococcus lactis subsp. cremoris Costarter Culture in Commercial Hard Cheese Production

This study evaluated in situ expression of the nisA gene by an indigenous, nisin A-producing (NisA+) Lactococcus lactis subsp. cremoris raw milk genotype, represented by strain M78, in traditional Greek Graviera cheeses under real factory-scale manufacturing and ripening conditions. Cheeses were produced with added a mixed thermophilic and mesophilic commercial starter culture (CSC) or with the CSC plus strain M78 (CSC+M78). Cheeses were sampled after curd cooking (day 0), fermentation of the unsalted molds for 24 h (day 1), brining (day 7), and ripening of the brined molds (14 to 15 kg each) for 30 days in a fully controlled industrial room (16.5°C; 91% relative humidity; day 37). Total RNA was directly extracted from the cheese samples, and the expression of nisA gene was evaluated by real-time reverse transcription PCR (qRT-PCR). Agar overlay and well diffusion bioassays were correspondingly used for in situ detection of the M78 NisA+ colonies in the cheese agar plates and antilisterial activity in whole-cheese slurry samples, respectively. Agar overlay assays showed good growth (>8 log CFU/g of cheese) of the NisA+ strain M78 in coculture with the CSC and vice versa. The nisA expression was detected in CSC+M78 cheese samples only, with its expression levels being the highest (16-fold increase compared with those of the control gene) on day 1, followed by significant reduction on day 7 and almost negligible expression on day 37. Based on the results, certain intrinsic and mainly implicit hurdle factors appeared to reduce growth prevalence rates and decrease nisA gene expression, as well as the nisin A-mediated antilisterial activities of the NisA+ strain M78 postfermentation. To our knowledge, this is the first report on quantitative expression of the nisA gene in a Greek cooked hard cheese during commercial manufacturing and ripening conditions by using a novel, rarely isolated, indigenous NisA+ L. lactis subsp. cremoris genotype as costarter culture.

Positive effects of sequential air ventilation on cooked hard Graviera cheese ripening in an industrial ripening room

An industrial ripening room used for Graviera cheese ripening was equipped in order to compare Continuous Ventilation (CV: full time air circulation) and Temperature based Sequential Ventilation (TSV) processes. Under controlled temperature and Relative Humidity of the ripening room, three reference 30-day ripening trials performed in CV mode were compared to three similar trials performed in TSV mode. TSV was monitored between 2 temperature set points. When the lower one (16 °C) was reached, ventilation was stopped; when the higher one (16.8 °C) was reached, ventilation restarted. Power consumption, cheese mass loss and cheese quality were determined as comparison criteria. The use of TSV involved important reductions on ventilation time (48%), energy consumption (42%), cheese mass loss (0.9%) leading to a positive impact on the economy of the ripening process. Additionally, as shown by microbiological, physicochemical and sensory analyses, TSV neither had negative nor significant positive effects on cheese quality.

Enhanced Control of Listeria monocytogenes by Enterococcus faecium KE82, a Multiple Enterocin–Producing Strain, in Different Milk Environments

Enterococcus faecium KE82, isolated from traditional Greek Graviera cheese, was identified in pure broth cultures in vitro as a multiple enterocin-producing bacterial strain possessing the structural entA, entB, and entP enterocin genes. E. faecium KE82 was further assessed for in situ antilisterial activity in raw milk (RM) and commercially thermized milk (TM; 63°C for 30 s) in the presence of the indigenous microbiota and in sterile raw milk (SRM; 121°C for 5 min) with or without the addition of two commercial starter culture (CSC) strains Streptococcus thermophilus and Lactococcus lactis . Growth of Listeria monocytogenes was completely inhibited in RM incubated at 37°C for 6 h, whereas the pathogen was significantly inactivated in RM+KE82 samples during further incubation at 18°C for 66 h. In contrast, L. monocytogenes levels increased by approximately 2 log CFU/ml in TM, but in TM+KE82 samples, pathogen growth was retarded during the first 6 h at 37°C followed by growth cessation and partial inactivation at 18°C. After 48 to 72 h, growth of L. monocytogenes in SRM+CSC samples decreased by 4 to 5 log CFU/ml compared with the SRM control, whereas additional 10-fold decreases in the pathogen were observed in SRM+CSC+KE82 samples. Reverse transcription PCR analysis of SRM+KE82 and SRM+CSC+KE82 samples confirmed that the entA and entB genes were transcribed, but entP gene transcription was not detected. All RM and SRM samples inoculated with E. faecium KE82 displayed strong in situ inhibitory activity against L. monocytogenes in well diffusion bioassays, whereas activity was weaker to undetectable in comparable or additional TM+KE82 samples; no milk sample without E. faecium KE82 had activity against L. monocytogenes . The findings of this study indicate that E. faecium KE82 is an antilisterial agent that could be used in traditional dairy foods because it concomitantly produces enterocins A and B in situ in milk.

