scholarly journals Antibody-based assay discriminates Zika virus infection from other flaviviruses

2017 ◽  
Vol 114 (31) ◽  
pp. 8384-8389 ◽  
Author(s):  
Angel Balmaseda ◽  
Karin Stettler ◽  
Raquel Medialdea-Carrera ◽  
Damaris Collado ◽  
Xia Jin ◽  
...  
Keyword(s):  
Zika Virus ◽  
Nonstructural Protein ◽  
Serological Test ◽  
Low Cost ◽  
Human Monoclonal Antibody ◽  
Cross Reactivity ◽  
Control Group ◽  
Rt Pcr ◽  
Mild Disease ◽  
Nonstructural Protein 1

Zika virus (ZIKV) is a mosquito-borne flavivirus that emerged recently as a global health threat, causing a pandemic in the Americas. ZIKV infection mostly causes mild disease, but is linked to devastating congenital birth defects and Guillain-Barré syndrome in adults. The high level of cross-reactivity among flaviviruses and their cocirculation has complicated serological approaches to differentially detect ZIKV and dengue virus (DENV) infections, accentuating the urgent need for a specific and sensitive serological test. We previously generated a ZIKV nonstructural protein 1 (NS1)-specific human monoclonal antibody, which we used to develop an NS1-based competition ELISA. Well-characterized samples from RT-PCR-confirmed patients with Zika and individuals exposed to other flavivirus infections or vaccination were used in a comprehensive analysis to determine the sensitivity and specificity of the NS1 blockade-of-binding (BOB) assay, which was established in laboratories in five countries (Nicaragua, Brazil, Italy, United Kingdom, and Switzerland). Of 158 sera/plasma from RT-PCR-confirmed ZIKV infections, 145 (91.8%) yielded greater than 50% inhibition. Of 171 patients with primary or secondary DENV infections, 152 (88.9%) scored negative. When the control group was extended to patients infected by other flaviviruses, other viruses, or healthy donors (n= 540), the specificity was 95.9%. We also analyzed longitudinal samples from DENV-immune and DENV-naive ZIKV infections and found inhibition was achieved within 10 d postonset of illness and maintained over time. Thus, the Zika NS1 BOB assay is sensitive, specific, robust, simple, low-cost, and accessible, and can detect recent and past ZIKV infections for surveillance, seroprevalence studies, and intervention trials.

Serological Differences after Acute Zika Virus Infections between Children and Adults: Implication for Use of a Serological Test

2021 ◽  
Author(s):  
Jurai Wongsawat ◽  
Patama Suttha ◽  
Sumalee Chanama ◽  
Somkid Srisopa ◽  
Nichapa Yonchoho ◽  
...  
Keyword(s):  
Zika Virus ◽  
Nonstructural Protein ◽  
Serological Test ◽  
Cross Reactivity ◽  
Virus Infections ◽  
Positive Real ◽  
Nonstructural Protein 1 ◽  
Age Related ◽  
Polymerase Chain ◽  
Post Onset

Information is limited regarding differential serological responses after acute Zika virus (ZIKV) infections and prevalence of cross-reactivity with anti-dengue virus (DENV) assays comparing children and adults. Early convalescent sera from a cohort of suspected mild DENV cases between December 2016 and September 2018 at Bamrasnaradura Infectious Diseases Institute in Thailand were tested for nonstructural protein 1 (NS1)–based anti-ZIKV IgM and IgG ELISAs (Euroimmun), and in-house anti-DENV IgM- and IgG-capture ELISAs. ZIKV cases were identified by positive real-time reverse transcriptase-polymerase chain reaction on urine. Sera from 26 (10 children and 16 adults) ZIKV and 237 (153 children and 74 adults) non-ZIKA cases collected at the median duration of 18 days (interquartile range [IQR] 18,19) post-onset of symptoms were tested. Comparing pediatric ZIKV to adult ZIKV cases, the mean anti-ZIKV IgM ratio was higher (2.12 versus 1.27 units, respectively; P = 0.07), whereas mean anti-ZIKV IgG ratio was lower (3.13 versus 4.24 units, respectively; P = 0.03). Sensitivity of anti-ZIKV IgM and specificity of anti-ZIKV IgG in pediatric ZIKV were higher than in adult ZIKV cases (80.0% versus 43.7% and 79.1% versus 43.2%, respectively). No cross-reactivity with anti-DENV IgM- and IgG-capture ELISA were reported in pediatric ZIKV cases in our study, whereas 25% and 12.5% were found in adult ZIKV cases, respectively. Age-related ZIKV serological differences have been observed. Positive NS1-based anti-ZIKV IgM and IgG ELISA at the early convalescent phase could be useful for ZIKV diagnosis in children, even in a dengue endemic setting.

