Proof of concept that melanoma nuclear count compares favourably with the benchmark histological prognostic feature, Breslow thickness.

AIMS: Breslow thickness (BT) is the most important histological prognostic feature for melanoma prognosis, but it only captures tumour size in one dimension. Adding a further measurement in a different axis has been shown to improve prognostic value. It seems reasonable that further prognostic value could be obtained by estimating the number of invasive melanoma cells using nuclear count. The aim of this study was to show proof of concept that nuclear count has prognostic value independent of BT. METHODS AND RESULTS: Melanoma cell nuclei were labelled with SRY-related HMG-box 10 (SOX10) protein, the sections scanned and StarDist machine-learning algorithm used to count nuclei in 102 cases of primary cutaneous melanoma. Prognostic value was assessed using survival analyses. Nuclear count correlated strongly with T category, BT and calculated tumour area (each P < 0.001), suggesting that it was a valid marker of melanoma burden. Nuclear count was a predictor for overall survival in univariable analysis [hazard ratio (HR) = 2.25, confidence interval (CI) = 1.66-3.06, P < 0.001] and multivariable analysis (HR = 2.60, CI = 1.59-4.24, P < 0.001). BT and ulceration were significant in univariable analyses, but not in multivariable models with nuclear count. Models containing nuclear count showed the best fit. Similar results were seen for melanoma-specific and metastasis-free survival. Nuclear count was able to stratify melanomas within a given T stage. CONCLUSIONS: This study demonstrated proof of concept that counting melanoma nuclei may be an improved measure of invasive tumour burden compared to BT. Future studies will need to refine methods of nuclear detection and also to confirm its prognostic value.

The High Mutational Sensitivity of ccdA Antitoxin Is Linked to Codon Optimality.

Deep mutational scanning studies suggest that synonymous mutations are typically silent and that most exposed, nonactive-site residues are tolerant to mutations. Here, we show that the ccdA antitoxin component of the Escherichia coli ccdAB toxin-antitoxin system is unusually sensitive to mutations when studied in the operonic context. A large fraction (∼80%) of single-codon mutations, including many synonymous mutations in the ccdA gene shows inactive phenotype, but they retain native-like binding affinity towards cognate toxin, CcdB. Therefore, the observed phenotypic effects are largely not due to alterations in protein structure/stability, consistent with a large region of CcdA being intrinsically disordered. E. coli codon preference and strength of ribosome-binding associated with translation of downstream ccdB gene are found to be major contributors of the observed ccdA mutant phenotypes. In select cases, proteomics studies reveal altered ratios of CcdA:CcdB protein levels in vivo, suggesting that the ccdA mutations likely alter relative translation efficiencies of the two genes in the operon. We extend these results by studying single-site synonymous mutations that lead to loss of function phenotypes in the relBE operon upon introduction of rarer codons. Thus, in their operonic context, genes are likely to be more sensitive to both synonymous and nonsynonymous point mutations than inferred previously.

Effect of Rose Syrup and Marigold Powder on the Physicochemical, Phytochemical, Sensorial and Storage Properties of Nutricereals and Milk-Based Functional Beverage.

ABSTACTAim: The present study aimed to investigate the effect of rose sirup and marigold powder on the physicochemical properties, bioactive potential, sensory acceptability and storage life of the nutricereals (finger millet, oats) and milk-based functional beverage (FB).Method: Preliminary trials were performed using different levels of rose sirup (8-14%) and marigold powder (0.40-0.55%) in the pre-standardized FB. The most acceptable concentration was selected on the basis of sensory analysis. Selected beverages were then subjected to the physicochemical analysis, assessment of bioactive compounds and FTIR characterization. The effect of flower extracts on the mineral content and storage life (4 ± 1 °C) of beverages was also studied. The significant difference in treatments was determined using Duncan's multiple range test, SPSS 25.0.Results: The best acceptable concentrations for rose sirup and marigold powder were 10% and 0.50%, respectively. A significant (p ≤ 0.05) decrease in the dietary fiber (6.50%) and ß-glucan (3.95%) content was observed on the addition of rose sirup. Significant (p ≤ 0.05) increase in the total phenols (119.18-145.23%), ß-carotene (0.37%), anthocyanins (78.82-230.58%) and antioxidant activity (4.98-7.17%) was observed on the addition of flower extracts. Strong peaks were observed in the regions of 3600-3200, 3000-2800 and 1700-1600 cm - 1 on FTIR characterization. A significant decrease in the mineral content of FB was also found on the addition of rose sirup. Rose flavored beverage had the highest overall acceptability (7.83 ± 0.23) and storage stability (50 days at refrigerated storage) among the prepared beverages.Conclusion: The addition of flower extracts significantly improved the acceptability of the prepared beverages. It not only improved the phytochemical profile but also had a substantial impact on storage stability.

Contribution of the Chromosomal ccdAB Operon to Bacterial Drug Tolerance.

