Evaluation of Two Serological Assays for Diagnosing Zika Virus Infection

ABSTRACT Measles is a highly contagious respiratory virus infection, with typical clinical symptoms including maculopapular rash, fever, cough, coryza, and conjunctivitis. Despite implementation of widespread vaccination programs throughout the world, the rates of global morbidity and mortality are still considerable. This study was performed to design a reliable indirect enzyme-linked immunosorbent assay (ELISA) to measure measles-specific immunoglobulin M (IgM). First, human IgM was purified, and then an anti-IgM antibody was produced in rabbits and purified in a multistep process. The rabbit IgG against human IgM was conjugated with peroxidase. Measles virus-infected Vero cells produced viral antigen. One hundred serum samples from infants of 9 to 18 months of age, mostly vaccinated, were evaluated for determining the presence of specific IgM antibodies against measles virus. The samples were also evaluated for neutralizing antibodies against measles virus by a microneutralization test (MNT). By comparing the results of the ELISA with those of MNT, it was demonstrated that ELISA had a sensitivity and specificity of 100 and 92%, respectively. On the other hand, when the results obtained by our ELISA system were compared with those of an imported measles virus IgM ELISA kit (EIAgen; Adaltis Italia SPa, Bologna, Italy), a high level of agreement was shown (k = 0.926).

Download Full-text

Serologic Testing for Zika Virus: Comparison of Three Zika Virus IgM-Screening Enzyme-Linked Immunosorbent Assays and Initial Laboratory Experiences

Journal of Clinical Microbiology ◽

10.1128/jcm.00580-17 ◽

2017 ◽

Vol 55 (7) ◽

pp. 2127-2136 ◽

Cited By ~ 58

Author(s):

Dane Granger ◽

Heather Hilgart ◽

Lori Misner ◽

Jaime Christensen ◽

Sarah Bistodeau ◽

...

Keyword(s):

Zika Virus ◽

Neutralizing Antibodies ◽

Enzyme Linked Immunosorbent Assay ◽

Reference Laboratory ◽

Zikv Infection ◽

Serologic Testing ◽

Igm Elisa ◽

Control And Prevention ◽

Laboratory Experiences ◽

Positive Agreement

ABSTRACT Serologic evaluation for Zika virus (ZIKV) infection currently includes an initial screen using an anti-ZIKV IgM antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) followed by supplemental testing of specimens with nonnegative results by a plaque reduction neutralization test (PRNT). We compared the performance characteristics of three ELISAs for the detection of IgM class antibodies to ZIKV, including the Centers for Disease Control and Prevention (CDC) Zika MAC-ELISA, the InBios ZIKV Detect MAC-ELISA, and the Euroimmun anti-Zika Virus IgM ELISA. Additionally, we present our initial experiences with ZIKV serologic testing from a national reference laboratory perspective. Using both retrospectively and prospectively collected specimens from patients with possible ZIKV infection, we show that the CDC and InBios MAC-ELISAs perform comparably to each other, with positive agreement, negative agreement, and interrater kappa values ranging from 87.5% to 93.1%, 95.7% to 98.5%, and 0.52 to 0.83, respectively. In contrast, comparison of the Euroimmun ZIKV ELISA to either the CDC or InBios MAC-ELISAs resulted in positive agreement, negative agreement, and interrater kappa values ranging from 17.9% to 42.9%, 91.7% to 98.6%, and 0.10 to 0.39, respectively. Among the 19 prospective samples submitted for PRNT, nine were negative, eight specimens had neutralizing antibodies to a flavivirus (unable to be identified), and one sample each was confirmed for ZIKV or dengue virus infection. This study highlights the ongoing challenges associated with serologic diagnosis of ZIKV infection. Although the availability of a commercial serologic test for ZIKV has greatly expanded the national capacity for such testing, the need to further characterize and improve these assays, particularly with regard to specificity, remains.

