We report a series of 2′-deoxyribonucleoside triphosphates bearing dicarba-nido-undecaborate (C2B9H111–), 3,3′-iron-bis(1,2-dicarbollide)− (FESAN, Fe(C2B9H11)22–) or ...3,3′-cobalt-bis(1,2-dicarbollide)− (COSAN, Co(C2B9H11)22–) groups prepared either through the Sonogashira cross-coupling or the CuAAC click reaction. The modified dN X TPs were substrates for KOD XL DNA polymerase in enzymatic synthesis of modified DNA through primer extension (PEX). The nido-carborane- and FESAN-modified nucleotides gave analytically useful oxidation signals in square-wave voltammetry and were used for redox labeling of DNA. The redox-modified DNA probes were prepared by PEX using tailed primers and were hybridized to electrode (gold or glassy carbon) containing capture oligonucleotides. The combination of nido-carborane- and FESAN-linked nucleotides with 7-ferrocenylethynyl-7-deaza-dATP and 7-deaza-dGTP allowed polymerase synthesis of DNA fully modified at all four nucleobases, and each of the redox labels gave four differentiable and ratiometric signals in voltammetry. Thus, the combination of these four redox labels constitutes the first fully orthogonal redox coding of all four canonical nucleobases, which can be used for determination of nucleobase composition of short DNA stretches in one simple PEX experiment with electrochemical readout.
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IJS, KILJ, NUK, PNG, UL, UM
To determine the associations between training load, baseline characteristics (e.g. age or previous injury) and rate of musculoskeletal injury and/or pain specifically within an Endurance Sporting ...Population (ESP).
Prospectively registered systematic review.
Eight electronic databases were searched by two independent reviewers. Studies were required to prospectively monitor both (i) training loads and (ii) musculoskeletal injury and/or pain for >3 months. Methodological quality and risk of bias were determined utilising the Critical Skills Appraisal Program (CASP). Reported effect sizes were categorised as small, medium or large.
Twelve endurance sport studies were eligible (running, triathlon, rowing). Increased injury and/or pain risk was associated with: (i) high total training distances per week/month (medium effect size) (ii) training frequency <2 sessions/week (medium effect size) and (iii) both low weekly (<2hours/week) and high monthly (large effect size) training durations. None of the studies reported internal training load data or acute:chronic workload ratios. Baseline characteristics found to increase the rate of injury and/or pain included: (i) a history of previous injury (medium effect size), (ii) age >45 years (small effect size), (iii) non-musculoskeletal comorbidities (large effect size), (iv) using older running shoes (small effect size) and (v) non-competitive behaviour.
This review identifies a range of external training load factors and baseline characteristics associated with an increased rate of injury and/or pain within ESPs. There is an absence of research relating to internal training loads and acute:chronic workload ratios in relation to rate of injury and/or pain within ESPs.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
The importance of ciguatoxins (CTXs) in seafood safety and their emerging occurrence in locations far away from tropical areas highlight the need for simple and low-cost methods for the sensitive and ...rapid detection of these potent marine toxins to protect seafood consumers. Herein, an electrochemical immunosensor for the detection of CTXs is presented. A sandwich configuration is proposed, using magnetic beads (MBs) as immobilization supports for two capture antibodies, with their combination facilitating the detection of CTX1B, CTX3C, 54-deoxyCTX1B, and 51-hydroxyCTX3C. PolyHRP-streptavidin is used for the detection of the biotinylated detector antibody. Experimental conditions are first optimized using colorimetry, and these conditions are subsequently used for electrochemical detection on electrode arrays. Limits of detection at the pg/mL level are achieved for CTX1B and 51-hydroxyCTX3C. The applicability of the immunosensor to the analysis of fish samples is demonstrated, attaining detection of CTX1B at contents as low as 0.01 μg/kg and providing results in correlation with those obtained using mouse bioassay (MBA) and cell-based assay (CBA), and confirmed by liquid chromatography coupled to high-resolution mass spectrometry (LC-ESI-HRMS). This user-friendly bioanalytical tool for the rapid detection of CTXs can be used to mitigate ciguatera risk and contribute to the protection of consumer health.
