Microbiological and chemical characterization of Fiore Sardo, a traditional Sardinian cheese made from ewe's milk

The purpose of this study was to assess the chemical and microbial characteristics of 12 batches of artisanal Fiore Sardo, a protected designation of origin (PDO) hard cheese made from raw ewe's milk without addition of starters, during maturation. High standard deviations were observed for moisture percentage, total solids percentage and NaCl percentage content, possibly owing to differences in manufacturing processes and/or milk composition. Total mesophilic bacteria varied between 10 log₁₀ cfu/g in 48-h-old cheese samples and 3 log₁₀ cfu/g in 9-month-old samples. Total coliforms and staphylococci showed the highest counts at 48 h of ripening then decreased significantly, dropping to levels below 2 log₁₀ cfu/g at 3 months of maturation. Lactic acid bacteria and enterococci were the dominant micro-organisms throughout maturation. They were mainly represented by the species Lactococcus lactis ssp. lactis, Enterococcus faecium, Lactobacillus plantarum and Lactobacillus casei group. Low levels of yeasts were detected throughout the maturation period of the cheese. Debaryomyces hansenii and Kluyveromyces lactis var. lactis were the prevalent yeast species isolated.     

Influence of starter and nonstarter on the formation of biogenic amine in goat cheese during ripening

Two commercial starters were investigated for their potential ability to decarboxylate amino acids during goat cheese ripening. Two batches of goat cheese were produced with identical pasteurized milk but different starter cultures. One of them contained Lactococcus lactis subsp. lactis and Lactococcus lactis subsp. cremoris and the other Lactococcus lactis subsp. lactis. The amine contents, microbial counts, proteolysis-related parameters, pH, total solids and salt content were studied in raw materials and cheeses. In raw materials, polyamines were the prevailing amines, whereas the main amines in cheeses were putrescine, tryptamine and, in particular, tyramine (94.59 mg/kg). Aerobic mesophilic microorganisms and Lactococcus counts increased throughout ripening, while Enterobacteriaceae were no longer detectable in cheese after 30 days of ripening. Amine concentration rose during cheese ripening in both batches. Moreover, the decarboxylase activity of microorganisms isolated from samples during cheese ripening was assayed and discussed.     

Microbiological changes throughout ripening of goat cheese made from raw,pasteurized and high-pressure-treated milk

The bacteriological quality during ripening of raw (RA), pasteurized (PA; 72°C, 15 s) and pressure-treated (PR; 500 MPa, 20°C, 15 min) goat milk assessed by enumeration of total bacteria, psychrotrophic bacteria, Enterobacteriaceae, lactobacilli, enterococci, Micrococcaceae and lactococci was evaluated. The high pressure treatment applied was as efficient as pasteurization in reducing the bacterial population of milk. Experimental cheeses were made from RA, PA and PR milks to study the microbial population during ripening. Lactobacilli and lactococci were the predominant microbiota present during ripening in all the cheeses. There were no differences in numbers of starter bacteria during ripening. However, lactobacilli counts for RA milk cheese were significantly higher than for PA and PR cheeses in all the ripening stages studied. Micrococcaceae and enterococci remained at a secondary level, and no differences were observed between cheeses at the end of ripening. On the other hand, the number of Enterobacteriaceae decreased during ripening, but faster in PR milk cheese than in PA and RA milk cheeses. The results of this study suggest that goat cheese made from PR milk had similar microbiological characteristics to PA milk cheeses.     

Modelling the fate of Listeria monocytogenes during manufacture and ripening of smeared cheese made with pasteurised or raw milk

The dynamics of the physicochemical characteristics of foods help to determine the fate of pathogens throughout processing. The aim of this study was to assess the behaviour of Listeria monocytogenes during cheesesmaking and ripening and to model the growth observed under the dynamic conditions of the cheese. A laboratory scale cheese was made in 4 independent replicates from pasteurised or raw cow's milk, artificially contaminated with L. monocytogenes. No growth of L. monocytogenes occurred during raw milk cheese-making, whereas growth did occur in pasteurised milk. During ripening, growth occurred in raw milk cheese, but inactivation occurred in pasteurised milk cheese. The behaviour observed for L. monocytogenes was modelled using a logistic primary model coupled with a secondary cardinal model, taking into account the effect of physicochemical conditions (temperature, pH, water activity and lactate). A novel statistical approach was proposed to assess the optimal growth rate of a microorganism from experiments performed in dynamic conditions. This complex model had an acceptable quality of fit on the experimental data. The estimated optimum growth rates can be used to predict the fate of L. monocytogenes during cheese manufacture in raw or pasteurized milk in different physicochemical conditions. The data obtained contributes to a better understanding of the potential risk that L. monocytogenes presents to cheese producers (growth on the product, if it is contaminated) and consumers (the presence of high numbers) and constitutes a very useful set of data for the completion of chain-based modelling studies.     