Download Full-text

High correlation between Zika virus NS1 antibodies and neutralizing antibodies in selected serum samples from normal healthy Thais

Scientific Reports ◽

10.1038/s41598-019-49569-0 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 3

Author(s):

Wannapa Sornjai ◽

Suwipa Ramphan ◽

Nitwara Wikan ◽

Prasert Auewarakul ◽

Duncan R. Smith

Keyword(s):

Southeast Asia ◽

Zika Virus ◽

Neutralizing Antibodies ◽

Neutralization Test ◽

Ns1 Protein ◽

Serum Samples ◽

Zikv Infection ◽

Ns1 Antigen ◽

Cross Neutralization ◽

Immunological Protection

Abstract Despite the widespread presence of the mosquito transmitted Zika virus (ZIKV) over much of Southeast Asia, the number of reported cases remains low. One possibility is that residents in Southeast Asia are immunologically protected, although the nature of any such protection remains unclear. This study sought to investigate the presence of antibodies directed to ZIKV NS1 protein in a selected sub-set of samples from a well characterized cohort of serum samples from normal, healthy Thais that had been previously characterized for the presence of neutralizing antibodies to ZIKV, DENV 1-4, and JEV. Because of similarities in molecular weight between the flavivirus E and NS1 proteins, an immunoblot system was established in which the NS1 antigen was not denatured, allowing detection of the dimer form of NS1, distinctly clear from the migration position of the E and NS1 monomer proteins. The results showed that antibodies to ZIKV NS1 protein were only detected in samples with ZIKV neutralizing antibodies (27/30 samples), and no sample (0/30) with a ZIKV plaque reduction neutralization test (PRNT)90 < 20 showed evidence of anti-ZIKV NS1 antibodies. The high correlation between the presence of ZIKV NS1 antibodies and ZIKV PRNT suggests that immunological protection against ZIKV infection in Thailand arises from prior exposure to ZIKV, and not through cross neutralization.

Download Full-text

Prevalence of Zika virus neutralizing antibodies in healthy adults in Vietnam during and after the Zika virus epidemic season: a longitudinal population-based survey

10.21203/rs.2.23060/v2 ◽

2020 ◽

Author(s):

Co Thach Nguyen ◽

Meng Ling Moi ◽

Thi Quynh Mai Le ◽

Thi Thu Thuy Nguyen ◽

Thi Bich Hau Vu ◽

...

Keyword(s):

Zika Virus ◽

Neutralizing Antibodies ◽

Neutralizing Antibody ◽

Fold Increase ◽

Immunoglobulin M ◽

Control Measures ◽

Serum Samples ◽

Zikv Infection ◽

Epidemic Season ◽

Low Levels

Abstract Background : Between 2016 and 2019, 265 cases of Zika virus (ZIKV) infection were reported in Vietnam, predominantly in southern Vietnam. In 2016, a case of ZIKV-associated microcephaly was confirmed in the Central Highlands, and several members of the infant’s family were confirmed to be infected with ZIKV. The study aims to determine the level of immunity to ZIKV in the general population of the ZIKV epidemic region. Methods: A total of 879 serum samples were collected from 801 participants between January 2017 and July 2018, during and after the ZIKV epidemic in Vietnam. The samples were tested for anti-ZIKV immunoglobulin M (IgM) and immunoglobulin G (IgG), and anti-dengue virus (DENV) IgG antibodies using enzyme-linked immunosorbent assays (ELISA). Plaque-reduction neutralization test (PRNT) for ZIKV was performed on all samples, and for DENV on the samples that ZIKV neutralizing antibody positive. Results: A total of 83 (10.3%) participants had anti-ZIKV IgM. Of the 83, 6 were confirmed to be ZIKV antibodies positive using PRNT and anti-ZIKV IgG ELISA. Of the 718 participants who were anti-ZIKV IgM negative, a further 3 cases were confirmed as positive for antibodies against ZIKV. Of the 9 participants with ZIKV infection, 5 lived in the same village as the infant with ZIKV-associated microcephaly and the other 4 lived in 2 neighboring communes. Repeat samples were collected from the 83 ZIKV IgM positive participants 1.5 years after the first collection. No new cases of ZIKV infection were detected. In addition, 2 of 3 participants with anti-ZIKV NS1 IgG demonstrated a 4- to 8-fold increase in ZIKV neutralizing antibody titer. Conclusions: ZIKV was present in the area around Krong Buk, with the rate of ZIKV-specific antibodies was 1.1% in the community since at least 2016. While the low levels of circulation together with low seroprevalence suggests a limited outbreak in the region, the results also reflect on low levels of protective immunity to Zika within the population. These results provide a better understanding of the current ZIKV epidemic status in the region and demonstrate a need for implementation of more effective ZIKV infection control measures.