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IJS, KILJ, NUK, PNG, UL, UM
Ostreopsis cf. ovata is a benthic microalga distributed in tropical and temperate regions worldwide which produces palytoxins (PlTXs). Herein, an electrochemical biosensor for the detection of this ...toxic microalga is described. The detection strategy involves isothermal recombinase polymerase amplification (RPA) of the target using tailed primers and a sandwich hybridisation assay on maleimide-coated magnetic beads immobilised on electrode arrays. The biosensor attained a limit of detection of 9 pg/μL of O. cf. ovata DNA (which corresponds to ~640 cells/L), with no interferences from two non-target Ostreopsis species (O. cf. siamensis and O. fattorussoi). The biosensor was applied to the analysis of planktonic and benthic environmental samples. Electrochemical O. cf. ovata DNA quantifications demonstrated an excellent correlation with other molecular methods (qPCR and colorimetric assays) and allowed the construction of a predictive regression model to estimate O. cf. ovata cell abundances. This new technology offer great potential to improve research, monitoring and management of O. cf. ovata and harmful algal blooms.
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•First electrochemical biosensor for the detection of Ostreopsis cf. ovata•The biosensor combines isothermal amplification with magnetic beads as supports•The application of the biosensor to environmental samples is successfully proved•Excellent correlation with other molecular techniques and light microscopy•A rapid, simple and inexpensive alternative tool for marine monitoring is provided
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Abstract Esophageal adenocarcinoma (EAC) is the eighth most common cancer worldwide, and approximately 15% of patients survive 5 years. Reflux disease (GERD) and Barrett’s esophagus (BE) are major ...risk factors for the development of EAC, and epidemiologic studies highlight a strong association with obesity. The immune, inflammatory and intracellular signaling changes resulting from chronic inflammation of the esophageal squamous epithelium are increasingly well characterized. In GERD and Barrett’s, an essential role for T-cells in the initiation of inflammation in the esophagus has been identified, and a balance between T-cell responses and phenotype may play an important role in disease progression. Obesity is a chronic low-grade inflammatory state, fueled by adipose tissue derived- inflammatory mediators such as IL-6, TNF-α and leptin, representing a novel area for targeted research. Additionally, reactive oxygen species (ROS) and receptor tyrosine kinase (RTK) activation may drive progression from esophagitis to EAC, and downstream signaling pathways employed by these molecules may be important. This review will explain the diverse range of mechanisms potentially driving and maintaining inflammation within the esophagus and explore both existing and future therapeutic strategies targeting the process.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
Karlodinium is a dinoflagellate responsible for fish-killing events worldwide. In Alfacs Bay (NW Mediterranean Sea), the presence of two Karlodinium species (K. veneficum and K. armiger) with ...different toxicities has been reported. This work presents a method that combines recombinase polymerase amplification (RPA) with an enzyme-linked oligonucleotide assay (ELONA) to identify, discriminate and quantify these two species. The system was characterised using synthetic DNA and genomic DNA, and the specificity was confirmed by cross-reactivity experiments. Calibration curves were constructed using 10-fold dilutions of cultured cells, attaining a limit of detection of around 50,000 cells/L, far below the Karlodinium spp. alert threshold (200,000 cells/L). Finally, the assay was applied to spiked seawater samples, showing an excellent correlation with the spiking levels and light microscopy counts. This approach is more rapid, specific and user-friendly than traditional microscopy techniques, and shows great promise for the surveillance and management of harmful algal blooms.
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•An assay for the detection of two Karlodinium species has been developed.•The assay is based on isothermal DNA amplification and a sandwich hybridisation.•The assay discriminates between Karlodinium veneficum and Karlodinium armiger.•Seawater samples spiked with Karlodinium cells have been analysed.•Excellent correlations between the assay and light microscopy have been obtained.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
The use of selective dry cow antimicrobial therapy requires precisely differentiating cows with an intramammary infection (IMI) from uninfected cows close to drying-off to enable treatment ...allocation. Milk somatic cell count (SCC) is an indicator of an inflammatory response in the mammary gland and is usually associated with IMI. However, SCC can also be influenced by cow-level variables such as milk yield, lactation number, and stage of lactation. In recent years, predictive algorithms have been developed to differentiate cows with IMI from cows without IMI based on SCC data. The objective of this observational study was to explore the association between SCC and subclinical IMI, taking cognizance of cow-level predictors on Irish seasonal spring calving, pasture-based systems. Additionally, the optimal test-day SCC cut-point (maximized sensitivity and