Characterisation of the microflora during ripening of a Norwegian semi-hard cheese with adjunct culture of propionic acid bacteria

The microflora of a semi-hard, washed curd, Norwegian cheese with an added adjunct culture of propionic acid bacteria (PAB) was investigated throughout ripening by phenotypic and physiological tests, API test and 16S rRNA sequencing. Cheeses were made at two commercial Norwegian dairies using different milk treatments (pasteurisation versus microfiltration plus pasteurisation) and the same type of starter cultures. Microflora in the cheese varied according to different plant site, milk treatment, and ripening time. PAB dominated the microflora throughout the ripening process. Leuconostoc spp., most probably from the starter, dominated among the isolates from the cheese using microfiltered and pasteurised milk; however, after 40 weeks of ripening non-starter lactic acid bacteria specie Lactobacillus casei/paracasei and Leuconostoc spp. dominated at the dairy using pasteurised milk. Cheese made at the two plants on two subsequent days showed almost identical microflora throughout ripening.     

Microbiological study of Manura, a hard cheese made from raw ovine milk in the Greek island Sifnos

Changes in the microbial flora of Manura, a raw ovine milk cheese, were studied during ripening. In general, the various microbial groups developed better on the cheese surface than in the interior, but red wine treatment had an inhibitory effect on their growth and microbial counts decreased ( P  < 0.05) more rapidly on the cheese surface than in the interior. NaCl and moisture of the cheese affected microbial levels significantly. Thus, Enterobacteriaceae and coliforms were reduced sharply ( P  < 0.05) during ripening on a straw bed (∼3 months) and they were not detected in mature cheese. Lactic acid bacteria predominated over the other microbial groups throughout ripening. Leuconostoc mesenteroides ssp. cremoris , Pediococcus pentosaceus and Lactobacillus paracasei ssp. paracasei , frequently found in maturing cheese, could be used as starters to make this cheese. Moreover, the lactic acid bacteria predominating in mature cheese, such as Weissella paramesenteroides , Lactobacillus bifermentans and Lactobacillus brevis , may contribute to cheese ripening through their biochemical activities.     

Effect of mesophilic lactobacilli and enterococci adjunct cultures on the final characteristics of a microfiltered milk Swiss-type cheese

The effect of four associations of adjunct cultures composed of mesophilic lactobacilli and enterococci, either solely or combined, on the microbiological, biochemical and sensory characteristics of Swiss-type cheese made using microfiltered cows’ milk and supplemented with propionibacteria was studied. The global pattern of growth was similar to that generally observed in raw milk cheese and interactions between microflora were highlighted during ripening. Enterococci, which negatively affected the survival of streptococci starters, seemed to play a limited role in the formation of volatile compounds, probably due to their low levels throughout ripening. On the contrary, mesophilic lactobacilli, which affected the evolution of propionibacteria, enterococci and L. delbrueckii subsp. lactis starter counts, modified free amino acid content, production of volatile compounds and organoleptic properties of mature cheese. This population appeared to be of major importance in the formation of cheese flavor as it was positively related to numerous potential flavor compounds such as alcohols and their corresponding esters, acetaldehyde and 4-methyl-4-heptanone. The original mesophilic lactobacilli present in milk could play an important role in the sensorial diversity of raw milk Swiss-type cheeses such as Comte.     

Effect of activating lactoperoxidase system in cheese milk on the quality of Saint‐Paulin cheese

Saint‐Paulin cheese was made from cow’s milk refrigerated at 4 °C for 72 h and preserved by the lactoperoxidase (LP) system. The effect of the LP system on the microbiological, physicochemical and biochemical properties of cheese over a ripening period of 23 days was investigated, using a control (C0), refrigerated LP‐inactivated cow’s milk (C1) and refrigerated LP‐activated cow’s milk (LPA). The LPA treatment showed the least contamination in flora count, particularly salt‐tolerant bacteria at the end of the ripening period. LPA cheese had significantly lower coliform, yeasts and mould counts (P < 0.05) than the other cheeses; this demonstrated the bacteriostatic effect of the LP system. The proteolysis results showed the least value for LPA cheese as compared with the two other samples, as determined by using sodium dodecyl sulphate‐polyacrylamide gel electrophoresis of casein fractions extracted from the three samples. The findings indicated that the preservation of cow’s cheese milk by the LP system can be used to improve the microbiological quality, inhibit psychotropic germs, correct the losses of soluble nitrogen fractions in the whey and conserve the cheese yield affected by refrigeration.     