Download Full-text

Prevalence of Zika virus neutralizing antibodies in healthy adults in Vietnam during and after the Zika virus epidemic season: a longitudinal population-based survey

10.21203/rs.2.23060/v3 ◽

2020 ◽

Author(s):

Co Thach Nguyen ◽

Meng Ling Moi ◽

Thi Quynh Mai Le ◽

Thi Thu Thuy Nguyen ◽

Thi Bich Hau Vu ◽

...

Keyword(s):

Zika Virus ◽

Neutralizing Antibodies ◽

Neutralizing Antibody ◽

Fold Increase ◽

Immunoglobulin M ◽

Control Measures ◽

Serum Samples ◽

Zikv Infection ◽

Epidemic Season ◽

Low Levels

Abstract Background : Between 2016 and 2019, 265 cases of Zika virus (ZIKV) infection were reported in Vietnam, predominantly in southern Vietnam. In 2016, a case of ZIKV-associated microcephaly was confirmed in the Central Highlands, and several members of the infant’s family were confirmed to be infected with ZIKV. The study aims to determine the level of immunity to ZIKV in the general population of the ZIKV epidemic region. Methods: A total of 879 serum samples were collected from 801 participants between January 2017 and July 2018, during and after the ZIKV epidemic in Vietnam. The samples were tested for anti-ZIKV immunoglobulin M (IgM) and immunoglobulin G (IgG), and anti-dengue virus (DENV) IgG antibodies using enzyme-linked immunosorbent assays (ELISA). Plaque-reduction neutralization test (PRNT) for ZIKV was performed on all samples, and for DENV on the samples that ZIKV neutralizing antibody positive. Results: A total of 83 (10.3%) participants had anti-ZIKV IgM. Of the 83, 6 were confirmed to be ZIKV antibodies positive using PRNT and anti-ZIKV IgG ELISA. Of the 718 participants who were anti-ZIKV IgM negative, a further 3 cases were confirmed as positive for antibodies against ZIKV. Of the 9 participants with ZIKV infection, 5 lived in the same village as the infant with ZIKV-associated microcephaly and the other 4 lived in 2 neighboring communes. Repeat samples were collected from the 83 ZIKV IgM positive participants 1.5 years after the first collection. No new cases of ZIKV infection were detected. In addition, 2 of 3 participants with anti-ZIKV NS1 IgG demonstrated a 4- to 8-fold increase in ZIKV neutralizing antibody titer. Conclusions: ZIKV was present in the area around Krong Buk, with the rate of ZIKV-specific antibodies was 1.1% in the community since at least 2016. While the low levels of circulation together with low seroprevalence suggests a limited outbreak in the region, the results also reflect on low levels of protective immunity to Zika within the population. These results provide a better understanding of the current ZIKV epidemic status in the region and demonstrate a need for implementation of more effective ZIKV infection control measures.

