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Background and purposeBenefit of early endovascular treatment (EVT) for ischemic stroke varies considerably among patients. The MR PREDICTS decision tool, derived from MR CLEAN (Multicenter Randomized Clinical Trial of Endovascular Treatment for Acute Ischemic Stroke in the Netherlands), predicts outcome and treatment benefit based on baseline characteristics. Our aim was to externally validate and update MR PREDICTS with data from international trials and daily clinical practice.MethodsWe used individual patient data from 6 randomized controlled trials within the HERMES (Highly Effective Reperfusion Evaluated in Multiple Endovascular Stroke Trials) collaboration to validate the original model. Then, we updated the model and performed a second validation with data from the observational MR CLEAN Registry. Primary outcome was functional independence (defined as modified Rankin Scale score 0–2) 3 months after stroke. Treatment benefit was defined as the difference between the probability of functional independence with and without EVT. Discriminative performance was evaluated using a concordance (C) statistic.ResultsWe included 1242 patients from HERMES (633 assigned to EVT, 609 assigned to control) and 3156 patients from the MR CLEAN Registry (all of whom underwent EVT within 6.5 hours). The C-statistic for functional independence was 0.74 (95% CI, 0.72–0.77) in HERMES and, after model updating, 0.80 (0.78–0.82) in the Registry. Median predicted treatment benefit of routinely treated patients (Registry) was 10.3% (interquartile range, 5.8%–14.4%). Patients with low (

IntroductionThe Meta VCI Map consortium performs meta-analyses on strategic lesion locations for vascular cognitive impairment using lesion-symptom mapping. Integration of data from different cohorts will increase sample sizes, to improve brain lesion coverage and support comprehensive lesion-symptom mapping studies.MethodsCohorts with available imaging on white matter hyperintensities or infarcts and cognitive testing were invited. We performed a pilot study to test the feasibility of multicenter data processing and analysis and determine the benefits to lesion coverage.ResultsForty-seven groups have joined Meta VCI Map (stroke n = 7800 patients; memory clinic n = 4900; population-based n = 14,400). The pilot study (six ischemic stroke cohorts, n = 878) demonstrated feasibility of multicenter data integration (computed tomography/magnetic resonance imaging) and achieved marked improvement of lesion coverage.DiscussionMeta VCI Map will provide new insights into the relevance of vascular lesion location for cognitive dysfunction. After the successful pilot study, further projects are being prepared. Other investigators are welcome to join.

Abstract Background Delirium is a common disorder in the early phase of stroke. Given the presumed cholinergic deficiency in delirium, we tested treatment with the acetylcholinesterase inhibitor rivastigmine. Methods This pilot study was performed within an epidemiological study. In 527 consecutive stroke patients presence of delirium was assessed during the first week with the confusion assessment method. Severity was scored with the delirium rating scale (DRS). Sixty-two patients developed a delirium in the acute phase of stroke. Only patients with a severe and persistent delirium (defined as a DRS of 12 or more for more than 24 hours) were enrolled in the present study. In total 26 fulfilled these criteria of whom 17 were treated with orally administered rivastigmine with a total dose between 3 and 12 mg a day. Eight patients could not be treated because of dysphagia and one because of early discharge. Results No major side effects were recorded. In 16 patients there was a considerable decrease in severity of delirium. The mean DRS declined from 14.8 on day one to 8.5 after therapy and 5.6 after tapering. The mean duration of delirium was 6.7 days (range; 2–17). Conclusion Rivastigmine is safe in stroke patients with delirium even after rapid titration. In the majority of patients the delirium improved after treatment. A randomized controlled trial is needed to establish the usefulness of rivastigmine in delirium after stroke. Trial registration Nederlands Trial Register NTR1395

