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Strain-dependent neutralization reveals antigenic variation of human parechovirus 3.

ABSTRACT: Human parechovirus 3 (HPeV3), a member of the Picornavirus family, is frequently detected worldwide. However, the observed seropositivity rates for HPeV3 neutralizing antibodies (nAbs) vary from high in Japan to low in the Netherlands and Finland. To study if this can be explained by technical differences or antigenic diversity among HPeV3 strains included in the serological studies, we determined the neutralizing activity of Japanese and Dutch intravenous immunoglobulin batches (IVIG), a rabbit HPeV3 hyperimmune polyclonal serum, and a human HPeV3-specific monoclonal antibody (mAb) AT12-015, against the HPeV3 A308/99 prototype strain and clinical isolates from Japan, the Netherlands and Australia, collected between 1989 and 2015. The rabbit antiserum neutralized all HPeV3 isolates whereas the neutralization capacity of the IVIG batches varied, and the mAb exclusively neutralized the A308/99 strain. Mapping of the amino acid variation among a subset of the HPeV3 strains on an HPeV3 capsid structure revealed that the majority of the surface-exposed amino acid variation was located in the VP1. Furthermore, amino acid mutations in a mAb AT12-015-resistant HPeV3 A308/99 variant indicated the location for potential antigenic determinants. Virus aggregation and the observed antigenic diversity in HPeV3 can explain the varying levels of nAb seropositivity reported in previous studies.

SUBMITTER: Karelehto E

PROVIDER: S-EPMC5608956 | biostudies-literature | 2017 Sep

REPOSITORIES: biostudies-literature

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Strain-dependent neutralization reveals antigenic variation of human parechovirus 3.

Karelehto Eveliina E van der Sanden Sabine S Geraets James A JA Domanska Aušra A van der Linden Lonneke L Hoogendoorn Dionne D Koen Gerrit G van Eijk Hetty H Shakeel Shabih S Beaumont Tim T de Jong Menno M Pajkrt Dasja D Butcher Sarah J SJ Wolthers Katja C KC

Scientific reports 20170921 1

Human parechovirus 3 (HPeV3), a member of the Picornavirus family, is frequently detected worldwide. However, the observed seropositivity rates for HPeV3 neutralizing antibodies (nAbs) vary from high in Japan to low in the Netherlands and Finland. To study if this can be explained by technical differences or antigenic diversity among HPeV3 strains included in the serological studies, we determined the neutralizing activity of Japanese and Dutch intravenous immunoglobulin batches (IVIG), a rabbit ...[more]

PMID: 28935894

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Project description:Since it was first recognized in 2004 that human parechoviruses (HPeV) are a significant cause of central nervous system and neonatal sepsis, their clinical importance, primarily in children, has started to emerge. Intravenous immunoglobulin treatment is the only treatment available in such life-threatening cases and has given moderate success. Direct inhibition of parechovirus infection using monoclonal antibodies is a potential treatment. We have developed two neutralizing monoclonal antibodies against HPeV1 and HPeV2, namely, AM18 and AM28, which also cross-neutralize other viruses. Here, we present the mapping of their epitopes using peptide scanning, surface plasmon resonance, fluorescence-based thermal shift assays, electron cryomicroscopy, and image reconstruction. We determined by peptide scanning and surface plasmon resonance that AM18 recognizes a linear epitope motif including the arginine-glycine-aspartic acid on the C terminus of capsid protein VP1. This epitope is normally used by the virus to attach to host cell surface integrins during entry and is found in 3 other viruses that AM18 neutralizes. Therefore, AM18 is likely to cause virus neutralization by aggregation and by blocking integrin binding to the capsid. Further, we show by electron cryomicroscopy, three-dimensional reconstruction, and pseudoatomic model fitting that ordered RNA interacts with HPeV1 VP1 and VP3. AM28 recognizes quaternary epitopes on the capsid composed of VP0 and VP3 loops from neighboring pentamers, thereby increasing the RNA accessibility temperature for the virus-AM28 complex compared to the virus alone. Thus, inhibition of RNA uncoating probably contributes to neutralization by AM28.Human parechoviruses can cause mild infections to severe diseases in young children, such as neonatal sepsis, encephalitis, and cardiomyopathy. Intravenous immunoglobulin treatment is the only treatment available in such life-threatening cases. In order to develop more targeted treatment, we have searched for human monoclonal antibodies that would neutralize human parechoviruses 1 and 2, associated with mild infections such as gastroenteritis and severe infections of the central nervous system, and thus allow safe treatment. In the current study, we show how two such promising antibodies interact with the virus, modeling the atomic interactions between the virus and the antibody to propose how neutralization occurs. Both antibodies can cause aggregation; in addition, one antibody interferes with the virus recognizing its target cell, while the other, recognizing only the whole virus, inhibits the genome uncoating and replication in the cell.

