The occurrence of ochratoxin A in blue cheese

Evidence for the occurrence of ochratoxin A in blue cheeses is reported for the first time. The development of an accurate and reliable procedure for the extraction of OTA from cheese, as well as the availability of a new sensitive HPLC-FLD method, has allowed us to determine ochratoxin A in complex matrices such as cheeses, even at very low levels (LOD in cheese: 0.02 μg/kg). A good linearity for the OTA concentration, between 0.2 and 2.5 μg/kg, was obtained and no matrix effect was observed in the same concentration range. The mean recovery for OTA was 97%, while the average RSD was 3%, within a spiking range of 0.5–2.0 μg/kg.     

蓝纹干酪加工工艺研究

以新鲜牛乳为原料，通过单因素实验和正交实验对蓝纹干酪的加工工艺进行了初步研究，确定了制备蓝纹干酪的工艺流程及参数。新鲜牛乳经添加发酵剂和凝乳酶，凝乳切割，加热，排乳清，入模，盐浴，打孔，经12～14d成熟即得到蓝白相闻的蓝纹干酪。pH值对凝乳质地和霉菌的生长具有显著的影响，添加凝乳酶的最佳pH值为6.1。凝块的加热温度为36℃，保证干酪的硬度，避免干酪打孔后孔隙复合及干酪坍塌。     

Yeast occurrence in Danish feta cheese

The yeast content in feta cheese from three different dairies was examined. Almost all samples contained yeasts in considerable numbers, 10⁴–10⁶cfu g⁻¹. A specific narrow yeast flora was found for each dairy.Torulaspora delbrueckiiwas the dominating species in feta from Dairy A. This strong-fermenting species was found to cause swelling of the cans.Debaryomyces hansenii,Candida sakeandKluyveromyces marxianuswere dominant in samples from Dairy B.Candida butyriandDebaromyces hanseniiwere found in Dairy C.Yarrowia lipolyticawas present in low numbers in cheese samples from all three dairies.Geotrichum candidumwas also commonly found although in low concentrations. Yeast occurrence was due to recontamination. Yeasts were found in high concentrations in production areas.     

Partial characterization of dextran-degrading enzyme obtained from blue cheese

Degradation of dextran beads was observed when the water-soluble fraction of a blue cheese extract was applied to the top of a Sephadex G-150 or G-200 column. This phenomenon suggests the presence of a specific enzyme that can hydrolyze dextran. After removal of casein components from the blue cheese fraction, ammonium sulfate treatment and gel filtration chromatography were performed to isolate the enzyme fraction. The enzymatic products were analyzed by thin-layer chromatography and gel filtration chromatography and identified as isomaltooligosaccharides. The isoelectric point of this enzyme fraction was approximately 4.9, as determined by isoelectric focusing using Rotofor, and the molecular weight of the fraction was 65 kDa, as estimated by sodium dodecyl sulfate (SDS)-PAGE. Optimum pH for enzymatic activity was 5.0 to 5.3. A partial N-terminal amino acid sequence of 20 residues was determined to be ATPDEWRSRSIYFMLTDRGA from an enzyme fraction further purified by ion-exchange chromatography and native PAGE. This sequence showed a maximum homology of 80% with alpha-amylase or Taka amylase that originated from various microorganisms.     