Texture and flavour development in Ras cheese made from raw and pasteurised milk

Texture, proteolysis and flavour development in Ras cheeses made from raw or pasteurised milk with two different thermophilic lactic cultures were monitored during ripening. Results showed that at day 1 of manufacture, the moisture content and pH were lower in raw milk cheese than in pasteurised milk cheeses. Levels of water-soluble nitrogen, casein breakdown, free amino groups and free fatty acids were higher in cheese made from raw milk than in that made from pasteurised milk. Textural characteristics, such as hardness, cohesiveness and chewines, increased in all treatments during the first 60 days of ripening due to the reduction in the moisture level during the second stage of salting (dry salting during the first 60 days of ripening). Cheese made from raw milk received the highest texture and flavour scores by panellists.     

Interrelationships among microbiological, physicochemical, and biochemical properties of Terrincho cheese, with emphasis on biogenic amines

Changes in the microbiological, physicochemical, and biochemical characteristics of Terrincho cheese as represented by native microflora, pH, water activity, soluble nitrogen fractions, free amino acids, and biogenic amines (e.g., ethylamine, dimethylamine, tryptamine, phenylethylamine, putrescine, cadaverine, histamine, tyramine, cystamine, and spermine) during ripening were monitored. Terrincho is a traditional Portuguese cheese manufactured from raw ewe's milk. The main groups of microorganisms (lactococci, lactobacilli, enterococci, pseudomonads, staphylococci, coliforms, yeasts, and molds) were determined following conventional microbiological procedures. Free amino acids and biogenic amines were determined by reverse-phase high-performance liquid chromatography, following extraction from the cheese matrix and derivatization with dabsyl chloride. The total content of free amino acids ranged from 1,730 mg/kg of dry matter at the beginning of the ripening stage to 5,180 mg/kg of dry matter by day 60 of ripening; such an increase was highly correlated with the increase of water-soluble nitrogen in total nitrogen, 12% trichloroacetic acid-soluble nitrogen in total nitrogen, and 5% phosphotungstic acid-soluble nitrogen in total nitrogen throughout ripening. Histamine was consistently present at very low levels, whereas putrescine, cadaverine, and tryptamine were the dominant biogenic amines and increased in concentration during ripening. Ethylamine, tryptamine, phenylethylamine, and cystamine reached maxima by 30 days of ripening and decreased thereafter. Significant correlations between amino acid precursors and corresponding biogenic amines, as well as between biogenic amines and microbial viable numbers, were observed.     

Effect of addition of native cultures on characteristics of Armada cheese manufactured with pasteurized milk: A preliminary study

Six batches of Armada cheese were produced, one from raw milk with no added starter, another from pasteurized milk with a commercial mesophilic starter and four from pasteurized milk with experimental starters. These starters included: Lactococcus lactis subsp. lactis and L. lactis subsp. lactis biovar. diacetylactis; or the same strains plus either Enterococcus raffinosus, Leuconostoc mesenteroides subsp. dextranicum or Lactobacillus plantarum, all isolated from Armada cheese made from raw milk. The highest counts of aerobic mesophilic bacteria and populations of lactic acid bacteria for all batches were reached in the first week of ripening. These counts declined later throughout the ripening, although not at identical rates in every batch. Counts of lactobacilli were significantly higher in cheeses made from raw milk and those inoculated with the lactobacilli strain than in the other batches, over the whole ripening period. Added native starters minimized growth of Enterobacteriaceae. Cheeses made with the starter containing the Enterococcus strain had the most favourable sensory attributes throughout ripening.     

The effects of carbon dioxide addition to cheese milk on the microbiological properties of Turkish White brined cheese

This study invest0igated the effect of CO₂ added to achieve three pH levels: pH 6.1, pH 6.2 and pH 6.3 for treatments X, Y, Z, respectively, on some microbiological properties of Turkish White (TW) brined cheese. For each pH, four batches of cheese were produced from: raw milk with no added carbon dioxide (UR), raw milk with carbon dioxide (TR), pasteurised milk with no carbon dioxide addition (UP) and pasteurised milk with carbon dioxide addition (TP). The microbiological analysis of TW brined cheeses was carried out for 90 days of maturation period. Total aerobic mesophilic bacteria, mesophilic lactic acid bacteria, yeasts and moulds and coliform group were determined in control and CO₂ treatment groups. Mesophilic bacteria count was determined as 5.14, 5.29, 5.67 log cfu/g for pH 6.1, 6.2 and 6.3, respectively, in CO₂‐treated raw milk cheeses. Yeasts and moulds reduction increased significantly by applying CO₂ (P < 0.01). For TW cheese samples, the most significant microbial inactivation was detected at sample groups of pH 6.1.     

