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Determining the epitope dominance on the capsid of a serotype SAT2 foot-and-mouth disease virus by mutational analyses.

ABSTRACT: Monoclonal-antibody (MAb)-resistant mutants were used to map antigenic sites on foot-and-mouth disease virus (FMDV), which resulted in the identification of neutralizing epitopes in the flexible ?G-?H loop in VP1. For FMDV SAT2 viruses, studies have shown that at least two antigenic sites exist. By use of an infectious SAT2 cDNA clone, 10 structurally exposed and highly variable loops were identified as putative antigenic sites on the VP1, VP2, and VP3 capsid proteins of SAT2/Zimbabwe (ZIM)/7/83 (topotype II) and replaced with the corresponding regions of SAT2/Kruger National Park (KNP)/19/89 (topotype I). Virus neutralization assays using convalescent-phase antisera raised against the parental virus, SAT2/ZIM/7/83, indicated that the mutant virus containing the TQQS-to-ETPV mutation in the N-terminal part of the ?G-?H loop of VP1 showed not only a significant increase in the neutralization titer but also an increase in the index of avidity to the convalescent-phase antisera. Furthermore, antigenic profiling of the epitope-replaced and parental viruses with nonneutralizing SAT2-specific MAbs led to the identification of two nonneutralizing antigenic regions. Both regions were mapped to incorporate residues 71 to 72 of VP2 as the major contact point. The binding footprint of one of the antigenic regions encompasses residues 71 to 72 and 133 to 134 of VP2 and residues 48 to 50 of VP1, and the second antigenic region encompasses residues 71 to 72 and 133 to 134 of VP2 and residues 84 to 86 and 109 to 11 of VP1. This is the first time that antigenic regions encompassing residues 71 to 72 of VP2 have been identified on the capsid of a SAT2 FMDV.Monoclonal-antibody-resistant mutants have traditionally been used to map antigenic sites on foot-and-mouth disease virus (FMDV). However, for SAT2-type viruses, which are responsible for most of the FMD outbreaks in Africa and are the most varied of all seven serotypes, only two antigenic sites have been identified. We have followed a unique approach using an infectious SAT2 cDNA genome-length clone. Ten structurally surface-exposed, highly varied loops were identified as putative antigenic sites on the VP1, VP2, and VP3 capsid proteins of the SAT2/ZIM/7/83 virus. These regions were replaced with the corresponding regions of an antigenically disparate virus, SAT2/KNP/19/89. Antigenic profiling of the epitope-replaced and parental viruses with SAT2-specific MAbs led to the identification of two unique antibody-binding footprints on the SAT2 capsid. In this report, evidence for the structural engineering of antigenic sites of a SAT2 capsid to broaden cross-reactivity with antisera is provided.

SUBMITTER: Opperman PA

PROVIDER: S-EPMC4135947 | biostudies-literature | 2014 Aug

REPOSITORIES: biostudies-literature

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Determining the epitope dominance on the capsid of a serotype SAT2 foot-and-mouth disease virus by mutational analyses.

Opperman Pamela A PA Rotherham Lia S LS Esterhuysen Jan J Charleston Bryan B Juleff Nicholas N Capozzo Alejandra V AV Theron Jacques J Maree Francois F FF

Journal of virology 20140514 15

<h4>Unlabelled</h4>Monoclonal-antibody (MAb)-resistant mutants were used to map antigenic sites on foot-and-mouth disease virus (FMDV), which resulted in the identification of neutralizing epitopes in the flexible βG-βH loop in VP1. For FMDV SAT2 viruses, studies have shown that at least two antigenic sites exist. By use of an infectious SAT2 cDNA clone, 10 structurally exposed and highly variable loops were identified as putative antigenic sites on the VP1, VP2, and VP3 capsid proteins of SAT2/ ...[more]

