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

Structural determination of oxofatty acids by charge-remote fragmentations

A strategy is described to locate the carbonyl position in oxofatty acids by utilizing charge-remote fragmentations of various molecular ions that are desorbed by fast atom bombardment (FAB). Oxofatty acids were cationized with alkali metal ions (Li+, Na+, K+, Rb+, and Cs+) to form [M+2Met − H]+ or alkaline earth metal ions (Mg2+, Ca2+, Sr2+ or Ba2+) to form [M+Met−H]+ in the gas phase. The cationized acids undergo charge-remote fragmentations upon high-energy activation, giving a product-ion pattern that has a gap corresponding to the oxo position and bordered by two high-intensity peaks. One of the peaks corresponds to an ion that is formed by the cleavage of the C-C bond β to the oxo position and proximal to the charge (β ion), whereas the other is formed from the cleavage of the C-C bond γ to the oxo position and distal to the charge (γ′ ion). The oxo position is easily determined by identifying the gap and the β and γ′ ions. Furthermore, there are two competing patterns of fragments in a CAD spectrum of an oxofatty acid or ester [M+Li]+ ion. These arise because Li+ attaches to either the oxo or the carboxylic end, as was confirmed by ab initio molecular orbital calculations. The results demonstrate that control of the fragmentation can be guided by an understanding of metal-ion affinities. Collisional activation of the anionic carboxylates gives results that are similar to those for positive ions, showing that the process is not related to the charge status. Collisional activation of [M+H]+ ions does not give structural information because the charge migrates, leading to charge-mediated fragmentations

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

Gravitational Radiation from First-Order Phase Transitions

It is believed that first-order phase transitions at or around the GUT scale will produce high-frequency gravitational radiation. This radiation is a consequence of the collisions and coalescence of multiple bubbles during the transition. We employ high-resolution lattice simulations to numerically evolve a system of bubbles using only scalar fields, track the anisotropic stress during the process and evolve the metric perturbations associated with gravitational radiation. Although the radiation produced during the bubble collisions has previously been estimated, we find that the coalescence phase enhances this radiation even in the absence of a coupled fluid or turbulence. We comment on how these simulations scale and propose that the same enhancement should be found at the Electroweak scale; this modification should make direct detection of a first-order electroweak phase transition easier.Comment: 7 pages, 7 figure

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

Burden of illness of trigeminal neuralgia among patients managed in a specialist center in England

BACKGROUND: Trigeminal neuralgia (TN) causes severe episodic, unilateral facial pain and is initially treated with antiepileptic medications. For patients not responding or intolerant to medications, surgery is an option. METHODS: In order to expand understanding of the pain-related burden of illness associated with TN, a cross-sectional survey was conducted of patients at a specialist center that utilizes a multidisciplinary care pathway. Participants provided information regarding their pain experience and treatment history, and completed several patient-reported outcome (PRO) measures. RESULTS: Of 129 respondents, 69/128 (54%; 1 missing) reported no pain in the past 4 weeks. However, 84 (65%) respondents were on medications, including 49 (38%) on monotherapy and 35 (27%) on polytherapy. A proportion of patients had discontinued at least one medication in the past, mostly due to lack of efficacy (n = 62, 48%) and side effects (n = 51, 40%). A total of 52 (40%) patients had undergone surgery, of whom 30 had microvascular decompression (MVD). Although surgery, especially MVD, provided satisfactory pain control in many patients, 29% of post-surgical patients reported complications, 19% had pain worsen or stay the same, 48% were still taking pain medications for TN, and 33% reported new and different facial pain. CONCLUSIONS: In most PRO measures, respondents with current pain interference had poorer scores than those without pain interference. In the Patient Global Impression of Change, 79% expressed improvement since beginning of treatment at this clinic. These results indicate that while the multidisciplinary approach can substantially alleviate the impact of TN, there remains an unmet medical need for additional treatment options

Researching the Aftermath of Slavery in Mainland East Africa: Methodological, Ethical, and Practical Challenges

This article examines ethical, practical, and methodological challenges in researching the aftermath of slavery in continental East Africa away from the coastal plantation belt. Interest in post-slavery there is recent and inspired by the apparent contrast with West Africa, where the issue is much more salient. The article explains this silence by highlighting politically-motivated avoidance of the issue in colonial sources and the preference of post-colonial historians for ‘useful’ pasts. Further, it questions the balance of successful integration and continuing marginalization reflected in the apparent obsolescence of slavery. It argues that tracing the trajectories of ex-slaves requires attention to all forms of social inequality and dependency, to the potential status implications for informants of speaking about slavery, and to the variety of terms and fields of meaning relevant to freedom, unfreedom and dependency. Recent research in this vein shows that slave antecedents remain a matter of aibu, shame, and that ex-slaves’ disappearance as a social category took lifelong efforts on their part. While the social valence of slave antecedents is relatively limited in mainland East Africa, slavery remains a problematic and painful heritage that demands great circumspection by researchers

Evidence for an Early High-Energy Afterglow Observed with BATSE from GRB980923

In this Letter, we present the first evidence in the BATSE data for a prompt high-energy (25-300 keV) afterglow component from a gamma-ray burst (GRB), GRB980923. The event consists of rapid variabilty lasting ~40 s followed by a smooth power law emission tail lasting ~400 s. An abrupt change in spectral shape is found when the tail becomes noticeable. Our analysis reveals that the spectral evolution in the tail of the burst mimics that of a cooling synchrotron spectrum, similar to the spectral evolution of the low-energy afterglows for GRBs. This evidence for a separate emission component is consistent with the internal-external shock scenario in the relativistic fireball picture. In particular, it illustrates that the external shocks can be generated during the gamma-ray emission phase, as in the case of GRB990123.Comment: 4 pages, 4 figures, accepted for publication in Astrophysical Journal Letter