One of the first identified and best-studied toxin-antitoxin (TA) systems in Escherichia coli is the F-plasmid-based CcdAB system. This system is involved in plasmid maintenance through postsegregational killing. More recently, ccdAB homologs have been found on the chromosome, including in pathogenic strains of E. coli and other bacteria. However, the functional role of chromosomal ccdAB genes, if any, has remained unclear. We show that both the native ccd operon of the E. coli O157 strain (ccdO157) and the ccd operon from the F plasmid (ccdF), when inserted on the E. coli chromosome, lead to protection from cell death under multiple antibiotic stress conditions through formation of persisters, with the O157 operon showing higher protection. While the plasmid-encoded CcdB toxin is a potent gyrase inhibitor and leads to bacterial cell death even under fully repressed conditions, the chromosomally encoded toxin leads to growth inhibition, except at high expression levels, where some cell death is seen. This was further confirmed by transiently activating the chromosomal ccd operon through overexpression of an active-site inactive mutant of F-plasmid-encoded CcdB. Both the ccdF and ccdO157 operons may share common mechanisms for activation under stress conditions, eventually leading to multidrug-tolerant persister cells. This study clearly demonstrates an important role for chromosomal ccd systems in bacterial persistence.IMPORTANCE A large number of free-living and pathogenic bacteria are known to harbor multiple toxin-antitoxin systems, on plasmids as well as on chromosomes. The F-plasmid CcdAB system has been extensively studied and is known to be involved in plasmid maintenance. However, little is known about the function of its chromosomal counterpart, found in several pathogenic E. coli strains. We show that the native chromosomal ccd operon of the E. coli O157 strain is involved in drug tolerance and confers protection from cell death under multiple antibiotic stress conditions. This has implications for generation of potential therapeutics that target these TA systems and has clinical significance because the presence of persisters in an antibiotic-treated population can lead to resuscitation of chronic infection and may contribute to failure of antibiotic treatment.

Molecular Determinants of Mutant Phenotypes, Inferred from Saturation Mutagenesis Data.

Understanding how mutations affect protein activity and organismal fitness is a major challenge. We used saturation mutagenesis combined with deep sequencing to determine mutational sensitivity scores for 1,664 single-site mutants of the 101 residue Escherichia coli cytotoxin, CcdB at seven different expression levels. Active-site residues could be distinguished from buried ones, based on their differential tolerance to aliphatic and charged amino acid substitutions. At nonactive-site positions, the average mutational tolerance correlated better with depth from the protein surface than with accessibility. Remarkably, similar results were observed for two other small proteins, PDZ domain (PSD95pdz3) and IgG-binding domain of protein G (GB1). Mutational sensitivity data obtained with CcdB were used to derive a procedure for predicting functional effects of mutations. Results compared favorably with those of two widely used computational predictors. In vitro characterization of 80 single, nonactive-site mutants of CcdB showed that activity in vivo correlates moderately with thermal stability and solubility. The inability to refold reversibly, as well as a decreased folding rate in vitro, is associated with decreased activity in vivo. Upon probing the effect of modulating expression of various proteases and chaperones on mutant phenotypes, most deleterious mutants showed an increased in vivo activity and solubility only upon over-expression of either Trigger factor or SecB ATP-independent chaperones. Collectively, these data suggest that folding kinetics rather than protein stability is the primary determinant of activity in vivo This study enhances our understanding of how mutations affect phenotype, as well as the ability to predict fitness effects of point mutations.

Challenges and Opportunities for Equity in US School Meal Programs: A Scoping Review of Qualitative Literature Regarding the COVID-19 Emergency.

The emergency school meals program provided free meals to children in the United States (US) during COVID-19-related school closures. This scoping review aims to synthesize the existing qualitative empirical evidence published between March 2020 and January 2023 on the operations and utilization of emergency school meal programs during the pandemic. Qualitative, US-based peer-reviewed literature was collected from three sources: (1) parent review of all federal nutrition assistance programs; (2) systematic search of four databases; and (3) manual search of grey literature. Identified scientific articles and grey literature reports (n = 183) were uploaded into Covidence and screened for duplicates and inclusion/exclusion criteria. Our final sample included 21 articles/reports, including 14 mixed methods and seven qualitative-only projects. Articles spanned all U.S. states. Articles had similar research questions to understand school meals and/or general food access during COVID-19, with an emphasis on long-term policy implications. Hybrid deductive/inductive analytic coding was used to analyze data, utilizing domains from the Getting to Equity Framework (GTE). GTE considers multi-level factors that influence nutrition behavior while centering more equitable pathways to improve nutrition security and reduce adverse health. Findings were sorted into two categories: operational challenges during the pandemic and solutions to address inequities in school meal distribution during and after the pandemic, particularly during school closures such as summers or future emergencies. Key challenges related to supply chain issues, safety, and balancing families' needs with limited staff capacity. Programs addressed equity by (a) reducing deterrents through federally issued waivers and increased communications which allowed the serving of meals by programs to families who previously did not have access, (b) building community capacity through collaborations and partnerships which allowed for increased distribution, and (c) preparing and distributing healthy options unless barriers in supply chain superseded the effort. This review highlights the importance of emergency school meal programs and provides insights into addressing challenges and promoting equity in future out-of-school times. These insights could be applied to policy and practice change to optimize program budgets, increase reach equitably, and improve access to nutritious meals among populations at highest risk for nutrition insecurity.