Lactic acid bacteria isolated from a hand-made blue cheese

Nearly 500 strains isolated from different media used to study the aerobic mesophilic and lactic acid flora of Valdeón cheese (a Spanish hand-made blue cheese) have been identified. Nearly 95% of aerobic mesophiles were lactic acid bacteria (LAB). From these, Enterococcus (40·4%) and Lactococcus (42·2%) were the dominant genera, with Lactobacillus (4·1%) and Leuconostoc (5·0%) being also found. The selectivity of the other media used was variable and this is discussed. Several species of Enterococcus were isolated from our samples (Ent. avium, Ent. faecium and Ent. durans), although one was outstanding (Ent. faecalis, 24·7% of the total of strains identified). The dominating LAB species found was Lact. lactis subsp. lactis (31·1%). Other LAB identified were Lact. raffinolactis, Lb. plantarum, Lb. casei, Leuc. mesenteroides subsp. dextranicum, Leuc. mesenteroides subsp. mesenteroides and Leuc. paramesenteroides. The evolution of the lactic-acid flora found during the manufacture and ripening of this variety of cheese showed a pattern marked by the dominance of lactococci and enterococci during the first stages and the substitution of lactococci by lactobacilli and leuconostoc from drying onwards which would be, along with enterococci, the major genera found in the cheese at the stage of consumption. From a technological point of view, the quantitative importance of certain species such as Lact. lactis and Ent. faecalis, as well as the presence of others, i.e. Lb. plantarum and Leuc. mesenteroides, suggest their possible use as starters in the industrial manufacture of this variety.     

蓝纹干酪成熟期间氨基酸的变化研究

采用HITACHIL—8800型氨基酸自动分析仪测定分析了自制蓝纹干酪成熟过程中的氨基酸组分与含量变化。成熟30～60d期间,所有氨基酸含量都呈现上升趋势,成熟90d时,门冬氨酸、谷氨酸、缬氨酸、蛋氨酸、亮氨酸、苯丙氨酸、精氨酸、脯氨酸几种氨基酸由于转化成其他物质含量有所降低。门冬氨酸降低0.07%,谷氨酸降低0.32%,缬氨酸降低0.09%,蛋氨酸降低0.14%,亮氨酸降低0.22%,苯丙氨酸降低0.02%,精氨酸降低0.07%,脯氨酸降低0.32%。成熟90d时,含量较高的是谷氨酸,亮氨酸,脯氨酸,赖氨酸、酪氨酸。     

活性艳蓝筒子纱色花的解决方法

分析活性艳蓝筒子染色花纱的原因，制定相应的工艺和预防措施。诸如清水化料，选用元明粉作促染剂；未固色前提高染液温度到70℃，降温至55℃再固色；固色初期加小苏打；分次加入碱剂；染后立即溢流洗水等。     

Comparative study of different methods for evaluating proteolysis in blue cheese

Summary Methods of evaluating proteolysis using different precipitants and nitrogen evaluation methods were assessed. The coefficient of determination (R ²) values relating the various methods and the ripening period were positive and greater than 0.87. Amino acid nitrogen in the fraction soluble in 12% trichloroacetic acid had the highest coefficient value (R ²=0.95). Of all the methods for evaluating nitrogen, dialysis had the highest value ofR ² with respect to water-soluble nitrogen determined using the Kjedahl method. However, dialysis is not applicable to blue cheeses with high mould contents. Cluster analysis was applied to the ripening index values and yielded groupings according to the method of nitrogen evaluation.

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Vergleichende Studie über verschiedene Methoden der Bewertung der Proteolyse im Blauschimmelkäse

Zusammenfassung Die Methoden zur Bewertung der Proteolyse verwenden verschiedene Methoden an Fällungsmitteln und der Stickstoffauswertung. Die Koeffizienten der Bestimmungswerte (R ²) bei verschiedenen Methoden und der Reifungsperiode waren positiv und größer als 0,87. Der Aminosäuren-Stickstoff in der löslichen Fraktion in der 12%igen Trichloressigsäure hatte den höchsten Wert (R ²=0,95). Von allen Methoden der Stickstoff-Bewertung hatte die Dialyse den höchstenR ²-Wert bezogen auf den mit der Kjeldahl-Methode erzielten wasserlöslichen Stickstoff; jedoch die Dialyse ist bei Blauschimmelkäse mit hohem Pilzgehalt nicht brauchbar. Die Cluster-Analyse war für den Reife-Index und die Gruppenausbeute für die Methoden der Stickstoff-Auswertung anwendbar.

