scholarly journals Development of a highly specific serodiagnostic ELISA for West Nile virus infection using subviral particles

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Keisuke Maezono ◽  
Shintaro Kobayashi ◽  
Koshiro Tabata ◽  
Kentaro Yoshii ◽  
Hiroaki Kariwa
Keyword(s):  
West Nile Virus ◽  
West Nile Virus Infection ◽  
Cross Reactivity ◽  
Terminal Region ◽  
Full Length ◽  
Enzyme Linked Immunosorbent Assay ◽  
E Proteins ◽  
West Nile ◽  
E Protein ◽  
Subviral Particles

AbstractWest Nile virus (WNV), a member of the Japanese encephalitis virus (JEV) serocomplex group, causes lethal encephalitis in humans and horses. Because serodiagnosis of WNV and JEV is hampered by cross-reactivity, the development of a simple, secure, and WNV-specific serodiagnostic system is required. The coexpression of prM protein and E protein leads to the secretion of subviral particles (SPs). Deletion of the C-terminal region of E protein is reported to affect the production of SPs by some flaviviruses. However, the influence of such a deletion on the properties and antigenicity of WNV E protein is unclear. We analyzed the properties of full-length E protein and E proteins lacking the C-terminal region as novel serodiagnostics for WNV infection. Deletion of the C-terminal region of E protein suppressed the formation of SPs but did not affect the production of E protein. The sensitivity of an enzyme-linked immunosorbent assay (ELISA) using the full-length E protein was higher than that using the truncated E proteins. Furthermore, in the ELISA using full-length E protein, there was little cross-reactivity with anti-JEV antibodies, and the sensitivity was similar to that of the neutralization test.

scholarly journals N-Linked Glycosylation of West Nile Virus Envelope Proteins Influences Particle Assembly and Infectivity

2005 ◽  
Vol 79 (21) ◽  
pp. 13262-13274 ◽  
Author(s):  
Sheri L. Hanna ◽  
Theodore C. Pierson ◽  
Melissa D. Sanchez ◽  
Asim A. Ahmed ◽  
Mariam M. Murtadha ◽  
...  
Keyword(s):  
West Nile Virus ◽  
Envelope Proteins ◽  
Glycosylation Site ◽  
Protein Glycosylation ◽  
E Proteins ◽  
Virus Envelope ◽  
West Nile ◽  
Particle Assembly ◽  
E Protein ◽  
The Impact

ABSTRACT West Nile virus (WNV) encodes two envelope proteins, premembrane (prM) and envelope (E). While the prM protein of all WNV strains contains a single N-linked glycosylation site, not all strains contain an N-linked site in the E protein. The presence of N-linked glycosylation on flavivirus E proteins has been linked to virus production, pH sensitivity, and neuroinvasiveness. Therefore, we examined the impact of prM and E glycosylation on WNV assembly and infectivity. Similar to other flaviviruses, expression of WNV prM and E resulted in the release of subviral particles (SVPs). Removing the prM glycosylation site in a lineage I or II strain decreased SVP release, as did removal of the glycosylation site in a lineage I E protein. Addition of the E protein glycosylation site in a lineage II strain that lacked this site increased SVP production. Similar results were obtained in the context of either reporter virus particles (RVPs) or infectious lineage II WNV. RVPs or virions bearing combinations of glycosylated and nonglycosylated forms of prM and E could infect mammalian, avian, and mosquito cells (BHK-21, QT6, and C6/36, respectively). Those particles lacking glycosylation on the E protein were modestly more infectious per genome copy on BHK-21 and QT6 cells, while this absence greatly enhanced the infection of C6/36 cells. Thus, glycosylation of WNV prM and E proteins can affect the efficiency of virus release and infection in a manner that is cell type and perhaps species dependent. This suggests a multifaceted role for envelope N-linked glycosylation in WNV biology and tropism.

scholarly journals Establishment of an Algorithm Using prM/E- and NS1-Specific IgM Antibody-Capture Enzyme-Linked Immunosorbent Assays in Diagnosis of Japanese Encephalitis Virus and West Nile Virus Infections in Humans

2015 ◽  
Vol 54 (2) ◽  
pp. 412-422 ◽  
Author(s):  
Jedhan U. Galula ◽  
Gwong-Jen J. Chang ◽  
Shih-Te Chuang ◽  
Day-Yu Chao
Keyword(s):  
West Nile Virus ◽  
Japanese Encephalitis Virus ◽  
Japanese Encephalitis ◽  
Cross Reactivity ◽  
Encephalitis Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Test Accuracy ◽  
West Nile ◽  
Igm Antibody ◽  
Antibody Capture