specificity) for IMI diagnosis was determined. A total of 2,074 cows across 21 spring calving dairy herds with an average monthly milk weighted bulk tank SCC of ≤200,000 cells/mL were enrolled in the study. Quarter-level milk sampling was carried out on all cows in late lactation (interquartile range = 240–261 d in milk) for bacteriological culturing. Bacteriological results were used to define cows with IMI when ≥1 quarter sample resulted in bacterial growth. Cow-level test-day SCC records were provided by the herd owners. The ability of the average, maximum, and last test-day SCC to predict infection were compared using receiver operator curves. Predictive logistic regression models tested included parity (primiparous or multiparous), yield at last test-day, and a standardized count of high SCC test-days. In total, 18.7% of cows were classified as having an IMI, with first-parity cows having a higher proportion of IMI (29.3%) compared with multiparous cows (16.1%). Staphylococcus aureus accounted for the majority of these infections. The last test-day SCC was the best predictor of infection with the highest area under the curve. The inclusions of parity, yield at last test-day, and a standardized count of high SCC test-days as predictors did not significantly improve the ability of last test-day SCC to predict IMI. The cut-point for last test-day SCC that maximized sensitivity and specificity was 64,975 cells/mL. This study indicates that in Irish seasonal pasture-based dairy herds with low bulk tank SCC, the last test-day SCC (interquartile range days in milk = 221–240) is the best predictor of IMI in late lactation.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
The use of isothermal nucleic acid amplification strategies to detect harmful algal blooms (HABs) is in its infancy. We describe recent advances in these systems and highlight the challenges for the ...achievement of simple, low-cost, compact, and portable devices for field applications.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Recombinase polymerase amplification (RPA) is a technique that is used to specifically amplify a target nucleic acid sequence. Unlike the polymerase chain reaction (PCR), RPA is performed at a ...constant temperature between 37 and 42°C. Therefore, it can be potentially used for the onsite detection of various pathogens when combined with DNA extraction and amplicon detection techniques. In this study, we prepared recombinant recombinase and single-stranded DNA-binding protein from T4 phage and used them to examine the effects of reaction conditions and additives on the efficiency of RPA. The results revealed that the optimal pH was 7.5–8.0, optimal potassium acetate concentration was 40–80 mM, and optimal reaction temperature was 37–45°C although dimethyl sulfoxide at 5% v/v and formamide at 5% v/v inhibited the reaction. Our results suggest that RPA could be conducted using a wider range of optimal reaction conditions than those required for PCR and that RPA is highly suitable for point-of-care use.
•Recombinase polymerase amplification (RPA) amplifies a target nucleic acid sequence.•We prepared recombinant recombinase and single-stranded DNA-binding protein.•We examined the effects of reaction conditions and additives on the RPA efficiency.•The optimal pH was 7.5–8.0, and optimal CH3COOK concentration was 40–80 mM.•Dimethyl sulfoxide at 5% v/v and formamide at 5% v/v inhibited the reaction.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
To examine the association between subjective health complaints, sleep quantity and new injury within an endurance sport population.
Prospective cohort study.
Ninety-five endurance sporting ...participants were recruited from running, triathlon, swimming, cycling and rowing disciplines. Over 52-week period participants submitted weekly data regarding subjective health complaints (SHCs) (cardiorespiratory, gastrointestinal and psychological/lifestyle), sleep quantity, training load and new injury episodes. Applying a 7- and 14-day lag period, a shared frailty model was used to explore new injury risk associations with total SHCs and sleep quantity.
92.6% of 95 participants completed all 52 weeks of data submission and the remainder of the participants completed ≥30 weeks. Seven-day lag psychological/lifestyle SHCs were significantly associated with new injury risk (Hazard ratio (HR)=1.32; CI 95%=1.01–1.72, p<0.04). In contrast, cardiorespiratory (HR=1.15; CI 95%=0.99–1.36, p=0.07) and gastrointestinal (HR=0.77; CI 95%=0.56–1.05, p=0.09) SHCs were not significantly associated with new injury risk. New injury risk had a significant increased association with 14-day lag <7h/day sleep quantity (HR=1.51; CI 95%=2.02–1.13, p<0.01) and a significant decreased association with >7h/day sleep quantity (HR=0.63, CI 95%=0.45–0.87, p<0.01. A secondary regression analysis demonstrated no significant association with total SHCs and training load factors (Relative Risk (RR)=0.08, CI 95%=0.04–0.21, p=0.20).
To minimise an increased risk of new injuries within an endurance sporting population, this study demonstrates that psychological/lifestyle subjective health complaints and sleep quantity should be considered. The study also highlights a lag period between low sleep quantity and its subsequent impact on new injury risk. No association was demonstrated between subjective health complaints, sleep quantity and training load factors.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
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