Table S1. Information of PRRSV sequences used in this paper.

Isolates (genes) / Accession no. / Year of isolation
HM-1 (Nsp2) / EU177102 / 2006
HM-1 (GP2a) / EU177103 / 2006
HM-1 (GP2b) / EU642597 / 2006
HM-1 (GP3) / EU177104 / 2006
HM-1 (GP4) / EU177105 / 2006
HM-1 (GP5) / EU177106 / 2006
HM-1 (M) / EU177107 / 2006
HM-1 (N) / EU255921 / 2006
GC-2 (Nsp2) / EU255919 / 2006
GC-2 (GP2a) / EU255922 / 2006
GC-2 (GP2b) / EU642598 / 2006
GC-2 (GP3) / EU255923 / 2006
GC-2 (GP4) / EU255922 / 2006
GC-2 (GP5) / EU177114 / 2006
GC-2 (M) / EU177115 / 2006
GC-2 (N) / EU177116 / 2006
GCH-3 (Nsp2) / EU669820 / 2007
GCH-3 (GP2a) / EU177117 / 2007
GCH-3 (GP2b) / EU642599 / 2007
GCH-3 (GP3) / EU177118 / 2007
GCH-3 (GP4) / EU177119 / 2007
GCH-3 (GP5) / EU177120 / 2007
GCH-3 (M) / EU177121 / 2007
GCH-3 (N) / EU177122 / 2007
LS-4 (Nsp2) / EU075304 / 2007
LS-4 (GP2a) / EU017510 / 2007
LS-4 (GP2b) / EU642600 / 2007
LS-4 (GP3) / EU017511 / 2007
LS-4 (GP4) / EU017512 / 2007
LS-4 (GP5) / EU075303 / 2007
LS-4 (M) / EU017513 / 2007
LS-4 (N) / EU017514 / 2007
HQ-5 (Nsp2) / EU255920 / 2007
HQ-5 (GP2a) / EU177108 / 2007
HQ-5 (GP2b) / EU642601 / 2007
HQ-5 (GP3) / EU177109 / 2007
HQ-5 (GP4) / EU177110 / 2007
HQ-5 (GP5) / EU439252 / 2007
HQ-5 (M) / EU177111 / 2007
HQ-5 (N) / EU177112 / 2007
HQ-6 (Nsp2) / EU653014 / 2007
HQ-6 (GP2a) / EU642603 / 2007
HQ-6 (GP2b) / EU642602 / 2007
HQ-6 (GP3) / EU366149 / 2007
HQ-6 (GP4) / EU255926 / 2007
HQ-6 (GP5) / EU255925 / 2007
HQ-6 (M) / EU439253 / 2007
HQ-6 (N) / EU255924 / 2007
ST-7 (Nsp2) / EU642604 / 2007
ST-7 (GP2a) / EU653015 / 2007
ST-7 (GP2b) / EU653016 / 2007
ST-7 (GP3) / EU439254 / 2007
ST-7 (GP4) / EU366150 / 2007
ST-7 (GP5) / EU366151 / 2007
ST-7 (M) / EU366152 / 2007
ST-7 (N) / EU439255 / 2007
BD-8 (Nsp2) / EU660956 / 2008
BD-8 (GP2a) / EU697884 / 2008
BD-8 (GP2b) / EU697885 / 2008
BD-8 (GP3) / EU660957 / 2008
BD-8 (GP4) / EU792475 / 2008
BD-8 (GP5) / EU660958 / 2008
BD-8 (M) / EU660959 / 2008
BD-8 (N) / EU660960 / 2008
VR-23321 / U87392 / 1992
CH-1a / AY032626 / 1999
BJ-4 / AF331831 / 2000
HB-1 / AY150312 / 2002
HB-2 / AY262352 / 2002
JA1422 / AY424271 / 2004
RespPRRS2 / AF066183 / 1998
Ingelvac2 / EF532801 / 2000
PrimePac2 / AF066384 / 1997
LV1,3 / M96262 / 1991
HN1 / AY457635 / 2003
XH-GD / EU624117 / 2007
JX143 / EU708726 / 2006
01NP14 / DQ056373 / 2001
EuroPRRSV1,2 / AY366525 / 1999
HPBEDV2 / EU236259 / 2007
Henan-1 / EU200962 / 2007
Jiangxi-3 / EU200961 / 2007
S1 / DQ459471 / 2006
WUH1 / EU187484 / 2007
SY0608 / EU144079 / 2007
GD / EU109503 / 2007
LN / EU109502 / 2007
SHH / EU106888 / 2007
BJsy06 / EU097707 / 2007
NX06 / EU097706 / 2007
HuN / EF517962 / 2007
HEB1 / EF112447 / 2006
HUB2 / EF112446 / 2006
JXA1 / EF112445 / 2006
JXwn06 / EF641008 / 2007
HUN4 / EF635006 / 2007
CC-1 / EF153486 / 2006
HUB1 / EF075945 / 2006

1 These two viral strains belong to Genotype I, and others belong to Genotype II; 2 These viral strains were isolated from North America; 3 This viral strain was isolated from Netherlands; 4 This viral strain was isolated from Thailand. The strains without footnote were isolated from China.

