Journal articles: 'Flavivirus diagnosis'

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Wee, Sheena, Asfa Alli-Shaik, Relus Kek, Hannah L. F. Swa, Wei-Ping Tien, Vanessa W. Lim, Yee-Sin Leo, Lee-Ching Ng, Hapuarachchige C. Hapuarachchi, and Jayantha Gunaratne. "Multiplex targeted mass spectrometry assay for one-shot flavivirus diagnosis." Proceedings of the National Academy of Sciences 116, no. 14 (March 18, 2019): 6754–59. http://dx.doi.org/10.1073/pnas.1817867116.

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Targeted proteomic mass spectrometry is emerging as a salient clinical diagnostic tool to track protein biomarkers. However, its strong analytical properties have not been exploited in the diagnosis and typing of flaviviruses. Here, we report the development of a sensitive and specific single-shot robust assay for flavivirus typing and diagnosis using targeted mass spectrometry technology. Our flavivirus parallel reaction monitoring assay (fvPRM) has the ability to track secreted flaviviral nonstructural protein 1 (NS1) over a broad diagnostic and typing window with high sensitivity, specificity, extendibility, and multiplexing capability. These features, pivotal and pertinent to efficient response toward flavivirus outbreaks, including newly emerging flavivirus strains, circumvent the limitations of current diagnostic assays.fvPRM thus carries high potential in positioning itself as a forerunner in delivering early and accurate diagnosis for disease management.

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Musso, Didier, and Philippe Desprès. "Serological Diagnosis of Flavivirus-Associated Human Infections." Diagnostics 10, no. 5 (May 14, 2020): 302. http://dx.doi.org/10.3390/diagnostics10050302.

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Arthropod-borne viruses (arboviruses) belonging to the Flavivirus genus of the Flaviviridae family, are a major public health threat in tropical and subtropical regions, and have recently become a medical concern in temperate zones. Most flaviviruses are classified as zoonotic viruses. Human flavivirus infections can be asymptomatic, responsible for unspecific symptoms in the first few days following infection, or responsible for severe complications potentially resulting in death. During the first days following symptom onset, laboratory diagnosis of acute human flavivirus infection is mainly based on molecular detection of the viral genome by RT-PCR methods, followed by the capture of specific antibodies using serological tests after the first week of infection. The detection of antibodies that have virus neutralizing activity can be used to confirm flavivirus infection. However, human flavivirus infections induce the production of cross-reactive antibodies, often making serology inconclusive. Indeed, serological diagnosis of flavivirus infection can be hampered by a patient’s history of flavivirus exposure, particularly in regions where multiple antigenically related flaviviruses co-circulate. We focus our mini review on conventional immunoassays that allow the diagnosis of major flavivirus-associated human infections in basic, routine and high-profile central health centers; and the interpretation of diagnostic serology tests for patients living within different epidemiological situations.

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Thibodeaux, Brett A., and John T. Roehrig. "Development of a Human-Murine Chimeric Immunoglobulin M Antibody for Use in the Serological Detection of Human Flavivirus Antibodies." Clinical and Vaccine Immunology 16, no. 5 (March 18, 2009): 679–85. http://dx.doi.org/10.1128/cvi.00354-08.

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ABSTRACT Current diagnosis of human flaviviral infections relies heavily on serological techniques such as the immunoglobulin M (IgM) antibody capture enzyme-linked immunosorbent assay (MAC-ELISA). Broad application of this assay is hindered by a lack of standardized human positive-control sera that react with the wide variety of flaviviruses that can cause human disease, e.g., dengue virus (DENV), West Nile virus (WNV), yellow fever virus (YFV), Japanese encephalitis virus (JEV), and St. Louis encephalitis virus (SLEV). We have created a human-murine chimeric antibody combining the variable regions of the broadly flavivirus cross-reactive murine monoclonal antibody (MAb) 6B6C-1 and the constant region of human IgM to produce a standardized reagent capable of replacing human positive-control sera in a MAC-ELISA for the diagnosis of all human flaviviral infections. The human-murine chimeric IgM antibody secreted from plasmid-transformed Sp2/0-Ag14 cells had a level of serological activity identical to that of 6B6C-1 as measured by ELISA, immunoblotting, and MAC-ELISA for multiple members of the flavivirus genus, including WNV, SLEV, YFV, DENV, and JEV.

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Beck, Cécile, Philippe Desprès, Sylvie Paulous, Jessica Vanhomwegen, Steeve Lowenski, Norbert Nowotny, Benoit Durand, et al. "A High-Performance Multiplex Immunoassay for Serodiagnosis of Flavivirus-Associated Neurological Diseases in Horses." BioMed Research International 2015 (2015): 1–13. http://dx.doi.org/10.1155/2015/678084.

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West Nile virus (WNV), Japanese encephalitis virus (JEV), and tick-borne encephalitis virus (TBEV) are flaviviruses responsible for severe neuroinvasive infections in humans and horses. The confirmation of flavivirus infections is mostly based on rapid serological tests such as enzyme-linked immunosorbent assays (ELISAs). These tests suffer from poor specificity, mainly due to antigenic cross-reactivity among flavivirus members. Robust diagnosis therefore needs to be validated through virus neutralisation tests (VNTs) which are time-consuming and require BSL3 facilities. The flavivirus envelope (E) glycoprotein ectodomain is composed of three domains (D) named DI, DII, and DIII, with EDIII containing virus-specific epitopes. In order to improve the serological differentiation of flavivirus infections, the recombinant soluble ectodomain of WNV E (WNV.sE) and EDIIIs (rEDIIIs) of WNV, JEV, and TBEV were synthesised using theDrosophilaS2 expression system. Purified antigens were covalently bonded to fluorescent beads. The microspheres coupled to WNV.sE or rEDIIIs were assayed with about 300 equine immune sera from natural and experimental flavivirus infections and 172 nonimmune equine sera as negative controls. rEDIII-coupled microspheres captured specific antibodies against WNV, TBEV, or JEV in positive horse sera. This innovative multiplex immunoassay is a powerful alternative to ELISAs and VNTs for veterinary diagnosis of flavivirus-related diseases.

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Chiou, Shyan-Song, Wayne D. Crill, Li-Kuang Chen, and Gwong-Jen J. Chang. "Enzyme-Linked Immunosorbent Assays Using Novel Japanese Encephalitis Virus Antigen Improve the Accuracy of Clinical Diagnosis of Flavivirus Infections." Clinical and Vaccine Immunology 15, no. 5 (March 12, 2008): 825–35. http://dx.doi.org/10.1128/cvi.00004-08.

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ABSTRACT The cross-reactive antibodies induced by flavivirus infections confound serodiagnosis and pathogenesis, especially in secondary infections caused by antigenically closely related yet distinct flaviviruses. The envelope (E) glycoprotein fusion peptide contains immunodominant cross-reactive determinants. Using a recombinant Japanese encephalitis virus (JEV) premembrane and E expression plasmid producing JEV virus-like particles (VLPs), dramatic reductions in cross-reactivity were produced by the G106K-L107D (KD) double-mutant VLP against a panel of flavivirus murine monoclonal antibodies. Human serum panels from patients with recent flavivirus infections were analyzed to compare the accuracy of JEV wild-type (WT) and KD VLPs as serodiagnostic antigens in enzyme-linked immunosorbent assays. Statistical analysis demonstrated significant differences in assay performances for accurate determination of current JEV infections between WT and KD antigens by detecting immunoglobulin M antibodies at a serum dilution of 1:4,000 (likelihood ratios = 2.74 [WT] and 22 [KD]). The application and continued development of cross-reactivity-reduced antigens should improve both flavivirus infection serodiagnosis and estimates of disease burden.

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Thibodeaux, Brett A., Amanda N. Panella, and John T. Roehrig. "Development of Human-Murine Chimeric Immunoglobulin G for Use in the Serological Detection of Human Flavivirus and Alphavirus Antibodies." Clinical and Vaccine Immunology 17, no. 10 (August 25, 2010): 1617–23. http://dx.doi.org/10.1128/cvi.00097-10.

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ABSTRACT Diagnosis of human arboviral infections relies heavily on serological techniques such as the immunoglobulin M (IgM) antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) and the indirect IgG ELISA. Broad application of these assays is hindered by the lack of standardized positive human control sera that react with a wide variety of flaviviruses (e.g., dengue, West Nile, yellow fever, Japanese encephalitis, Saint Louis encephalitis, and Powassan viruses), or alphaviruses (e.g., Eastern equine encephalitis, Western equine encephalitis, Venezuelan equine encephalitis, and chikungunya viruses) that can cause human disease. We have created human-murine chimeric monoclonal antibodies (cMAbs) by combining the variable regions of flavivirus (6B6C-1) or alphavirus (1A4B-6) broadly cross-reactive murine MAbs (mMAbs) with the constant region of human IgG1. These cMAbs may be used as standardized reagents capable of replacing human infection-immune-positive control sera in indirect IgG ELISA for diagnosis of all human flaviviral or alphaviral infections. The IgG cMAbs secreted from plasmid-transformed Sp2/0-Ag14 cells had serological activity identical to that of the parent mMAbs, as measured by ELISA using multiple flaviviruses or alphaviruses.