Download Full-text

Cross-Reaction, Enhancement, and Neutralization Activity of Dengue Virus Antibodies against Zika Virus: A Study in the Mexican Population

Journal of Immunology Research ◽

10.1155/2019/7239347 ◽

2019 ◽

Vol 2019 ◽

pp. 1-14 ◽

Cited By ~ 7

Author(s):

Mayra R. Montecillo-Aguado ◽

Alfredo E. Montes-Gómez ◽

Julio García-Cordero ◽

Josselin Corzo-Gómez ◽

Héctor Vivanco-Cid ◽

...

Keyword(s):

Dengue Virus ◽

Acute Phase ◽

Zika Virus ◽

Neutralizing Antibodies ◽

Cross Reactivity ◽

Health Concern ◽

Serum Samples ◽

Zikv Infection ◽

Antibody Dependent Enhancement ◽

Homologous Antigen

Zika virus (ZIKV), an emerging mosquito-borne flavivirus, has quickly spread in many regions around the world where dengue virus (DENV) is endemic. This represents a major health concern, given the high homology between these two viruses, which can result in cross-reactivity. The aim of this study was to determine the cross-reacting antibody response of the IgM and IgG classes against the recombinant envelope protein of ZIKV (rE-ZIKV) in sera from patients with acute-phase infection of different clinical forms of dengue, i.e., dengue fever (DF) and dengue hemorrhagic fever (DHF) (before the arrival of ZIKV in Mexico 2010), as well as acute-phase sera of ZIKV patients, together with the implications in neutralization and antibody-dependent enhancement. Differences in IgM responses were observed in a number of DF and DHF patients whose sera cross-reacted with the rE-ZIK antigen, with 42% recognition between acute-phase DHF and ZIKV but 27% recognition between DF and ZIKV. Regarding IgG antibodies, 71.5% from the DF group showed cross-reactivity to rE-ZIKV in contrast with 50% and only 25% of DHF and ZIKV serum samples, respectively, which specifically recognized the homologous antigen. The DHF group showed more enhancement of ZIKV infection of FCRγ-expressing cells compared to the DF group. Furthermore, the DHF group also showed a higher cross-neutralizing ability than that of DF. This is the first report where DF and DHF serum samples were evaluated for cross-reactivity against Zika protein and ZIKV. Furthermore, DENV serum samples cross-protect against ZIKV through neutralizing antibodies but at the same time mediate antibody-dependent enhancement in the sequential ZIKV infection.

Download Full-text

Prevalence of Zika virus neutralizing antibodies in healthy adults in Vietnam during and after the Zika virus epidemic season: a longitudinal population-based survey

10.21203/rs.2.23060/v1 ◽

2020 ◽

Author(s):

Co Thach Nguyen ◽

Meng Ling Moi ◽

Thi Quynh Mai Le ◽

Thi Thu Thuy Nguyen ◽

Thi Bich Hau Vu ◽

...

Keyword(s):