PMID: 24829347

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Project description:Foot-and-mouth disease virus (FMDV), particularly strains of the O and SAT serotypes, is notoriously unstable. Consequently, vaccines derived from heat-labile SAT viruses have been linked to the induction of immunity with a poor duration and hence require more frequent vaccinations to ensure protection. In silico calculations predicted residue substitutions that would increase interactions at the interpentamer interface, supporting increased stability. We assessed the stability of the 18 recombinant mutant viruses in regard to their growth kinetics, antigenicity, plaque morphology, genetic stability, and temperature, ionic, and pH stability by using Thermofluor and inactivation assays in order to evaluate potential SAT2 vaccine candidates with improved stability. The most stable mutant for temperature and pH stability was the S2093Y single mutant, while other promising mutants were the E3198A, L2094V, and S2093H single mutants and the F2062Y-H2087M-H3143V triple mutant. Although the S2093Y mutant had the greatest stability, it exhibited smaller plaques, a reduced growth rate, a change in monoclonal antibody footprint, and poor genetic stability properties compared to those of the wild-type virus. However, these factors affecting production can be overcome. The addition of 1 M NaCl was found to further increase the stability of the SAT2 panel of viruses. The S2093Y and S2093H mutants were selected for future use in stabilizing SAT2 vaccines.IMPORTANCE Foot-and-mouth disease virus (FMDV) causes a highly contagious acute vesicular disease in cloven-hoofed livestock and wildlife. The control of the disease by vaccination is essential, especially at livestock-wildlife interfaces. The instability of some serotypes, such as SAT2, affects the quality of vaccines and therefore the duration of immunity. We have shown that we can improve the stability of SAT2 viruses by mutating residues at the capsid interface through predictive modeling. This is an important finding for the potential use of such mutants in improving the stability of SAT2 vaccines in countries where FMD is endemic, which rely heavily on the maintenance of the cold chain, with potential improvement to the duration of immune responses.

Project description:Foot-and-mouth disease (FMD) is a highly contagious viral disease affecting cloven-hoofed animals that causes a significant economic burden globally. Vaccination is the most effective FMD control strategy. However, FMD virus (FMDV) particles are prone to dissociate when appropriate physical or chemical conditions are unavailable, such as an incomplete cold chain. Such degraded vaccines result in compromised herd vaccination. Therefore, thermostable FMD particles are needed for use in vaccines. This study generated thermostable FMDV mutants (M3 and M10) by serial passages at high temperature, subsequent amplification, and purification. Both mutants contained an alanine-to-threonine mutation at position 13 in VP1 (A1013T), although M3 contained 3 additional mutations. The selected mutants showed improved stability and immunogenicity in neutralizing antibody titers, compared with the wild-type (wt) virus. The sequencing analysis and cryo-electron microscopy showed that the mutation of alanine to threonine at the 13th amino acid in the VP1 protein (A1013T) is critical for the capsid stability of FMDV. Virus-like particles containing A1013T (VLPA1013T) also showed significantly improved stability to heat treatment. This study demonstrated that Thr at the 13th amino acid of VP1 could stabilize the capsid of FMDV. Our findings will facilitate the development of a stable vaccine against FMDV serotype O. IMPORTANCE Foot-and-mouth disease (FMD) serotype O is one of the global epidemic serotypes and causes significant economic loss. Vaccination plays a key role in the prevention and control of FMD. However, the success of vaccination mainly depends on the quality of the vaccine. Here, the thermostable FMD virus (FMDV) mutants (M3 and M10) were selected through thermal screening at high temperatures with improved stability and immunogenicity compared with the wild-type virus. The results of multisequence alignment and cryo-electron microscopy (cryo-EM) analysis showed that the Thr substitution at the 13th amino acid in the VP1 protein is critical for the capsid stability of FMDV. For thermolabile type O FMDV, this major discovery will aid the development of its thermostable vaccine.

Dataset Information

Determining the epitope dominance on the capsid of a serotype SAT2 foot-and-mouth disease virus by mutational analyses.

Publications

Determining the epitope dominance on the capsid of a serotype SAT2 foot-and-mouth disease virus by mutational analyses.

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