Background: The increasing burden of dengue virus on public health due to more explosive and frequent outbreaks highlights the need for improved surveillance and control. Genomic surveillance of dengue virus not only provides important insights into the emergence and spread of genetically diverse serotypes and genotypes, but it is also critical to monitor the effectiveness of newly implemented control strategies. Here, we present DengueSeq, an amplicon sequencing protocol, which enables whole-genome sequencing of all four dengue virus serotypes., Results: We developed primer schemes for the four dengue virus serotypes, which can be combined into a pan-serotype approach. We validated both approaches using genetically diverse virus stocks and clinical specimens that contained a range of virus copies. High genome coverage (>95%) was achieved for all genotypes, except DENV2 (genotype VI) and DENV 4 (genotype IV) sylvatics, with similar performance of the serotype-specific and pan-serotype approaches. The limit of detection to reach 70% coverage was 10-100 RNA copies/μL for all four serotypes, which is similar to other commonly used primer schemes. DengueSeq facilitates the sequencing of samples without known serotypes, allows the detection of multiple serotypes in the same sample, and can be used with a variety of library prep kits and sequencing instruments., Conclusions: DengueSeq was systematically evaluated with virus stocks and clinical specimens spanning the genetic diversity within each of the four dengue virus serotypes. The primer schemes can be plugged into existing amplicon sequencing workflows to facilitate the global need for expanded dengue virus genomic surveillance., (© 2024. The Author(s).)

Dengue is the most prevalent mosquito-borne viral disease in humans, and cases are continuing to rise globally. In particular, islands in the Caribbean have experienced more frequent outbreaks, and all four dengue virus (DENV) serotypes have been reported in the region, leading to hyperendemicity and increased rates of severe disease. However, there is significant variability regarding virus surveillance and reporting between islands, making it difficult to obtain an accurate understanding of the epidemiological patterns in the Caribbean. To investigate this, we used travel surveillance and genomic epidemiology to reconstruct outbreak dynamics, DENV serotype turnover, and patterns of spread within the region from 2009-2022. We uncovered two recent DENV-3 introductions from Asia, one of which resulted in a large outbreak in Cuba, which was previously under-reported. We also show that while outbreaks can be synchronized between islands, they are often caused by different serotypes. Our study highlights the importance of surveillance of infected travelers to provide a snapshot of local introductions and transmission in areas with limited local surveillance and suggests that the recent DENV-3 introductions may pose a major public health threat in the region., (© 2024. The Author(s).)

During May 2022-April 2023, dengue virus serotype 3 was identified among 601 travel-associated and 61 locally acquired dengue cases in Florida, USA. All 203 sequenced genomes belonged to the same genotype III lineage and revealed potential transmission chains in which most locally acquired cases occurred shortly after introduction, with little sustained transmission.

Background: The use of meta-analysis to aggregate the results of multiple studies has increased dramatically over the last 40 years. For homogeneous meta-analysis, the Mantel-Haenszel technique has typically been utilized. In such meta-analyses, the effect size across the contributing studies of the meta-analysis differs only by statistical error. If homogeneity cannot be assumed or established, the most popular technique developed to date is the inverse-variance DerSimonian and Laird (DL) technique (DerSimonian and Laird, in Control Clin Trials 7(3):177-88, 1986). However, both of these techniques are based on large sample, asymptotic assumptions. At best, they are approximations especially when the number of cases observed in any cell of the corresponding contingency tables is small., Results: This research develops an exact, non-parametric test for evaluating statistical significance and a related method for estimating effect size in the meta-analysis of k 2 × 2 tables for any level of heterogeneity as an alternative to the asymptotic techniques. Monte Carlo simulations show that even for large values of heterogeneity, the Enhanced Bernoulli Technique (EBT) is far superior at maintaining the pre-specified level of Type I Error than the DL technique. A fully tested implementation in the R statistical language is freely available from the author. In addition, a second related exact test for estimating the Effect Size was developed and is also freely available., Conclusions: This research has developed two exact tests for the meta-analysis of dichotomous, categorical data. The EBT technique was strongly superior to the DL technique in maintaining a pre-specified level of Type I Error even at extremely high levels of heterogeneity. As shown, the DL technique demonstrated many large violations of this level. Given the various biases towards finding statistical significance prevalent in epidemiology today, a strong focus on maintaining a pre-specified level of Type I Error would seem critical. In addition, a related exact method for estimating the Effect Size was developed., (© 2021. The Author(s).)

