Isolation of casein for stable isotope ratio analysis of butter, cheese, and milk powder.

RATIONALE: Stable isotope ratio analysis (SIRA) is commonly used for the authentication of dairy commodities, providing evidence to support the geographical origin and production background of products. We set out to optimise methods for the isolation of a common constituent (casein) from three dairy commodities, which would permit easier inter- and intra-commodity comparisons following SIRA. METHODS: Three published methods for isolation of protein (from cheese, milk, and butter) were adapted to yield protein (casein) fractions from commercial cheddar cheese, whole milk powder (WMP), and butter samples with a high degree of purity for subsequent SIRA. The casein fractions isolated underwent elemental analysis (H, C, and N), protein determination, and some also underwent SIRA of O and S. Two-way analysis of variance and Tukey post hoc comparisons tested differences between methods. RESULTS: For each product, an optimised casein isolation method was chosen based on the C/N ratio and protein content. An optimum solvent lipid extraction (petroleum spirit-diethyl ether (2:1)) and casein precipitation method was chosen for cheddar cheese casein. A final solvent lipid extraction (heptane-isopropanol (3:2)) was necessary for WMP and butter casein extraction. δ13 C and δ2 H values validated the methods' abilities to remove contaminating lipid and isolate pure casein. CONCLUSIONS: Casein of high purity, for subsequent SIRA, can be isolated from cheddar cheese, WMP, and butter following modifications of previously published methods.

A comparison of the revised Delirium Rating Scale (DRS-R98) and the Memorial Delirium Assessment Scale (MDAS) in a palliative care cohort with DSM-IV delirium.

OBJECTIVE: Assessment of delirium is performed with a variety of instruments, making comparisons between studies difficult. A conversion rule between commonly used instruments would aid such comparisons. The present study aimed to compare the revised Delirium Rating Scale (DRS-R98) and Memorial Delirium Assessment Scale (MDAS) in a palliative care population and derive conversion rules between the two scales. METHOD: Both instruments were employed to assess 77 consecutive patients with DSM-IV delirium, and the measures were repeated at three-day intervals. Conversion rules were derived from the data at initial assessment and tested on subsequent data. RESULTS: There was substantial overall agreement between the two scales [concordance correlation coefficient (CCC) = 0.70 (CI 95 = 0.60-0.78)] and between most common items (weighted κ ranging from 0.63 to 0.86). Although the two scales overlap considerably, there were some subtle differences with only modest agreement between the attention (weighted κ = 0.42) and thought process (weighted κ = 0.61) items. The conversion rule from total MDAS score to DRS-R98 severity scores demonstrated an almost perfect level of agreement (r = 0.86, CCC = 0.86; CI 95 = 0.79-0.91), similar to the conversion rule from DRS-R98 to MDAS. SIGNIFICANCE OF RESULTS: Overall, the derived conversion rules demonstrated promising accuracy in this palliative care population, but further testing in other populations is certainly needed.

Verifying origin claims on dairy products using stable isotope ratio analysis and random forest classification.

Scientifically underpinning geographic origin claims will improve consumer trust in food labels. Stable isotope ratio analysis (SIRA) is an analytical technique that supports origin verification of food products based on naturally occurring differences in isotopic compositions. SIRA of five relevant elements (C, H, N, O, S) was conducted on casein isolated from butter (n = 60), cheese (n = 96), and whole milk powder (WMP) (n = 41). Samples were divided into four geographic regions based on their commercial origin: Ireland (n = 79), Europe (n = 67), Australasia (n = 29) and USA (n = 22). A random forest machine learning model built using δ13C, δ2H, δ15N, δ18O and δ34S values of all products (n = 197) accurately (88% model accuracy rate) predicted the region of origin with class accuracy of 95% for Irish, 84% for European, 71% for Australasia, and 94% for US products.

CD4+ T cells are homeostatic regulators during Mtb reinfection.

Immunological priming - either in the context of prior infection or vaccination - elicits protective responses against subsequent Mycobacterium tuberculosis (Mtb) infection. However, the changes that occur in the lung cellular milieu post-primary Mtb infection and their contributions to protection upon reinfection remain poorly understood. Here, using clinical and microbiological endpoints in a non-human primate reinfection model, we demonstrate that prior Mtb infection elicits a long-lasting protective response against subsequent Mtb exposure and that the depletion of CD4+ T cells prior to Mtb rechallenge significantly abrogates this protection. Leveraging microbiologic, PET-CT, flow cytometric, and single-cell RNA-seq data from primary infection, reinfection, and reinfection-CD4+ T cell depleted granulomas, we identify differential cellular and microbial features of control. The data collectively demonstrate that the presence of CD4+ T cells in the setting of reinfection results in a reduced inflammatory lung milieu characterized by reprogrammed CD8+ T cell activity, reduced neutrophilia, and blunted type-1 immune signaling among myeloid cells, mitigating Mtb disease severity. These results open avenues for developing vaccines and therapeutics that not only target CD4+ and CD8+ T cells, but also modulate innate immune cells to limit Mtb disease.

CD8+ lymphocytes are critical for early control of tuberculosis in macaques.

