Estuarine ecosystems face increasing anthropogenic pressures, necessitating effective monitoring methods to mitigate their impacts on the biodiversity they harbour. The use of environmental DNA (eDNA) based detection methods is increasingly recognized as a promising tool to complement other, potentially invasive monitoring techniques. Integrating such eDNA analyses into monitoring frameworks for large ecosystems is still challenging and requires a deeper understanding of the scale and resolution at which eDNA patterns may offer insights in species presence and community composition space and time. The Scheldt estuary, characterized by its diverse habitats and complex currents, is one of the largest Western European tidal river systems. Until now, it remains challenging to obtain accurate information on fish communities living in and migrating through this ecosystem, consequently confining our knowledge to specific locations. To explore the potential of eDNA based monitoring, we simultaneously combine stow net fishing with eDNA metabarcoding, to assess spatiotemporal shifts in the Scheldt estuary\'s fish communities. In total, we detected 71 fish species in the estuary using eDNA metabarcoding, partly overlapping with historic fish community data gathered at the different study locations and in contrast to only 42 species using stow net fishing during the same survey period. Community compositions found by both detection methods varied among sampling locations, driven by a clear correlation to the salinity gradient. Limited effects of sampling depth and tide were observed on the eDNA metabarcoding data, allowing a significant reduction of the eDNA sampling effort for future eDNA fish monitoring campaigns in this study system. Our results further demonstrate that seasonal shifts in fish species occurrence can be detected using eDNA metabarcoding. Combining eDNA metabarcoding and stow net fishing further enhances our understanding of this vital waterway\'s diverse fish populations, allowing a higher resolution and more efficient monitoring strategy.

Pain intensity is the most commonly used outcome domain in pain clinical trials. To minimize the chances of type II error (ie, concluding that a treatment does not have beneficial effects, when in fact it does), the measure of pain intensity used should be sensitive to changes produced by effective pain treatments. Here we sought to identify the combination of pain intensity ratings that would balance the need for reliability and validity against the need to minimize assessment burden. We conducted secondary analyses using data from a completed 4-arm clinical trial of psychological pain treatments (N = 164 adults). Current, worst, least, and average pain intensity in the past 24 hours were assessed 4 times before and after treatment using 0 to 10 numerical rating scale-11. We created a variety of composite scores using these ratings and evaluated their reliability (Cronbach\'s alphas) and validity (ie, associations with a gold standard score created by averaging 16 ratings and sensitivity for detecting between-group differences in treatment efficacy). We found that for each measure, reliability increased as the number of ratings used to create the measures increased and that ratings from 3 or more days were needed to have adequately strong associations with the gold standard. Regarding sensitivity, the findings suggest that composite scores made up of ratings from 4 days are needed to maximize the chances of detecting treatment effects, especially with smaller sample sizes. In conclusion, using data from 3 or 4 days of assessment may be the best practice. PERSPECTIVE: Composite scores made up of at least 3 days of pain ratings appear to be needed to maximize reliability and validity while minimizing the assessment burden. TRIAL REGISTRATION: clinicaltrials.gov NCT01800604.

Preclinical evaluation of drugs in animals helps researchers to select potentially informative clinical laboratory markers for human trials. To assess the utility of animal thrombin generation (TG) assay, we studied the sensitivity of animal plasmas to triggers of TG, human Tissue Factor (TF), and Activated Factor XI (FXIa). Pooled human, mouse, rat, guinea pig, rabbit, bovine, sheep, and goat plasmas were used in this study. TF- or FXIa-triggered TG and clotting were measured via fluorescence and optical density, respectively. Thrombin peak height (TPH) and time (TPT), clot time (CT), and fibrin clot density (FCD) were all analyzed. The trigger low and high sensitivity borders (LSB and HSB) for each assay parameter were defined as TF and FXIa concentrations, providing 20 and 80% of the maximal parameter value, unless the baseline (no trigger) value exceeded 20% of the maximal, in which case, LSB was derived from 120% of baseline value. Normal human samples demonstrated lower TPH HSB than most of the animal samples for both TF and FXIa. Animal samples, except mice, demonstrated lower TPT LSB for FXIa versus humans. Most rodent and rabbit samples produced baseline TG in the absence of TG triggers that were consistent with the pre-activation of blood coagulation. FCD was not sensitive to both TF and FXIa in either of the plasmas. Animal plasmas have widely variable sensitivities to human TF and FXIa, which suggests that optimization of trigger concentration is required prior to test use, and this complicates the extrapolation of animal model results to humans.

Necessity for finding improved intervention in many legacy therapeutic areas are of high priority. This has the potential to decrease the expense of medical care and poor outcomes for many patients. Typically, clinical efficacy is the primary evaluating criteria to measure any beneficial effect of a treatment. Albeit, there could be situations when several other factors (e.g. side-effects, cost-burden, less debilitating, less intensive, etc.) which can permit some slightly less efficacious treatment options favorable to a subgroup of patients. This often leads to non-inferiority (NI) testing. NI trials may or may not include a placebo arm due to ethical reasons. However, when included, the resulting three-arm trial is more prudent since it requires less stringent assumptions compared to a two-arm placebo-free trial. In this article, we consider both Frequentist and Bayesian procedures for testing NI in the three-arm trial with binary outcomes when the functional of interest is risk difference. An improved Frequentist approach is proposed first, which is then followed by a Bayesian counterpart. Bayesian methods have a natural advantage in many active-control trials, including NI trial, as it can seamlessly integrate substantial prior information. In addition, we discuss sample size calculation and draw an interesting connection between the two paradigms.

