Assessment and Control of Foodborne Pathogens in Ireland

End of Project ReportConsumers are increasingly demanding food that is free from pathogens, but with less preservatives and additives. As a response to these conflicting demands, current trends in the food industry include minimal processing, and the investigation of alternative inhibitors for use in foods. Additionally, the manufacture of an increasing range of novel foods, and the inclusion of non-dairy ingredients into dairy products, and vice versa, poses additional dangers with respect to safety. Furthermore, the dramatic increase in incidence of food-borne illness internationally, as a result of contamination with food-borne pathogens such as Listeria monocytogenes, is a cause of considerable consumer concern. Bacteriocins are inhibitory peptides produced by a number of Lactic Acid Bacteria which are capable of killing other bacteria. These natural inhibitors have widespread applications in the preservation of foods, since they can kill a number of pathogenic and spoilage bacteria. The broad spectrum bacteriocin Lacticin 3147 (discovered in a previous project and patented - see DPRC No. 3) is produced by Lactococcus lactis subsp. lactis DPC3147, a food-grade strain, similar to strains used for commercial cheese manufacture. Lacticin 3147 is effective in the inhibition of all Gram positive bacteria tested including the food pathogens Listeria monocytogenes and Staphylococcus aureus and food spoilage bacteria such as Clostridia and Bacillus species. As part of this project the bacteriocin Lacticin 3147 was assessed as a food preservative for improving food safety via inhibition of pathogenic organisms. Thus the project plan followed a "twin-track" approach to assessing and controlling the food safety aspects of Irish food. The first of these was designed to investigate the current safety status of Irish dairy products. The second approach involved an attempt to exploit natural antimicrobial substances, including Lacticin 3147, to protect foods from pathogenic bacteria.Department of Agriculture, Food and the Marin

Therapeutic Potential of a Novel Vitamin D3 Oxime Analogue, VD1-6, with CYP24A1 Enzyme Inhibitory Activity and Negligible Vitamin D Receptor Binding

Abstract: The regulation of vitamin D3 actions in humans occurs mainly through the Cytochrome P450 24-hydroxylase (CYP24A1) enzyme activity. CYP24A1 hydroxylates both 25-hydroxycholecalciferol (25(OH)D3) and 1,25-dihydroxycholecalciferol (1,25(OH)2D3), which is the first step of vitamin D catabolism. An abnormal status of the upregulation of CYP24A1 occurs in many diseases, including chronic kidney disease (CKD). CYP24A1 upregulation in CKD and diminished activation of vitamin D3 contribute to secondary hyperparathyroidism (SHPT), progressive bone deterioration, and soft tissue and cardiovascular calcification. Previous studies have indicated that CYP24A1 inhibition may be an effective strategy to increase endogenous vitamin D activity and decrease SHPT. This study has designed and synthesized a novel C-24 O-methyloxime analogue of vitamin D3 (VD1-6) to have specific CYP24A1 inhibitory properties. VD1-6 did not bind to the vitamin D receptor (VDR) in concentrations up to 10-7 M, assessed by a VDR binding assay. The absence of VDR binding by VD1-6 was confirmed in human embryonic kidney HEK293T cultures through the lack of CYP24A1 induction. However, in silico docking experiments demonstrated that VD1-6 was predicted to have superior binding to CYP24A1, when compared to that of 1,25(OH)2D3. The inhibition of CYP24A1 by VD1-6 was also evident by the synergistic potentiation of 1,25(OH)2D3-mediated transcription and reduced 1,25(OH)2D3 catabolism over 24 h. A further indication of CYP24A1 inhibition by VD1-6 was the reduced accumulation of the 24,25(OH)D3, the first metabolite of 25(OH)D catabolism by CYP24A1. Our findings suggest the potent CYP24A1 inhibitory properties of VD1-6 and its potential for testing as an alternative therapeutic candidate for treating SHPT.Ali K. Alshabrawy, Yingjie Cui, Cyan Sylvester, Dongqing Yang, Emilio S. Petito, Kate R. Barratt, Rebecca K. Sawyer, Jessica K. Heatlie, Ruhi Polara, Matthew J. Sykes, Gerald J. Atkins, Shane M. Hickey, Michael D. Wiese, Andrea M. Stringer, Zhaopeng Liu, and Paul H. Anderso

Legumes as food ingredient: characterization, processing, and applications

Editores: Jiménez-López, José Carlos (CSIC); Clemente, Alfonso (CSIC

Prediction of Prostate Cancer Biochemical and Clinical Recurrence Is Improved by IHC-Assisted Grading Using Appl1, Sortilin and Syndecan-1.