Project description:BackgroundHuman parechoviruses (HPeVs) are among the most frequently detected picornaviruses in humans. HPeVs are usually associated with mild gastrointestinal and respiratory symptoms with the exception of HPeV3 which causes neonatal sepsis and CNS infection. Previous studies showed various results in culturing different HPeV genotypes, inducing only a low cytopathic effect (CPE).MethodsIn vitro growth characteristics of the different HPeV genotypes in a range of 10 different cell lines are scored with CPE and measured in the supernatant by real time PCR. In the optimal cell line for each genotype a standard neutralization assay with the available HPeV antibodies (Abs) was performed and scored by CPE and measured by real time PCR.ResultsAll six HPeV types were able to replicate on the RD99, A549, and Vero cell lines. HPeV1 was the only genotype able to replicate on all cell lines. Most efficient growth of HPeV1, 2, 4, 5, and 6 was shown on the HT29 cell line, while HPeV3 was unable to replicate on HT29. In all cases viral replication could be measured by real time PCR before CPE appeared. The polyclonal Abs available against HPeV1, 2, 4 and 5 all showed neutralization of their respective genotype after 7 days with inhibition of >60% in real time PCR and full inhibition of CPE, although cross-neutralization is shown. Replication of HPeV3 could only be inhibited by 12% by the anti-HPeV3 (aHPeV3) Ab and no inhibition of CPE was shown after 7 days.ConclusionWhen replication is monitored by PCR, growth of HPeV genotypes 1 to 6 is supported by most of the cell lines tested, where viral replication is measured before appearance of CPE. A combination of HT29 and Vero cells would therefore support replication of all culturable HPeV types, so viral replication could be detected by PCR within 3 days for all genotypes.In addition, we showed efficient neutralization for HPeV1, 2, 4, 5, while cross- neutralization was shown between these types, indicating possible common neutralizing epitopes. For HPeV3 no efficient (cross-) neutralization was shown, indicating different neutralizing epitopes for HPeV3 compared to the other HPeV genotypes.

Project description:BackgroundChikungunya virus (CHIKV) is a re-emerging mosquito-borne virus which causes epidemics of fever, severe joint pain and rash. Between 2005 and 2010, the East/Central/South African (ECSA) genotype was responsible for global explosive outbreaks across India, the Indian Ocean and Southeast Asia. From late 2013, Asian genotype CHIKV has caused outbreaks in the Americas. The characteristics of cross-antibody efficacy and epitopes are poorly understood.Methodology/principal findingsWe characterized human immune sera collected during two independent outbreaks in Malaysia of the Asian genotype in 2006 and the ECSA genotype in 2008-2010. Neutralizing capacity was analyzed against representative clinical isolates as well as viruses rescued from infectious clones of ECSA and Asian CHIKV. Using whole virus antigen and recombinant E1 and E2 envelope glycoproteins, we further investigated antibody binding sites, epitopes, and antibody titers. Both ECSA and Asian sera demonstrated stronger neutralizing capacity against the ECSA genotype, which corresponded to strong epitope-antibody interaction. ECSA serum targeted conformational epitope sites in the E1-E2 glycoprotein, and E1-E211K, E2-I2T, E2-H5N, E2-G118S and E2-S194G are key amino acids that enhance cross-neutralizing efficacy. As for Asian serum, the antibodies targeting E2 glycoprotein correlated with neutralizing efficacy, and I2T, H5N, G118S and S194G altered and improved the neutralization profile. Rabbit polyclonal antibody against the N-terminal linear neutralizing epitope from the ECSA sequence has reduced binding capacity and neutralization efficacy against Asian CHIKV. These findings imply that the choice of vaccine strain may impact cross-protection against different genotypes.Conclusion/significanceImmune serum from humans infected with CHIKV of either ECSA or Asian genotypes showed differences in binding and neutralization characteristics. These findings have implications for the continued outbreaks of co-circulating CHIKV genotypes and effective design of vaccines and diagnostic serological assays.

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Strain-dependent neutralization reveals antigenic variation of human parechovirus 3.

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Strain-dependent neutralization reveals antigenic variation of human parechovirus 3.

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