     

Electronic nose: New tool in modelling the ripening of Danish blue cheese

Changes during maturation of Danish blue cheese were analysed with a commercial electronic nose (e-nose) system. Features of the signals of the chemical sensors when in contact with the cheese headspace were modelled based on multivariate analysis. Chemical analyses performed on the same cheeses were used to indicate these changes. E-nose measurements were then used to model and predict the chemical changes that are strongly related to the quality parameters of this cheese. Prediction models were then tested on cheeses from the same producer. Selection of sensors and data pretreatment makes the modeling of the cheeses maturation and consequently the prediction of quality characteristics possible with the e-nose system.     

Taxonomical and technological characteristics of Saccharomyces spp. associated with blue veined cheese

In blue veined cheeses, the dominant yeast species in most cases is Debaryomyces hansenii. Saccharomyces spp. occurs less frequently, but they can be found in some blue veined cheeses. In the present study, the taxonomy of Saccharomyces spp. associated to blue veined cheeses was studied and comparisons made to type strains of Saccharomyces spp. and starter cultures of Saccharomyces spp. used in other food fermentations. Phenotypically, the cheese strains were referred to the Saccharomyces sensu stricto complex and were further identified as S. cerevisiae. Genotypically, the Saccharomyces spp. investigated were similar although chromosomal polymorphism were observed. Concerning the technological characteristics, they were similar in assimilation and fermentation of the residual sugars and organic acids naturally found in cheese. The investigated yeasts were also similar in their lipolytic activity being able to hydrolyse tributyrin and low chain (C:8), but not C:14 fatty acids. However, they differed in their tolerance to NaCl with the blue cheese strains showing a higher tolerance. The cheese strain S. cerevisiae FB 7 was the only yeast capable of degrading casein. It mainly degraded the alpha(s1)-casein and the beta(alpha2)-casein components. It was also the only isolate stimulating the development of Penicillium roqueforti in cheese agar imitating the conditions in blue veined cheese. The stimulation of P. roqueforti was most pronounced for the least proteolytic strain of P. roqueforti examined.     

Free and total amino acids in the non-protein fraction of an artisan blue cheese during ripening

Summary Changes in the free amino acid content and total amino acid content of the fraction soluble in 5% phosphotungstic acid from three batches of blue cheeses were studied by HPLC over the ripening period. The correlation coefficients between the free and total amino acids and ripening time were positive and higher than 0.85 for most amino acids. Non-protein amino acids showed the lowest correlations with ripening. Principal component analysis performed on the data showed ripening time to be the main cause of variation between the samples analyzed, explaining 60% of the variance; differences between batches were the second most important source of variation (15%). The ripening stage can be approximately estimated from the concentration of certain amino acids.

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Freie und gesamte Aminosäuren in der Nicht-Protein-Fraktion eines Blaukäses während der Reifung

Zusammenfassung Während des Reifungsprozesses bei drei Partien Blauschimmelkäse wurde der Verlauf des Gehaltes an freien und gesamten Aminosäuren der in 5% iger Phosphorwolframsäure löslichen Fraktion mittels HPLC beobachtet. Die Korrelationskoeffizienten zwischen dem Gehalt an freien und Gesamt-Aminosäuren während der Reifungszeit waren positiv und größer als 0,85 für die meisten Aminosäuren. Ein geringerer Korrelationsgrad im Verlauf der Reifung war bei den Nicht-Protein-Aminosäuren zu beobachten. Die Anwendung der Faktor-Analyse wies darauf hin, daß die Reifung die wichtigste Ursache der Unterschiede zwischen den analytischen Proben war (60% der Varianz) und die zweite Ursache für die Varianz (15%) der Unterschied zwischen den Partien. Der Reifungsgrad kann in etwa aus der Konzentration bestimmter Aminosäuren geschätzt werden.