Introduction: Whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can be detected in semen and transmitted sexually is a vital question that has, thus far, been inconclusive. Prior studies, with limited numbers, have included men in various stages of infection with most in the recovery phase of the illness. The timing of test results and severity of illness has made recruiting study participants a significant challenge. Our pilot study will examine semen from men with a recent diagnosis of COVID-19 as well as those in the convalescent phase to determine if SARS-CoV-2 can be detected and its relationship, if any, with the severity of the disease., Methods: Eighteen men with a median age of 32 (range, 24-57) who tested positive for COVID-19 by rt-PCR analysis were enrolled and provided a semen sample. The study group demonstrated symptoms of COVID-19 ranging from asymptomatic to moderate and none required hospitalization. Samples were subjected to viral RNA extraction and then processed by real-time RT-PCR using the US Centers for Disease Control and Prevention (CDC, USA) panel of 2019-Novel Coronavirus (2019-nCoV) primers and probes to detect the presence of SARS-CoV-2 RNA., Results: Length of time from diagnosis to providing a specimen ranged from 1 to 28 days (median, 6 days). Fifteen participants were symptomatic and three were asymptomatic, including recovering men, at the time of semen collection. No SARS-CoV-2 was detected in any of the semen samples., Conclusion: Based on these preliminary results and consistent with prior findings, we suggest SARS-CoV-2 is not present in semen during the acute or convalescent phase of COVID-19.

Metagenomic sequencing has the potential to transform microbial detection and characterization, but new tools are needed to improve its sensitivity. Here we present CATCH, a computational method to enhance nucleic acid capture for enrichment of diverse microbial taxa. CATCH designs optimal probe sets, with a specified number of oligonucleotides, that achieve full coverage of, and scale well with, known sequence diversity. We focus on applying CATCH to capture viral genomes in complex metagenomic samples. We design, synthesize, and validate multiple probe sets, including one that targets the whole genomes of the 356 viral species known to infect humans. Capture with these probe sets enriches unique viral content on average 18-fold, allowing us to assemble genomes that could not be recovered without enrichment, and accurately preserves within-sample diversity. We also use these probe sets to recover genomes from the 2018 Lassa fever outbreak in Nigeria and to improve detection of uncharacterized viral infections in human and mosquito samples. The results demonstrate that CATCH enables more sensitive and cost-effective metagenomic sequencing.

How viruses evolve within hosts can dictate infection outcomes; however, reconstructing this process is challenging. We evaluate our multiplexed amplicon approach, PrimalSeq, to demonstrate how virus concentration, sequencing coverage, primer mismatches, and replicates influence the accuracy of measuring intrahost virus diversity. We develop an experimental protocol and computational tool, iVar, for using PrimalSeq to measure virus diversity using Illumina and compare the results to Oxford Nanopore sequencing. We demonstrate the utility of PrimalSeq by measuring Zika and West Nile virus diversity from varied sample types and show that the accumulation of genetic diversity is influenced by experimental and biological systems.

Background: The use of meta-analysis to aggregate multiple studies has increased dramatically over the last 30 years. For meta-analysis of homogeneous data where the effect sizes for the studies contributing to the meta-analysis differ only by statistical error, the Mantel-Haenszel technique has typically been utilized. If homogeneity cannot be assumed or established, the most popular technique is the inverse-variance DerSimonian-Laird technique. However, both of these techniques are based on large sample, asymptotic assumptions and are, at best, an approximation especially when the number of cases observed in any cell of the corresponding contingency tables is small., Results: This paper develops an exact, non-parametric test based on a maximum likelihood test statistic as an alternative to the asymptotic techniques. Further, the test can be used across a wide range of heterogeneity. Monte Carlo simulations show that for the homogeneous case, the ML-NP-EXACT technique to be generally more powerful than the DerSimonian-Laird inverse-variance technique for realistic, smaller values of disease probability, and across a large range of odds ratios, number of contributing studies, and sample size. Possibly most important, for large values of heterogeneity, the pre-specified level of Type I Error is much better maintained by the ML-NP-EXACT technique relative to the DerSimonian-Laird technique. A fully tested implementation in the R statistical language is freely available from the author., Conclusions: This research has developed an exact test for the meta-analysis of dichotomous data. The ML-NP-EXACT technique was strongly superior to the DerSimonian-Laird technique in maintaining a pre-specified level of Type I Error. As shown, the DerSimonian-Laird technique demonstrated many large violations of this level. Given the various biases towards finding statistical significance prevalent in epidemiology today, a strong focus on maintaining a pre-specified level of Type I Error would seem critical.

