Virus Research 136 (2008) 50–57
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Virus Research
journal homepage: www.elsevier.com/locate/virusres
Enhanced immune responses of mice inoculated recombinant adenoviruses
expressing GP5 by fusion with GP3 and/or GP4 of PRRS virus
Wenming Jiang, Ping Jiang
∗
, Xianwei Wang, Yufeng Li, Yijun Du, Xinglong Wang
Key Laboratory of Animal Diseases Diagnostic and Immunology, College of Veterinary Medicine, Nanjing Agricultural University, Ministry of Agriculture, Nanjing 210095, China
article info
Article history:
Received 12 November 2007
Received in revised form 7 April 2008
Accepted 18 April 2008
Available online 6 June 2008
Keywords:
PRRSV
GP3
GP4
GP5
Recombinant adenovirus
Immune responses
abstract
Porcine reproductive and respiratory syndrome (PRRS) is one of the most important causes of eco-
nomic losses of the swine industry. PRRS virus (PRRSV) infection poses a challenge to current vaccination
strategies. In this study, three replication-defective adenovirus recombinants expressing fusion protein
GP3–GP5, GP4–GP5, or GP3–GP4–GP5 were developed as potential vaccine against PRRSV in a mouse
model. Six groups of BALB/c mice (24 mice per group) were inoculated subcutaneously twice at 2-week
intervals with above mentioned recombinants and other adenoviruses expressing single GP3, GP4, or
GP5 protein. The results showed that the mice inoculated with recombinant adenoviruses developed
PRRSV-specific antibodies, cellular immune response by 2 weeks post-boost-immunization. However,
mice immunized with recombinant adenoviruses rAd-GP3–GP5, rAd-GP4–GP5, and rAd-GP3–GP4–GP5
developed significantly higher titers of neutralizing antibodies to PRRSV and produced stronger lympho-
cyte proliferation responses compared tomice immunized with rAd-GP3,rAd-GP4 or rAd-GP5 alone. It was
also found that mice immunized with rAd-GP3–GP5 and rAd-GP3–GP4–GP5 were primed for significant
higher levels of anti-PRRSV CTL responses than mice immunized with rAd-GP3 and rAd-GP5. These find-
ings suggested that the recombinant adenoviruses expressing fusion proteins GP3–GP5 or GP3–GP4–GP5
might be an attractive candidate vaccine for preventing PRRSV infection.
© 2008 Elsevier B.V. All rights reserved.
1. Introduction
Porcine reproductive and respiratory syndrome (PRRS) is one
of the most economically significant viral diseases in the swine
industry (Polson et al., 1992; Dee et al., 1997). It is characterized
by reproductive problems in sows such as poor farrowing rates,
premature farrowings, and increased stillbirths, as well as respira-
tory problems in piglets such as pneumonia and atrophic rhinitis
(Bilodeau et al., 1991; Wensvoort et al., 1992). Infection of PRRSV
also predisposes pigs to infection by bacterial and viral pathogens
such as Steptococcus suis, Haemophilus parasuis, Mycoplasma
hyopneumoniae, Actinobaccillus pleuropneumoniae, Salmonella spp.,
Pasteurella multocida, and swine influenza virus (Benfield et al.,
1992; Groschup et al., 1993; Zeman et al., 1993; Galina et al., 1994;
Done and Paton, 1995; Kawashima et al., 1996). PRRS virus (PRRSV)
is an enveloped, positive-stranded RNA virus that is a member of
the order Nidovirales, family Arteriviridae. The genome of PRRSV
is about 15 kb in length and consists of nine open reading frames
(ORFs) (Conzelmann et al., 1993; Snijder et al., 1999; Wu et al.,
2001). The ORFs 3–6 encode the four structural proteins of viri-
∗
Corresponding author. Tel.: +86 25 84395504; fax: +86 25 84396640.
E-mail address: jiangp@njau.edu.cn (P. Jiang).
ons which are the glycoprotein GP3 (42–45 kDa), the glycoprotein
GP4 (31–35 kDa), the envelope glycoprotein GP5 (24–26 kDa) and
non-glycosylated membrane protein M (18–19 kDa) (Meulenberg
et al., 1995; Murtaugh et al., 1995). The GP3 protein is highly gly-
cosylated potentially by seven N-linked oligosaccharides and has a
single N-terminal hydrophobic domain (Meulenberg et al., 1995).
Whereas the GP3 protein was shown to be incorporated into viri-
ons of European-type strains (van Nieuwstadtet al., 1996), for North
American strains, its structural nature has been questioned (Gonin
et al., 1998; Mardassi et al., 1998). This discrepancy might be real
or due to different analytical procedures. The GP4 protein is a puta-
tive class I integral membrane protein with an N-terminal signal
sequence and a C-terminal membrane anchor, containing four pre-
dicted N-glycosylation sites (Meulenberg et al., 1995). GP5 is a
glycosylated transmembrane protein. It has a putative N-terminal
signal peptide of approximately 25 amino acids, followed by an
ectodomain of approximately 40 residues with a variable num-
ber of potential N-linked glycosylation sites, a long hydrophobic
region of about 60 residues that is presumed to span the membrane
one to three times, and a hydrophilic C terminus of approximately
70 amino acids (Meulenberg et al., 1995; Mardassi et al., 1996;
Balasuriya and MacLachlan, 2004). These PRRSV structural proteins
are closely associated, GP5 and M proteins being associated in the
form of heterodimers (Mardassi et al., 1996; Delputte et al., 2002).
0168-1702/$ – see front matter © 2008 Elsevier B.V. All rights reserved.
doi:10.1016/j.virusres.2008.04.016