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Khristunova, Ekaterina, Elena Dorozhko, Elena Korotkova, Bohumil Kratochvil, Vlastimil Vyskocil, and Jiri Barek. "Label-Free Electrochemical Biosensors for the Determination of Flaviviruses: Dengue, Zika, and Japanese Encephalitis." Sensors 20, no. 16 (August 16, 2020): 4600. http://dx.doi.org/10.3390/s20164600.

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A highly effective way to improve prognosis of viral infectious diseases and to determine the outcome of infection is early, fast, simple, and efficient diagnosis of viral pathogens in biological fluids. Among a wide range of viral pathogens, Flaviviruses attract a special attention. Flavivirus genus includes more than 70 viruses, the most familiar being dengue virus (DENV), Zika virus (ZIKV), and Japanese encephalitis virus (JEV). Haemorrhagic and encephalitis diseases are the most common severe consequences of flaviviral infection. Currently, increasing attention is being paid to the development of electrochemical immunological methods for the determination of Flaviviruses. This review critically compares and evaluates recent research progress in electrochemical biosensing of DENV, ZIKV, and JEV without labelling. Specific attention is paid to comparison of detection strategies, electrode materials, and analytical characteristics. The potential of so far developed biosensors is discussed together with an outlook for further development in this field.

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Taylor, Carmel, Russell Simmons, and Ina Smith. "Development of Immunoglobulin M Capture Enzyme-Linked Immunosorbent Assay To Differentiate Human Flavivirus Infections Occurring in Australia." Clinical Diagnostic Laboratory Immunology 12, no. 3 (March 2005): 371–74. http://dx.doi.org/10.1128/cdli.12.3.371-374.2005.

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ABSTRACT We report the development of a flavivirus immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay (MAC-ELISA) which improves the determination of an infecting flavivirus serotype over that by current serological methods. A panel of 165 IgM-positive sera from flavivirus patients with specific diagnostic results was tested by the flavivirus MAC-ELISA using a panel of 10 antigens. For 134 of these sera (81.2%), the highest reactivity was demonstrated against the infecting virus, which was consistent with the original diagnostic result. Specific antibody reactions inconsistent with the original diagnosis were found for six sera (3.6%). In our experience, the flavivirus-serotyping ELISA provides a rapid and accurate alternative to other serological tests, such as hemagglutination inhibition, for the specific diagnosis of flavivirus infections.

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Chu, Charleen T., David N. Howell, Joel C. Morgenlander, Christine M. Hulette, Roger E. McLendon, and Sara E. Miller. "Electron Microscopic Diagnosis of Human Flavivirus Encephalitis." American Journal of Surgical Pathology 23, no. 10 (October 1999): 1217. http://dx.doi.org/10.1097/00000478-199910000-00006.

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Landry, Marie Louise, and Kirsten St. George. "Laboratory Diagnosis of Zika Virus Infection." Archives of Pathology & Laboratory Medicine 141, no. 1 (October 20, 2016): 60–67. http://dx.doi.org/10.5858/arpa.2016-0406-sa.

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Context.—The rapid and accurate diagnosis of Zika virus infection is an international priority. Objective.—To review current recommendations, methods, limitations, and priorities for Zika virus testing. Data Sources.—Sources include published literature, public health recommendations, laboratory procedures, and testing experience. Conclusions.—Until recently, the laboratory diagnosis of Zika infection was confined to public health or research laboratories that prepared their own reagents, and test capacity has been limited. Furthermore, Zika cross-reacts serologically with other flaviviruses, such as dengue, West Nile, and yellow fever. Current or past infection, or even vaccination with another flavivirus, will often cause false-positive or uninterpretable Zika serology results. Detection of viral RNA during acute infection using nucleic acid amplification tests provides more specific results, and a number of commercial nucleic acid amplification tests have received emergency use authorization. In addition to serum, testing of whole blood and urine is recommended because of the higher vial loads and longer duration of shedding. However, nucleic acid amplification testing has limited utility because many patients are asymptomatic or present for testing after the brief period of Zika shedding has passed. Thus, the greatest need and most difficult challenge is development of accurate antibody tests for the diagnosis of recent Zika infection. Research is urgently needed to identify Zika virus epitopes that do not cross-react with other flavivirus antigens. New information is emerging at a rapid pace and, with ongoing public-private and international collaborations and government support, it is hoped that rapid progress will be made in developing robust and widely applicable diagnostic tools.

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Delfin-Riela, Triana, Martín Rossotti, Romina Alvez-Rosado, Carmen Leizagoyen, and Gualberto González-Sapienza. "Highly Sensitive Detection of Zika Virus Nonstructural Protein 1 in Serum Samples by a Two-Site Nanobody ELISA." Biomolecules 10, no. 12 (December 9, 2020): 1652. http://dx.doi.org/10.3390/biom10121652.

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The Zika virus was introduced in Brazil in 2015 and, shortly after, spread all over the Americas. Nowadays, it remains present in more than 80 countries and represents a major threat due to some singularities among other flaviviruses. Due to its easy transmission, high percentage of silent cases, the severity of its associated complications, and the lack of prophylactic methods and effective treatments, it is essential to develop reliable and rapid diagnostic tests for early containment of the infection. Nonstructural protein 1 (NS1), a glycoprotein involved in all flavivirus infections, is secreted since the beginning of the infection into the blood stream and has proven to be a valuable biomarker for the early diagnosis of other flaviviral infections. Here, we describe the development of a highly sensitive nanobody ELISA for the detection of the NS1 protein in serum samples. Nanobodies were selected from a library generated from a llama immunized with Zika NS1 (ZVNS1) by a two-step high-throughput screening geared to identify the most sensitive and specific nanobody pairs. The assay was performed with a sub-ng/mL detection limit in the sera and showed excellent reproducibility and accuracy when validated with serum samples spiked with 0.80, 1.60, or 3.10 ng/mL of ZVNS1. Furthermore, the specificity of the developed ELISA was demonstrated using a panel of flavivirus’ NS1 proteins; this is of extreme relevance in countries endemic for more than one flavivirus. Considering that the nanobody sequences are provided, the assay can be reproduced in any laboratory at low cost, which may help to strengthen the diagnostic capacity of the disease even in low-resource countries.

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Nunes, Jannyce G. C., Bruno T. D. Nunes, Chao Shan, Adriana F. Moraes, Tais R. Silva, Maria H. R. de Mendonça, Liliane L. das Chagas, et al. "Reporter Virus Neutralization Test Evaluation for Dengue and Zika Virus Diagnosis in Flavivirus Endemic Area." Pathogens 10, no. 7 (July 3, 2021): 840. http://dx.doi.org/10.3390/pathogens10070840.

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Reporter virus neutralization test (RVNT) has been used as an alternative to the more laborious and time-demanding conventional PRNT assay for both DENV and ZIKV. However, few studies have investigated how these techniques would perform in epidemic areas with the circulation of multiple flavivirus. Here, we evaluate the performance of ZIKV and DENV Rluc RVNT and ZIKV mCh RVNT assays in comparison to the conventional PRNT assay against patient sera collected before and during ZIKV outbreak in Brazil. These samples were categorized into groups based on (1) acute and convalescent samples according to the time of disease, and (2) laboratorial diagnostic results (DENV and ZIKV RT-PCR and IgM-capture ELISA). Our results showed that DENV Rluc assay presented 100% and 78.3% sensitivity and specificity, respectively, with 93.3% accuracy, a similar performance to the traditional PRNT. ZIKV RVNT90, on the other hand, showed much better ZIKV antibody detection performance (around nine-fold higher) when compared to PRNT, with 88% clinical sensitivity. Specificity values were on average 76.8%. Even with these results, however, ZIKV RVNT90 alone was not able to reach a final diagnostic conclusion for secondary infection in human samples due to flavivirus cross reaction. As such, in regions where the flavivirus differential diagnosis represents a challenge, we suggest the establishment of a RVNT panel including other flaviviruses circulating in the region, associated with the other serological techniques such as IgM ELISA and the investigation of seroconversion, in order to help define an accurate diagnostic conclusion using serology.

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Loroño-Pino, M. A., J. A. Farfan-Ale, B. J. Blitvich, J. L. Beebe, R. G. Jarman, and B. J. Beaty. "Evaluation of an Epitope-Blocking Enzyme-Linked Immunosorbent Assay for the Diagnosis of West Nile Virus Infections in Humans." Clinical and Vaccine Immunology 16, no. 5 (March 25, 2009): 749–55. http://dx.doi.org/10.1128/cvi.00393-08.

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ABSTRACT An epitope-blocking enzyme-linked immunosorbent assay (b-ELISA) was evaluated for the diagnosis of West Nile virus (WNV) infections in humans. Sera from patients diagnosed with WNV infections from an outbreak in 2003 in Colorado and from patients diagnosed with dengue virus infections from Mexico and Thailand were tested with the b-ELISA. The b-ELISAs were performed using the WNV-specific monoclonal antibody (MAb) 3.1112G and the flavivirus-specific MAb 6B6C-1. Although the WNV-specific b-ELISA was effective in diagnosing WNV infections in humans from Colorado, it was not efficacious for diagnosing WNV infections in serum specimens from Mexico and Thailand. In serum specimens from patients from Colorado, the WNV b-ELISA and the WNV plaque reduction neutralization test showed an overall agreement of 91%. The sensitivity and specificity of the WNV b-ELISA were 89% and 92%, respectively, with a false-positive rate of 5%, based on receiver operating characteristic analysis. In contrast, false-positive rate results in specimens from the countries of Mexico and Thailand, where flaviviruses are endemic, were 79% and 80%, presumably due to the presence of antibodies resulting from previous dengue virus infections in Mexico and/or Japanese encephalitis virus infections or vaccination in Thailand. Thus, in regions where people have experienced previous or multiple flavivirus infections, the use of the b-ELISA for WNV diagnosis is contraindicated.

