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Pink discolouration in cheese
Jonnala, Bhagya Rekha Yeluri
University College Cork
Pink discoloration defect impacts on a wide range of ripened cheese varieties, including Swiss, Cheddar, and Italian-type cheeses. The problem manifests either near the surface of the cheese block as a uniform pink border, or is distributed sporadically within the cheese block. The pink colour develops in the cheese during ripening, usually at least 2 months after manufacture and often leads to product downgrade. Although much international research has focused on trying to elucidate the causes of this defect, recent research within this group has focused on the presence of Thermus thermophilus and its potential to cause the defect, possibly through its ability to produce carotenoids. In this thesis, a wide range of approaches, including pilot-scale cheese manufacture, UHPLC-DAD-QTof mass spectrometry, shotgun metagenomic sequencing, and exploiting the antimicrobial activity of biopreservative nisin were used to determine the role of Thermus in causing pink defect and to investigate potential control measures. The thesis begins with a review of relevant literature relating to the use of high throughput sequencing to investigate the cheese microbiome, with a particular focus on factors of relevance to industry. In particular, the review highlights the influence of biotic and abiotic factors on the cheese microbiome, bacterial functional potential, biodiversity, spoilage and also previously overlooked bacteria present in cheese. The first research chapter shows the varying potential of different Thermus strains to cause pink defect in Swiss-type cheese. Cheese manufactured at pilot scale with Thermus thermophilus HB27 strain results in significantly higher redness levels in cheese in comparison to control cheeses (with no Thermus) while other strains of T. scotoductus SE1, and T. thermophilus DPC6866 (a strain isolated from pink defect cheese was used as positive control) did not significantly influence redness levels in cheese. In the second research chapter, carotenoids present on the surface of smear-ripened cheeses such as Ashbrook, Charloe, Taleggio, and Limburger were characterised using UHPLC-DAD-APCI-MS/MS. The most dominant carotenoid was β-carotene followed by other provitamin A carotenoids and carotenoids of bacterial origin like decaprenoxanthin, sarcinaxanthin and echinenone. Analysis of the core extracts of these cheeses showed β-carotene and very low levels of cryptoxanthin-like carotenoids. In the third research chapter, metagenomic sequencing of and carotenoid extraction from pink defect cheeses sourced from three companies was undertaken to determine microbiome variance among pink defect cheese in comparison to control cheeses (with no defect). Higher counts of environmental microbes were detected in the pink defect cheeses from all three companies, when compared to their controls. A unique carotenoid absent from the control cheese but present in a pink defect cheese of company A was identified, this compound had a UV-Visible spectrum similar to that of zeaxanthin. In the final research chapter, the biopreservative nisin A and 20 bioengineered derivatives thereof were tested, for their bioactivity and minimum inhibitory concentrations, against six Thermus strains. Notably, the pure peptides nisin A M17Q and nisin A M21F were shown to have antimicrobial activity against all of the Thermus strains and may offer a means of controlling Thermus in cheese. Overall this research further adds to the evidence that Thermus is a major factor in forming the pink defect in cheese, provides an optimized protocol for carotenoid extraction from a pink cheese matrix, analyses microbiome variance across a number of pink defect cheeses, as well as identifying nisin derivatives with enhanced antimicrobial activity against six Thermus strains, which possibly can be used as biopreservatives in cheese for the prevention of Thermus growth. Ultimately this study highlights the role of Thermus, a carotenoid producing environmental bacteria, in causing the pink defect in cheese.
Cheese , Pink , Discolouration
Jonnala, B. R. Y. 2019. Pink discolouration in cheese. PhD Thesis, University College Cork.