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

peer-reviewedThe 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.This work was funded by the Department of Agriculture, Food and the Marine under the Food Institutional Research Measure. Daniel J. O’Sullivan is in receipt of a Teagasc Walsh Fellowship, Grant Number:2012205

Proteolysis as a function of distance from surface to centre in a smear-ripened Irish farmhouse cheese

This study focused on proteolysis in an Irish farmhouse smear-ripened cheese by serial slicing (0.41 mm/slice) the first 2 cm from surface towards the centre of the cheese. Ureapolyacrylamide gel electrophoretograms confirmed higher proteolysis in the outer layers than at the centre. Free amino acid (FAA) analysis confirmed decrease in proteolytic activity from surface to centre. Peptides produced at depths 0.41 mm and 20.5 mm were 720 and 427 from αs1-casein; 691 and 337 from αs2-casein; 807 and 453 from β-casein; 180 and 109 from κ-casein. The study confirms higher proteolytic activity at surface due to action of enzymes of the smear microbiota, than at the centre of cheese and identified the agents responsible for production of many peptides

Temporal and spatial differences in microbial composition during the manufacture of a Continental-type cheese

peer-reviewedWe sought to determine if the time, within a production day, that a cheese is manufactured has an influence on the microbial community present within that cheese. To facilitate this, 16S rRNA amplicon sequencing was used to elucidate the microbial community dynamics of brine salted Continental-type cheese in cheeses produced early and late in the production day. Differences in microbial composition of the core and rind of the cheese were also investigated. Throughout ripening, it was apparent that late production day cheeses had a more diverse microbial population than their early day equivalents. Spatial variation between the cheese core and rind was also noted in that cheese rinds were found to initially have a more diverse microbial population but thereafter the opposite was the case. Interestingly, the genera Thermus, Pseudoalteromonas and Bifidobacterium, not routinely associated with a Continental-type cheese produced from pasteurised milk were detected. The significance, if any, of the presence of these genera will require further attention. Ultimately, the use of high throughput sequencing has facilitated a novel and detailed analysis of the temporal and spatial distribution of microbes in this complex cheese system and established that the period during a production cycle at which a cheese is manufactured can influence its microbial composition.This work was funded by the Department of Agriculture, Food and the Marine under the Food Institutional Research Measure through the ‘Cheeseboard 2015’ project. Daniel J. O’Sullivan is in receipt of a Teagasc Walsh Fellowship, Grant Number: 201220

High-throughput DNA sequencing to survey bacterial histidine and tyrosine decarboxylases in raw milk cheeses

peer-reviewedBackground The aim of this study was to employ high-throughput DNA sequencing to assess the incidence of bacteria with biogenic amine (BA; histamine and tyramine) producing potential from among 10 different cheeses varieties. To facilitate this, a diagnostic approach using degenerate PCR primer pairs that were previously designed to amplify segments of the histidine (hdc) and tyrosine (tdc) decarboxylase gene clusters were employed. In contrast to previous studies in which the decarboxylase genes of specific isolates were studied, in this instance amplifications were performed using total metagenomic DNA extracts. Results Amplicons were initially cloned to facilitate Sanger sequencing of individual gene fragments to ensure that a variety of hdc and tdc genes were present. Once this was established, high throughput DNA sequencing of these amplicons was performed to provide a more in-depth analysis of the histamine- and tyramine-producing bacteria present in the cheeses. High-throughput sequencing resulted in generation of a total of 1,563,764 sequencing reads and revealed that Lactobacillus curvatus, Enterococcus faecium and E. faecalis were the dominant species with tyramine producing potential, while Lb. buchneri was found to be the dominant species harbouring histaminogenic potential. Commonly used cheese starter bacteria, including Streptococcus thermophilus and Lb. delbreueckii, were also identified as having biogenic amine producing potential in the cheese studied. Molecular analysis of bacterial communities was then further complemented with HPLC quantification of histamine and tyramine in the sampled cheeses. Conclusions In this study, high-throughput DNA sequencing successfully identified populations capable of amine production in a variety of cheeses. This approach also gave an insight into the broader hdc and tdc complement within the various cheeses. This approach can be used to detect amine producing communities not only in food matrices but also in the production environment itself.This work was funded by the Department of Agriculture, Food and the Marine under the Food Institutional Research Measure through the ‘Cheeseboard 2015’ project. Daniel J. O’Sullivan is in receipt of a Teagasc Walsh Fellowship, Grant Number: 2012205

Strategic Alliances & Customer Impact: A Case Study Of Community Hospitals

A strategic alliance (SA) is a mutually beneficial long-term formal relationship formed between two or more parties to pursue a set of agreed upon goals or to meet a critical business need while remaining independent organizations.  It is a synergistic arrangement whereby two or more organizations agree to cooperate in the carrying out of a business activity where each brings different strengths and capabilities to the arrangement.  The social structure of alliances has been considered previously (Gulati 1995, et al.), so instead of discussing the social structure relative to alliance partners, this paper looks at the relationship between the dyad alliance entity and its customer(s).  This newer aspect is particularly important when there are differences in trust and culture to consider (Das & Teng 1998) between alliance partners.  Other considerations include authority, governance and structure, conflict, and the make-up of the strategic alliance, its partners, and the customer(s).&nbsp