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Wilson, Heather L., Thomas Tran, Julian Druce, Myrielle Dupont-Rouzeyrol, and Michael Catton. "Neutralization Assay for Zika and Dengue Viruses by Use of Real-Time-PCR-Based Endpoint Assessment." Journal of Clinical Microbiology 55, no. 10 (August 9, 2017): 3104–12. http://dx.doi.org/10.1128/jcm.00673-17.

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ABSTRACTThe global spread and infective complications of Zika virus (ZKV) and dengue virus (DENV) have made them flaviviruses of public health concern. Serological diagnosis can be challenging due to antibody cross-reactivity, particularly in secondary flavivirus infections or when there is a history of flavivirus vaccination. The virus neutralization assay is considered to be the most specific assay for measurement of anti-flavivirus antibodies. This study describes an assay where the neutralization endpoint is measured by real-time PCR, providing results within 72 h. It demonstrated 100% sensitivity (24/24 ZKV and 15/15 DENV) and 100% specificity (11/11 specimens) when testing well-characterized sera. In addition, the assay was able to determine the correct DENV serotype in 91.7% of cases. The high sensitivity and specificity of the real-time PCR neutralization assay makes it suitable to use as a confirmatory test for sera that are reactive in commercial IgM/IgG enzyme immunoassays. Results are objective and the PCR-based measurement of the neutralization endpoint lends itself to automation so that throughput may be increased in times of high demand.

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Stiasny, Karin, Stefan Malafa, Stephan W. Aberle, Iris Medits, Georgios Tsouchnikas, Judith H. Aberle, Heidemarie Holzmann, and Franz X. Heinz. "Different Cross-Reactivities of IgM Responses in Dengue, Zika and Tick-Borne Encephalitis Virus Infections." Viruses 13, no. 4 (March 31, 2021): 596. http://dx.doi.org/10.3390/v13040596.

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Flaviviruses circulate worldwide and cause a number of medically relevant human diseases, such as dengue, Zika, yellow fever, and tick-borne encephalitis (TBE). Serology plays an important role in the diagnosis of flavivirus infections, but can be impeded by antigenic cross-reactivities among flaviviruses. Therefore, serological diagnosis of a recent infection can be insufficiently specific, especially in areas where flaviviruses co-circulate and/or vaccination coverage against certain flaviviruses is high. In this study, we developed a new IgM assay format, which is well suited for the specific diagnosis of TBE, Zika and dengue virus infections. In the case of TBE and Zika, the IgM response proved to be highly specific for the infecting virus. In contrast, primary dengue virus infections induced substantial amounts of cross-reactive IgM antibodies, which is most likely explained by structural peculiarities of dengue virus particles. Despite the presence of cross-reactive IgM, the standardized nature and the quantitative read-out of the assay even allowed the serotype-specific diagnosis of recent dengue virus infections in most instances.

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Khongwichit, Sarawut, Sirikwan Libsittikul, Sutee Yoksan, Prasert Auewarakul, Yupin Suputtamongkol, and Duncan R. Smith. "Retrospective screening of acute undifferentiated fever serum samples with universal flavivirus primers." Journal of Infection in Developing Countries 9, no. 07 (July 30, 2015): 760–64. http://dx.doi.org/10.3855/jidc.5866.

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Introduction: Fever is a common symptom of many tropical diseases and in many cases the etiologic agent remains unidentified as a consequence of either the etiologic agent not being part of routine diagnostic screening or as a consequence of false negatives on standard diagnostic tests. Methodology: This study screened a well characterized panel of 274 serum samples collected on day of admission from adult patients with acute undifferentiated fever admitted to a hospital in Nakhon Ratchasima, Thailand by RT-PCR using pan-flavivirus degenerate primers. Results: Subsequent clinical diagnosis was achieved for 38 of the patients, and included 19 cases of dengue fever. RT-PCR screening identified seven positive samples (2.5%) which were revealed by sequence analysis to be dengue virus 1 (2 cases), dengue virus 2 (2 cases) and dengue virus 3 (3 cases). Only 5 out of 19 (26%) serum samples from patients subsequently diagnosed with dengue were positive, but 2 samples which clinically remained undiagnosed were shown to be positive for dengue virus. Sequence analysis suggested that the dengue virus 3 cases occurred as a result of importation of a strain of dengue from India or China. No other flaviviruses were identified. Conclusions: No evidence was found of other flaviviruses besides dengue circulating in this population. Despite improved diagnostic tests, cases of dengue are still evading correct diagnosis.

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Kushwaha, Nikhal, Vipin Kesharwani, and Pankaj Kumar Jaiswal. "A GLOBAL CONCERN ON ZIKA VIRUS: TRANSMISSION, DIAGNOSIS, PREVENTION, AND TREATMENT." Journal of Drug Delivery and Therapeutics 8, no. 5 (September 11, 2018): 136–40. http://dx.doi.org/10.22270/jddt.v8i5.1972.

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Zika virus is a mosquito-transmitted flavivirus belongs to family Flaviviridae which becomes the focus of an ongoing pandemic and public health emergency all around the world. Zika virus has two lineages African and Asian. Mosquito-borne flavivirus is thought to replicate initially in dendritic cell and then spread to lymph nodes and then to the bloodstream. Zika virus was initially recognized in Uganda in 1947 in Monkeys through a method that observed yellow fever. It was later distinguished in people in 1952 in Uganda and the United Republic of Tanzania. The explosions of the zika virus disease have been recorded in Africa, The Americas, Asia, and The Pacific. Gillian-Berre syndrome and congenital malformation (microcephaly) suspected to be linked with Zika virus. The virus can only be confirmed through laboratory test on blood or other body fluids, such as urine, saliva or semen. No specific antiviral treatment for Zika virus disease exists. Treatment is aimed at relieving symptoms with rest, fluid and medications. WHO/PAHO encourages the countries to establish and maintain Zika Virus infections, detection, clinical management and community assurances strategies to reduce transmission of the virus. The future of Zika Virus spreading to other parts of the world is still unknown. Keywords: Zika Virus, flavivirus, Mosquito, Vaccine, Treatment, Microcephaly, WHO/PAHO.

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Espinoza, Daniel, Yerun Zhu, and Matthew H. Collins. "LB18. Understanding Zika-Specific Immunity for Prevention and Protection." Open Forum Infectious Diseases 6, Supplement_2 (October 2019): S1001. http://dx.doi.org/10.1093/ofid/ofz415.2501.

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Abstract Background Flavivirus infection represents a major public health problem, with over 2 billion people at risk for one or more infections each year. There are no licensed antiviral treatments for flaviviruses, and effective vaccines are lacking for several. Accurate diagnosis of flaviviruses is also complicated by serologic cross-reactivity. Thus, there is an urgent need to develop public health tools and interventions to reduce the burden of flavivirus infection worldwide. Methods We recently isolated two potently neutralizing human monoclonal antibodies (NmAb) against Zika, which exhibit activity against multiple strains of Zika but do not bind the closely related dengue virus. Mapping studies revealed that the NmAb target distinct epitopes on the envelope protein (E). A competition ELISA (ZE-BOB) was developed with the two NmAb to test the hypotheses that neutralizing Ab responses in the general population would target the same epitopes on Zika E as the novel NmAb. Results We found that competitive Ab responses were detected in 19 of 19 (100%) convalescent sera from travelers with confirmed Zika infection. We tested a panel of 20 sera from individuals with either primary or secondary prior dengue infection and found that 18/20 (90%) contained IgG reactive with Zika virus; however, 0/20 (0%) exhibited ZE-BOB activity. These results indicate that the epitopes targeted by these two Zika NmAb are consistently immunogenic humans infected by Zika. Additionally, these epitopes elicit type-specific Ab to Zika, providing a basis for development of simple serodiagnostic assays with utility in epidemiologic studies as well as in the clinical setting. Because NAb are key determinants of long-term protection against future flavivirus infection, we further hypothesized that the results of the ZE-BOB assay may simultaneously provide a correlate of immunity, which would be a critical tool for vaccine development. In comparing the 50% neutralization titer against Zika with the magnitude of competition by ZE-BOB, there was no correlation in these values, but sera tended to be positive in both or neither assay. Conclusion Therefore, the ZE-BOB assay constitutes a novel tool that is widely deployable for purposes ranging from clinical diagnosis to epidemiologic monitoring to vaccine development for Zika. Disclosures All authors: No reported disclosures.

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Roldán, Julieta S., Alejandro Cassola, and Daniela S. Castillo. "Development of a novel NS1 competitive enzyme-linked immunosorbent assay for the early detection of Zika virus infection." PLOS ONE 16, no. 8 (August 17, 2021): e0256220. http://dx.doi.org/10.1371/journal.pone.0256220.

