The microbial profile of cheese is a primary determinant of cheese quality. Microorganisms can contribute to aroma and taste defects, form biogenic amines, cause gas and secondary fermentation defects, and can contribute to cheese pinking and mineral deposition issues. These defects may be as a result of seasonality and the variability in the composition of the milk supplied, variations in cheese processing parameters, as well as the nature and number of the non-starter microorganisms which come from the milk or other environmental sources. Such defects can be responsible for production and product recall costs and thus represent a significant economic burden for the dairy industry worldwide. Traditional non-molecular approaches are often considered biased and have inherently slow turnaround times. Molecular techniques can provide early and rapid detection of defects that result from the presence of specific spoilage microbes and, ultimately, assist in enhancing cheese quality and reducing costs. Here we review the DNA-based methods that are available to detect/quantify spoilage bacteria, and relevant metabolic pathways in cheeses and, in the process, highlight how these strategies can be employed to improve cheese quality and reduce the associated economic burden on cheese processors.

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Nucleic acid-based approaches to investigate microbial-related cheese quality defects

Nomenclature of the Proteins of Cows’ Milk—Sixth Revision

The principal pathways for the formation of flavour compounds in cheese (glycolysis, lipolysis and proteolysis) are reviewed. Depending on variety, microflora and ripening conditions, lac- tate may be metabolized by a number of pathways to various compounds which contribute to cheese flavour or off-flavours. Citrate metabolism by citrate-positive lactococci or Leuconostoc spp. is important in certain varieties (e.g., Dutch cheeses). Lipolysis results directly in the formation of flavour compounds by liberating free fatty acids (FFA). FFA may also be metabolized to alkan-2-ones and fatty acid lactones. Proteolysis of the caseins to a range of small- and intermediate-sized peptides and free amino acids (FAA) probably only contributes to the background flavour of most cheese varieties, but FAA are important precursors for a range of poorly-understood catabolic reactions which produce volatile compounds essential for flavour.

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Biochemical pathways for the production of flavour compounds in cheeses during ripening: A review

Lactic acid bacteria (LAB) constitute a group of gram-positive bacteria united by a constellation of morphological, metabolic, and physiological characteristics The general description of the bacteria included in the group is gram-positive, nonsporing, nonrespiring cocci or rods, which produce lactic acid as the major end product during the fermentation of carbohydrates The LAB term is intimately associated with bacteria involved in food and feed fermentation, including related bacteria normally associated with the (healthy) mucosal surfaces of humans and animals The boundaries of the group have been subject to some controversy, but historically the genera Lactobacillus, Leuconostoc, Pediococcus, and Streptococcus form the core of the group Taxonomic revisions of these genera and the description of new genera mean that LAB could, in their broad physiological definition, comprise around 20 genera However, from a practical, food-technology point of view, the following genera are considered the principal LAB: Aerococcus, Carnobacterium, Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Oenococcus, Pediococcus, Streptococcus, Tetragenococcus, Vagococcus, and Weissella The genus Bifidobacterium, often considered in the same context as the genuine lactic acid bacteria and sharing some of their typical features, is phylogenetically unrelated and has a unique mode of sugar fermentation The classification of lactic acid bacteria into different genera is largely based on morphology, mode of glucose fermentation, growth at different temperatures, configuration of the lactic acid produced, ability to grow at high salt concentrations, and acid or alkaline tolerance Chemotaxonomic markers such as fatty acid composition and constituents of the cell wall are also used in classification In addition, the present taxonomy relies partly on true phylogenetic relationships,

Lactic acid bacteria: classification and physiology.