Variability in pain-related outcomes can hamper assay sensitivity of chronic pain clinical trials. Expectations of outcome in such trials may account for some of this variability, and thereby impede development of novel pain treatments. Measurement of participants\' expectations prior to initiating study treatment (active or placebo) is infrequent, variable, and often unvalidated. Efforts to optimize and standardize measurement, analysis, and management of expectations are needed. In this Focus Article, we provide an overview of research findings on the relationship between baseline expectations and pain-related outcomes in clinical trials of pharmacological and nonpharmacological pain treatments. We highlight the potential benefit of adjusting for participants\' expectations in clinical trial analyses and draw on findings from patient interviews to discuss critical issues related to measurement of expectations. We conclude with suggestions regarding future studies focused on better understanding the utility of incorporating these measures into clinical trial analyses. PERSPECTIVE: This focus article provides an overview of the relationship between participants\' baseline expectations and pain-related outcomes in the setting of clinical trials of chronic pain treatments. Systematic research focused on the measurement of expectations and the impact of adjusting for expectations in clinical trial analyses may improve assay sensitivity.

BACKGROUND: The efficacy of analgesics in controlling orthodontic pain: a systematic review and meta- analysis. Cheng C, Xie T, Wang J. BMC Oral Health 2020; 20:259.
BACKGROUND: The systematic review was funded by grants from the National Natural Science Foundation of China (No. 81771114 and No. 81970967). The authors have no actual or potential conflicts of interest.
METHODS: Systematic review with meta-analysis of data.

The present study aimed to determine whether current commercial immunoassays are adequate for detecting anti-Omicron antibodies. We analyzed the anti-SARS-CoV-2 antibody response of 23 unvaccinated individuals 1-2 months after an Omicron infection. All blood samples were tested with a live virus neutralization assay using a clinical Omicron BA.1 strain and four commercial SARS-CoV-2 immunoassays. We assessed three anti-Spike immunoassays (SARS-CoV-2 IgG II Quant [Abbott S], Wantaï anti-SARS-CoV-2 antibody ELISA [Wantaï], Elecsys Anti-SARS-CoV-2 S assay [Roche]) and one anti-Nucleocapsid immunoassay (Abbott SARS-CoV-2 IgG assay [Abbott N]). Omicron neutralizing antibodies were detected in all samples with the live virus neutralization assay. The detection rate of the Abbott S, Wantai, Roche, and Abbott N immunoassays were 65.2%, 69.6%, 86.9%, and 91.3%, respectively. The sensitivities of Abbott S and Wantai immunoassays were significantly lower than that of the live virus neutralization assay (p = 0.004, p = 0.009; Fisher\'s exact test). Antibody concentrations obtained with anti-S immunoassays were correlated with Omicron neutralizing antibody concentrations. These data provide clinical evidence of the loss of performance of some commercial immunoassays to detect antibodies elicited by Omicron infections. It highlights the need to optimize these assays by adapting antigens to the circulating SARS-CoV-2 strains.

For indications where only unstable reference treatments are available and use of placebo is ethically justified, three-arm \"gold standard\" designs with an experimental, reference and placebo arm are recommended for non-inferiority trials. In such designs, the demonstration of efficacy of the reference or experimental treatment is a requirement. They have the disadvantage that only little can be concluded from the trial if the reference fails to be efficacious. To overcome this, we investigate novel single-stage, adaptive test strategies where non-inferiority is tested only if the reference shows sufficient efficacy and otherwise δ $$ \\delta $$ -superiority of the experimental treatment over placebo is tested. With a properly chosen superiority margin, δ $$ \\delta $$ -superiority indirectly shows non-inferiority. We optimize the sample size for several decision rules and find that the natural, data driven test strategy, which tests non-inferiority if the reference\'s efficacy test is significant, leads to the smallest overall and placebo sample sizes. We proof that under specific constraints on the sample sizes, this procedure controls the family-wise error rate. All optimal sample sizes are found to meet this constraint. We finally show how to account for a relevant placebo drop-out rate in an efficient way and apply the new test strategy to a real life data set.

According to the ICH S7B guideline, drug candidates are screened for hERG block prior to first-in-human testing to predict the likelihood of delayed repolarization associated with a rare, but life-threatening, ventricular tachyarrhythmia. The new ICH E14 Q&As guideline allows hERG results to be used in later clinical development for decision-making (Q&As 5.1 and 6.1). To pursue this path, the hERG assay should be conducted following the new ICH S7B Q&A 2.1 guideline, which calls for best practice considerations of the recording temperature, voltage protocol, stimulation frequency, recording/data quality, and concentration verification. This study investigated hERG block by cisapride, dofetilide, terfenadine, sotalol, and E-4031 - positive controls commonly used to demonstrate assay sensitivity - using the manual whole cell patch clamp method and an action potential-like voltage protocol presented at 0.2 Hz. Recordings were conducted at room and near physiological temperature. Drug concentrations were measured using samples collected during real patch clamp experiments and satellite experiments. Results showed temperature effects for E-4031, terfenadine, and sotalol, but not cisapride and dofetilide. Cisapride and terfenadine showed substantial concentration losses, largely due to nonspecific binding to the perfusion apparatus. Using concentrations measured from the real and satellite experiments to assess block potencies yielded comparable results, indicating that satellite sample collection may be viable for drugs with nonspecific binding concerns only. In summary, this study provides block potencies for 5 hERG positive controls, and serves as a case study for hERG assays conducted, and results illustrated in accordance with the new ICH E14/S7B Q&As.

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