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Zika virus (ZIKV) is a flavivirus that has emerged as a global health threat after the 2015 outbreak in the Americas, where devastating congenital defects were documented. There are currently no vaccines to prevent ZIKV infections nor commercially available clinical diagnostic tests demonstrated to identify ZIKV without cross-reactive interference of related flaviviruses. Early diagnosis is critical when treating symptomatic patients and in preventing ZIKV transmission. In this context, the development of sensitive and accurate diagnostic methods are urgently needed for the detection of ZIKV acute infection. The aim of this study consisted of obtaining monoclonal antibodies (mAbs) against denatured monomeric ZIKV Nonstructural protein 1 (ZNS1), a useful diagnostic marker for flavivirus early detection, in order to develop a highly specific and sensitive ZNS1 indirect competitive ELISA (icELISA). The production of hybridomas secreting ZNS1 mAbs was carried out through immunizations with denatured monomeric ZNS1. We selected 1F5 and 6E2 hybridoma clones, which recognized the heat-denatured ZNS1 hexameric form by indirect ELISA. Cross-reaction studies indicated that these mAbs specifically bind to a ZNS1 linear epitope, and that they do not cross-react with the NS1 protein from other related flaviviruses. The 1F5 mAb enabled the development of a sensitive and reproducible icELISA to detect and quantify small amounts of ZNS1 disease marker in heat-denatured human sera. Here, we establish a reliable 1F5 based-icELISA that constitutes a promising diagnostic tool for control strategies and the prevention of ZIKV propagation.

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Santos, Adriano, Paulo R. Bueno, and Jason J. Davis. "A dual marker label free electrochemical assay for Flavivirus dengue diagnosis." Biosensors and Bioelectronics 100 (February 2018): 519–25. http://dx.doi.org/10.1016/j.bios.2017.09.014.

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Martinez Viedma, Maria del Pilar, Nurgun Kose, Leda Parham, Angel Balmaseda, Guillermina Kuan, Ivette Lorenzana, Eva Harris, James E. Crowe Jr., and Brett E. Pickett. "Peptide arrays of three collections of human sera from patients infected with mosquito-borne viruses." F1000Research 8 (November 7, 2019): 1875. http://dx.doi.org/10.12688/f1000research.20981.1.

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Background: Global outbreaks caused by emerging or re-emerging arthropod-borne viruses (arboviruses) are becoming increasingly more common. These pathogens include the mosquito-borne viruses belonging to the Flavivirus and Alphavirus genera. These viruses often cause non-specific or asymptomatic infection, which can confound viral prevalence studies. In addition, many acute phase diagnostic tests rely on the detection of viral components such as RNA or antigen. Standard serological tests are often not reliable for diagnosis after seroconversion and convalescence due to cross-reactivity among flaviviruses. Methods: In order to contribute to development efforts for mosquito-borne serodiagnostics, we incubated 137 human sera on individual custom peptide arrays that consisted of over 866 unique peptides in quadruplicate. Our bioinformatics workflow to analyze these data incorporated machine learning, statistics, and B-cell epitope prediction. Results: Here we report the results of our peptide array data analysis, which revealed sets of peptides that have diagnostic potential for detecting past exposure to a subset of the tested human pathogens including Zika virus. These peptides were then confirmed using the well-established ELISA method. Conclusions: These array data, and the resulting peptides can be useful in diverse efforts including the development of new pan-flavivirus antibodies, more accurate epitope mapping, and vaccine development against these viral pathogens.

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Martinez Viedma, Maria del Pilar, Nurgun Kose, Leda Parham, Angel Balmaseda, Guillermina Kuan, Ivette Lorenzana, Eva Harris, James E. Crowe Jr., and Brett E. Pickett. "Peptide arrays of three collections of human sera from patients infected with mosquito-borne viruses." F1000Research 8 (February 17, 2020): 1875. http://dx.doi.org/10.12688/f1000research.20981.2.

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Background: Global outbreaks caused by emerging or re-emerging arthropod-borne viruses (arboviruses) are becoming increasingly more common. These pathogens include the mosquito-borne viruses belonging to the Flavivirus and Alphavirus genera. These viruses often cause non-specific or asymptomatic infection, which can confound viral prevalence studies. In addition, many acute phase diagnostic tests rely on the detection of viral components such as RNA or antigen. Standard serological tests are often not reliable for diagnosis after seroconversion and convalescence due to cross-reactivity among flaviviruses. Methods: In order to contribute to development efforts for mosquito-borne serodiagnostics, we incubated 137 human sera on individual custom peptide arrays that consisted of over 866 unique peptides in quadruplicate. Our bioinformatics workflow to analyze these data incorporated machine learning, statistics, and B-cell epitope prediction. Results: Here we report the results of our peptide array data analysis, which revealed sets of peptides that have diagnostic potential for detecting past exposure to a subset of the tested human pathogens including Zika virus. These peptides were then confirmed using the well-established ELISA method. Conclusions: These array data, and the resulting peptides can be useful in diverse efforts including the development of new pan-flavivirus antibodies, more accurate epitope mapping, and vaccine development against these viral pathogens.

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Martinez Viedma, Maria del Pilar, Nurgun Kose, Leda Parham, Angel Balmaseda, Guillermina Kuan, Ivette Lorenzana, Eva Harris, James E. Crowe Jr., and Brett E. Pickett. "Peptide arrays incubated with three collections of human sera from patients infected with mosquito-borne viruses." F1000Research 8 (February 28, 2020): 1875. http://dx.doi.org/10.12688/f1000research.20981.3.

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Background: Global outbreaks caused by emerging or re-emerging arthropod-borne viruses (arboviruses) are becoming increasingly more common. These pathogens include the mosquito-borne viruses belonging to the Flavivirus and Alphavirus genera. These viruses often cause non-specific or asymptomatic infection, which can confound viral prevalence studies. In addition, many acute phase diagnostic tests rely on the detection of viral components such as RNA or antigen. Standard serological tests are often not reliable for diagnosis after seroconversion and convalescence due to cross-reactivity among flaviviruses. Methods: In order to contribute to development efforts for mosquito-borne serodiagnostics, we incubated 137 human sera on individual custom peptide arrays that consisted of over 866 unique peptides in quadruplicate. Our bioinformatics workflow to analyze these data incorporated machine learning, statistics, and B-cell epitope prediction. Results: Here we report the results of our peptide array data analysis, which revealed sets of peptides that have diagnostic potential for detecting past exposure to a subset of the tested human pathogens including Zika virus. These peptides were then confirmed using the well-established ELISA method. Conclusions: These array data, and the resulting peptides can be useful in diverse efforts including the development of new pan-flavivirus antibodies, more accurate epitope mapping, and vaccine development against these viral pathogens.

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Qing, Min, Zhi-ming Yuan, and Pei-Yong Shi. "A virus-type specific serological diagnosis of flavivirus infection using virus-like particles." Virologica Sinica 24, no. 2 (April 2009): 136–45. http://dx.doi.org/10.1007/s12250-009-3023-6.

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Furuya, Andrea K. M., Danielle Hunt, Kirsten St George, Alan P. Dupuis, Laura D. Kramer, Pei-Yong Shi, and Susan Wong. "Use of the immunoglobulin G avidity assay to differentiate between recent Zika and past dengue virus infections." Clinical Science 133, no. 7 (April 2019): 859–67. http://dx.doi.org/10.1042/cs20180874.

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Abstract Zika (ZIKV) and dengue (DENV) virus infections elicit a robust but cross-reactive antibody response against the viral envelope protein, while antibody responses against non-structural proteins (NS) are more virus specific. Building on this premise, we have previously developed a flavivirus multiplex microsphere immunoassay (MIA) for the serologic diagnosis of ZIKV and DENV infections. This assay significantly improved diagnostic accuracy; however, MIA could not differentiate more recent from past infections, which still represents a major diagnostic challenge. Therefore, an immunoglobulin G (IgG) based avidity assay was developed and its diagnostic performance evaluated. Specimens from New York State residents were submitted to the Wadsworth Center New York State Department of Health (NYSDOH) for routine clinical testing by Zika IgM ELISA and plaque reduction neutralization test (PRNT). Using our previously developed flavivirus MIA as a platform, we developed an IgG avidity assay to discriminate recent ZIKV from past DENV infections. Zika IgM positive specimens had an average Zika IgG avidity index of 14.8% (95% CI: 11.0–18.4%), while Zika IgM negative but flavivirus MIA and PRNT positive samples had an average Zika IgG avidity index of 34.9% (95% CI: 31.1–38.7%). Specimens positive for dengue antibodies by flavivirus MIA and PRNT had an average dengue IgG avidity index of 68.7% (95% CI: 62.7–75.0%). The IgG avidity assay accurately distinguished recent ZIKV from past DENV infections in patients who traveled to dengue endemic regions. This assay could be very useful in patients with high risk of Zika complications such as pregnant women and monitoring immune responses in vaccine trials.

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Akinshina, Yu A., V. F. Larichev, S. G. Mardanly, A. M. Butenko, N. V. Khutoretskaya, and M. A. Saifullin. "Application of immunoassay systems for diagnosis of dengue fever." Epidemiology and Infectious Diseases 20, no. 4 (August 15, 2015): 8–12. http://dx.doi.org/10.17816/eid40851.

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As a result of the application of experimental test systems "ELISA-IgM-dengue", "ELISA-IgM-WN" and "ELISA-IgM-TBE" when examining sera from patients with dengue fever (DF), West Nile fever (WNF) and tick-borne encephalitis (TBE) there was established the possibility of a clear differential diagnosis of these related flavivirus infections. Application of the ELISA-IgG test systems fail to provide such an opportunity due to cause of pronounced cross-reactions of group-IgG antibodies. In the article there are presented data on dynamics of IgM and IgG antibodies in patients with dengue fever. Detection of specific IgG in the first days of the disease may indicate to a secondary nature of infection and the possibility of the development of hemorrhagic syndrome.