The functional role of CD8+ lymphocytes in tuberculosis remains poorly understood. We depleted innate and/or adaptive CD8+ lymphocytes in macaques and showed that loss of all CD8α+ cells (using anti-CD8α antibody) significantly impaired early control of Mycobacterium tuberculosis (Mtb) infection, leading to increased granulomas, lung inflammation, and bacterial burden. Analysis of barcoded Mtb from infected macaques demonstrated that depletion of all CD8+ lymphocytes allowed increased establishment of Mtb in lungs and dissemination within lungs and to lymph nodes, while depletion of only adaptive CD8+ T cells (with anti-CD8ß antibody) worsened bacterial control in lymph nodes. Flow cytometry and single-cell RNA sequencing revealed polyfunctional cytotoxic CD8+ lymphocytes in control granulomas, while CD8-depleted animals were unexpectedly enriched in CD4 and γδ T cells adopting incomplete cytotoxic signatures. Ligand-receptor analyses identified IL-15 signaling in granulomas as a driver of cytotoxic T cells. These data support that CD8+ lymphocytes are required for early protection against Mtb and suggest polyfunctional cytotoxic responses as a vaccine target.

Nationwide Trends in COVID-19 Cases and SARS-CoV-2 RNA Wastewater Concentrations in the United States.

Wastewater-based epidemiology has emerged as a promising technology for population-level surveillance of COVID-19. In this study, we present results of a large nationwide SARS-CoV-2 wastewater monitoring system in the United States. We profile 55 locations with at least six months of sampling from April 2020 to May 2021. These locations represent more than 12 million individuals across 19 states. Samples were collected approximately weekly by wastewater treatment utilities as part of a regular wastewater surveillance service and analyzed for SARS-CoV-2 RNA concentrations. SARS-CoV-2 RNA concentrations were normalized to pepper mild mottle virus, an indicator of fecal matter in wastewater. We show that wastewater data reflect temporal and geographic trends in clinical COVID-19 cases and investigate the impact of normalization on correlations with case data within and across locations. We also provide key lessons learned from our broad-scale implementation of wastewater-based epidemiology, which can be used to inform wastewater-based epidemiology approaches for future emerging diseases. This work demonstrates that wastewater surveillance is a feasible approach for nationwide population-level monitoring of COVID-19 disease. With an evolving epidemic and effective vaccines against SARS-CoV-2, wastewater-based epidemiology can serve as a passive surveillance approach for detecting changing dynamics or resurgences of the virus.

Developing a guidance resource for managing delirium in patients with COVID-19.

As the COVID-19 pandemic escalates worldwide, it is apparent that many patients with more severe illness will also experience delirium. These patients pose a particular challenge in the application of optimal care due to issues with infectious risk, respiratory compromise and potential interactions between medications that can be used to manage delirium with antiviral and other treatments used for COVID-19. We describe a guidance resource adapted from existing guidelines for delirium management that has been tailored to the specific challenge of managing delirium in patients with COVID-19 infection. Issues around the assessment and treatment of these patients are examined and distilled into a simple (one-paged guidance resource that can assist clinicians in managing suspected delirium.

Higher rates of disengagement among young adults attending a general adult community mental health team: Time to consider a youth-specific service?

AIM: The initial onset of mental illness occurs most frequently in adolescence or early adulthood. In order to inform the development of mental health services tailored for youth, we sought to compare the characteristics of young (18-25 years old) and older (over 25 years old) adults following referral to a general adult community mental health team. METHODS: All individuals referred to a Dublin-based community mental health team and offered an appointment between January 1 and December 31, 2016 were included in the study. Information in relation to engagement patterns, demographic characteristics and clinical characteristics was collected. RESULTS: A total of 298 appointments were offered during the study period among which 94 (31.6%) were for young adults. Significant differences in demographic and clinical characteristics between the two age groups were evident. Young adults were significantly less likely to have been prescribed psychotropic medication at the point of referral (63% vs. 82% respectively, χ2 = 12.30, p < .001). Older adults were four times more likely to demonstrate a good level of early engagement in treatment than young adults (AOR 4.00, 95% CI 1.11-14.37, p = .03). CONCLUSIONS: Young adults had distinct clinical needs and a lower level of engagement in the early stage of treatment compared with their older counterparts in this community team. Further research and stakeholder consultation is needed to more clearly identify the issues in relation to patient engagement. These insights will help to inform the development of youth-specific community mental health services.

Delirium and depression: inter-relationship and clinical overlap in elderly people.

Delirium and depression are complex neuropsychiatric syndromes common in the elderly and are associated with poor health-care outcomes. Accurate diagnosis is essential to the provision of optimum health care for individuals with these conditions but is complicated by substantial clinical overlap in symptoms and comorbidities. A careful assessment of the patient's symptoms, including their context and time course, is needed for accurate diagnosis. Previous depression is common in patients with delirium and depressive illness is a recognised sequelae of delirium. The two syndromes seem to be caused by similar pathophysiological mechanisms, involving disturbances in stress and inflammatory responses, monoaminergic and melatonergic signalling, which point to new avenues for therapeutic intervention. Improved methods to assess delirium and depression in populations at high risk by virtue of their age, diminished cognitive reserve and physical frailty is a key target to achieve improved health-care outcomes in elderly individuals.