     

Mycotoxin-forming ability of two Penicillium roqueforti strains in blue moldy tulum cheese ripened at various temperatures

Isolated and identified toxigenic and nontoxigenic Penicillium roqueforti (PR) strains from moldy tulum cheeses were inoculated into tulum cheeses made in the laboratory and ripened at 5 and 12 degrees C. Mycotoxin (patulin, penicillic acid, PR toxin, and roquefortine) formation in the control and mold-inoculated cheeses were detected by thin-layer chromatography on the first through fourth months of ripening. Patulin, penicillic acid, and PR toxin were not detected in the experimental cheeses. Only roquefortine was detected in cheese inoculated with the toxigenic strain of the mold and ripened at 5 and 12 degrees C on the third and first months of ripening, respectively. Toxin in cheeses ripened at 5 and 12 degrees C was 2.1 to 2.4 and 2.1 to 3.8 mg/kg cheese, respectively.     

The occurrence of Listeria monocytogenes in cheese from a manufacturer associated with a case of listeriosis

A case of listeriosis was associated with the consumption of a soft cheese produced in England. Goats cheese and other products from the same food manufacturer were examined for the presence of Listeria over the following 11 months. Listeria monocytogenes was isolated from 16 of 25 cheese samples on retail sale, 12 of 24 cheese samples obtained directly from the factory, and from shelving within the plant. Phage-typing of 68 isolates of L. monocytogenes from cheese samples and the factory showed that 66 (97%) were indistinguishable from the strain isolated from the patient's cerebrospinal fluid and stool. L. monocytogenes was not isolated from seven goats milk or two yoghurt samples. Listeria innocua was isolated from 10 cheese samples, two of which contained no other species of Listeria. Levels of L. monocytogenes shortly after production were low (<10/g), but were higher (10⁵–10⁷ cfu/g) in six of the 16 cheese samples obtained from retail outlets. Multiplication of L. monocytogenes was demonstrated in cheeses contaminated at the factory and held at 4°C in the laboratory.     

Identification of the Pseudomonas fluorescens group as being responsible for blue pigment on fresh cheese

New cases of blue cheese discoloration has led to recent research to identify the causal agent and factors that favor blue pigment appearing. Nonetheless, very few reports have described the source of contamination and the measurements to eradicate the microbiological source on cheese farms by determining the relation between blue discoloration on fresh cheese and the Pseudomonas fluorescens group. Thus, 60 samples from a cheese farm (cheese, equipment surfaces, tap water, and raw and pasteurized milk) were analyzed by phenotypical, MALDI-TOF, 16S rRNA sequencing and pulsed-field gel electrophoresis tests to determine the causal agent. The results obtained by pulsed-field gel electrophoresis with restriction enzymes XbaI and SpeI confirmed tap water as the initial contaminated source. The above-mentioned result was essential to avoid Pseudomonas contamination due to the most residual microorganisms being inactivated through a new disinfection program.     

The occurrence of Staphylococcus aureus on a farm with small-scale production of raw milk cheese

In recent years, the small-scale production of raw milk products has increased in Norway, and there is some concern that such foods may pose a risk of staphylococcal food poisoning to consumers. The aim of the study was to evaluate potential sources of contamination of raw milk cheese with Staphylococcus aureus on a bovine dairy farm with small-scale production. Samples for bacteriological analyses (n = 144) were collected from the animals, the environment, processing equipments, from humans, and from cheeses at various stages of production. Staphylococcus aureus was isolated from 10 of 11 cows, the farmer, equipment, the environment, and the cheese. Seventy-five Staph. aureus isolates were genotyped by pulsed-field gel electrophoresis, tested for enterotoxin (SE) production by reversed passive latex agglutination, for SE genes by multiplex polymerase chain reaction, and for penicillin resistance by the cloverleaf method. Five different pulsotypes were identified and SE gene fragments were identified in 11 isolates, but no isolates produced SE or were penicillin resistant. Staphylococcus aureus was found throughout the farm, and appeared to be spread with the milk to the environment, equipment, and to products. One pulsotype dominated and was identified from most sample sites on the farm. Raw milk products are vulnerable to contamination with Staph. aureus. Strategies to reduce the occurrence of Staph. aureus in bulk milk are of particular importance on farms where milk is used for raw milk products.     