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Lorch, Matías Sebastián, María Soledad Collado, Marcelo Horacio Argüelles, Rosana Paola Rota, Lorena Ivana Spinsanti, Mario Enrique Lozano, and Sandra Elizabeth Goñi. "Production of recombinant NS1 protein and its possible use in encephalitic flavivirus differential diagnosis." Protein Expression and Purification 153 (January 2019): 18–25. http://dx.doi.org/10.1016/j.pep.2018.08.008.

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Bedin, Frederic, Laurent Boulet, Elodie Voilin, Gerald Theillet, Agnes Rubens, and Christine Rozand. "Paper-based point-of-care testing for cost-effective diagnosis of acute flavivirus infections." Journal of Medical Virology 89, no. 9 (March 28, 2017): 1520–27. http://dx.doi.org/10.1002/jmv.24806.

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Nagar, Pradeep Kumar, Deepali Savargaonkar, and Anupkumar R. Anvikar. "Detection of Dengue Virus-Specific IgM and IgG Antibodies through Peptide Sequences of Envelope and NS1 Proteins for Serological Identification." Journal of Immunology Research 2020 (August 4, 2020): 1–8. http://dx.doi.org/10.1155/2020/1820325.

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Dengue is an acute febrile illness caused by positive-sense single-stranded RNA virus, belonging to the family Flaviviridae and genus Flavivirus. Transmission of virus among the individuals occurred by blood-feeding Aedes mosquitoes. This virus has four serotypes differentiated on the basis of antibody neutralization assay. At present, there is no particular treatment or vaccine candidate available for dengue infection. Approximately 3.9 billion human populations are at risk of dengue virus (DENV) infection. Thus, precise diagnosis of dengue at the early stage is very essential for disease control and effective therapy in order to treat or prevent severe complications. Indeed, the accurate diagnosis of DENV remains a problem because of low detection accuracy along with high testing price. Sensitivity and specificity of available kits vary from test to test, and cross-reactivity with other Flavivirus is a challenging issue for diagnosis. In this study, linear epitopes of envelope (E) and NS1 proteins were identified to diagnose the DENV. Whole protein sequences of E and NS1 of DENV were obtained from UniProtKB database. On the basis of algorithm prediction from DNASTAR, BCEPRED, and IEDB data resources, twelve peptides of E (EP1 to EP12) and eight peptides of NS1 (NS1-1 to NS1-8) were selected, which were common in all serotypes. Sequence homologies of peptides with other Flavivirus were checked by Multiple Sequence Alignment Tool ClustalX2. Peptide sequences were synthesized chemically by solid-phase peptide synthesis technique. Dengue-specific IgM and IgG (secondary response) antibodies in the patient’s antisera were tested with the peptides using ELISA protocol. Peptides EP1, EP2, EP4, EP7, EP10, and EP12 of E protein and NS1-1, NS1-3, NS1-4, NS1-7, and NS1-8 of NS1 protein were considered the best immunoreactive peptides with the sensitivity (73.33-96.66%) and specificity (82.14-100%). Such peptides together can be used to construct the multiple antigen peptides (MAP) or multiplexed microbeads for designing a precise, cost-effective, and easy-to-make peptide-based immunodiagnostic kit for DENV detection.

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Nguyen, Ngoc Minh, Bao Tuan Duong, Mudsser Azam, Truong Thai Phuong, Hyun Park, Phung Thi Bich Thuy, and Seon-Ju Yeo. "Diagnostic Performance of Dengue Virus Envelope Domain III in Acute Dengue Infection." International Journal of Molecular Sciences 20, no. 14 (July 15, 2019): 3464. http://dx.doi.org/10.3390/ijms20143464.

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Dengue, one of the most prevalent illnesses caused by dengue viruses that are members of the genus Flavivirus, is a significant global health problem. However, similar clinical symptoms and high antigenic homologies with other Flaviviruses in the endemic area pose difficulties for differential diagnosis of dengue from other arbovirus infections. Here, we investigated four types of recombinant envelope protein domain III (DV-rED III) derived from four dengue virus (DENV) serotypes for diagnostic potential in detecting IgM in acute phase (mainly 2–3 days after onset of fever). Each independent DV-1, -3, and -4-rED III-ELISA showed less than 60% sensitivity, but the combined results of DV-1, -3, and -4-rED III-ELISA led to sensitivity of 81.82% (18/22) (95% CI, 59.72 to 94.81) and 100% specificity (46/46) (95% CI, 92.29 to 100.00) as each antigen compensated the other antigen-derived negative result. In conclusion, the independent combination of data derived from each recombinant antigen (DV1-, DV3-, and DV4-rED III) showed comparable efficacy for the detection of IgM in patients with acute-phase dengue infection.

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Chiuya, Tatenda, Daniel K. Masiga, Laura C. Falzon, Armanda D. S. Bastos, Eric M. Fèvre, and Jandouwe Villinger. "A survey of mosquito-borne and insect-specific viruses in hospitals and livestock markets in western Kenya." PLOS ONE 16, no. 5 (May 28, 2021): e0252369. http://dx.doi.org/10.1371/journal.pone.0252369.

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Aedes aegypti and Culex pipiens complex mosquitoes are prolific vectors of arboviruses that are a global threat to human and animal health. Increased globalization and ease of travel have facilitated the worldwide dissemination of these mosquitoes and the viruses they transmit. To assess disease risk, we determined the frequency of arboviruses in western Kenyan counties bordering an area of high arboviral activity. In addition to pathogenic viruses, insect-specific flaviviruses (ISFs), some of which are thought to impair the transmission of specific pathogenic arboviruses, were also evaluated. We trapped mosquitoes in the short and long rainy seasons in 2018 and 2019 at livestock markets and hospitals. Mosquitoes were screened for dengue, chikungunya and other human pathogenic arboviruses, ISFs, and their blood-meal sources as determined by high-resolution melting analysis of (RT-)PCR products. Of 6,848 mosquitoes collected, 89% were trapped during the long rainy season, with A. aegypti (59%) and Cx. pipiens sensu lato (40%) being the most abundant. Most blood-fed mosquitoes were Cx. pipiens s.l. with blood-meals from humans, chicken, and sparrow (Passer sp.). We did not detect dengue or chikungunya viruses. However, one Culex poicilipes female was positive for Sindbis virus, 30 pools of Ae. aegypti had cell fusing agent virus (CFAV; infection rate (IR) = 1.27%, 95% CI = 0.87%-1.78%); 11 pools of Ae. aegypti had Aedes flavivirus (AeFV; IR = 0.43%, 95% CI = 0.23%-0.74%); and seven pools of Cx. pipiens s.l. (IR = 0.23%, 95% CI = 0.1%-0.45%) and one pool of Culex annulioris had Culex flavivirus. Sindbis virus, which causes febrile illness in humans, can complicate the diagnosis and prognosis of patients with fever. The presence of Sindbis virus in a single mosquito from a population of mosquitoes with ISFs calls for further investigation into the role ISFs may play in blocking transmission of other arboviruses in this region.

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Bergamaschi, Greta, Enrico M. A. Fassi, Alessandro Romanato, Ilda D'Annessa, Maria Teresa Odinolfi, Dario Brambilla, Francesco Damin, et al. "Computational Analysis of Dengue Virus Envelope Protein (E) Reveals an Epitope with Flavivirus Immunodiagnostic Potential in Peptide Microarrays." International Journal of Molecular Sciences 20, no. 8 (April 18, 2019): 1921. http://dx.doi.org/10.3390/ijms20081921.

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The mosquito-borne viral disease caused by the Dengue virus is an expanding global threat. Diagnosis in low-resource-settings and epidemiological surveillance urgently requires new immunoprobes for serological tests. Structure-based epitope prediction is an efficient method to design diagnostic peptidic probes able to reveal specific antibodies elicited in response to infections in patients’ sera. In this study, we focused on the Dengue viral envelope protein (E); computational analyses ranging from extensive Molecular Dynamics (MD) simulations and energy-decomposition-based prediction of potentially immunoreactive regions identified putative epitope sequences. Interestingly, one such epitope showed internal dynamic and energetic properties markedly different from those of other predicted sequences. The epitope was thus synthesized as a linear peptide, modified for chemoselective immobilization on microarrays and used in a serological assay to discriminate Dengue-infected individuals from healthy controls. The synthetic epitope probe showed a diagnostic performance comparable to that of the full antigen in terms of specificity and sensitivity. Given the high level of sequence identity among different flaviviruses, the epitope was immune-reactive towards Zika-infected sera as well. The results are discussed in the context of the quest for new possible structure-dynamics-based rules for the prediction of the immunoreactivity of selected antigenic regions with potential pan-flavivirus immunodiagnostic capacity.

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Tan, Li Kiang, Wing Yan Wong, Hui Ting Yang, Roland G. Huber, Peter J. Bond, Lee Ching Ng, Sebastian Maurer-Stroh, and Hapuarachchige Chanditha Hapuarachchi. "Flavivirus Cross-Reactivity to Dengue Nonstructural Protein 1 Antigen Detection Assays." Diagnostics 10, no. 1 (December 24, 2019): 11. http://dx.doi.org/10.3390/diagnostics10010011.

