Specimen sharing for epidemic preparedness: Building a virtual biorepository system from local governance to global partnerships.

We present a framework for a federated, virtual biorepository system (VBS) with locally collected and managed specimens, as a 'global public good' model based on principles of equitable access and benefit sharing. The VBS is intended to facilitate timely access to biological specimens and associated data for outbreak-prone infectious diseases to accelerate the development and evaluation of diagnostics, assess vaccine efficacy, and to support surveillance and research needs. The VBS is aimed to be aligned with the WHO BioHub and other specimen sharing efforts as a force multiplier to meet the needs of strengthening global tools for countering epidemics. The purpose of our initial research is to lay the basis of the collaboration, management and principles of equitable sharing focused on low- and middle-income country partners. Here we report on surveys and interviews undertaken with biorepository-interested parties to better understand needs and barriers for specimen access and share examples from the ZIKAlliance partnership on the governance and operations of locally organized biorepositories.

Early diagnostic indicators of dengue versus other febrile illnesses in Asia and Latin America (IDAMS study): a multicentre, prospective, observational study.

BACKGROUND: Improvements in the early diagnosis of dengue are urgently needed, especially in resource-limited settings where the distinction between dengue and other febrile illnesses is crucial for patient management. METHODS: In this prospective, observational study (IDAMS), we included patients aged 5 years and older with undifferentiated fever at presentation from 26 outpatient facilities in eight countries (Bangladesh, Brazil, Cambodia, El Salvador, Indonesia, Malaysia, Venezuela, and Viet Nam). We used multivariable logistic regression to investigate the association between clinical symptoms and laboratory tests with dengue versus other febrile illnesses between day 2 and day 5 after onset of fever (ie, illness days). We built a set of candidate regression models including clinical and laboratory variables to reflect the need of a comprehensive versus parsimonious approach. We assessed performance of these models via standard measures of diagnostic values. FINDINGS: Between Oct 18, 2011, and Aug 4, 2016, we recruited 7428 patients, of whom 2694 (36%) were diagnosed with laboratory-confirmed dengue and 2495 (34%) with (non-dengue) other febrile illnesses and met inclusion criteria, and were included in the analysis. 2703 (52%) of 5189 included patients were younger than 15 years, 2486 (48%) were aged 15 years or older, 2179 (42%) were female and 3010 (58%) were male. Platelet count, white blood cell count, and the change in these variables from the previous day of illness had a strong association with dengue. Cough and rhinitis had strong associations with other febrile illnesses, whereas bleeding, anorexia, and skin flush were generally associated with dengue. Model performance increased between day 2 and 5 of illness. The comprehensive model (18 clinical and laboratory predictors) had sensitivities of 0·80 to 0·87 and specificities of 0·80 to 0·91, whereas the parsimonious model (eight clinical and laboratory predictors) had sensitivities of 0·80 to 0·88 and specificities of 0·81 to 0·89. A model that includes laboratory markers that are easy to measure (eg, platelet count or white blood cell count) outperformed the models based on clinical variables only. INTERPRETATION: Our results confirm the important role of platelet and white blood cell counts in diagnosing dengue, and the importance of serial measurements over subsequent days. We successfully quantified the performance of clinical and laboratory markers covering the early period of dengue. Resulting algorithms performed better than published schemes for distinction of dengue from other febrile illnesses, and take into account the dynamic changes over time. Our results provide crucial information needed for the update of guidelines, including the Integrated Management of Childhood Illness handbook. FUNDING: EU's Seventh Framework Programme. TRANSLATIONS: For the Bangla, Bahasa Indonesia, Portuguese, Khmer, Spanish and Vietnamese translations of the abstract see Supplementary Materials section.

Diagnostic performance of anti-Zika virus IgM, IgAM and IgG ELISAs during co-circulation of Zika, dengue, and chikungunya viruses in Brazil and Venezuela.