Occurrence of foodborne pathogens in Irish farmhouse cheese

Food safety is a critical factor in the production of farmhouse cheese. In Ireland the varieties of farmhouse cheese produced reflect a much broader range than those produced commercially and some of these cheese varieties are associated with greater microbiological risk. These include cheese produced from unpasteurised milk and soft ripened cheese such as mould or smear-ripened cheeses which have high pH and relatively short ripening times. The aim of this study was to determine the microbiological quality of farmhouse cheeses in Ireland. Three hundred and fifty one cheese samples, from 15 cheese producers, were analysed for microbiological quality on a monthly basis throughout the year. The analyses included enumeration of Escherichia coli, Staphylococcus aureus and Listeria monocytogenes (using the relevant agars) and enrichment for L. monocytogenes. The cheeses selected were produced from ovine, caprine and bovine milk. Both unpasteurised and pasteurised milk cheeses were sampled and these included hard, semi-hard and soft cheeses, internal/external mould-ripened and smear-ripened cheeses and the cheeses represented different geographic regions. Of the cheeses tested, 94% were free of L. monocytogenes, all were within the EU limits for E. coli and only one cheese variety had S. aureus levels above the recommended numbers for the first 6 months of the year. Due to a modified production process the numbers were within the guidelines for the second six months. The results indicate that Irish farmhouse cheeses are of a high microbiological quality.     

Changes in the microbiological and chemical characteristics of an artisanal, low-fat cheese made from raw ovine milk during ripening

Changes in the microbial flora of batzos cheese made from raw ovine milk were studied during ripening. Lactic acid bacteria and Enterobacteriaceae were the predominant groups of micro-organisms. Cheeses manufactured in summer had higher microbial counts than those made in spring, with the exception of staphylococci. Nevertheless, Enterobacteriaceae and coliforms decreased more rapidly in cheese made in summer and counts at the end of storage were lower than those in spring cheese.
Enterococci predominated in the ripened curd of cheese made in spring, whereas lactobacilli were the most abundant lactic acid bacteria in cheese made in summer. Enterococcus faecium was the predominant species in spring, and Lactobacillus paracasei ssp. paracasei predominated in cheese made in summer. The pH of the cheeses was > 5.0 throughout ripening, and NaCl-in-moisture content (> 8.0%) permitted the growth and survival of salt-tolerant micro-organisms. α_s1-Casein degraded at a faster rate than β-casein; both caseins were hydrolysed more rapidly in spring than in summer. The free amino acid content became higher in summer cheese (566.24–3460.25 µg/g of glycine equivalent) than in spring cheese because of the progress of ripening. Moreover, the milk fat of the cheese was degraded more in the summer than in the spring. The results suggest that there could be advantages to using starter cultures and improving the level of hygiene during milk and cheese production in order to eliminate undesirable micro-organisms and standardize cheese quality.     

The effect of refrigerated storage of raw milk on the physicochemical and microbiological quality of Tunisian semihard Gouda‐type cheese during ripening

A Tunisian semihard Gouda‐type cheese made from milk kept at 4 °C for 24, 48, 72 and 96 h was monitored during 45 days of ripening. The effect of milk refrigeration on the evolution of physicochemical parameters in relation to the quantitative variation of the microbial population during ripening of Gouda‐type cheese was investigated. Microbiological and physicochemical analyses were performed on raw milk and cheese samples after curding, 2, 9, 16, 23, 30, 37 and 45 days of ripening time. The raw milk kept under refrigeration at 4 °C for 96 h showed the highest microbial count and proteolysis level. The duration of storage significantly reduced the cheese yield as a result of important solubilisation casein in proteoses‐peptones. Results of different nitrogenous fractions by Kjeldahl method showed enzymatic hydrolysis products of casein whose intensity depended on the maturing stage as well as the refrigeration time. Besides the evident action of the plasmin, original milk protease, on the hydrolysis of casein in soluble fractions, the proteolysis of cheese caseins is also initiated by proteolytic action of the chymosin and extracellular heat‐resistant proteases notably produced by the same psychrotrophic microflora. Lactic acid bacteria starters that constitute the dominant microflora of this type of cheese are also considered as aroma precursors.     