Gleason scoring is used within a five-tier risk stratification system to guide therapeutic decisions for patients with prostate cancer. This study aimed to compare the predictive performance of routine H&E or biomarker-assisted ISUP (International Society of Urological Pathology) grade grouping for assessing the risk of biochemical recurrence (BCR) and clinical recurrence (CR) in patients with prostate cancer. This retrospective study was an assessment of 114 men with prostate cancer who provided radical prostatectomy samples to the Australian Prostate Cancer Bioresource between 2006 and 2014. The prediction of CR was the primary outcome (median time to CR 79.8 months), and BCR was assessed as a secondary outcome (median time to BCR 41.7 months). The associations of (1) H&E ISUP grade groups and (2) modified ISUP grade groups informed by the Appl1, Sortilin and Syndecan-1 immunohistochemistry (IHC) labelling were modelled with BCR and CR using Cox proportional hazard approaches. IHC-assisted grading was more predictive than H&E for BCR (C-statistic 0.63 vs. 0.59) and CR (C-statistic 0.71 vs. 0.66). On adjusted analysis, IHC-assisted ISUP grading was independently associated with both outcome measures. IHC-assisted ISUP grading using the biomarker panel was an independent predictor of individual BCR and CR. Prospective studies are needed to further validate this biomarker technology and to define BCR and CR associations in real-world cohorts.Jessica M. Logan ... Lisa M. Butler ... Douglas A. Brooks ... et al

Social learning in LEADER: Exogenous, endogenous and hybrid evaluation in rural development

This paper considers the relationship between the centralised exogenous, institutions and the embedded, endogenous institutions of rural governance in Europe through an examination the evaluation procedures of the European LEADER programme. LEADER is presented in the literature as progressive in terms of innovation and stakeholder engagement. Yet while the planning and management of LEADER embraces heterogeneity and participation, programmatic evaluation is centralised and held at arms length from delivery organisations. The paper reviews previous efforts to improve evaluation in LEADER and considers alternative strategies for evaluation, contrasting LEADER practice with participatory evaluation methodologies in the wider international context. Can evaluation in itself be valuable as a mode of social learning and hence a driver for endogenous development in rural communities in Europe? The paper concludes by examining the challenges in producing a hybrid form of evaluation which accommodates endogenous and exogenous values

A 3,4-dimethoxy-1,8-naphthalimide for lipid droplet imaging in live and fixed cells

Lipid droplets (LDs) are found in most eukaryotic cells and in addition to serving as lipid reservoirs, these highly dynamic organelles play fundamental roles in cell metabolism and growth. The study of LDs and their in- teractions with other cellular compartments provides new insights into normal cell biology and disease processes such as virus packaging, metabolic disorders, and cancer progression. Here we report the synthesis and comprehensive evaluation of two readily prepared, low molecular weight 3,4-dimethoxy-1,8-naphthalimides, as selective LD stains. The newly synthesised 1,8-naphthalimide derivative (DMN-LD), demonstrated impressive versatility compared to currently available options, by staining LDs in both live and fixed cells, as well as a three- dimensional spheroid, using both single and two-photon excitation. This new imaging agent has the potential to further unravel the complex biology of LDs, which are essential to cell survival, and when altered, underpin many disease states.Ian R.D. Johnson, Elley E. Rudebeck, Martin J. Sweetman, Alexandra Sorvina, Trent D. Ashton, Frederick M. Pfeffer, Douglas A. Brooks, Shane M. Hicke

LC-MS/MS analysis of vitamin D2 metabolites in human serum using a salting-out based liquid-liquid extraction and DAPTAD derivatization

Available online 12 March 2021LC-MS/MS has recently emerged as the best-practice for simultaneous analysis of vitamin D metabolites. We have developed and validated an LC-MS/MS method for simultaneous quantification of 25(OH)D3, 24,25(OH)2D3, and 3-epi-25(OH)D3 in human serum. These three metabolites were extracted from 50 μL of serum by acetonitrile protein precipitation followed by salting-out of acetonitrile. DAPTAD (4-(4′-dimethylaminophenyl)-1,2,4-triazoline-3,5-dione) was used to derivatize the extracted metabolites and their deuterated isotope internal standards. Chromatographic separation was achieved on a UPLC C18 column (Waters® ACQUITY 100 × 2.1 mm, 1.7 µm) utilizing 0.1% formic acid and acetonitrile as mobile phases. Limits of quantification were 1 ng/mL for 25(OH)D3 and 0.1 ng/mL for 24,25(OH)D3 and 3-epi-25(OH)D3. In-house and external Vitamin D External Quality Assessment Scheme (DEQAS) quality control sample analysis revealed satisfactory method accuracy. Within-analytical batch and between analytical batches precision were <15%. Extraction recovery for the three analytes were all ˃ 85% and all showed adequate autosampler, bench-top and freeze–thaw stability. Inter-methodological comparison of 25(OH)D3 results in patient serum samples revealed systematic and proportional differences between our method and DiaSorin® Liaison immunoassay, however a good agreement with an independent LC-MS/MS method was found.Ali K.Alshabrawy, Amanda Bergamin, Deepti K.Sharma, Shane M.Hickey, Doug A.Brooks, Peter O'Loughlin, Michael D. Wiese, Paul H. Anderso