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Development of an Enzyme-Linked Immunosorbent Assay To Detect Antibodies Targeting Recombinant Envelope Protein 2 of
Marcílio Jorge Fumagalli1, William Marciel de Souza2, Marília Farignoli Romeiro2
1Virology Research Center, Ribeirão Preto Medical School of University of São Paulo, Ribeirão Preto, São Paulo, Brazil marcilio_jorge@hotmail.com.
Abstract:
Mayaro virus (MAYV) is a neglected arthropod-borne virus (arbovirus) antigenically clustered into the Semliki Forest complex group of Alphavirus genus (Togaviridae family), maintained in an unclear zoonotic cycle involving mosquitoes from Haemagogus genus as the main vector. The genome is composed of a positive single-stranded RNA of 11.5 kb in length, which contains two genes that encode four nonstructural (nsP1 to nsP4) and five structural (C, E3, E2, 6K, and E1) proteins. In the present study, we have developed an enzyme-linked immunosorbent assay (ELISA) using as antigen the recombinant envelope protein 2 of MAYV produced in an Escherichia coli system (rE2-MAYV ELISAs). A panel of 68 human serum samples from suspected arboviral cases was analyzed and titrated for anti-MAYV IgM and IgG antibody detection. The rE2-MAYV ELISA detected 33.8% (23/68) IgG-positive samples, demonstrating 100% sensitivity and 78.95% specificity compared to the MAYV-specific 50% plaque reduction neutralization assay. In addition, the positive MAYV-neutralizing samples showed high titers of detection by rE2-MAYV ELISA, suggesting a highly sensitive test. The rE2-MAYV ELISA also detected 42.5% (29/68) IgM-positive samples, of which 13.8% (4/29) presented high-avidity interactions with rE2-MAYV. Cross-reactivity was observed with Chikungunya virus (CHIKV)-specific murine antibody sample but not with CHIKV-specific human and other Alphavirus murine antibodies. In short, we have developed a rapid, simple, specific, and sensitive MAYV rE2-ELISA, and our preliminary results show its potential applicability to diagnosis of MAYV infections.
Insights
A new Mayaro virus (MAYV) enzyme-linked immunosorbent assay (ELISA) using recombinant envelope protein 2 (rE2-MAYV) shows high sensitivity and specificity for detecting MAYV antibodies in human serum. This diagnostic tool aids in identifying Mayaro virus infections.
Area of Science:
- Virology
- Immunology
- Diagnostic Assay Development
Background:
- Mayaro virus (MAYV) is a neglected arbovirus in the Alphavirus genus, posing a public health concern due to its unclear zoonotic transmission cycle.
- Current diagnostic methods for MAYV infection can be limited, necessitating the development of more efficient and reliable detection tools.
- MAYV's genome encodes structural proteins, including envelope protein 2 (E2), a potential target for antibody-based diagnostics.
Purpose of the Study:
- To develop and evaluate a novel enzyme-linked immunosorbent assay (ELISA) for detecting Mayaro virus (MAYV)-specific IgM and IgG antibodies.
- To utilize the recombinant envelope protein 2 of MAYV (rE2-MAYV) produced in an Escherichia coli system as the antigen in the developed ELISA.
- To assess the sensitivity, specificity, and potential cross-reactivity of the rE2-MAYV ELISA for diagnosing MAYV infections.
Main Methods:
- Production of recombinant MAYV envelope protein 2 (rE2-MAYV) in an Escherichia coli expression system.
- Development of an enzyme-linked immunosorbent assay (ELISA) utilizing rE2-MAYV as the antigen.
- Analysis of 68 human serum samples from suspected arboviral cases to detect anti-MAYV IgM and IgG antibodies using the rE2-MAYV ELISA.
Main Results:
- The rE2-MAYV ELISA demonstrated 100% sensitivity and 78.95% specificity for detecting IgG antibodies compared to the plaque reduction neutralization assay.
- A significant percentage of samples (33.8% IgG, 42.5% IgM) tested positive for MAYV antibodies, with high titers observed in neutralizing samples.
- Limited cross-reactivity was noted with Chikungunya virus (CHIKV)-specific murine antibodies, but not with human CHIKV or other Alphavirus antibodies.
Conclusions:
- A rapid, simple, specific, and sensitive rE2-MAYV ELISA has been successfully developed for Mayaro virus diagnosis.
- The developed ELISA shows promising applicability for the clinical diagnosis of Mayaro virus infections.
- Further validation is warranted, but the assay holds potential for improved arbovirus surveillance and patient management.
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