BACKGROUND: Serological diagnosis of Zika virus (ZIKV) infection is challenging because of the antibody cross-reactivity among flaviviruses. At the same time, the role of Nucleic Acid Testing (NAT) is limited by the low proportion of symptomatic infections and the low average viral load. Here, we compared the diagnostic performance of commercially available IgM, IgAM, and IgG ELISAs in sequential samples during the ZIKV and chikungunya (CHIKV) epidemics and co-circulation of dengue virus (DENV) in Brazil and Venezuela. METHODOLOGY/PRINCIPAL FINDINGS: Acute (day of illness 1-5) and follow-up (day of illness ≥ 6) blood samples were collected from nine hundred and seven symptomatic patients enrolled in a prospective multicenter study between June 2012 and August 2016. Acute samples were tested by RT-PCR for ZIKV, DENV, and CHIKV. Acute and follow-up samples were tested for IgM, IgAM, and IgG antibodies to ZIKV using commercially available ELISAs. Among follow-up samples with a RT-PCR confirmed ZIKV infection, anti-ZIKV IgAM sensitivity was 93.5% (43/46), while IgM and IgG exhibited sensitivities of 30.3% (10/33) and 72% (18/25), respectively. An additional 24% (26/109) of ZIKV infections were detected via IgAM seroconversion in ZIKV/DENV/CHIKV RT-PCR negative patients. The specificity of anti-ZIKV IgM was estimated at 93% and that of IgAM at 85%. CONCLUSIONS/SIGNIFICANCE: Our findings exemplify the challenges of the assessment of test performance for ZIKV serological tests in the real-world setting, during co-circulation of DENV, ZIKV, and CHIKV. However, we can also demonstrate that the IgAM immunoassay exhibits superior sensitivity to detect ZIKV RT-PCR confirmed infections compared to IgG and IgM immunoassays. The IgAM assay also proves to be promising for detection of anti-ZIKV seroconversions in sequential samples, both in ZIKV PCR-positive as well as PCR-negative patients, making this a candidate assay for serological monitoring of pregnant women in future ZIKV outbreaks.

Study protocol for the multicentre cohorts of Zika virus infection in pregnant women, infants, and acute clinical cases in Latin America and the Caribbean: the ZIKAlliance consortium.

BACKGROUND: The European Commission (EC) Horizon 2020 (H2020)-funded ZIKAlliance Consortium designed a multicentre study including pregnant women (PW), children (CH) and natural history (NH) cohorts. Clinical sites were selected over a wide geographic range within Latin America and the Caribbean, taking into account the dynamic course of the ZIKV epidemic. METHODS: Recruitment to the PW cohort will take place in antenatal care clinics. PW will be enrolled regardless of symptoms and followed over the course of pregnancy, approximately every 4 weeks. PW will be revisited at delivery (or after miscarriage/abortion) to assess birth outcomes, including microcephaly and other congenital abnormalities according to the evolving definition of congenital Zika syndrome (CZS). After birth, children will be followed for 2 years in the CH cohort. Follow-up visits are scheduled at ages 1-3, 4-6, 12, and 24 months to assess neurocognitive and developmental milestones. In addition, a NH cohort for the characterization of symptomatic rash/fever illness was designed, including follow-up to capture persisting health problems. Blood, urine, and other biological materials will be collected, and tested for ZIKV and other relevant arboviral diseases (dengue, chikungunya, yellow fever) using RT-PCR or serological methods. A virtual, decentralized biobank will be created. Reciprocal clinical monitoring has been established between partner sites. Substudies of ZIKV seroprevalence, transmission clustering, disabilities and health economics, viral kinetics, the potential role of antibody enhancement, and co-infections will be linked to the cohort studies. DISCUSSION: Results of these large cohort studies will provide better risk estimates for birth defects and other developmental abnormalities associated with ZIKV infection including possible co-factors for the variability of risk estimates between other countries and regions. Additional outcomes include incidence and transmission estimates of ZIKV during and after pregnancy, characterization of short and long-term clinical course following infection and viral kinetics of ZIKV. STUDY REGISTRATIONS: clinicaltrials.gov NCT03188731 (PW cohort), June 15, 2017; clinicaltrials.gov NCT03393286 (CH cohort), January 8, 2018; clinicaltrials.gov NCT03204409 (NH cohort), July 2, 2017.

Spatial Dynamics of Chikungunya Virus, Venezuela, 2014.