scholarly journals The immunodominant antibody response to Zika virus NS1 protein is characterized by cross-reactivity to self

2021 ◽  
Vol 218 (9) ◽  
Author(s):  
Cecilia B. Cavazzoni ◽  
Vicente B.T. Bozza ◽  
Tostes C.V. Lucas ◽  
Luciana Conde ◽  
Bruno Maia ◽  
...  
Keyword(s):  
Immune Response ◽  
B Cell ◽  
Humoral Immune Response ◽  
Zika Virus ◽  
Nonstructural Protein ◽  
Cross Reactivity ◽  
Ns1 Protein ◽  
Humoral Immune ◽  
Nonstructural Protein 1 ◽  
Autoreactive Antibodies

Besides antigen-specific responses to viral antigens, humoral immune response in virus infection can generate polyreactive and autoreactive antibodies. Dengue and Zika virus infections have been linked to antibody-mediated autoimmune disorders, including Guillain-Barré syndrome. A unique feature of flaviviruses is the secretion of nonstructural protein 1 (NS1) by infected cells. NS1 is highly immunogenic, and antibodies targeting NS1 can have both protective and pathogenic roles. In the present study, we investigated the humoral immune response to Zika virus NS1 and found NS1 to be an immunodominant viral antigen associated with the presence of autoreactive antibodies. Through single B cell cultures, we coupled binding assays and BCR sequencing, confirming the immunodominance of NS1. We demonstrate the presence of self-reactive clones in germinal centers after both infection and immunization, some of which present cross-reactivity with NS1. Sequence analysis of anti-NS1 B cell clones showed sequence features associated with pathogenic autoreactive antibodies. Our findings demonstrate NS1 immunodominance at the cellular level as well as a potential role for NS1 in ZIKV-associated autoimmune manifestations.

scholarly journals Highly Sensitive Detection of Zika Virus Nonstructural Protein 1 in Serum Samples by a Two-Site Nanobody ELISA

Biomolecules ◽  
2020 ◽  
Vol 10 (12) ◽  
pp. 1652
Author(s):  
Triana Delfin-Riela ◽  
Martín Rossotti ◽  
Romina Alvez-Rosado ◽  
Carmen Leizagoyen ◽  
Gualberto González-Sapienza
Keyword(s):  
Zika Virus ◽  
High Throughput Screening ◽  
Nonstructural Protein ◽  
Low Cost ◽  
Blood Stream ◽  
Ns1 Protein ◽  
Serum Samples ◽  
Nonstructural Protein 1 ◽  
Highly Sensitive ◽  
Highly Sensitive Detection

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.

Nanovaccine based on self-assembling nonstructural protein 1 boosts antibody responses to Zika virus

2021 ◽  
Vol 32 ◽  
pp. 102334
Author(s):  
Marianna Teixeira Pinho Favaro ◽  
Monica Josiane Rodrigues-Jesus ◽  
Alexia Adrianne Venceslau-Carvalho ◽  
Rúbens Prince Dos Santos Alves ◽  
Lennon Ramos Pereira ◽  
...  
Keyword(s):  
Zika Virus ◽  
Nonstructural Protein ◽  
Antibody Responses ◽  
Self Assembling ◽  
Nonstructural Protein 1

scholarly journals Comparison of Four Serological Methods and Two Reverse Transcription-PCR Assays for Diagnosis and Surveillance of Zika Virus Infection

2018 ◽  
Vol 56 (3) ◽  
Author(s):  
Angel Balmaseda ◽  
José Victor Zambrana ◽  
Damaris Collado ◽  
Nadezna García ◽  
Saira Saborío ◽  
...  
Keyword(s):  
Reverse Transcription ◽  
Zika Virus ◽  
Clinical Manifestations ◽  
Cross Reactivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Emerging Infections ◽  
Rt Pcr ◽  
Convalescent Phase ◽  
Reverse Transcription Pcr ◽  
Pcr Assays

