Research Article: Development of a Blocking ELISA Using a Monoclonal Antibody to a Dominant Epitope in Non-Structural Protein 3A of Foot-and-Mouth Disease Virus, as a Matching Test for a Negative-Marker Vaccine

Date Published: January 20, 2017

Publisher: Public Library of Science

Author(s): Yuanfang Fu, Pinghua Li, Yimei Cao, Na Wang, Pu Sun, Qian Shi, Xincheng Ji, Huifang Bao, Dong Li, Yingli Chen, Xingwen Bai, Xueqing Ma, Jing Zhang, Zengjun Lu, Zaixin Liu, Joanne M Devlin.


Foot-and-mouth disease (FMD) is a devastating animal disease. Strategies for differentiation of infected from vaccinated animals (DIVA) remain very important for controlling disease. Development of an epitope-deleted marker vaccine and accompanying diagnostic method will improve the efficiency of DIVA. Here, a monoclonal antibody (Mab) was found to recognize a conserved “AEKNPLE” epitope spanning amino acids 109–115 of non-structural protein (NSP) 3A of foot-and-mouth disease virus (FMDV; O/Tibet/CHA/99 strain), which could be deleted by a reverse-genetic procedure. In addition, a blocking ELISA was developed based on this Mab against NSP 3A, which could serve as a matching test for a negative-marker vaccine. The criterion of this blocking ELISA was determined by detecting panels of sera from different origins. The serum samples with a percentage inhibition (PI) equal or greater than 50% were considered to be from infected animals, and those with <50% PI were considered to be from non-infected animals. This test showed similar performance when compared with other 2 blocking ELISAs based on an anti-NSP 3B Mab. This is the first report of the DIVA test for an NSP antibody based on an Mab against the conserved and predominant “AEKNPLE” epitope in NSP 3A of FMDV.

Partial Text

Foot-and-mouth disease (FMD) is a highly contagious viral disease that affects many cloven-hoofed animals. The causative agent is the FMD virus (FMDV), which is a single-stranded, positive-sense RNA virus belonging to the Aphthovirus genus in the Picornaviridae family. The FMDV genome contains a single open reading frame (ORF) that encodes structural proteins (SPs; VP1, VP2, VP3, and VP4) and non-structural proteins (NSPs; L, 2A, 2B, 2C, 3A, 3B, 3C, and 3D). The NSPs are relatively conserved among 7 distinct FMDV serotypes [1, 2].

In developing countries, the control and eradication of foot-and-mouth disease mainly relies upon vaccination. However, how to effectively distinguish infected animals from vaccinated animals is an important challenge because this is the basis for detecting and culling potentially virus carrier animals. A major problem stems from the unpurified vaccine antigen with residual NSPs, which will interfere with DIVA test based on detecting NSP antibodies. Although the sensitivity of the NSP-based DIVA test is decreased under vaccinated condition, it remains the most useful method for evaluating the infection status on a herd level [20–22]. In China, it is not officially required that residual NSPs be removed from the FMD vaccine antigen, although virus suspension culture and antigen production techniques have been improved greatly to reduce the level of NSP. Using an unpurified vaccine causes further problems for disease serological surveillance and evaluation of the infection status of animals. Thus, research on negative-marker FMD virus vaccines and its companion DIVA test are important for reducing vaccine costs and resolving the above difficulties for DIVA purposes.




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