General Population ◽

Zika Virus ◽

Neutralizing Antibodies ◽

Neutralizing Antibody ◽

Immunoglobulin M ◽

Control Measures ◽

Serum Samples ◽

Zikv Infection ◽

Epidemic Season ◽

Low Levels

Abstract Background: Between 2016 and 2019, 265 cases of Zika virus (ZIKV) infection were reported in Vietnam, predominantly in southern Vietnam. In 2016, a case of ZIKV-associated microcephaly was confirmed in the Central Highlands, and several members of the infant’s family were confirmed to be infected with ZIKV. However, data on the seroprevalence of ZIKV antibodies in the general population is limited. The purpose of this survey was to determine the level of immunity to ZIKV in the general population of the ZIKV epidemic region.Methods: A total of 879 serum samples were collected from 801 participants between January 2017 and July 2018, during and after the ZIKV epidemic in Vietnam. The samples were tested for anti-ZIKV immunoglobulin M (IgM) and immunoglobulin G (IgG), and anti-dengue virus (DENV) IgG antibodies using enzyme-linked immunosorbent assays (ELISA). Plaque-reduction neutralization test (PRNT) for ZIKV was performed on all samples, and for DENV on the samples that ZIKV neutralizing antibody positive.Results: A total of 83 (10.3%) participants had anti-ZIKV IgM. Of the 83, 6 were confirmed to be ZIKV positive using PRNT and anti-ZIKV IgG ELISA. Of the 718 participants who were anti-ZIKV IgM negative, a further 3 cases were confirmed to be ZIKV positive. Of the 9 participants with ZIKV infection, 5 lived in the same village as the infant with ZIKV-associated microcephaly and the other 4 lived in 2 neighboring communes. Repeat samples were collected from the 83 ZIKV IgM positive participants 1.5 years after the first collection. No new cases of ZIKV infection were detected. In addition, 2 of 3 participants with anti-ZIKV NS1 IgG demonstrated a 4- to 8-fold increase in ZIKV neutralizing antibody titer.Conclusion: ZIKV was present in the area around Krong Buk since at least 2016. While the low levels of circulation together with low seroprevalence suggests a limited outbreak in the region, the results also reflect on low levels of protective immunity to Zika within the population. These results provide a better understanding of the current ZIKV epidemic status in the region and demonstrate a need for implementation of more effective ZIKV infection control measures.

Download Full-text

Evaluation of ELISA-Based Multiplex Peptides for the Detection of Human Serum Antibodies Induced by Zika Virus Infection across Various Countries

Viruses ◽

10.3390/v13071319 ◽

2021 ◽

Vol 13 (7) ◽

pp. 1319

Author(s):

Maria del Pilar Martinez Viedma ◽

Stephen Panossian ◽

Kennedy Gifford ◽

Kimberly García ◽

Isis Figueroa ◽

...

Keyword(s):

Zika Virus ◽

Clinical Symptoms ◽

Sampling Period ◽

Public Health Issue ◽

Diagnostic Challenge ◽

Zikv Infection ◽

Past Infection ◽

Neurological Sequelae ◽

Methods Development ◽

Specificity And Sensitivity

Zika virus (ZIKV) is a mosquito-borne Flavivirus with a positive-sense RNA genome, which are generally transmitted through the bite of an infected Aedes mosquito. ZIKV infections could be associated with neurological sequelae that, and otherwise produces similar clinical symptoms as other co-circulating pathogens. Past infection with one member of the Flavivirus genus often induces cross-reactive antibodies against other flaviruses. These attributes complicate the ability to differentially diagnose ZIKV infection from other endemic mosquito-borne viruses, making it both a public health issue as well as a diagnostic challenge. We report the results from serological analyses using arbovirus-specific peptides on 339 samples that were previously collected from 6 countries. Overall, we found that our multiplexed peptide-based ELISA was highly efficient for identifying ZIKV antibodies as early as 2 weeks post infection, and that it correlates with microneutralization, plaque reduction neutralization tests (PRNTs) and commercial tests for ZIKV in previously characterized samples. We observed that seropositivity varied by patient cohort, reflecting the sampling period in relation to the 2015–2016 ZIKV outbreak. This work evaluates the accuracy, specificity, and sensitivity of our peptide-based ELISA method for detecting ZIKV antibodies from geographically diverse regions. These findings can contribute to ongoing serological methods development and can be adapted for use in future studies.

Download Full-text

Embryonic Stage of Congenital Zika Virus Infection Determines Fetal and Postnatal Outcomes in Mice

Viruses ◽

10.3390/v13091807 ◽

2021 ◽

Vol 13 (9) ◽

pp. 1807

Author(s):

Eri Nakayama ◽

Yasuhiro Kawai ◽

Satoshi Taniguchi ◽

Jessamine E. Hazlewood ◽

Ken-ichi Shibasaki ◽

...

