Virus Research 136 (2008) 157–165
Contents lists available at ScienceDirect
Virus Research
journal homepage: www.elsevier.com/locate/virusres
RNA transcription analysis and completion of the genome sequence
of yellow head nidovirus
Nusra Sittidilokratna
a,b,1
, Sirintip Dangtip
b
, Jeff A. Cowley
c
, Peter J. Walker
d,∗
a
National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency (NSTDA), Phathumthani 12120, Thailand
b
Center of Excellence for Shrimp Molecular Biology and Biotechnology, Faculty of Science, Mahidol University, Rama VI Road, Phyathai, Bangkok 10400, Thailand
c
CSIRO Livestock Industries, Queensland Bioscience Precinct, 306 Carmody Road, St. Lucia, Queensland 4067, Australia
d
CSIRO Livestock Industries, Australian Animal Health Laboratory (AAHL), 5 Portarlington Road, Geelong, Victoria 3220, Australia
article info
Article history:
Received 18 February 2008
Received in revised form 30 April 2008
Accepted 3 May 2008
Available online 11 June 2008
Keywords:
YHV
Genome sequence
Transcription
Shrimp
Penaeus monodon
abstract
Yellow head virus (YHV) is a pathogen of the black tiger shrimp (Penaeus monodon) and, with gill-
associated virus (GAV), is one of two known invertebrate nidoviruses. We describe sequences of the
large replicase gene (ORF1a) and 5
- and 3
-terminal UTRs, completing the 26,662 nt sequence of the YHV
genome. ORF1a (12,219 nt) encodes a ∼462,662 Da polypeptide containing a putative 3C-like protease and
a putative papain-like protease with the canonical C/H catalytic dyad and ␣ +  fold. The read-through
pp1ab polyprotein contains putative uridylate-specific endoribonuclease and ribose-2
-O-methyl trans-
ferase domains, and an exonuclease domain incorporating unusual dual Zn
2+
-binding fingers. Upstream of
ORF1a, the 71 nt 5
-UTR shares 82.4% identity with the 68 nt 5
-UTR of GAV. The 677 nt 3
-terminal region
contains a single 60 nt ORF, commencing 298 nt downstream of ORF3, that is identical to N-terminal
coding region of the 249 nt GAV ORF4. Northern blots using RNA from YHV-infected shrimp and probes
directed at ORF1a, ORF1b, ORF2 and ORF3 identified a nested set of 3
-coterminal RNAs comprising the
full-length genomic RNA and two sub-genomic (sg) mRNAs. Intergenic sequences upstream of ORF2 and
ORF3 share high identity with GAV, particularly in the conserved domains predicted to mediate sgmRNA
transcription.
Crown Copyright © 2008 Published by Elsevier B.V. All rights reserved.
1. Introduction
Yellow head virus (YHV) is a highly virulent pathogen of the
black tiger shrimp (Penaeus monodon). It can cause rapidly accu-
mulating mortalities in aquaculture ponds, resulting in total crop
loss within several days of the first signs of disease. Yellow head dis-
ease first emerged in central Thailand in 1990 (Limsuwan, 1991). It
has since been reported in most major shrimp producing countries
in Asia and remains a disease of serious concern globally (Walker
et al., 2001; OIE, 2007). YHV is now known to be one of several
closely related genotypes.These include gill-associated virus (GAV),
a less virulent virus that was first detected in Australiain association
with a condition named mid-crop mortality syndrome, and at least
four other genotypes identified only in healthy P. monodon shrimp
from Southeast Asia, India and East Africa (Walker et al., 2001;
Wijegoonawardane et al., 2004; Wijegoonawardane et al., 2008).
∗
Corresponding author. Tel.: +61 3 5227 5400; fax: +61 3 5227 5000.
E-mail address: Peter.Walker@csiro.au (P.J. Walker).
1
Current Address: CSIRO Livestock Industries, Australian Animal Health Labora-
tory (9AAHL), 5 Portarlington Road, Geelong, Victoria, 3220, Australia.
YHV is a positive-sense (+) ssRNA virus that is currently clas-
sified with GAV as a member of the species Gill-associated virus,
genus Okavirus, family Roniviridae in the order Nidovirales (Walker
et al., 2005). The enveloped virions are rod-shaped (approximately
70 nm × 180 nm) with prominent surface projections and an inter-
nal helical nucleocapsid (Wongteerasupaya et al., 1995; Nadala
et al., 1997). Virions contain three structural proteins compris-
ing two transmembrane glycoproteins (gp116 and gp64) and a
nucleoprotein (p20) (Jitrapakdee et al., 2003; Sittidilokratna et
al., 2006). For GAV, the 26,235 nt genome has been shown to be
3
-polyadenylated, and contain five open reading frames (ORFs)
bounded by 5
- and 3
-UTRs. ORF1a and ORF1b are expressed from
genome-length mRNA and overlap (Cowley et al., 2000, 2002).
ORF1a encodes a long polyprotein (pp1a) that includes a 3C-like
protease domain. ORF1b is translated only following a −1 ribosomal
frame-shift at a predicted pseudoknot structure in the ORF overlap
region and encodes enzymes of the replication complex (Cowley et
al., 2000; Ziebuhr et al., 2003). GAV ORF2 and ORF3 are expressed
from sub-genomic (sg)mRNAs and encode the nucleoprotein and
the virion envelope glycoprotein, respectively (Cowley and Walker,
2002; Cowley et al., 2004). Downstream of ORF3 is a 638 nt region
which includes a 249-nt open reading frame (ORF4) that has been
0168-1702/$ – see front matter. Crown Copyright © 2008 Published by Elsevier B.V. All rights reserved.
doi:10.1016/j.virusres.2008.05.008