MICROBIOLOGICAL STUDY OF ARZÚA CHEESE (NW SPAIN) THROUGHOUT CHEESEMAKING AND RIPENING

Changes during production and ripening in the microbial flora of 11 batches of Arzúa, a soft cheese made from raw cow's milk, were investigated. The following microbial groups were counted on the surface and interior of the cheese: total viable count (TVC), lactic acid bacteria (LAB), halotolerant flora, enterococci, proteolytic enterococci, staphylococci, Staphylococcus aureus, Enterobacteriaceae, faecal coliforms, molds, yeasts, Listeria spp. and (in milk) Brucella spp. pH and water activity were also determined. TVC and LAB were, generally, more than 9 log (cfu/g). Enterococci counts increased gradually, reaching values in excess of 6 log units. Halotolerant flora and staphylococci remained practically constant throughout ripening, at 6–8 and 5–7 log units, respectively. Maximum Enterobacteriaceae and faecal coliform counts exceeded 7 and 6 log units, respectively. Brucella spp. were not detected in any of the milk samples. Listeria spp. were detected in four batches, and Listeria monocytogenes in two.     

The effect of wild lactic acid bacteria on the production of goat’s milk soft cheese

A blend of four wild strains of lactic acid bacteria, Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus, Lactobacillus paracasei subsp. paracasei and Enterococcus faecalis, was used as starter to produce a goat’s milk soft cheese. The cheese was analysed microbiologically, physicochemically and organoleptically through a 30‐day period of ripening. Counts of the starter cultures increased in the first 24 h and then remained stable during ripening. Yeasts and coliforms were found at low numbers initially and gradually decreased to undetectable levels. Micrococcal counts were high throughout ripening. The end product was a soft cheese characterised by a mild, aromatic taste as well as a smooth structure.     

Adjunct starter properties affect characteristic features of Swiss-type cheeses

A large number of microorganisms, both starter microorganisms and non-starter lactic acid bacteria originating from the base milk, or from various contamination sources during cheese manufacture, is associated with cheese ripening and the formation of flavour, texture and aroma. Under controlled conditions, Emmental and Bergk?se, a Gruyère-type cheese variety, were produced from pasteurised milk with standard starters and defined strains of facultatively heterofermentative lactobacilli (FHL), and partly with addition of a defined mixture of enterococci. Lactobacillus casei subsp. casei and L. rhamnosus (two strains each) were selected with respect to their potential for the utilisation of citric acid and ribose as sole energy source. The FHL developed up to 10(8) cfu/g within the first weeks of ripening, and viable counts in mature cheeses were 10(7) cfu/g, independent of the cheese variety. Bergk?se made with addition of L. rhamnosus strains showed a more pronounced proteolysis, resulting in reduced firmness and elasticity values of the cheese body, and FHL strains able to utilise citric acid improved the appearance of the cheeses by increasing the number of small eyes to the desired level. In Emmental cheese, the citric acid (+) strains reduced the intensity of propionic acid formation as the FHL apparently competed with the propionibacteria, and enterococci disappeared completely during maturation. Although further work is needed the study shows that, depending on the cheese variety, particular properties of FHL adjunct starters significantly affect important quality attributes of the resulting cheeses.     

Determination of bovine lactoferrin concentrations in cheese with specific monoclonal antibodies

In order to determine bovine lactoferrin concentration in cheese, bovine lactoferrin-specific monoclonal antibodies have been raised and an ELISA has been developed to determine lactoferrin concentrations in milk, whey and experimental soft, semi-hard and Swiss-type cheeses made with raw or pasteurised milk. The lactoferrin concentration in cheese was shown to depend on the cheese-making process, with higher values in Swiss-type and semi-hard cheeses than in soft cheeses. Furthermore, Western-blotting analysis of lactoferrin in cheese showed that this protein stayed intact throughout ripening in raw milk cheese, whereas it was partially hydrolysed in cheeses made with pasteurised milk. Based on these observations, we propose that cheese may constitute a natural dairy source of lactoferrin beneficial to health.     

Free fatty acids and oxidative changes of a raw goat milk cheese through maturation

Free fatty acids (FFA) and lipid and protein oxidation changes were studied throughout maturation process of a raw goat milk cheese with protected designation of origin. Cheeses were analyzed at 4 different times of maturation, at 1, 30, 60, and 90 d. All FFA significantly increased during maturation and the relative increase was higher for long-chain than medium- or short-chain FFA. At the end of maturation, oleic (C18:1 n9), butyric (C4:0), and palmitic (C16:0) acids were the most abundant. The higher levels of short-chain fatty acids (SCFA) regarding total FFA obtained at the end of Ibores cheese ripening compared with other raw goat milk cheeses, highlight the notable role of SCFA on the flavor of this cheese owing to their low-odor thresholds. Lipid oxidation values significantly increased during maturation process but low levels of malondialdehyde were reported; however, protein oxidation did not significantly change during ripening.