Since chikungunya virus emerged in the Caribbean region in late 2013, ≈45 countries have experienced chikungunya outbreaks. We described and quantified the spatial and temporal events after the introduction and propagation of chikungunya into an immunologically naive population from the urban north-central region of Venezuela during 2014. The epidemic curve (n = 810 cases) unraveled within 5 months with a basic reproductive number of 3.7 and a radial spread traveled distance of 9.4 km at a mean velocity of 82.9 m/day. The highest disease diffusion speed occurred during the first 90 days, and space and space-time modeling suggest the epidemic followed a particular geographic pathway with spatiotemporal aggregation. The directionality and heterogeneity of transmission during the first introduction of chikungunya indicated existence of areas of diffusion and elevated risk for disease and highlight the importance of epidemic preparedness. This information will help in managing future threats of new or reemerging arboviruses.

Applied shotgun metagenomics approach for the genetic characterization of dengue viruses.

Dengue virus (DENV), an arthropod-borne virus, has rapidly spread in recent years. DENV diagnosis is performed through virus serology, isolation or molecular detection, while genotyping is usually done through Sanger sequencing of the envelope gene. This study aimed to optimize the use of shotgun metagenomics and subsequent bioinformatics analysis to detect and type DENV directly from clinical samples without targeted amplification. Additionally, presence of DENV quasispecies (intra-host variation) was revealed by detecting single nucleotide variants. Viral RNA was isolated with or without DNase-I treatment from 17 DENV (1-4) positive blood samples. cDNA libraries were generated using either a combination of the NEBNext® RNA to synthesize cDNA followed by Nextera XT DNA library preparation, or the TruSeq RNA V2 (TS) library preparation kit. Libraries were sequenced using both the MiSeq and NextSeq. Bioinformatic analysis showed complete ORFs for all samples by all approaches, but longer contigs and higher sequencing depths were obtained with the TS kit. No differences were observed between MiSeq and NextSeq sequencing. Detection of multiple DENV serotypes in a single sample was feasible. Finally, results were obtained within three days with associated reagents costs between €130-170/sample. Therefore, shotgun metagenomics is suitable for identification and typing of DENV in a clinical setting.

Clinical evaluation of dengue and identification of risk factors for severe disease: protocol for a multicentre study in 8 countries.

BACKGROUND: The burden of dengue continues to increase globally, with an estimated 100 million clinically apparent infections occurring each year. Although most dengue infections are asymptomatic, patients can present with a wide spectrum of clinical symptoms ranging from mild febrile illness through to severe manifestations of bleeding, organ impairment, and hypovolaemic shock due to a systemic vascular leak syndrome. Clinical diagnosis of dengue and identification of which patients are likely to develop severe disease remain challenging. This study aims to improve diagnosis and clinical management through approaches designed a) to differentiate between dengue and other common febrile illness within 72 h of fever onset, and b) among patients with dengue to identify markers that are predictive of the likelihood of evolving to a more severe disease course. METHOD/DESIGN: This is a prospective multi-centre observational study aiming to enrol 7-8000 participants aged ≥ 5 years presenting with a febrile illness consistent with dengue to outpatient health facilities in 8 countries across Asia and Latin America. Patients presenting within 72 h of fever onset who do not exhibit signs of severe disease are eligible for the study. A broad range of clinical and laboratory parameters are assessed daily for up to 6 days during the acute illness, and also at a follow up visit 1 week later. DISCUSSION: Data from this large cohort of patients, enrolled early with undifferentiated fever, will be used to develop a practical diagnostic algorithm and a robust clinical case definition for dengue. Additionally, among patients with confirmed dengue we aim to identify simple clinical and laboratory parameters associated with progression to a more severe disease course. We will also investigate early virological and serological correlates of severe disease, and examine genetic associations in this large heterogeneous cohort. In addition the results will be used to assess the new World Health Organization classification scheme for dengue in practice, and to update the guidelines for "Integrated Management of Childhood Illness" used in dengue-endemic countries. TRIAL REGISTRATION: NCT01550016. Registration Date: March 7, 2012.

The effect of montelukast in a model of gouty arthritis induced by sodium monourate crystals.