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Dengue virus (DENV) and Zika virus (ZIKV) are flaviviruses of public health relevance. Both viruses circulate in the same endemic settings and acute infections generally manifest similar symptoms. This highlights the importance of accurate diagnosis for clinical management and outbreak control. One of the commonly used acute diagnostic markers for flaviviruses is nonstructural protein 1 (NS1). However, false positives due to antigenic cross-reactivity have been reported between DENV and ZIKV infections when using DENV NS1 antigen (NS1 Ag) detection assays in acute cases. Therefore, we investigated the lowest detectable virus titres and cross-reactivity of three commercial dengue NS1 Ag rapid assays and two ELISAs for different flaviviruses. Our results showed that substantially high viral titres of ZIKV, Kunjin virus (KUNV) and yellow fever virus (YFV) are required to give false-positive results when using DENV NS1 rapid detection assays. Commercial DENV NS1 ELISAs did not react with ZIKV and YFV. In comparison, tested assays detected DENV at a significantly low virus titre. Given the relatively low viral loads reported in clinical samples, our findings suggest that commercially available dengue NS1 Ag detection assays are less likely to generate false-positive results among clinical samples in areas where multiple flaviviruses cocirculate.

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Barth, O. M., S. Majerowicz, L. P. Menasce, and H. G. Schatzmayr. "Detection of viral infection by immunofluorescence in formalin-fixed tissues, pretreated with trypsin." Memórias do Instituto Oswaldo Cruz 83, no. 2 (June 1988): 207–12. http://dx.doi.org/10.1590/s0074-02761988000200010.

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The presence of viral antigen in sections from formalin-fixed and paraffin-embedded human tissues was demonstrated by trypsin digestion followed by direct or indirect immunofluorescence. The specimens may be used for retrospective diagnosis. The immunofluorescence technique has to be adapted to the suspected virus infection on the basis of previous histopathology study. Variations of trypsin concentration time and temperature of incubation, expose different viral antigens and have to be previously tested for each unknown system. For measles virus detection in lung a stronger digestion has to be applied as compared to adenovirus or respiratory disease viruses in the same tisue. Flavivirus in liver tissue needs a weaker digestion. The reproducibility of the method makes it useful as a routine technique in diagnosis of virus infection.

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Chávez, Juliana Helena, Jaqueline Raymondi Silva, Alberto Anastacio Amarilla, and Luiz Tadeu Moraes Figueiredo. "Domain III peptides from flavivirus envelope protein are useful antigens for serologic diagnosis and targets for immunization." Biologicals 38, no. 6 (November 2010): 613–18. http://dx.doi.org/10.1016/j.biologicals.2010.07.004.

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Kumar, Deepak, Rajesh Kumar Verma, Amit Singh, Manoj Kumar, Dharmendra Prasad Singh, Richa Pandey, and Kiran Krishnappa. "Evaluation of NS1, IgM ELISA and RT-PCR in diagnosis of dengue fever." International Journal of Research in Medical Sciences 6, no. 7 (June 25, 2018): 2440. http://dx.doi.org/10.18203/2320-6012.ijrms20182832.

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Background: Dengue fever is a mosquito borne disease caused by flavivirus. Its cases are increasing in India with increasing mortality rate year by year hence, prompt and accurate diagnosis is necessary to prevent morbidity and mortality.Methods: In this study we enrolled 125 clinically suspected cases of dengue. All the collected samples were processed for RT-PCR, NS1 and IgM ELISA. We evaluated NS1 antigen ELISA alone, and combination of NS1 and IgM ELISA against RT-PCR.Results: Among 125 clinically suspected case 67 were positive by RT-PCR and 58 were negative. Sensitivity and Specificity of NS1 ELISA and NS1 with IgM ELISA (in combination) against RT-PCR were 83.58%, 94.82% and 95.55%, 79.31% respectively. (p<0.001).Conclusions: The NS1 ELISA alone was sufficient to detect acute phase of dengue fever, although, combination of NS1 and IgM proved to be most appropriate method for detection of acute as well as late phase of dengue fever.

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Martin, Denise A., Brad J. Biggerstaff, Becky Allen, Alison J. Johnson, Robert S. Lanciotti, and John T. Roehrig. "Use of Immunoglobulin M Cross-Reactions in Differential Diagnosis of Human Flaviviral Encephalitis Infections in the United States." Clinical and Vaccine Immunology 9, no. 3 (May 2002): 544–49. http://dx.doi.org/10.1128/cdli.9.3.544-549.2002.

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ABSTRACT To define the virus specificity of the immunoglobulin M (IgM) antibody-capture enzyme-linked immunosorbent assay (MAC-ELISA) among the medically important members of the Japanese encephalitis (JE) virus serocomplex of flaviviruses, 103 IgM-positive human serum samples from patients with confirmed West Nile (WN) virus, St. Louis encephalitis (SLE) virus, or JE virus infections were assembled and simultaneously tested against all three viral antigens in a standardized MAC-ELISA. Of the serum samples tested, 96 (93%) showed higher positive-to-negative absorbance ratios (P/Ns) with the infecting virus antigen compared to those obtained with the other two virus antigens. Of the seven specimens with higher P/Ns with heterologous virus antigens, six were from patients with SLE virus infections (the serum samples had higher levels of reactivity with WN virus antigen) and one was from a patient with a JE virus infection (this serum sample also had a higher level of reactivity with WN virus antigen). Not surprisingly, similar virus specificity was observed with WN virus-elicited IgM in cerebrospinal fluid. As shown in previous studies, a subset of these specimens was even less reactive in the MAC-ELISA with dengue virus, a member of a different flavivirus serocomplex. The degree of virus cross-reactivity did not appear to be related to days postonset, at least during the first 40 days of infection. Infections with WN virus could be correctly distinguished from infections with SLE virus on the basis of the observed anti-viral IgM cross-reactivities alone 92% of the time. Infections with SLE virus resulted in antibody that was more cross-reactive, so identification of SLE virus as the infecting agent by use of MAC-ELISA cross-reactivity alone was more problematic.

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Chao, Day-Yu, Jedhan Ucat Galula, Wen-Fan Shen, Brent S. Davis, and Gwong-Jen J. Chang. "Nonstructural Protein 1-Specific Immunoglobulin M and G Antibody Capture Enzyme-Linked Immunosorbent Assays in Diagnosis of Flaviviral Infections in Humans." Journal of Clinical Microbiology 53, no. 2 (December 10, 2014): 557–66. http://dx.doi.org/10.1128/jcm.02735-14.

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IgM antibody- and IgG antibody-capture enzyme-linked immunosorbent assays (MAC/GAC-ELISAs) targeted at envelope protein (E) of dengue viruses (DENV), West Nile virus, and Japanese encephalitis virus (JEV) are widely used as serodiagnostic tests for presumptive confirmation of viral infection. Antibodies directed against the flavivirus nonstructural protein 1 (NS1) have been proposed as serological markers of natural infections among vaccinated populations. The aim of the current study is to optimize an IgM and IgG antibody-capture ELISA (MAC/GAC-ELISA) to detect anti-NS1 antibodies and compare it with anti-E MAC/GAC-ELISA. Plasmids to express premembrane/envelope (prM/E) or NS1 proteins of six medically important flaviviruses, including dengue viruses (DENV-1 to DENV-4), West Nile virus (WNV), and Japanese encephalitis virus (JEV), were constructed. These plasmids were used for the production of prM/E-containing virus-like particles (VLPs) and secreted NS1 (sNS1) from COS-1 cells. Archived clinical specimens from patients with confirmed DENV, JEV, and WNV infections, along with naive sera, were subjected to NS1-MAC/GAC-ELISAs before or after depletion of anti-prM/E antibodies by preabsorption with or without VLPs. Human serum specimens from previously confirmed DENV infections showed significantly enhanced positive-to-negative (P/N) ratios for NS1-MAC/GAC-ELISAs after the depletion of anti-prM/E antibodies. No statistical differences in sensitivities and specificities were found between the newly developed NS1- and VLP-MAC/GAC-ELISAs. Further application of the assays to WNV- and JEV-infected serum panels showed similar results. A novel approach to perform MAC/GAC-ELISAs for NS1 antibody detection was successfully developed with great potential to differentiate antibodies elicited by the tetravalent chimeric yellow fever-17D/dengue vaccine or DENV infection.

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Piyasena, Thisun B. H., Yin X. Setoh, Jody Hobson-Peters, Natalie A. Prow, Helle Bielefeldt-Ohmann, Alexander A. Khromykh, David Perera, Mary J. Cardosa, Peter D. Kirkland, and Roy A. Hall. "Differential Diagnosis of Flavivirus Infections in Horses Using Viral Envelope Protein Domain III Antigens in Enzyme-Linked Immunosorbent Assay." Vector-Borne and Zoonotic Diseases 17, no. 12 (December 2017): 825–35. http://dx.doi.org/10.1089/vbz.2017.2172.

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Modo, Emmanuel, Favour Okoro, Blessing Orji, and Rex-Clovis Njoku. "Zika virus, the way forward: Nigeria as case study." International Journal of Scientific World 5, no. 2 (June 27, 2017): 96. http://dx.doi.org/10.14419/ijsw.v5i2.7900.