Keyword(s):

Zika Virus ◽

Neutralizing Antibodies ◽

Neonatal Death ◽

Motor Behavior ◽

Wide Spectrum ◽

Fetal Loss ◽

Congenital Infection ◽

Sensory Function ◽

Zikv Infection ◽

Wide Range

Zika virus (ZIKV) infection during pregnancy causes a wide spectrum of congenital abnormalities and postnatal developmental sequelae such as fetal loss, intrauterine growth restriction (IUGR), microcephaly, or motor and neurodevelopmental disorders. Here, we investigated whether a mouse pregnancy model recapitulated a wide range of symptoms after congenital ZIKV infection, and whether the embryonic age of congenital infection changed the fetal or postnatal outcomes. Infection with ZIKV strain PRVABC59 from embryonic day 6.5 (E6.5) to E8.5, corresponding to the mid-first trimester in humans, caused fetal death, fetal resorption, or severe IUGR, whereas infection from E9.5 to E14.5, corresponding to the late-first to second trimester in humans, caused stillbirth, neonatal death, microcephaly, and postnatal growth deficiency. Furthermore, 4-week-old offspring born to dams infected at E12.5 showed abnormalities in neuropsychiatric state, motor behavior, autonomic function, or reflex and sensory function. Thus, our model recapitulated the multiple symptoms seen in human cases, and the embryonic age of congenital infection was one of the determinant factors of offspring outcomes in mice. Furthermore, maternal neutralizing antibodies protected the offspring from neonatal death after congenital infection at E9.5, suggesting that neonatal death in our model could serve as criteria for screening of vaccine candidates.

Download Full-text

High Mobility Group Box 1: a Novel Host Restriction Factor in Zika Virus Replication

10.21203/rs.3.rs-929310/v1 ◽

2021 ◽

Author(s):

Kim-Ling Chin ◽

Nurhafiza binti Zainal ◽

Sing-Sin Sam ◽

Pouya Hassandarvish ◽

Rafidah Lani ◽

...

Keyword(s):

Zika Virus ◽

Severe Disease ◽

High Mobility ◽

High Mobility Group ◽

Enzyme Linked Immunosorbent Assay ◽

Dependent Manner ◽

Zikv Infection ◽

Hmgb1 Level ◽

Antiviral Effects ◽

Huh7 Cells

Abstract Neonatal microcephaly and adult Guillain-Barré syndrome are severe complications of Zika virus (ZIKV) infection. The robustly induced inflammatory cytokine expressions in ZIKV-infected patients may constitute a hallmark for severe disease. In the present study, the potential role of high mobility group box 1 protein (HMGB1) in ZIKV infection was investigated. HMGB1 protein expression was determined by the enzyme-linked immunosorbent assay (ELISA) and immunoblot assay. HMGB1’s role in ZIKV infection was also explored using treatment with dexamethasone, an immunomodulatory drug. Antiviral effects of dexamethasone treatment on both wild-typed (WT) and HMGB1-knockdown (shHMGB1) Huh7 cells were determined by the focus-forming assay. Results showed that the Huh7 cells were highly susceptible to ZIKV infection. The infection was found to induce HMGB1 nuclear-to-cytoplasmic translocation, resulting in a >99% increase in the cytosolic HMGB1 expression at 72h.p.i. The extracellular HMGB1 level was elevated in a time- and multiplicity of infection (MOI)- dependent manner. Dexamethasone 150 µM treatment of the ZIKV-infected cells reduced HMGB1 extracellular release in a dose-dependent manner, with a maximum reduction of 71 ± 5.84% (p < 0.01). The treatment also reduced virus titers by over 83 ± 0.50% (p < 0.01). The antiviral effects, however, was not observed in the dexamethasone-treated HMGB1-knockdown cells, suggesting the importance of the intracellular HMGB1 in ZIKV infection. Overall, these results suggest that translocation of HMGB1 occurred during ZIKV infection and inhibition of the translocation reduced ZIKV replication. These findings highlight the potential of developing therapeutics against ZIKV infection by affecting the translocation of HMGB1 from the nucleus to the cytoplasm.

Download Full-text