ABSTRACTZika virus (ZIKV) is a mosquito-borne flavivirus that is responsible for recent explosive epidemics in the Americas. Notably, ZIKV infection during pregnancy has been found to cause congenital birth defects, including microcephaly, and ZIKV has been associated with Guillain-Barré syndrome in adults. Diagnosis and surveillance of Zika in the Americas have been challenging due to similar clinical manifestations and extensive antibody cross-reactivity with endemic flaviviral diseases, such as dengue. We evaluated four serological and two reverse transcription-PCR (RT-PCR) methods in acute-phase (mean day, 1.8), early-convalescent-phase (mean day, 16.7), and late-convalescent-phase (mean, ~7 months) samples from the same individuals in a long-term pediatric cohort study in Nicaragua. Well-characterized samples from 301 cases of Zika, dengue, or non-Zika, nondengue febrile illnesses were tested. Compared to a composite reference, an in-house IgM antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) and the NIAID-Biodefense and Emerging Infections (BEI) MAC-ELISA measuring IgM yielded sensitivities of 94.5% and 70.1% and specificities of 85.6% and 82.8%, respectively. The NS1 blockade-of-binding ELISA measuring anti-ZIKV NS1 antibody levels yielded sensitivities of 85.0% and 96.5% and specificities of 91.4% and 92.6% at early and late convalescence, respectively. An inhibition ELISA detecting total anti-ZIKV antibodies had sensitivity and specificity values of 68.3% and 58.3% for diagnosis and 94.0% and 98.6% for measuring annual infection incidence. Finally, the ZCD and Trioplex real-time RT-PCR assays detecting Zika, chikungunya, and dengue viruses both yielded a sensitivity of 96.1% and specificity of 100%. Together, these assays resolve the urgent need for diagnostic and surveillance tools for countries affected by Zika virus infections.

scholarly journals Attenuation of Zika Virus by Passage in Human HeLa Cells

Vaccines ◽  
2019 ◽  
Vol 7 (3) ◽  
pp. 93 ◽  
Author(s):  
Li ◽  
Collins ◽  
Widen ◽  
Davis ◽  
Kaiser ◽  
...  
Keyword(s):  
Hela Cell ◽  
Hela Cells ◽  
Zika Virus ◽  
Infected Animal ◽  
Vaccine Development ◽  
Nonstructural Protein ◽  
A549 Cells ◽  
Wild Type ◽  
Nonstructural Protein 1

Zika virus (ZIKV) is a mosquito-borne Flavivirus. Previous studies have shown that mosquito-transmitted flaviviruses, including yellow fever, Japanese encephalitis, and West Nile viruses, could be attenuated by serial passaging in human HeLa cells.  Therefore, it was hypothesized that wild-type ZIKV would also be attenuated after HeLa cell passaging. A human isolate from the recent ZIKV epidemic was subjected to serial HeLa cell passaging, resulting in attenuated in vitro replication in both Vero and A549 cells. Additionally, infection of AG129 mice with 10 plaque forming units (pfu) of wild-type ZIKV led to viremia and mortality at 12 days, whereas infection with 103 pfu of HeLa-passage 6 (P6) ZIKV led to lower viremia, significant delay in mortality (median survival: 23 days), and increased cytokine and chemokine responses.  Genomic sequencing of HeLa-passaged virus identified two amino acid substitutions as early as HeLa-P3: pre-membrane E87K and nonstructural protein 1 R103K. Furthermore, both substitutions were present in virus harvested from HeLa-P6-infected animal tissue. Together, these data show that, similarly to other mosquito-borne flaviviruses, ZIKV is attenuated following passaging in HeLa cells. This strategy can be used to improve understanding of substitutions that contribute to attenuation of ZIKV and be applied to vaccine development across multiple platforms.

scholarly journals Equine Arteritis Virus Does Not Induce Interferon Production in Equine Endothelial Cells: Identification of Nonstructural Protein 1 as a Main Interferon Antagonist

2014 ◽  
Vol 2014 ◽  
pp. 1-13 ◽  
Author(s):  
Yun Young Go ◽  
Yanhua Li ◽  
Zhenhai Chen ◽  
Mingyuan Han ◽  
Dongwan Yoo ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Nonstructural Protein ◽  
Critical Role ◽  
Equine Arteritis Virus ◽  
Luciferase Reporter ◽  
Type I ◽  
Mrna Levels ◽  
Type I Ifn ◽  
Rt Pcr ◽  
Nonstructural Protein 1

The objective of this study was to investigate the effect of equine arteritis virus (EAV) on type I interferon (IFN) production. Equine endothelial cells (EECs) were infected with the virulent Bucyrus strain (VBS) of EAV and expression of IFN-βwas measured at mRNA and protein levels by quantitative real-time RT-PCR and IFN bioassay using vesicular stomatitis virus expressing the green fluorescence protein (VSV-GFP), respectively. Quantitative RT-PCR results showed that IFN-βmRNA levels in EECs infected with EAV VBS were not increased compared to those in mock-infected cells. Consistent with quantitative RT-PCR, Sendai virus- (SeV-) induced type I IFN production was inhibited by EAV infection. Using an IFN-βpromoter-luciferase reporter assay, we subsequently demonstrated that EAV nsps 1, 2, and 11 had the capability to inhibit type I IFN activation. Of these three nsps, nsp1 exhibited the strongest inhibitory effect. Taken together, these data demonstrate that EAV has the ability to suppress the type I IFN production in EECs and nsp1 may play a critical role to subvert the equine innate immune response.

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