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Zika Virus is a flavivirus that is responsible for an unprecedented current epidemic. It is an emerging mosquito borne virus that is a (+) sense single-stranded RNA virus from the Flaviviridae virus family, Flavivirus genre and the Zika virus specie. Zika virus bears a complex genome and virion structure. Its reproductive cycle in host cell involves virion endocytosis that allows entrance into host cell. Zika virus infection gives symptoms that are usually mild and last for few days and infection can be easily transferred from infected mosquitoes, from a pregnant woman to her fetus, sexual intercourse, blood transfusion, and laboratory exposure. Several methods are made available for the diagnosis of this infection and several ways to prevent this infection such as elimination and control of mosquito, prevention of mosquito bites and public awareness about Zika and mosquitoes. No vaccines are available for treatment yet but common medications are advised for symptoms. The virus has been associated with fetal microcephaly in humans. Information is provided for the epidemiology of Zika virus to evaluate the level of risk for people who may be planning to travel to or are recently returning from areas with possible local transmission. In the African region this is done considering a number of factors and cases in the past. The rapid development of a safe and effective vaccine, genetically modified mosquitoes, use of bacteria, and other innovations are currently worked on to curb the spread of Zika virus.

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Tai, Dar-Fu, Chung-Yin Lin, Tzong-Zeng Wu, Jyh-Hsiung Huang, and Pei-Yun Shu. "Artificial Receptors in Serologic Tests for the Early Diagnosis of Dengue Virus Infection." Clinical Chemistry 52, no. 8 (August 1, 2006): 1486–91. http://dx.doi.org/10.1373/clinchem.2005.064501.

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Abstract Background: Because of the range and nonspecificity of clinical presentations of dengue virus infections, we felt there was a need to create diagnostic tests. We used artificial receptors for the virus to develop serologic assays to detect dengue virus infection. Methods: We coated a quartz crystal microbalance (QCM) with molecularly imprinted polymers specific for nonstructural protein 1 of flavivirus. These artificial receptors were specifically created on a QCM chip by polymerization of monomers and were cross-linked in the presence of the epitope site of nonstructural protein 1. We tested serum samples from patients with confirmed cases of dengue reported to the Center for Disease Control in Taipei. Samples were diluted 100-fold; no other sample pretreatment was used. The QCM response was compared with results of monoclonal ELISA. Results: QCM signals were >15 Hz in 18 of 21 (86%) of dengue samples and in 0 of 16 control samples. The correlation (r2) of the QCM response and the ELISA result was 0.73. Within-run and run-to-run imprecisions (CV) were 4%–28% and 10%–32%, respectively. Conclusions: The described assay offers a serologic technique for diagnosis of early viremia. The results illustrate the potential of well-organized polymers on the highly sensitive sensor system for diagnostic and biotechnological applications.

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Mammen, Shoba, Aiswarya Nair, Santhosh Kumar, Zayina Zonderveni, A. T. Prabhakar, Turaka Vijay Prakash, Sanjith Aaron, Mathew Alexander, Anand Zachariah, and Asha Mary Abraham. "Clinical Features of Four West Nile Virus Cases and Its Molecular Characterization from a South Indian Tertiary Care Hospital." Case Reports in Infectious Diseases 2020 (July 16, 2020): 1–6. http://dx.doi.org/10.1155/2020/1315041.

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West Nile virus (WNV) is currently a significant reemerging virus of the 21st century. It belongs to the family Flaviviridae and genus Flavivirus. Although it is primarily transmitted by the Culex spp of mosquitoes, other routes of transmission are also well defined. Of eight lineages described, Lineage 1a has been reported from many parts of South India and is known to cause neuroinvasive illness. Many tests and serological techniques have been described to diagnose WNV infection such as complement fixation, neutralization, heamagglutination inhibition, ELISA, and PCR for molecular confirmation. The latter far outweighs the limitations inherent in the other tests. WNV infection is being reported from Vellore for the first time after 1968. This paper aims to describe four cases of WNV infection causing central nervous system manifestations with its molecular characterization. West Nile virus infection was diagnosed with the available molecular techniques such as PCR and sequencing, which emphasizes the need for considering West Nile virus in the differential diagnosis of acute meningoencephalitis and the wider availability of molecular diagnostic tests.

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Ribeiro Junior, Marcelo Augusto Fontanelle, Vinicius Cunha Rodrigues, Celia Ya Dan Feng, Alexander Trong Minh Nguyen, Giovana El Khouri Bechara, and Raíssa Reis de Moura. "Yellow Fever." Revista de Medicina 97, no. 4 (December 18, 2018): 407–14. http://dx.doi.org/10.11606/issn.1679-9836.v97i4p407-414.

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Since January 2017, there have been at least 1563 suspected cases of Yellow Fever, 629 confirmed cases and 232 confirmed deaths. Yellow fever is a viral hemorrhagic disease endemic to the tropical parts of Africa and South America. At the present time, it has presented a significant increase in its incidence in Brazil, with important repercussions and impacts on the public health. This review paper outlines the causes of yellow fever, as well as the disease epidemiology, progression, diagnosis, treatment and prevention. We conclude by reporting on the current epidemic in Brazil and future directions for research. Method: Data from Pubmed, SciELO, Medline and government sources concerning Yellow Fever were used, dating from 2002 to 2018. In the collection of the data the following descriptors were used: Yellow-fever, Aedes, Arbovirus and Flavivirus.

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Zhang, Liding, Xuewei Du, Congjie Chen, Zhixin Chen, Li Zhang, Qinqin Han, Xueshan Xia, Yuzhu Song, and Jinyang Zhang. "Development and Characterization of Double-Antibody Sandwich ELISA for Detection of Zika Virus Infection." Viruses 10, no. 11 (November 15, 2018): 634. http://dx.doi.org/10.3390/v10110634.

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Zika virus (ZIKV) is an emerging mosquito-transmitted flavivirus that can cause severe disease, including congenital birth defect and Guillain−Barré syndrome during pregnancy. Although, several molecular diagnostic methods have been developed to detect the ZIKV, these methods pose challenges as they cannot detect early viral infection. Furthermore, these methods require the extraction of RNA, which is easy to contaminate. Nonstructural protein 1 (NS1) is an important biomarker for early diagnosis of the virus, and the detection methods associated with the NS1 protein have recently been reported. The aim of this study was to develop a rapid and sensitive detection method for the detection of the ZIKV based on the NS1 protein. The sensitivity of this method is 120 ng mL−1 and it detected the ZIKV in the supernatant and lysates of Vero and BHK cells, as well as the sera of tree shrews infected with the ZIKV. Without the isolation of the virus and the extraction of the RNA, our method can be used as a primary screening test as opposed to other diagnosis methods that detect the ZIKV.

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Akhras, Sami, Marie-Luise Herrlein, Fabian Elgner, Thomas Holzhauser, and Eberhard Hildt. "ZIKV Envelope Domain-Specific Antibodies: Production, Purification and Characterization." Viruses 11, no. 8 (August 13, 2019): 748. http://dx.doi.org/10.3390/v11080748.

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Infection with Zika virus (ZIKV) came first to public attention after it was found to be associated with congenital microcephaly during the outbreak in Brazil (2015–2016). Diagnosis of ZIKV suffers from extensive cross-reactivity with other Flaviviruses, which are circulating in many ZIKV epidemic areas. Due to the fatal outcome of ZIKV infection during pregnancy, detailed knowledge about neutralizing and non-neutralizing epitopes is crucial for the development of robust detection systems of protective antibodies. Therefore, additional information about ZIKV immunogenicity and antibody response is required. In this project, we report the production, purification and characterization of six different polyclonal antibodies against ZIKV envelope (E) protein. The produced antibodies bind to isolated ZIKV E protein as well as to the surface of ZIKV particles, interestingly without being potently neutralizing. Surface plasmon resonance measurement showed that these antibodies bind with high affinity to ZIKV E protein. Epitope mapping revealed that the epitopes are distributed among the three ZIKV E domains with seven binding sites. These identified binding sites overlap only partially with the previously described epitopes recognized by neutralizing antibodies, which is in accordance with their lack of potent neutralizing activity. Additionally, these antibodies showed neither cross-reactivity nor potent neutralizing activity against West Nile virus, a related flavivirus. The gained set of data helps to extend our understanding about the distribution of neutralizing and non-/weak-neutralizing epitopes in ZIKV E protein, and provides a rationale for ZIKV vaccine design and development of robust detection assays for neutralizing antibodies.

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Silva, Monaíse M. O., Laura B. Tauro, Mariana Kikuti, Rosângela O. Anjos, Viviane C. Santos, Thaiza S. F. Gonçalves, Igor A. D. Paploski, et al. "Concomitant Transmission of Dengue, Chikungunya, and Zika Viruses in Brazil: Clinical and Epidemiological Findings From Surveillance for Acute Febrile Illness." Clinical Infectious Diseases 69, no. 8 (December 18, 2018): 1353–59. http://dx.doi.org/10.1093/cid/ciy1083.