Study of Association of Serum Prolactin Levels with Insulin Resistance in Type 2 Diabetes Mellitus Patients.

Objective: Prolactin (PRL) regulates glucose metabolism and insulin sensitivity. The study aimed to evaluate the role of PRL in glucose homeostasis and its association with insulin resistance in patients with diabetes mellitus (DM). Methods and Materials: This cross-sectional, observational study included 100 patients (25-60 years) with T2DM. Primary information including demographics, anthropometric measurements, and biochemical measures (complete blood count, glucose parameters, liver and kidney function test, lipid profile, thyroid function test, serum fasting insulin levels, serum PRL levels) was collected. Results: A total of 100 patients, 50 men and 50 women (25 premenopausal and 25 postmenopausal), were enrolled in this study. The correlation between serum cholesterol and PRL was found to be statistically non-significant (P = 0.129) in men and significant (P = 0.041) in women. There was an inverse relationship between fasting plasma glucose and serum PRL levels in both men (r = -0.88; P < 0.0001) and women patients (r = -0.768; P < 0.0001). Negative correlation between postprandial plasma glucose and PRL was found to be statistically significant (r = -0.398; P = 0.048) in postmenopausal women. The comparison in both men and women indicated an inverse correlation between serum PRL and glycated haemoglobin levels. There was a significant negative correlation between homeostasis model assessment-estimated insulin resistance (HOMA-IR) and PRL levels in both men (r = -0.362; P = 0.039) and women patients (r = -0.362; P = 0.003). Homeostasis model assessment of ß cell function (HOMA-ß), which directly correlates with residual pancreatic beta cell function, was positively correlated with prolactin levels, irrespective of gender and menopausal status of female subjects. Conclusion: Serum PRL levels correlate with improved glycaemic control.

Effect of household processing on nutritional and antinutritional composition, mineral-mineral ratios, and functional properties of Colocasia leaves.

Colocasia leaves are high in nutrients and other phytochemicals but their utilization remains limited due to a lack of awareness. Higher content of anti-nutritional factors like oxalic and tannic acid in Colocasia leaves limit nutrient availability. In the present study, the effect of four household procedures viz. soaking (8-12 h), microwave heating (2-6 min), cooking (30-60 min), and blanching (1-3 min), followed by sun drying, was studied on the nutritional, antinutritional and functional properties of Colocasia leaves. A significant increase in crude fibre (25.7%-29.65%), and protein (4.33-15.6%) content was found in all the treatments except for the microwave treatment. A significant decrease in fat (5.7-31.4%), ash (20.34-28.22%), oxalic acid (27.07-35.32%), and tannic acid (up to 96%) was also found in various treatments. Among the minerals, a significant increase was reported for calcium (up to 16.38%), and iron (up to 5.9%). The highest mineral retention was found in soaked samples. The soaked and cooked samples also had a higher Ca: Mg ratio. A significant change in functional properties was also found. FTIR peaks suggested no significant qualitative effect occurred on phytochemical or physicochemical characteristics. Cluster analysis showed that cooking was second to soaking in terms of overall quality which were most comparable to the control. Cooking efficiently reduced the antinutritional factors, however, a significant loss of nutrients and functional properties was also observed. Therefore, the soaking of Colocasia leaves for 8-10 h is recommended as the best practice before their food applications.

Rare complications of chicken pox in immunocompetent children: Acute respiratory distress syndrome.

Chickenpox is a highly contagious disease caused by varicella zoster virus. Fever and papulovesicular rash are hallmarks of this disease. These manifestations are self-liming and complete recovery is seen in most cases. We report two cases of chickenpox infection where the rare and atypical manifestations of acute respiratory distress syndrome with pleural effusion were seen.

COVID-19 presenting as acute adominal pain: A case series.

The coronavirus infection presents primarily as a respiratory illness, however, extra-pulmonary manifestations are known to occur, including gastrointestinal manifestations. Hereby, we report three cases of the COVID-19 infection who presented with acute-onset abdominal pain during illness. All three patients had respiratory symptoms suggestive of COVID-19 and abdominal symptoms consistent with acute pancreatitis, acute cholecystitis, and acute appendicitis. All three patients improved in terms of acute abdominal pain; however, the overall clinical course, the three illnesses were variable because of differences in underlying organ involment and pathophysiology.