The front-line assay for the presumptive serodiagnosis of acute Japanese encephalitis virus (JEV) and West Nile virus (WNV) infections is the premembrane/envelope (prM/E)-specific IgM antibody-capture enzyme-linked immunosorbent assay (MAC-ELISA). Due to antibody cross-reactivity, MAC-ELISA-positive samples may be confirmed with a time-consuming plaque reduction neutralization test (PRNT). In the present study, we applied a previously developed anti-nonstructural protein 1 (NS1)-specific MAC-ELISA (NS1-MAC-ELISA) on archived acute-phase serum specimens from patients with confirmed JEV and WNV infections and compared the results with prM/E containing virus-like particle-specific MAC-ELISA (VLP-MAC-ELISA). Paired-receiver operating characteristic (ROC) curve analyses revealed no statistical differences in the overall assay performances of the VLP- and NS1-MAC-ELISAs. The two methods had high sensitivities of 100% but slightly lower specificities that ranged between 80% and 100%. When the NS1-MAC-ELISA was used to confirm positive results in the VLP-MAC-ELISA, the specificity of serodiagnosis, especially for JEV infection, was increased to 90% when applied in areas where JEV cocirculates with WNV, or to 100% when applied in areas that were endemic for JEV. The results also showed that using multiple antigens could resolve the cross-reactivity in the assays. Significantly higher positive-to-negative (P/N) values were consistently obtained with the homologous antigens than those with the heterologous antigens. JEV or WNV was reliably identified as the currently infecting flavivirus by a higher ratio of JEV-to-WNV P/N values or vice versa. In summary of the above-described results, the diagnostic algorithm combining the use of multiantigen VLP- and NS1-MAC-ELISAs was developed and can be practically applied to obtain a more specific and reliable result for the serodiagnosis of JEV and WNV infections without the need for PRNT. The developed algorithm should provide great utility in diagnostic and surveillance activities in which test accuracy is of utmost importance for effective disease intervention.

scholarly journals Development of Monoclonal Antibodies to West Nile Virus and Their Application in Immunohistochemistry

2012 ◽  
Vol 19 (11) ◽  
pp. 1853-1858 ◽  
Author(s):  
Jiro Hirota ◽  
Shinya Shimizu ◽  
Tomoyuki Shibahara ◽  
Takashi Isobe ◽  
Manabu Yamada ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
West Nile Virus ◽  
Japanese Encephalitis ◽  
Nonstructural Protein ◽  
Cross Reactivity ◽  
Encephalitis Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
West Nile ◽  
Nonstructural Protein 1 ◽  
Formalin Fixed Paraffin Embedded

ABSTRACTWest Nile virus (WNV) is endemic throughout Africa, Eurasia, America, and Australia and has important implications for avian, horse, and human health. In these regions, dead birds are monitored for the presence of WNV through immunohistochemistry (IHC) and PCR. However, a number of the tools for IHC are inadequate owing to their cross-reactivity to other Japanese encephalitis serogroup viruses. Here we have established eight monoclonal antibodies (MAbs) to WNV. Four of them bound to the envelope protein, three of them bound to nonstructural protein 1 (NS1), and one bound to precursor membrane protein (prM), as shown by Western blot analysis. The anti-NS1 MAbs and the anti-prM MAb did not cross-react with Japanese encephalitis virus (JEV), Murray valley encephalitis virus, or St. Louis encephalitis virus in an indirect enzyme-linked immunosorbent assay. One NS1-specific MAb, SHW-32B1, and the previously reported NS1-specific MAb, SHW-7A11, were shown by IHC to specifically detect the cytoplasm of degenerated cells in the heart and brain of a WNV-infected goose. Neither of these MAbs were shown by IHC to cross-react with degenerated cells in the brain of a JEV-infected pig. These MAbs are the first reported anti-NS1 MAbs that can be used for WNV-specific IHC using formalin-fixed, paraffin-embedded sections. They may be useful for WNV research and surveillance.

scholarly journals Immunological Cross-Reactivity to Dengue Virus among Persons with Neuroinvasive West Nile Virus Infection

2022 ◽  
Author(s):  
Vanessa N Raabe ◽  
Muktha S Natrajan ◽  
Christopher M Huerta ◽  
Yongxian Xu ◽  
Lilin Lai ◽  
...  
Keyword(s):  
West Nile Virus ◽  
Virus Infection ◽  
Dengue Virus ◽  
West Nile Virus Infection ◽  
Cross Reactivity ◽  
Igg Antibodies ◽  
West Nile ◽  
Dengue Viruses ◽  
Antibody Dependent Enhancement

Antibody dependent enhancement has been well described between Zika and dengue viruses, but is poorly characterized between West Nile and dengue viruses. We demonstrate that neuroinvasive West Nile virus infection leads to the development of non-neutralizing, cross-reactive IgG antibodies to dengue and Zika viruses capable of causing antibody dependent enhancement in vitro of dengue virus and leads to the formation of flavivirus cross-reactive memory B cells in some patients.

scholarly journals Seroprevalence and risk factors of West Nile virus infection in veterinarians and horses in Northern Palestine

2021 ◽  
pp. 1241-1245
Author(s):  
Ibrahim Alzuheir ◽  
Adnan Fayyad ◽  
Nasr Jalboush ◽  
Rosemary Abdallah ◽  
Sameeh Abutarbush ◽  
...  
Keyword(s):  
Risk Factors ◽  
West Nile Virus ◽  
Horse Serum ◽  
Age Groups ◽  
West Nile Virus Infection ◽  
Enzyme Linked Immunosorbent Assay ◽  
West Nile Fever ◽  
Serum Samples ◽  
West Nile ◽  
And Control

