Assay for and replication of Karshi (mammalian tick-borne flavivirus group) virus in mice

Little is known about the replication of Karshi virus, a member of the mammalian tick-borne flavivirus group, in its rodent hosts. Therefore, we developed a quantitative real-time RT-PCR assay and measured the amount of viral RNA in selected tissues of infected Swiss Webster mice. Two-day-old mice were highly susceptible, with 100% fatality 9 to 12 days after infection, whereas infection in 9-day-old mice was less virulent, with death occurring only rarely. In nearly all cases, mice inoculated when 2 days old contained similar numbers of viral genome equivalents from blood and liver samples from any given mouse, with titers declining after day 7. In contrast, the amount of viral RNA in the brain began to rise rapidly 4 days after exposure, peaked at about 6 days after virus exposure (titer of \u3e 1013 genome equivalents/g), and remained at that level until euthanasia or death. Viral profiles were similar in needle-inoculated or tick-exposed mice. Copyright © 2008 by The American Society of Tropical Medicine and Hygiene

Rapid design and fielding of four diagnostic technologies in Sierra Leone, Thailand, Peru, and Australia: Successes and challenges faced introducing these biosensors

Febrile illnesses are among the most common reasons for visits to hospitals and clinics worldwide. Since fevers can arise from a wide range of diseases, identifying the causative pathogen is essential not only for effective personal treatment but also for early detection of outbreaks. The Defense Threat Reduction Agency (DTRA) tasked a coalition of commercial, academic, and government researchers with moving diagnostic technology concepts from ideation to field use as rapidly as possible using scientifically sound evaluations. DTRA's 24 Month Challenge program examined >30 technologies before fielding four technologies on four continents. >10,000 in field test results were recorded. Here we discuss our tiered evaluation system to assess candidate technologies developed by commercial partners and the process of field testing those technologies at various front-line clinics in Sierra Leone, Thailand, Peru, and Australia. We discuss successes and challenges for introducing two multiplexed lateral flow immunoassay (LFI) tests that detect malaria, dengue fever, melioidosis, and the plague. Additionally we discuss the use of a LFI reader that assisted the interpretation of the assay, communicated results to a data cloud, and greatly facilitated reach-back support. Lastly, we discuss the concurrent field testing of a multiplexed PCR assay on the FilmArray platform, which had an assay pouch specially designed for the 24 Month Challenge. Either standard-of-care or gold-standard testing were run alongside our fielded technologies to benchmark their performance. Keywords: Diagnostic device, Lateral flow immunoassay, Nested PCR, Field testing, Malaria, Dengue feve

U-BIOPRED clinical adult asthma clusters linked to a subset of sputum omics

Background: Asthma is a heterogeneous disease in which there is a differential response to asthma treatments. This heterogeneity needs to be evaluated so that a personalized management approach can be provided. Objectives: We stratified patients with moderate-to-severe asthma based on clinicophysiologic parameters and performed an omics analysis of sputum. Methods: Partition-around-medoids clustering was applied to a training set of 266 asthmatic participants from the European Unbiased Biomarkers for the Prediction of Respiratory Diseases Outcomes (U-BIOPRED) adult cohort using 8 prespecified clinic-physiologic variables. This was repeated in a separate validation set of 152 asthmatic patients. The clusters were compared based on sputum proteomics and transcriptomics data. Results: Four reproducible and stable clusters of asthmatic patients were identified. The training set cluster T1 consists of patients with well-controlled moderate-to-severe asthma, whereas cluster T2 is a group of patients with late-onset severe asthma with a history of smoking and chronic airflow obstruction. Cluster T3 is similar to cluster T2 in terms of chronic airflow obstruction but is composed of nonsmokers. Cluster T4 is predominantly composed of obese female patients with uncontrolled severe asthma with increased exacerbations but with normal lung function. The validation set exhibited similar clusters, demonstrating reproducibility of the classification. There were significant differences in sputum proteomics and transcriptomics between the clusters. The severe asthma clusters (T2, T3, and T4) had higher sputum eosinophilia than cluster T1, with no differences in sputum neutrophil counts and exhaled nitric oxide and serum IgE levels. Conclusion: Clustering based on clinicophysiologic parameters yielded 4 stable and reproducible clusters that associate with different pathobiological pathways