Behavior of artificial listerial contamination in model Greek Graviera cheeses manufactured with the indigenous nisin A‐producing strain Lactococcus lactis subsp. cremoris M104 as costarter culture

AbstractGrowth of Listeria monocytogenes during processing of traditional Greek Graviera cheese is inhibited, but the pathogen may survive ripening. Therefore, this study used Lactococcus lactis subsp. cremoris M104, a nisin A‐producing (NisA+) raw milk isolate, to enhance inactivation of a nonpathogenic Listeria cocktail contaminated in model Graviera mini cheeses. Cheeses were manufactured from thermized milk with a commercial starter culture (CSC) or the CSC plus strain M104 (CSC + M104), ripened at 18 °C and 90% RH for 20 days and stored at 4 °C in vacuum for 60 days. Listeria populations declined 10‐fold in all fermenting cheeses but then survived with little death during ripening and storage. NisA+ M104 colonies were prevalent and the nisA gene was detected, but nisin activity was weak to undetectable, in all CSC + M104 cheeses. Thus, Graviera cheese processing should be optimized to ensure nisA gene expression by strain M104 at levels sufficient to increase inactivation of L. monocytogenes.Practical applicationsThis study provided important preliminary evidence that the in situ NisA+ antilisterial activity of L. lactis subsp. cremoris M104 during Graviera cheese fermentation increased by reducing the curd cooking temperature from 48 °C to 42 °C. This indicated the feasibility to increase nisin production and activity by suitably modifying selected technological factors during traditional cooked hard cheese processing. Apart from its bioprotective potential, this NisA+ L. lactis subsp. cremoris genotype (represented by the indigenous strains M104 and M78) possesses desirable sugar fermentation capabilities and enzymatic activities similar to those of the best performing industrial L. lactis or L. cremoris strains in commercial starters (Parapouli et al., ). Since 2015 this novel lactococcal genotype has been applied as hand‐made, costarter culture for the production of commercial Greek Graviera cheeses characterized by specific flavor characteristics and improved textural properties than counterpart cheeses produced with CSCs only (Pappas Bros. collaborating cheese plant, Epirus; private communication).

“Graviera Naxou and Graviera Kritis Greek PDO cheeses: Discrimination based on microbiological and physicochemical criteria and volatile organic compounds profile”

The purpose of the present investigation was to study the microbiological and physicochemical characteristics of Graviera cheese from raw milk, Graviera Naxou and Graviera Kritis, made in each of the respective islands, in order to discriminate those two types of cheeses. Totally, 103 Non Starter Lactic Acid Bacteria (NSLAB) isolates were obtained from fresh cheeses and after grouping by SDS-PAGE, selected strains were identified by 16S rRNA gene sequencing analysis. In Graviera Naxou leuconostocs and Lactobacillus brevis predominated. In Graviera Kritis lactococci predominated while enterococci and leuconostocs constituted a significant part. Mature Graviera Kritis cheeses had lower (P<0.05) moisture, salt-in-moisture, aminoacid content and caseins (CN) degradation. NSLAB from Graviera Naxou had stronger mean acidifying ability at 24h than Kritis’s, exhibited higher proteolysis, but lower antagonistic activity against undesirable bacteria. The profile of total microbiota by Denaturing Gradient Gel Electrophoresis (DGGE-PCR) suggested differences in the cheeses of the two islands, while profiles of NSLAB were more similar but not their composition and evolution. Each island had its own volatile organic compounds (VOCs) profile. Acids, esters and alcohols were the most abundant groups of VOCs. There were compounds common for both cheese types and others discriminant for each area. There were also common terpenes between pasture samples and the cheeses. The overall findings indicate that Graviera Naxou and Kritis cheeses differ considerably in respect of microbiological and physicochemical criteria, as well as VOCs content.

Behavior of Staphylococcus aureus in Culture Broth, in Raw and Thermized Milk, and during Processing and Storage of Traditional Greek Graviera Cheese in the Presence or Absence of Lactococcus lactis subsp. cremoris M104, a Wild, Novel Nisin A–Producing Raw Milk Isolate