Background and Aim: West Nile fever (WNF) is a neurotropic, mosquito-borne disease affecting humans and domesticated animals, caused by a member of the genus Flavivirus. Over the last decades, this virus has been responsible for several cases of illness in humans and animals. The current epidemiological status of WNF in horses is insufficient, and in veterinarians, as an occupational hazard is unknown. This study aimed to investigate and determine the seroprevalence and risk factors for WNF in veterinarians and horses in Palestine. Materials and Methods: In this study, serum samples from 100 veterinarians and 87 horses were collected between August 2020 and September 2020 from different cities of Northern Palestine. West Nile virus (WNV) antibodies were detected using an enzyme-linked immunosorbent assay. Results: Our results showed that 60.9% of the horse serum samples were positive in all investigated cities. In horses, location is a risk factor for the seropositivity for WNF, whereas age, sex, breed, and intended use of the horses, were not associated with increased WNF seropositivity. In veterinarians, 23.0% of the serum samples were positive. Positive samples were detected in all locations, age groups, experience length, and work sectors. However, the seropositivity for WNF was not influenced by these variables. Conclusion: The results revealed that WNV circulates in most regions of Palestine. Our results will help determine the risk of infection in animals and humans and control WNV transmission. Surveillance studies on humans, vectors, and animals are needed to better define endemic areas.

scholarly journals New Sensitive Competitive Enzyme-Linked Immunosorbent Assay Using a Monoclonal Antibody against Nonstructural Protein 1 of West Nile Virus NY99

2011 ◽  
Vol 19 (2) ◽  
pp. 277-283 ◽  
Author(s):  
Jiro Hirota ◽  
Yoshihiro Shimoji ◽  
Shinya Shimizu
Keyword(s):  
Monoclonal Antibody ◽  
West Nile Virus ◽  
Neutralization Test ◽  
Nonstructural Protein ◽  
Cross Reactivity ◽  
Encephalitis Virus ◽  
The Other ◽  
Enzyme Linked Immunosorbent Assay ◽  
West Nile ◽  
Nonstructural Protein 1

ABSTRACTAn anti-West Nile virus (anti-WNV) monoclonal antibody, SHW-7A11, was developed for competitive enzyme-linked immunosorbent assays (c-ELISAs). SHW-7A11 reacted with nonstructural protein 1 in Western blot analysis. SHW-7A11 was relatively specific for the WNV strain NY99 and recognized Kunjin and Eg101 strains in indirect ELISAs. Two c-ELISAs were developed for sera diluted 10 and 100 times and named c-ELISA10 and c-ELISA100, respectively. Both c-ELISAs detected antibodies against WNV NY99 and Kunjin strains. Little cross-reactivity was observed for antibodies against Japanese encephalitis virus and St. Louis encephalitis virus in these assays. Using the cutoff point for the St. Louis encephalitis virus, all WNV-infected chickens were found to be positive on day 21 after infection in both c-ELISAs. On the other hand, all infected chickens were found to be positive on day 35 after infection in a virus neutralization test. Our newly developed SHW-7A11-based c-ELISA can detect WNV infection with sera diluted 10 to 100 times. Therefore, this c-ELISA can be used for WNV serosurveillance of chickens and wild birds.

scholarly journals Isolation and Characterization of Human Monoclonal Antibodies from Individuals Infected with West Nile Virus

2006 ◽  
Vol 80 (14) ◽  
pp. 6982-6992 ◽  
Author(s):  
Mark Throsby ◽  
Cecile Geuijen ◽  
Jaap Goudsmit ◽  
Arjen Q. Bakker ◽  
Jehanara Korimbocus ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
West Nile Virus ◽  
Viral Envelope ◽  
Phage Library ◽  
Enzyme Linked Immunosorbent Assay ◽  
West Nile ◽  
Isolation And Characterization ◽  
E Protein ◽  
Domain Iii

ABSTRACT Monoclonal antibodies (MAbs) neutralizing West Nile Virus (WNV) have been shown to protect against infection in animal models and have been identified as a correlate of protection in WNV vaccine studies. In the present study, antibody repertoires from three convalescent WNV-infected patients were cloned into an scFv phage library, and 138 human MAbs binding to WNV were identified. One hundred twenty-one MAbs specifically bound to the viral envelope (E) protein and four MAbs to the premembrane (prM) protein. Enzyme-linked immunosorbent assay-based competitive-binding assays with representative E protein-specific MAbs demonstrated that 24/51 (47%) bound to domain II while only 4/51 (8%) targeted domain III. In vitro neutralizing activity was demonstrated for 12 MAbs, and two of these, CR4374 and CR4353, protected mice from lethal WNV challenge at 50% protective doses of 12.9 and 357 μg/kg of body weight, respectively. Our data analyzing three infected individuals suggest that the human anti-WNV repertoire after natural infection is dominated by nonneutralizing or weakly neutralizing MAbs binding to domain II of the E protein, while domain III-binding MAbs able to potently neutralize WNV in vitro and in vivo are rare.

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