Economic significance of biofilms: a multidisciplinary and cross-sectoral challenge.

The increasing awareness of the significance of microbial biofilms across different sectors is continuously revealing new areas of opportunity in the development of innovative technologies in translational research, which can address their detrimental effects, as well as exploit their benefits. Due to the extent of sectors affected by microbial biofilms, capturing their real financial impact has been difficult. This perspective highlights this impact globally, based on figures identified in a recent in-depth market analysis commissioned by the UK's National Biofilms Innovation Centre (NBIC). The outputs from this analysis and the workshops organised by NBIC on its research strategic themes have revealed the breath of opportunities for translational research in microbial biofilms. However, there are still many outstanding scientific and technological challenges which must be addressed in order to catalyse these opportunities. This perspective discusses some of these challenges.

Reverse-Transcription Loop-Mediated Isothermal Amplification Has High Accuracy for Detecting Severe Acute Respiratory Syndrome Coronavirus 2 in Saliva and Nasopharyngeal/Oropharyngeal Swabs from Asymptomatic and Symptomatic Individuals.

Previous studies have described reverse-transcription loop-mediated isothermal amplification (RT-LAMP) for the rapid detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in nasopharyngeal/oropharyngeal swab and saliva samples. This multisite clinical evaluation describes the validation of an improved sample preparation method for extraction-free RT-LAMP and reports clinical performance of four RT-LAMP assay formats for SARS-CoV-2 detection. Direct RT-LAMP was performed on 559 swabs and 86,760 saliva samples and RNA RT-LAMP on extracted RNA from 12,619 swabs and 12,521 saliva samples from asymptomatic and symptomatic individuals across health care and community settings. For direct RT-LAMP, overall diagnostic sensitivity (DSe) was 70.35% (95% CI, 63.48%-76.60%) on swabs and 84.62% (95% CI, 79.50%-88.88%) on saliva, with diagnostic specificity of 100% (95% CI, 98.98%-100.00%) on swabs and 100% (95% CI, 99.72%-100.00%) on saliva, compared with quantitative RT-PCR (RT-qPCR); analyzing samples with RT-qPCR ORF1ab CT values of ≤25 and ≤33, DSe values were 100% (95% CI, 96.34%-100%) and 77.78% (95% CI, 70.99%-83.62%) for swabs, and 99.01% (95% CI, 94.61%-99.97%) and 87.61% (95% CI, 82.69%-91.54%) for saliva, respectively. For RNA RT-LAMP, overall DSe and diagnostic specificity were 96.06% (95% CI, 92.88%-98.12%) and 99.99% (95% CI, 99.95%-100%) for swabs, and 80.65% (95% CI, 73.54%-86.54%) and 99.99% (95% CI, 99.95%-100%) for saliva, respectively. These findings demonstrate that RT-LAMP is applicable to a variety of use cases, including frequent, interval-based direct RT-LAMP of saliva from asymptomatic individuals who may otherwise be missed using symptomatic testing alone.

Preliminary optimisation of a simplified sample preparation method to permit direct detection of SARS-CoV-2 within saliva samples using reverse-transcription loop-mediated isothermal amplification (RT-LAMP).

We describe the optimisation of a simplified sample preparation method which permits rapid and direct detection of SARS-CoV-2 RNA within saliva, using reverse-transcription loop-mediated isothermal amplification (RT-LAMP). Treatment of saliva samples prior to RT-LAMP by dilution 1:1 in Mucolyse™, followed by dilution in 10 % (w/v) Chelex© 100 Resin and a 98 °C heat step for 2 min enabled detection of SARS-CoV-2 RNA in positive saliva samples. Using RT-LAMP, SARS-CoV-2 RNA was detected in as little as 05:43 min, with no amplification detected in 3097 real-time reverse transcription PCR (rRT-PCR) negative saliva samples from staff tested within a service evaluation study, or for other respiratory pathogens tested (n = 22). Saliva samples can be collected non-invasively, without the need for skilled staff and can be obtained from both healthcare and home settings. Critically, this approach overcomes the requirement for, and validation of, different swabs and the global bottleneck in obtaining access to extraction robots and reagents to enable molecular testing by rRT-PCR. Such testing opens the possibility of public health approaches for effective intervention during the COVID-19 pandemic through regular SARS-CoV-2 testing at a population scale, combined with isolation and contact tracing.