Non-steroidal anti-inflammatory drugs (NSAIDS) are the first line of therapy in acute gouty arthritis. NSAIDs inhibit the cyclooxygenase pathway, but not the lipooxygenase activity and can have many adverse effects and thus have a limited effect on the control of inflammation in this disease. In this work we studied the effect of montelukast on the cellular inflammatory infiltrate in a model of murine arthritis induced by sodium monourate crystals (SMU), using a subcutaneous air cavity (air pouch) in BALB/c mice. Seven groups of BALB/c mice (n = 4) were distributed into five experimental groups and two inflammatory control groups, a positive and a negative one. Previous to SMU exposure, the experimental groups received montelukast (1 and 0.01 mg/Kg/w) and/or indomethacine (2.5 mg/Kg/w), followed by administration of SMU in the air pouch. The total and differential counts of inflammatory cells were analyzed after 2, 6, 12 and 24 hours. Montelukast, significantly reduced the total number of cells (p < 0.05), with a predominant impact on polymorphonuclear over mononuclear cells, especially after 12 hours of the medication. The montelukast/indometacine combination showed an additive effect. Our data show that montelukast has an anti-inflammatory effect in the model of gouty arthritis. Consequently, anti-leukotrienes could represent a new and effective therapy, either isolated or combined with conventional therapy of gouty arthritis.

The effect of montelukast in a model of gouty arthritis induced by sodium monourate crystals / Efecto de montelukast en un modelo de artritis gotosa inducido por cristales de monourato de sodio

Non-steroidal anti-inflammatory drugs (NSAIDS) are the first line of therapy in acute gouty arthritis. NSAIDs inhibit the cyclooxygenase pathway, but not the lipooxygenase activity and can have many adverse effects and thus have a limited effect on the control of inflammation in this disease. In this work we studied the effect of montelukast on the cellular inflammatory infiltrate in a model of murine arthritis induced by sodium monourate crystals (SMU), using a subcutaneous air cavity (air pouch) in BALB/c mice. Seven groups of BALB/c mice (n = 4) were distributed into five experimental groups and two inflammatory control groups, a positive and a negative one. Previous to SMU exposure, the experimental groups received montelukast (1 and 0.01 mg/Kg/w) and/or indomethacine (2.5 mg/Kg/w), followed by administration of SMU in the air pouch. The total and differential counts of inflammatory cells were analyzed after 2, 6, 12 and 24 hours. Montelukast, significantly reduced the total number of cells (p<0.05), with a predominant impact on polymorphonuclear over mononuclear cells, especially after 12 hours of the medication. The montelukast/indometacine combination showed an additive effect. Our data show that montelukast has an anti-inflammatory effect in the model of gouty arthritis. Consequently, anti-leukotrienes could represent a new and effective therapy, either isolated or combined with conventional therapy of gouty arthritis.

En artritis gotosa aguda las drogas antiinflamatorias no esteroideas son la primera línea terapéutica. Este tratamiento no es satisfactorio porque inhibe la ciclooxigenasa sin modificar la actividad de la lipooxigenasa, y puede acompañarse de numerosos efectos adversos. Investigamos el efecto de montelukast sobre el infiltrado celular inflamatorio en un modelo de artritis múrida inducida por cristales de monourato de sodio (MUS) en el modelo experimental de la bolsa de aire (air pouch). Siete grupos de ratones BALB/c (n = 4) fueron distribuidos en cinco grupos experimentales y dos grupos controles inflamatorios: positivo y negativo. Los grupos experimentales recibieron, montelukast (1 y 0,01 mg/Kg/p) y/o indometacina (2,5 mg/Kg/p) por vía oral, previo a la administración de MUS en la bolsa del aire. El conteo absoluto y diferencial de las células inflamatorias fue analizado después de 2, 6, 12 y 24 horas de tratamiento. El tratamiento con montelukast redujo significativamente el número total de células presentes en el infiltrado inflamatorio (p < 0,05), con un efecto mayor sobre polimorfonucleares que sobre las células mononucleares, y con un máximo efecto a las 12 horas después de la administración del medicamento. La combinación montelukast/indometacina mostró un efecto aditivo. Los resultados demuestran que montelukast tiene un efecto antiinflamatorio en el modelo de la artritis gotosa. Por lo tanto, los anti-leucotrienos podrían representar una nueva y eficaz terapia, aislada o en combinación con la terapéutica convencional, para la artritis gotosa.