This study was conducted to evaluate the behavior of Staphylococcus aureus during processing, ripening, and storage of traditional Greek Graviera cheese in accordance with European Union Regulation 1441/2007 for coagulase-positive staphylococci in thermized milk cheeses. Lactococcus lactis subsp. cremoris M104, a wild, novel nisin A-producing (NisA+) strain, also was evaluated as an antistaphylococcal adjunct. A three-strain cocktail of enterotoxigenic (Ent+) S. aureus increased by approximately 2 log CFU/ml when co-inoculated (at approximately 3 log CFU/ml) in thermized Graviera cheese milk (TGCM; 63°C for 30 s) with commercial starter culture (CSC) and/or strain M104 at approximately 6 log CFU/ml and then incubated at 37°C for 3 h. However, after 6 h at 37°C, significant retarding effects on S. aureus growth were noted in the order TGCM + M104 > TGCM + CSC = TGCM + CSC + M104 > TGCM. Additional incubation of TGCM cultures at 18°C for 66 h resulted in a 1.2-log reduction (P < 0.05) of S. aureus populations in TGCM + M104. The Ent + S. aureus cocktail did not grow but survived during ripening and storage when inoculated (at approximately 3 log CFU/g) postcooking into Graviera mini cheeses prepared from TGCM + CSC or TGCM + CSC + M104, ripened at 18°C and 90% relative humidity for 20 days, and stored at 4°C in vacuum packages for 2 months. A rapid 10-fold decrease (P < 0.05) in S. aureus populations occurred within the first 24 h of cheese fermentation. Reductions of S. aureus were greater by approximately 0.4 log CFU/g in CSC + M104 than in CSC only cheeses, concomitantly with the presence of NisA + M104 colonies and nisin-encoding genes in the CSC plus M104 cheeses and their corresponding microbial consortia only. A high level of selective survival of a naturally nisin-resistant EntC z S. aureus strain from the cocktail was noted in CSC + M104 cheeses and in coculture with the NisA + M104 strain in M-17 broth. In conclusion, although S. aureus growth inhibition is assured during Graviera cheese ripening, early growth of the pathogen during milk curdling and curd cooking operations may occur. Nisin-resistant S. aureus strains that may contaminate Graviera cheese milks postthermally may be difficult to control even by the application of the NisA + L. lactis subsp. cremoris strain M104 as a bioprotective adjunct culture.

Probiotic and Technological Properties of Facultatively Heterofermentative Lactobacilli from Greek Traditional Cheeses

Nineteen isolates of facultatively heterofermentative lactobacilli from Feta, Graviera, and Kasseri cheeses were identified by SDS-PAGE of whole-cell proteins as L. paracasei subsp. paracasei (12 strains) and L. plantarum (7 strains) and differentiated at strain level by RAPD-PCR. Properties of technological interest, such as acidification ability, proteolytic activity, and enzyme activities, were also studied. The test strains exhibited a low acidification activity, with significant interstrain differences after growth in milk for 24 h. They were also characterized by different casein breakdown ability, with around 50% of them accumulating amino acids at low amounts in the milk. Lactobacilli isolates differed in respect of enzyme activities, with β-galactosidase being the strongest activity found. Their probiotic potential was evaluated with in vitro studies on the resistance to low pH, bile salts, and pancreatin. The isolates from Feta showed a better survival than those from Kasseri and Graviera at low pH and viable cells were detected even after 3 h at pH 2.0. All strains tolerated bile salts at 0.3% and retained viability in the presence of pancreatin at 0.1%. Different patterns of antibacterial activities were recorded. The strains inhibited preferentially LAB species and some of them clostridia, E. coli O44 and B. cereus. Distinguished strains are promising probiotic candidates as adjuncts and deserve further studies.

Effect of several MAP compositions on the microbiological and sensory properties of Graviera cheese

The shelf life of Graviera cheese, a full fat cheese produced in Heraklion (Crete Greece), was investigated. Graviera cheese was stored at 4 °C for up to 90 days in polyamide packages under three different modified atmosphere compositions. Control cheeses were packaged in air whereas MAP mixtures were MAP 1: 40% CO 2/55% N 2/5% O 2, MAP 2: 60% CO 2/40% N 2 and MAP 3: 50% CO 2/50% N 2. Sampling of product was carried out every 10 days to investigate its sensory quality and microbiological characteristics. Ten trained panelists participated in the sensory panel to evaluate the cheeses for external appearance (color, texture), taste, and flavor in a scale from 1 to 10 (1 very poor, 10 very good). The microbiological analysis revealed that there were no colonies of Staphylococcus aureus and Listeria monocytogenes whereas both Escherichia coli and Total Viable Counts (TVC) increased strongly in control samples but were inhibited under all MAP compositions.

