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Molecular Pathogens
MP2011, Vol.2, No.1
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usually focus on sequencing the part or all of the
genome region coding for the outer capsid protein
VP1. If the isolate with more than 85% nt sequence
identity has been placed within group or topotype,
which tends to be restricted in their geographic
distribution (Knowles and Samuel, 2003). FMDV type
O is the worldwide pandemic virus serotype, which
can be grouped into eight topotypes that is Cathay,
Middle East-South Asia (ME-SA), South-East Asia
(SEA), Europe-South America (Euro-SA), Indonesia
-
1
(ISA
-
1), Indonesia
-
2 (ISA
-
2), East Africa (EA), and
West Africa (WA), based on 15% nucleotide differ-
ences (Samuel and Knowles, 2001). In addition, the
FMDVs still exist in different evolutionary sub-
lineage within the topotypes or groups.
Recently, methodologies for FMDV vaccine strain
selection mainly rely on serological approaches (Paton
et al., 2005). The determination of FMDV nucleotide
sequences and phylogenetic analysis is the supplement
approaches for selecting vaccine strain to use in FMD
control program (Reeve et al., 2010). In 1997, a
devastating outbreak of foot-and-mouth disease, with
atypical virulence which produced high morbidity and
mortality in swine but did not affect cattle, occurred in
Taiwan (Dunn and Donaldson, 1997). We isolated a
FMDV strain with similar biological characteristics as
Tanwan’s isolates in Monitoring Station of Border
Region of Yunnan Province. Previously, the biological
characteristics and immunogenicity of this isolate
were studied. In this paper, we presented the
full-length genome sequence of this isolate and
analyzed its genetic characteristics in order to provide
a ground for selecting the vaccine candidate strain in
FMD prevention and control.
1 Results
1.1 Amplifying the whole genome of O/YM/YN/2000
isolate
In order to sequence whole genome of the O/YM/YN/
2000 isolate, seven primer sets were used for
amplifying the whole genome and then cDNA was
synthesized with the 6
nt random primer. Seven
fragments (S1, S2, F2, F3, Fa, F4b and F5) spanning
whole O/YM/YN/2000 genome were generated by
PCR. The expected sizes of 374 bp, 637 bp, 1 107 bp
2 169 bp, 2 454 bp, 1 571 bp and 1 533 bp in length
were synthesized in this research (Figure 1).
Figure 1 The fragments amplified from FMDV strain O/YM/
YN/2000 by RT-PCR
Note: M1: DNAMarker DL2000; M2: 500 bp DNAMarker
1.2 Sequencing whole genome of O/YM/YN/2000
The whole genome of O/YM/YN/2000 strain is
8 133
nt in length, which includes a 1 053
nt 5´
-
UTR
(including 18
nt continuous poly C tract), a 6 969
nt
length open reading frame (ORF) encoding a polyprotein
of 2 323 amino acids (aa) residues or 2 294 aa due to
two alternative initiation sites separated by 84
nt, a 93
nt
3´
-
UTR and 18
nt poly A tail. Table 1 presented the
genomic organization of O/YM/YN/2000 (nucleotides
and deduced amino acids). The genome sequence has
been deposited in to the Genbank database of the
NCBI with Accession Number HQ412603.
1.3 Genomic comparative analysis between O/YM/YN/
2000 with other known strains
The nucleotide and deduced aa sequences of each
region (5´
-
UTR, L, VP4, VP2, VP3, VP1, 2A, 2B, 2C,
3A, 3B, 3C, 3D and 3´
-
UTR) of O/YM/YN/2000 and
other FMDV reference strains were compared in this
research (Table 1). Regarding to coding region, the
sequence of non-structural proteins is much more
conserved than that of the structural proteins in the
FMDV genome. Among non-structural protein coding
regions, 2A (93.7%~100%), 2B (90.2%~100%) and
3D (94.2%~99.4%) are the most highly conserved
region. While in the structural protein regions,
VP1
gene is highly variation (52.1%~93.9%) identity in
nucleotide sequences),
VP4
shows the lowest
sequence identity (91.6%~99.2%). In the non-
structural protein coding regions, L (61.2%~95.3%
identity in aa sequence) and 3A (68.2%~96.6%) exist
more diversity. Among all the compared regions,
5´
-
UTR, VP1 and 3Awould be the lowest conservative.