Variants in the genes encoding TNF-α, IL-10, and GSTP1 influence the effect of α-tocopherol on inflammatory cell responses in healthy men.

BACKGROUND: Despite evidence of antioxidant effects of vitamin E in vitro and in animal studies, large, randomized clinical trials have not substantiated a benefit of vitamin E in reducing inflammation in humans. An individual's genetic background may affect the response to α-tocopherol supplementation, but this has rarely been investigated. OBJECTIVE: The aim of this study was to explore the role of genetic polymorphisms on changes in LPS-stimulated inflammatory cytokine production from peripheral blood mononuclear cells (PBMCs) after α-tocopherol supplementation. DESIGN: A total of 160 healthy, middle-aged male volunteers (mean age: 52.7 y) were given dietary supplements of either 75 IU (low dose; n = 57) or 600 IU (high dose; n = 103) α-tocopherol/d for 6 wk. The production of TNF-α and IL-1ß, -6, and -10 by PBMCs after LPS stimulation was measured at baseline and after 6 wk. Polymorphisms in 15 genes involved in inflammation or responses to oxidative stress were characterized in the subjects. RESULTS: The ability of α-tocopherol to affect TNF-α production by LPS-stimulated PBMCs was influenced by the TNFA -238 polymorphism (P = 0.016). The ability of α-tocopherol to affect IL-6 production was influenced by the GSTP1 313 polymorphism (P = 0.019). The ability of α-tocopherol to affect IL-1ß production was influenced by the IL10 -592 and -1082 polymorphisms (P = 0.025 and P = 0.016, respectively). CONCLUSIONS: In healthy control subjects, the effect of α-tocopherol supplementation on the production of inflammatory cytokines appears to be dependent on an individual's genotype. These genotype-specific differences may help explain some of the discordant results in studies that used vitamin E.

Epigenetic gene promoter methylation at birth is associated with child's later adiposity.

OBJECTIVE: Fixed genomic variation explains only a small proportion of the risk of adiposity. In animal models, maternal diet alters offspring body composition, accompanied by epigenetic changes in metabolic control genes. Little is known about whether such processes operate in humans. RESEARCH DESIGN AND METHODS: Using Sequenom MassARRAY we measured the methylation status of 68 CpGs 5' from five candidate genes in umbilical cord tissue DNA from healthy neonates. Methylation varied greatly at particular CpGs: for 31 CpGs with median methylation ≥5% and a 5-95% range ≥10%, we related methylation status to maternal pregnancy diet and to child's adiposity at age 9 years. Replication was sought in a second independent cohort. RESULTS: In cohort 1, retinoid X receptor-α (RXRA) chr9:136355885+ and endothelial nitric oxide synthase (eNOS) chr7:150315553+ methylation had independent associations with sex-adjusted childhood fat mass (exponentiated regression coefficient [ß] 17% per SD change in methylation [95% CI 4-31], P = 0.009, n = 64, and ß = 20% [9-32], P < 0.001, n = 66, respectively) and %fat mass (ß = 10% [1-19], P = 0.023, n = 64 and ß =12% [4-20], P = 0.002, n = 66, respectively). Regression analyses including sex and neonatal epigenetic marks explained >25% of the variance in childhood adiposity. Higher methylation of RXRA chr9:136355885+, but not of eNOS chr7:150315553+, was associated with lower maternal carbohydrate intake in early pregnancy, previously linked with higher neonatal adiposity in this population. In cohort 2, cord eNOS chr7:150315553+ methylation showed no association with adiposity, but RXRA chr9:136355885+ methylation showed similar associations with fat mass and %fat mass (ß = 6% [2-10] and ß = 4% [1-7], respectively, both P = 0.002, n = 239). CONCLUSIONS: Our findings suggest a substantial component of metabolic disease risk has a prenatal developmental basis. Perinatal epigenetic analysis may have utility in identifying individual vulnerability to later obesity and metabolic disease.