Genome-based analysis of Pseudomonas paracarnis RQ057, a strain responsible for blue discoloration spoilage in processed cheese

A strain of Pseudomonas causing blue discoloration was isolated from processed cheese ‘Requeijão em Barra’ and studied by whole-genome sequencing. The strain RQ057 was identified as Pseudomonas paracarnis using the TYGS web server and also by in silico DNA–DNA hybridisation and ANIb calculation. Pyoverdine biosynthetic clusters, integrative and conjugative elements (ICE), prophage regions and genomic islands (GIs) were predicted. Furthermore, we observed that copies of genes related to the tryptophan biosynthetic pathway (trpCBAFD), which are exclusive to strains causing blue discoloration, are contained in the region identified as ICE and which was also identified in GIs. This genomic region also contains genes for transposase, type IV transport system (T4SS) and pili construction. Thereby, we can assume that this region contains potential conjugative-transposon-like mobile genetic elements and it has plasmid-like conjugative properties.     

The occurrence of Listeria monocytogenes in cheese from a manufacturer associated with a case of listeriosis

A case of listeriosis was associated with the consumption of a soft cheese produced in England. Goats cheese and other products from the same food manufacturer were examined for the presence of Listeria over the following 11 months. Listeria monocytogenes was isolated from 16 of 25 cheese samples on retail sale, 12 of 24 cheese samples obtained directly from the factory, and from shelving within the plant. Phage-typing of 68 isolates of L. monocytogenes from cheese samples and the factory showed that 66 (97%) were indistinguishable from the strain isolated from the patient's cerebrospinal fluid and stool. L. monocytogenes was not isolated from seven goats milk or two yoghurt samples. Listeria innocua was isolated from 10 cheese samples, two of which contained no other species of Listeria. Levels of L. monocytogenes shortly after production were low (<10/g), but were higher (10⁵–10⁷ cfu/g) in six of the 16 cheese samples obtained from retail outlets. Multiplication of L. monocytogenes was demonstrated in cheeses contaminated at the factory and held at 4°C in the laboratory.     

The metabolism of fatty acids, methyl ketones and secondary alcohols by Penicillium roqueforti in blue cheese slurries.

A slurry system in which Blue cheese flavour was produced in six days was developed and used to study the metabolism of fatty acids, methyl ketones and secondary alcohols by Penicillium roqueforti. It was found that fatty acids inhibited lipolysis but that far greater quantities of methyl ketones were produced when fatty acids were added to the slurries than when fatty acids were produced by the lipolytic activity of P. roqueforti. Methyl ketones and secondary alcohols added to Blue cheese slurries in quantities ten times as great as those found in mature Blue cheese were broken down within 7 days And the presence of fatty acids did not appreciably inhibit this breakdown.     

Microbiological changes in cottage cheese varieties during storage at +7°C

Studies have been made of microbiological changes in cottage cheese varieties during storage at 7°C. Commercially prepared products obtained directly from three manufacturers were at an initial pH in the range 4·6 to 5·1 and differed significantly in their content of sorbic acid/sorbate and of viable lactic streptococci. In many of the varieties that did not contain sorbic acid/sorbate multiplication occurred of Pseudomonas spp., Enterobacteriaceae and yeasts; spoilage occurred in some batches due to formation of a surface film of Pseudomonas fluorescens or of surface colonies of Sporobolomyces roseus and Trichosporon sp. at times equal to, or slightly greater than the recommended storage life. Where high numbers of viable lactic streptococci were present in varieties immediately after manufacture, no marked decrease in pH or increase in titratable acidity was observed during storage at 7°C. In varieties that contained sorbic acid at a concentration higher than approximately 500 mg kg^?1 little increase in numbers of bacteria or yeasts was observed during storage at 7°C. Decrease in quality due to separation of whey occurred in some products from each manufacturer, but no clear correlation was observed between separation of whey and pH or the number of lactic streptococci present.Bacteria of public health significance, Escherichia coli type I, Staphylococcus aureus and Vibrio spp. were not detected in these products.Fifty-nine samples produced by six manufacturers, purchased from retail outlets, were examined on the sell-by date; the results confirmed the conclusions drawn from studies during storage.