Rennet-coagulated cheeses are ripened for periods ranging from about two weeks to two or more years depending on variety. During ripening, microbiological and biochemical changes occur that result in the development of the flavour and texture characteristic of the variety. Biochemical changes in cheese during ripening may be grouped into primary (lipolysis, proteolysis and metabolism of residual lactose and of lactate and citrate) or secondary (metabolism of fatty acids and of amino acids) events. Residual lactose is metabolized rapidly to lactate during the early stages of ripening. Lactate is an important precursor for a series of reactions including racemization, oxidation or microbial metabolism. Citrate metabolism is of great importance in certain varieties. Lipolysis in cheese is catalysed by lipases from various source, particularly the milk and cheese microflora, and, in varieties where this coagulant is used, by enzymes from rennet paste. Proteolysis is the most complex biochemical event that occurs during ripening and is catalysed by enzymes from residual coagulant, the milk (particularly plasmin) and proteinases and peptidases from lactic acid bacteria and, in certain varieties, other microorganisms that are encouraged to grow in or on the cheese. Secondary reactions lead to the production of volatile flavour compounds and pathways for the production of flavour compounds from fatty acids and amino acids are also reviewed.

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Biochemistry of Cheese Ripening

A number of Cheddar cheese samples of different age, pH and moisture content have been examined rheologically and electrophoretically to determine whether the progressive changes in cheese texture were related to casein proteolysis. The force-compression curves obtained by crushing cubes of cheese between small flat plates at constant speed were different for the different cheese samples and were affected by the moisture content, pH and extent of αsl-casein proteolysis that had taken place in the cheese. These results support a model of cheese micro structure in which an extensive network involving αsl- casein molecules traverses the cheese and as the cheese ripens, chymosin cleavage of αsl-casein weakens the protein network. Such a model explains the rapid decrease in Cheddar cheese yield-force that occurs during the early stages of ripening.

Rheological Evaluation of Maturing Cheddar Cheese

In dilute buffers ⋟ pH 5·8, chymosin hydrolysed bovine αs1-casein to αs1-I, αs1-II and αs1-III/αs1-IV in a sequential manner while at pH 4·6 αs1-casein was hydrolysed to αs1-I which was then hydrolysed to αs1-V. In the presence of 5 % (w/v) NaCl at pH 5·2, αs1-casein was hydrolysed to αs1-I which was then hydrolysed to αs1-VII and αs1-VIII. αs1-I, αs1-II and αs1-III/αs1-IV were isolated by chromatography on cellulose phosphate followed by preparative slab-gel electrophoresis; αs1-V was isolated by repeated preparative slab-gel electrophoresis and αs1-VII by gel filtration on Sephadex G-150 followed by preparative slab-gel electrophoresis. The mol. wts of the peptides, estimated by gel filtration on Sephadex G-100, were 21000, 17600, 15600, 19900 and 14600 for αs1-I, αs1-II, αs1-III/αs1-IV and αs1-V and αs1-VII respectively. Characterization of the peptides by amino acid, phosphorus and terminal residue analysis showed that they probably consisted of segments of the αs1-casein chain as follows: αs1-I: residues 24/25–199; αs1-II: residues 24/25–169; αs1-III/αs1-IV: residues 24/25–149–150; αs1-V: residues 29/33–199; αs1-VII: residues 56–179. Peptide bonds close to phosphate residues on the αs1-casein chain were not hydrolysed by chymosin at high pH values (⋟ 5·8) when the phosphate groups were charged, but became available for hydrolysis when the reaction pH was reduced. Proteolytic specificity was also modified by NaCl.

Proteolytic specificity of chymosin on bovine αs1-casein

Chesse flavor is a manifestation of complex interactions of volatile and non-volatile flavor-active compounds plus tactual perception. Numerous agents, including lactic acid bacteria, procece the flavou sensations. The effect of lactic acid bacteria is more dominant in cheese varieties with limited growth of secondary flora. This review describes the indirect and direct impacts of lactic acid bacteria in cheese with emphasis on carbohydrate fermentation, changes in oxidation-reduction potential, interactions with non-starter bacteria, autolysis, proteolytic and peptidolytic activities, transport of metabolites and flavor production.

Norman F. Olson

Papers

Rheological Evaluation of Maturing Cheddar Cheese

Proteolytic specificity of chymosin on bovine αs1-casein

The impact of lactic acid bacteria on cheese flavor

A Melting Test for Pasteurized Process Cheese Spreads

Effects of salt concentration and freezing on mozzarella cheese texture