1 INTRODUCTION
Porcine epidemic diarrhea virus (PEDV), a member of the genusAlphacoronavirus in the family Coronaviridae of the orderNidovirales , is a main etiology causing porcine epidemic diarrhea
(PED) which is a devastating disease characterized by acute watery
diarrhea, vomiting, dehydration, and high
mortality
in neonatal piglets. PEDV was first reported in England and Belgium
(Wood, 1977, Pensaert and de Bouck, 1978) in the late 1970s. Outbreaks
of PED in China were
sporadic
(Chen et al., 2010) until late 2010, when PEDV variant strains-caused
PED broke out and spread throughout the country (Wang et al., 2013; Sun
et al., 2012). PEDV has also been reported in the US (Stevenson et al.,
2013) and other countries in North America, including Canada and Mexico
(Mole, 2013, Ojkic et al., 2015, Trujillo-Ortega et al., 2016). PEDV has
caused considerable economic damage to the global pig industry.
The PEDV genome is a single-stranded positive –sense RNA with
approximately 28kb in length, and encodes four structural proteins which
include spike (S), nucleocapsid (N), envelop (E), and membrane (M)
protein. Among these proteins, the S protein is on the surface of the
PEDV virion that plays a pivotal role in regulating interactions with
specific host cell receptors to mediate viral entry, inducing
neutralizing antibodies in the natural host and determining antigenicity
and virulence (Li et al., 2020,
Suzuki
et al., 2018, Lv et al., 2016, Cruz et al, 2008, Sun et al.,2008, Park
et al., 2007) . The S protein is often used to evaluate the genetic
diversity of PEDV strains (Li et al., 2012; Lee et al.,2014). Based on
the phylogenetic analysis of the S gene, the current reported PEDV
strains could be genetically divided into two genotypes: G1 and G2. PEDV
G1 and G2 can be further divided into G1a, G1b, and G2a, G2b subtypes,
respectively. The G1 contains classical strains, including the prototype
strain CV777 and historical vaccine strains. The G2 contains the
post-2010 global epidemic isolates (Jung at al., 2020, Won et al.,
2020).
The fact of increased outbreaks of G2 PEDV and the failure of G1 PEDV
strain (CV777)-based vaccine in China since 2010
implied
that the difference of antigenicity between G1 and G2 PEDV strains.
Identification of factors contributing to the antigenicity difference
between G1 and G2 PEDV strains will lead to the development of an
effective PEDV vaccine with better protection against prevalent genotype
of PEDV. As amino acid mutations that distinguish the two PEDV genotypes
were mostly located in the N-terminal domain (NTD) (aa 1-380) of S
protein, the objective of this study is to determine the role of the NTD
of S protein in resulting antigenicity difference between PEDV G1 and
G2. The NTD of the S protein from CV777 vaccine strain (G1) and
CH/ZMDZY/11 strain (G2) was expressed in E. coli , respectively.
Polyclonal antibodies (PAbs) against genotype-specific S proteins were
prepared by immunizing BALB/c mice using purified S proteins.
Antigenicity was systematically compared by detection of PAbs against
two genotype PEDV strains and purified S proteins using Western blot,
indirect enzyme-linked immunosorbent assay (ELISA), indirect
immunofluorescence assay (IFA), and serum cross-neutralization assay
(SN).