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Abstract Background Since their emergence in the Americas, chikungunya (CHIKV) and Zika (ZIKV) viruses co-circulate with dengue virus (DENV), hampering clinical diagnosis. We investigated clinical and epidemiological characteristics of arboviral infections during the introduction and spread of CHIKV and ZIKV through northeastern Brazil. Methods Surveillance for arboviral diseases among febrile patients was performed at an emergency health unit of Salvador, Brazil, between September 2014 and July 2016. We interviewed patients to collect data on symptoms, reviewed medical records to obtain the presumptive diagnoses, and performed molecular and serological testing to confirm DENV, CHIKV, ZIKV, or nonspecific flavivirus (FLAV) diagnosis. Results Of 948 participants, 247 (26.1%) had an acute infection, of which 224 (23.6%) were single infections (DENV, 32 [3.4%]; CHIKV, 159 [16.7%]; ZIKV, 13 [1.4%]; and FLAV, 20 [2.1%]) and 23 (2.4%) coinfections (DENV/CHIKV, 13 [1.4%]; CHIKV/FLAV, 9 [0.9%]; and DENV/ZIKV, 1 [0.1%]). An additional 133 (14.0%) patients had serological evidence for a recent arboviral infection. Patients with ZIKV presented with rash and pruritus (69.2% each) more frequently than those with DENV (37.5% and 31.2%, respectively) and CHIKV (22.9% and 14.7%, respectively) (P < .001 for both comparisons). Conversely, arthralgia was more common in CHIKV (94.9%) and FLAV/CHIKV (100.0%) than in DENV (59.4%) and ZIKV (53.8%) (P < .001). A correct presumptive clinical diagnosis was made for 9%–23% of the confirmed patients. Conclusions Arboviral infections are frequent causes of febrile illness. Coinfections are not rare events during periods of intense, concomitant arboviral transmission. Given the challenge to clinically distinguish these infections, there is an urgent need for rapid, point-of-care, multiplex diagnostics.

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Islam, Md Daharul, SM Tajdit Rahman, Khaleda Akhter, Md Azizul Hoque, Anannya Roy, and Adiba Tarannum. "Zika Virus: A Clinical Review." KYAMC Journal 7, no. 1 (August 29, 2017): 719–25. http://dx.doi.org/10.3329/kyamcj.v7i1.33766.

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Zika virus is a flavivirus related to Dengue virus, yellow fever virus and West Nile virus. It is considered an emerging arbovirus transmitted by mosquito of the genus Aedes. Its first description took place in 1947 in the Zika Forest in Uganda, isolated on Rhesus monkey used as bait to study the yellow fever virus. Clinical picture is characterized as a 'dengue-like' syndrome, with abrupt onset of fever; and an early onset of evanescent rash, often pruritic. Occasionally the disease has been associated with Guillain-Barré syndrome. The diagnosis can be performed by PCR or by IgG and IgM antibodies detection. No specific treatment or vaccine is available for Zika virus disease. Treatment is generally supportive. Control measures are same for dengue and chikungunya based mostly on health education and vector control.KYAMC Journal Vol. 7, No.-1, Jul 2016, Page 719-725

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Naidu, Y. Thathayya, and R. Kiranmai. "A Study on Role of Ultrasonography in the Diagnosis of Dengue Fever." Asian Journal of Medical Radiological Research 8, no. 1 (July 5, 2020): 139–43. http://dx.doi.org/10.47009/ajmrr.2020.8.1.25.

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Background: Dengue Fever is an acute mosquito transmitted viral infection caused by one of the 4 serotypes of the genus flavivirus which has become a major international public health problem. The diagnosis of DF is often delayed owing to time taken for availability of serology test results. Moreover, this test is expensive and not widely available. Ultrasonography (USG) is a cheap, rapid and widely available non-invasive imaging method. In recent years several studies concluded that Ultrasonography of the chest and abdomen can be an important adjunct to clinical profile in diagnosis of DF and diagnosis can be made early in the course of the disease compared with other modes of diagnosis. The aim of the study is to demonstrate the ultrasound findings of dengue fever and also evaluate the specificity of gall bladder findings in dengue fever. Subjects and Methods: We conducted a prospective study in Govt Medical College& Hospital, Srikakulam and Konaseema Institute of Medical Sciences, Amalapuram, A.P. Study included 50 patients referred to the department of Radio-Diagnosis and Imaging for Ultrasonography with clinical suspicion of dengue fever, during a period of July 2018 to December 2019. USG of the abdomen, pelvis and chest was performed in all cases and findings were noted. Dengue serology was performed later and all the ultrasound findings were correlated with dengue serology. Results: In our study of 50 patients all the patients studied were diagnosed with dengue fever based on dengue serology. In our study, 100% of our patients diagnosed with DF (by dengue serology) showed gall bladder wall thickening, 88% showed splenomegaly, 44% showed ascites. Pleural effusion was present in 30% of which 66.66% of pleural effusion was bilateral and the rest 33.33% was right sided. Isolated left sided pleural effusion was not found in our study. Hepatomegaly was present in 28% of our patients. In our study mortality and complications from dengue fever were not seen. Conclusion: Ultrasound findings in dengue fever are gall bladder wall thickening, splenomegaly, ascites, pleural effusion and hepatomegaly. In an area where DF is an epidemic, when Ultrasonography shows gall bladder wall thickening in a febrile patient with thrombocytopenia DF should be suggested On Ultrasonography, when there is gall bladder wall thickening, splenomegaly, ascites, and pleural effusion in a febrile patient with thrombocytopenia in a DF epidemic area a diagnosis of DF should be considered in a differential diagnosis until proved otherwise.

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Lustig, Yaniv, Hana Zelena, Giulietta Venturi, Marjan Van Esbroeck, Camilla Rothe, Cecilia Perret, Ravit Koren, Shiri Katz-Likvornik, Ella Mendelson, and Eli Schwartz. "Sensitivity and Kinetics of an NS1-Based Zika Virus Enzyme-Linked Immunosorbent Assay in Zika Virus-Infected Travelers from Israel, the Czech Republic, Italy, Belgium, Germany, and Chile." Journal of Clinical Microbiology 55, no. 6 (April 5, 2017): 1894–901. http://dx.doi.org/10.1128/jcm.00346-17.

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ABSTRACT Serological diagnosis of Zika virus is challenging due to high cross-reactivity of Zika virus with other flavivirus antibodies. Recently, a Zika NS1-based enzyme-linked immunosorbent assay (ELISA) was developed and shown to be highly specific for Zika antibody detection; however, sensitivity was evaluated for only a small number of confirmed Zika-infected patients. In this study, we measured the sensitivity and kinetics of Zika IgM and IgG antibodies using the Zika NS1-based ELISA in 105 samples from 63 returning travelers infected with Zika virus (proven by PCR or neutralization assay) from Israel, Czech Republic, Italy, Belgium, Germany, and Chile. Zika virus IgM was detected from 2 to 42 days post-symptom onset (PSO) with an overall sensitivity of 79% in the first month and 68% until 2 months PSO, while IgG antibodies were detected from 5 days to 3 years PSO with 79% sensitivity. Interestingly, significant differences in IgM sensitivity and IgM detection period were observed between Israeli and European/Chilean Zika-infected travelers, adding to the complexity of Zika infection diagnosis and suggesting that other diagnostic methods should be complemented to reduce false-negative results.

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Kazachinskaya, E. I., D. V. Shan’shin, and A. V. Ivanova. "Zika Fever: Development of Diagnostics, Prevention and Treatment." Problems of Particularly Dangerous Infections, no. 2 (July 3, 2019): 6–13. http://dx.doi.org/10.21055/0370-1069-2019-2-6-13.

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This review is devoted to the analysis of the literature data on the development of tools for diagnostics of Zika fever and detection of etiological agent – Zika virus (ZIKV) belonging to the Flaviviridae family. Preventive vaccines and antiviral drugs are also considered. RT-PCR method is critical for confirmation of Zika fever diagnosis. ZIKV RNA may be detected in blood serum, saliva, amniotic and cerebrospinal fluids, urine, semen, vaginal and cervical secretions. The duration of viremia in case of Zika fever is short; therefore the presence of ZIKV RNA in urine and sperm for up to 26 and 80 days, respectively, extends the time interval for the detection of this pathogen. Detection of IgM antibodies by serological methods is not a good reason to confirm a recent infection, since antibodies of this class, specific to flaviviruses, circulate in the bloodstream for more than 12 weeks. The IgM show high diagnostic value in confirmation of congenital infection only. There is a problem of differential diagnostics of flavivirus infections caused by antigenically related viruses that are dangerous for humans, for instance, Dengue, Yellow fever, West Nile fever viruses, tick-borne and Japanese encephalitis viruses. It is associated with the similarity of their genomes and, consequently, similar antigenic structure of viral proteins, structural glycoprotein E in particular. More reliable results can be obtained by using the nonstructural glycoprotein NS1, produced by molecular biology methods, as an antigen for the detection of specific antibodies. This viral protein can also be used in serological tests, as a clinical indicator in case of acute Zika fever. Forty five types of candidate vaccines against ZIKV, such as inactivated, live attenuated, recombinant, peptide, recombinant DNA and RNA-based, virus-vector and virus-like particle ones were designed and studied. It was established that their protective efficacy is mediated by induced antibodies, specific to structural glycoprotein E which initiates receptor binding and fusion with the membranes of infected cells. Currently, there is no licensed preparation for treating patients with flaviviral infections. Various drugs are screened, both with known antiviral effect and approved for use in clinical practice, and new compounds that inhibit the penetration of viral particles into host cells (structural glycoprotein E being the target) and virus replication (targets are NS5, NS2B nonstructural proteins).

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Journal articles on the topic 'Flavivirus diagnosis'

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