Effect of sex and dietary fat intake on the fatty acid composition of phospholipids and triacylglycerol in rat heart.

Variations in the fatty acid composition of lipids in the heart alter its function and susceptibility to ischaemic injury. We investigated the effect of sex and dietary fat intake on the fatty acid composition of phospholipids and triacylglycerol in rat heart. Rats were fed either 40 or 100g/kg fat (9:1 lard:soybean oil) from weaning until day 105. There were significant interactive effects of sex and fat intake on the proportions of fatty acids in heart phospholipids, dependent on phospholipid classes. 20:4n-6, but not 22:6n-3, was higher in phospholipids in females than males fed a low, but not a high, fat diet. There was no effect of sex on the composition of triacylglycerol. These findings suggest that sex is an important factor in determining the incorporation of dietary fatty acids into cardiac lipids. This may have implications for sex differences in susceptibility to heart disease.

Maternal protein restriction with or without folic acid supplementation during pregnancy alters the hepatic transcriptome in adult male rats.

Feeding pregnant rats a protein-restricted (PR) diet induces altered expression of candidate genes in the liver of the adult offspring, which can be prevented by supplementation of the PR diet with folic acid (PRF). We investigated the effect of maternal nutrition during pregnancy on the liver transcriptome in their adult male offspring. Pregnant rats were fed control, PR or PRF diets. Male offspring were killed on day 84. The liver transcriptome was analysed by microarray (six livers per maternal dietary group) followed by post hoc analysis of relative mRNA levels and gene ontology. These results were confirmed for selected genes by real-time RT-PCR. There were 311 genes that differed significantly ( >or= 1.5-fold change; P < 0.05) between PR offspring (222 increased) and control offspring, while 191 genes differed significantly between PRF offspring (forty-five increased) compared with offspring of control dams. There were sixteen genes that were significantly altered in both PR and PRF offspring compared with controls. Ion transport, developmental process, and response to reactive oxygen species (RROS) and steroid hormone response (SHR) ontologies were altered in PR offspring. Folic acid supplementation prevented changes within RROS and SHR response pathways, but not in ion transport or developmental process. There was no effect of maternal PR on mRNA expression of imprinted genes. Insulin 1 and Pleckstrin homology-like domain family A member 2 were increased significantly in PRF compared with PR offspring. The present findings show that the pattern of induced changes in the adult liver transcriptome were dependent on maternal protein and folic acid intakes during pregnancy.

Folic acid supplementation during the juvenile-pubertal period in rats modifies the phenotype and epigenotype induced by prenatal nutrition.

Prenatal nutritional constraint is associated with increased risk of metabolic dysregulation in adulthood contingent on adult diet. In rats, folic acid supplementation of a protein-restricted (PR) diet during pregnancy prevents altered phenotype and epigenotype in the offspring induced by the PR diet. We hypothesized that increasing folic acid intake during the juvenile-pubertal (JP) period would reverse the effects of a maternal PR diet on the offspring. Rats were fed a control (C) or PR diet during pregnancy and AIN93G during lactation. Offspring were weaned on d 28 onto diets containing 1 mg [adequate folate (AF)] or 5 mg [folic acid-supplemented (FS)] folic acid/kg feed. After 28 d, all offspring were fed a high-fat (18% wt:wt) diet and killed on d 84. As expected, offspring of PR dams fed the AF diet had increased fasting plasma triglyceride (TAG) and beta-hydroxybutyrate (betaHB) concentrations. The FS diet induced increased weight gain, a lower plasma betaHB concentration, and increased hepatic and plasma TAG concentration compared with AF offspring irrespective of maternal diet. PPARalpha and glucocorticoid receptor promoter methylation increased in liver and insulin receptor promoter methylation decreased in liver and adipose tissue in FS compared with AF offspring, with reciprocal changes in mRNA expression irrespective of maternal diet. These findings show that increased folic acid intake during the JP period did not simply reverse the phenotype induced by the maternal diet. This may represent a period of plasticity when specific nutrient intakes may alter the phenotype of the offspring through epigenetic changes in specific genes.