[Clinical and pathological features in Venezuelan children with IgA nephropathy]. / Características clínico patológicas y evolución de la nefropatía por IgA en una serie hospitalaria de niños venezolanos.

IgA nephropathy (IgAN) is the most common glomerulonephritis in humans worldwide; its prevalence and prognosis vary according with geographical areas. The incidence is higher in adults under 30 years of age and in children, it occurs more frequently in patients between 3 and 10 years. Hematuria is the predominant manifestation at presentation of the disease and 20-40% of the cases progress to terminal chronic renal disease. Renal biopsies were performed in 426 children during the period 1980-2002, of them, 12 cases corresponded to IgAN. The clinico-pathological characteristics and evolution of patients were evaluated during an average of 3.85 years. Mean age of patients was 6.2 years, and it was more frequent in males. Hematuria and proteinuria were found in 100% of cases and proteinuria of nephrotic range in 75%. Hypertriglyceridemia and hypercholesterolemia in 91%, arterial hypertension in 50% and acute renal failure at presentation in 25%. The predominant histopathological patterns (WHO) were II and III, deposits of mesangial IgA, IgG and C3 were observed in all cases and C4 deposits in 25%. 41.7% of cases had complete remission, 41.7% maintained normal renal function with persistent proteinuria and 16% progressed to terminal chronic renal failure. The actuarial survival of patients was 100% at 3 years, 87% at 4 years and 76% at 8 years. Two patients died during the period of study, at 3.5 and 8.5 years. The variability of presentation of IgA nephropathy was confirmed in this study, which could be attributable to geographical differences, racial influences and clinicopathological features related to sanitary conditions. Despite of the frequency of bad prognosis characteristics at presentation of IgAN in our series, the evolution was similar to reports of other groups.

Características clínico patológicas y evolución de la nefropatía por IgA en una serie hospitalaria de niños venezolanos / Clinical and pathological features in Venezuela children with IgA nephropathy

La nefropatía por IgA (NigA) es la enfermedad glomerular más común en humanos; su prevalencia y evolución varían según la región geográfica. Su incidencia es mayor en menores de 30 años y en niños ocurre más frecuentemente, entre los 3 y 10 años; la hematuria es la manifestación predominante de inicio y evolución a nefropatía terminal en 20 a 40 por ciento de los casos. En el presente estudio, de un total de 426 biopsias renales realizadas entre 1980 y 2002, 12 casos correspondieron a NIgA . Se evaluaron las características clínico-patológicas y evolución de los pacientes durante un promedio de 3,85 años. La edad promedio fue de 6,2 años, con predominio de varones. Cien por ciento de los casos presentó hematuria y proteinuria y 75 por ciento proteinuria de rango nefrótico. La hipertrigliceridemia e hipercolesterolemia se observó en 91 por ciento, hipertensión arterial en 50 por ciento en IRA, al inicio de la enfermedad, en 25 por ciento. Predominaron los grados histológicos (OMS) II y III, y se observaron depósitos mesangiales de IgA, IgG y C3 en todos los pacientes, y de C4 en 25 por ciento. Se observó remisión total en 41,7 por ciento, proteinuria persistente con función renal de filtración normal en 41,7 por ciento y 16,7 por ciento progreso a enfermedad renal crónica. La sobrevida actuarial de los pacientes fue de 100 por ciento a los 3 años, 87 por ciento a los cuatro años y 76 por ciento a los 8 años, con dos muertes durante el estudio, una a los 3,5 años y la otra a los 8,5 años, después del diagnóstico. Se confirmó la variabilidad en la forma de presentación de NIgA, que puede ser atribuida a diferencias geográficas y raciales y a factores clínico-patológicos relacionados con condiciones sanitarias. A pesar de la alta frecuencia de factores de mal pronóstico, como forma de inicio de NIgA, en este estudio la evolución fue similar a la reportada por otros grupos.