Quality Evaluation of Grated Graviera Cheese Stored at 4 and 12°C using Active and Modified Atmosphere Packaging

AbstractThe effect of active [oxygen absorber (OA) combined with an ethanol emitter (EE)] and modified atmosphere (100% Nitrogen) packaging in combination with a high barrier experimental polyethylene terephthalate‐silicon oxides//low density polyethelene film on shelf‐life extension of grated Graviera cheese stored at 4 and 12°C was investigated. Microbiological (total viable counts, Pseudomonads, lactic acid bacteria, Enterobacteriaceae and Yeasts/Moulds), physico‐chemical (pH, thiobarbituric acid and colour) and sensory (odour and taste) changes occurring in the product were monitored as a function of treatment and storage time (10 weeks). Sensory shelf‐life was approximately 1, 1.5, 4.5, 6, 9 and at least 10 weeks for control samples (12 and 4°C), for N2 packaged samples (12 and 4°C) and samples packaged with the OA + EE (12 and 4°C), respectively. At the point of sensory rejection, yeasts and moulds increased from 2.00 to 3.60 and 5.55 log CFU/g for control samples stored at 4 and 12°C, respectively. Similarly, for samples stored under nitrogen, yeasts and moulds reached 2.00 and 2.32 log CFU/g at 4 and 12°C, respectively. Yeasts and moulds in samples with the OA + EE remained below 2 log CFU/g throughout the entire storage period, irrespective of storage temperature. pH varied between 5.72 and 6.49 depending on specific treatment. Malondialdehyde absorbance ranged between 0.05 for fresh samples and 0.79 mg/kg at the time of sensory rejection of samples. Light parameters L and b decreased while parameter a increased during storage, reflecting a gradual discolouration of all samples and especially those stored at 12°C.

FTIR-based polyphasic identification of lactic acid bacteria isolated from traditional Greek Graviera cheese

This study used a combination of phenotypic, physical (Fourier Transformed Infra-Red [FTIR] spectroscopy) and molecular (RFLP and SSCP analysis of 16S rRNA genes) methods to identify the lactic acid bacteria (LAB) flora present in traditional Greek Graviera cheese after five weeks of ripening. A total of 300 isolates collected from high dilution plates of TSAYE (incubated at 30 °C), M-17 (22 °C) and M-17 (42 °C) agar media were clustered by FTIR and then representative strains of each cluster were cross-identified blindly by all methods. Based on their FTIR spectra, 282 isolates were LAB grouped in 28 clusters. The LAB species identified and their prevalence in the cheese samples were: Lactobacillus casei/ paracasei (68.8%), Lactobacillus plantarum (19.5%), Streptococcus thermophilus (8.9%), Enterococcus faecium (2.1%), and Lactococcus lactis (0.7%). Also, Staphylococcus equorum (11 isolates), Corynebacterium sp. (5 isolates) and Brevibacterium sp. (1 isolate) were recovered from TSAYE. Comparative identification results showed that phenotypic and molecular methods were in mutual agreement as regards the LAB species identified. The present polyphasic identification approach based on rapid FTIR screening of 10-fold more isolates than a previous classical identification approach allowed or improved detection of few sub-dominant species; however the predominant LAB species in the cheese samples were the same with both approaches.

Microbial stability and safety of traditional Greek Graviera cheese: characterization of the lactic acid bacterial flora and culture-independent detection of bacteriocin genes in the ripened cheeses and their microbial consortia.

The microflora of four batches of traditional Greek Graviera cheese was studied at 5 weeks of ripening, and 200 lactic acid bacteria (LAB) isolates were phenotypically characterized and screened for antilisterial bacteriocins. The cheeses were also analyzed for organic acids by high-performance liquid chromatography and for the potential presence of 25 known LAB bacteriocin genes directly in cheese and their microbial consortia by PCR. All batches were safe according to the European Union regulatory criteria for Listeria monocytogenes, Salmonella, enterobacteria, and coagulase-positive staphylococci. The cheese flora was dominated by nonstarter Lactobacillus casei/paracasei (67.5%) and Lactobacillus plantarum (16.3%) strains, whereas few Streptococcus thermophilus (3.8%), Lactococcus lactis subsp. lactis (0.6%), and Leuconostoc (1.9%) organisms were present. Enterococcus faecium (9.4%) and Enterococcus durans (0.6%) were isolated among the dominant LAB from two batches; however, enterococci were present in all batches at 10- to 100-fold lower populations than mesophilic lactobacilli. Sixteen E. faecium isolates produced antilisterial enterocins. In accordance, enterocin B gene was detectable in all cheeses and enterocin P gene was present in one cheese, whereas the consortia of all cheeses contained at least two of the enterocin A, B, P, 31, L50A, and L50B genes. Plantaricin A gene was also amplified from all cheeses. Mean concentrations of lactic, acetic, citric, and propionic acids in the ripened cheeses exceeded 1.5% in total, of which approximately 0.9% was lactate. Thus, organic acid contents constitute an important hurdle factor for inhibiting growth of pathogens in traditional Graviera cheese products, with LAB bacteriocins, mainly enterocins, potentially contributing to increased cheese safety.