Oncogenic transcription factor FOXQ1 has been implicated in promotion of multiple transformed phenotypes in carcinoma cells. Recently, we have characterized FOXQ1 as a melanoma tumor suppressor that acts via repression of N-cadherin gene, and invasion and metastasis. Here we report that FOXQ1 induces differentiation in normal and transformed melanocytic cells at least partially via direct transcriptional activation of MITF gene, melanocytic lineage-specific regulator of differentiation. Importantly, we demonstrate that pigmentation induced in cultured melanocytic cells and in mice by activation of cAMP/CREB1 pathway depends in large part on FOXQ1. Moreover, our data reveal that FOXQ1 acts as a critical mediator of BRAF V600E -dependent regulation of MITF levels, thus providing a novel link between two major signal transduction pathways controlling MITF and differentiation in melanocytic cells.

Zika virus (ZIKV) is causing an unprecedented epidemic linked to severe congenital abnormalities. In July 2016, mosquito-borne ZIKV transmission was reported in the continental United States; since then, hundreds of locally acquired infections have been reported in Florida. To gain insights into the timing, source, and likely route(s) of ZIKV introduction, we tracked the virus from its first detection in Florida by sequencing ZIKV genomes from infected patients and Aedes aegypti mosquitoes. We show that at least 4 introductions, but potentially as many as 40, contributed to the outbreak in Florida and that local transmission is likely to have started in the spring of 2016-several months before its initial detection. By analysing surveillance and genetic data, we show that ZIKV moved among transmission zones in Miami. Our analyses show that most introductions were linked to the Caribbean, a finding corroborated by the high incidence rates and traffic volumes from the region into the Miami area. Our study provides an understanding of how ZIKV initiates transmission in new regions.

Although the recent Zika virus (ZIKV) epidemic in the Americas and its link to birth defects have attracted a great deal of attention, much remains unknown about ZIKV disease epidemiology and ZIKV evolution, in part owing to a lack of genomic data. Here we address this gap in knowledge by using multiple sequencing approaches to generate 110 ZIKV genomes from clinical and mosquito samples from 10 countries and territories, greatly expanding the observed viral genetic diversity from this outbreak. We analysed the timing and patterns of introductions into distinct geographic regions; our phylogenetic evidence suggests rapid expansion of the outbreak in Brazil and multiple introductions of outbreak strains into Puerto Rico, Honduras, Colombia, other Caribbean islands, and the continental United States. We find that ZIKV circulated undetected in multiple regions for many months before the first locally transmitted cases were confirmed, highlighting the importance of surveillance of viral infections. We identify mutations with possible functional implications for ZIKV biology and pathogenesis, as well as those that might be relevant to the effectiveness of diagnostic tests.

For decades, human infections with Zika virus (ZIKV), a mosquito-transmitted flavivirus, were sporadic, associated with mild disease, and went underreported since symptoms were similar to other acute febrile diseases. Recent reports of severe disease associated with ZIKV have greatly heightened awareness. It is anticipated that ZIKV will continue to spread in the Americas and globally where competent Aedes mosquito vectors are found. Dengue virus (DENV), the most common mosquito-transmitted human flavivirus, is both well-established and the source of outbreaks in areas of recent ZIKV introduction. DENV and ZIKV are closely related, resulting in substantial antigenic overlap. Through antibody-dependent enhancement (ADE), anti-DENV antibodies can enhance the infectivity of DENV for certain classes of immune cells, causing increased viral production that correlates with severe disease outcomes. Similarly, ZIKV has been shown to undergo ADE in response to antibodies generated by other flaviviruses. We tested the neutralizing and enhancing potential of well-characterized broadly neutralizing human anti-DENV monoclonal antibodies (HMAbs) and human DENV immune sera against ZIKV using neutralization and ADE assays. We show that anti-DENV HMAbs, cross-react, do not neutralize, and greatly enhance ZIKV infection in vitro . DENV immune sera had varying degrees of neutralization against ZIKV and similarly enhanced ZIKV infection. Our results suggest that pre-existing DENV immunity may enhance ZIKV infection in vivo and may lead to increased disease severity. Understanding the interplay between ZIKV and DENV will be critical in informing public health responses and will be particularly valuable for ZIKV and DENV vaccine design and implementation strategies.