Table S2. Primers used for RT-PCR.

Gene / Bases1 / Primer / Sequence / Genome position1
Nsp2 / 1340-3886 / F2.1 / 5′-GTTTGGCAGTCACAAGTGGTA-3′ / 1314-1335
R2.1 / 5′-CAGTTTCAGCAAGTCCTCCTC -3′ / 2512-2533
F2.2 / 5′-GGCTAACGCCCAGACGACTTC-3′ / 2532-2553
R2.2 / 5′-AACAGGGAGATGGGAGACGAG-3′ / 3892-3913
Gp2, E / 12073-12843,12078-12299 / 2F / 5′-GGATTACAATGATGCGTTTC-3′ / 11951-11971
2R / 5′-GTAGATCTCTGTGGCTGCTT-3′ / 12877-12897
Gp3 / 12696-13460 / 3F / 5′-CAGGGTCAAATGTAACCATAGTG-3′ / 12596-12619
3R / 5′-GTGATGGTAACATACACGGGT-3′ / 13465-13488
Gp4 / 13241-13777 / 4F / 5′-CCGTGTTTCAGACCTATTACC-3′ / 13183-13204
4R / 5′-CAAGCATTTCTCCAACATACT-3′ / 13785-13806
Gp5 / 13788-14390 / 5F / 5′-GTATGTTGGAGAAATGCTTGACC-3′ / 13786-13809
5R / 5′-CTAAGGACGACCCCATTGTTCCG-3′ / 14367-14390
M / 14375-14899 / 6F / 5′- ATGGGGTCGTCCTTAGATG-3′ / 14375-14394
6R / 5′-TGCTTGCCGTTGTTATTTG-3′ / 14893-14912
N / 14889-15260 / 7F / 5′-ATGCCAAATAACAACGGCAAG-3′ / 14889-14910
7R / 5′-TCATGCTGAGGGTGATGCTGTG-3′ / 15239-15261

1 The positions were based on VR-2332.

Table S3. AA divergence of ORF5 of 8 new isolates comparing to the standard strain VR-2332 and BJ-4.

Strain / 8 new isolates versus VR-2332 / 8 new isolates versus BJ-4
HM-1 / G9C S16F C24Y F25L A29V D34N S35N L39I N44K N58Q / G9C Q13R S16F C24Y F25L A27V A29V D34N S35N L39I N44K
S66T A92G V94A F101Y V102Y T121I F127L A137S I161V / N58Q S66T A92G V94A F101Y V102Y C109S T121I F127L A137S
R164G V185A I189L / G151R I161V R164G V185A I189L
GC-2 / G9R C11R S16F C24Y F25L A29V D34N S35N L39I N44K / G9R C11R Q13R S16F C24Y F25L A27V A29V D34N S35N L39I
N58Q S66T P71S A92G V94A F101Y V102Y T121I F127P / N44K N58Q S66T P71S A92G V94A F101Y V102Y C109S T121I
A137S I161V R164G V185A I189L / F127P A137S G151R I161V R164G V185A I189L
GCH-3 / G9C S16F C24Y F25L A29V D34N S35N L39I N44K N58Q / G9C Q13R S16F C24Y F25L A27V A29V D34N S35N L39I N44K
S66T A92G V94A F101Y V102Y T121I F127L A137S I161V / N58Q S66T A92G V94A F101Y V102Y C109S T121I F127L A137S
R164G V185A I189L / G151R I161V R164G V185A I189L
LS-4 / G9C S16F C24Y F25L A29V D34N S35N L39I N58Q S66T / G9C Q13R S16F C24Y F25L A27V A29V D34N S35N L39I N58Q
A92G V94A F101Y V102Y T121I F127L A137S I161V R164G / S66T A92G V94A F101Y V102Y C109S T121I F127L A137S G151R
V185A I189L / I161V R164G V185A I189L
HQ-5 / G9C S16F C24Y F25L A29V D34N L39I N58Q S66T A92G / G9C Q13R S16F C24Y F25L A27V A29V D34N L39I N58Q S66T
V94A F101Y V102Y T121I F127L A137S I161V R164G V185A / A92G V94A F101Y V102Y C109S T121I F127L A137S G151R I161V
I189L / R164G V185A I189L
HQ-6 / G9C S16F C24Y F25L A29V D34N S35N L39I N44K N58Q / G9C Q13R S16F C24Y F25L A27V A29V D34N S35N L39I N44K
S66T A92G V94A F101Y V102Y T121I F127L A137S D147G / N58Q S66T A92G V94A F101Y V102Y C109S T121I F127L A137S
I161V R164G V185A I189L / D147G G151R I161V R164G V185A I189L
ST-7 / G9C S16F C24Y F25L A29V D34N S35N L39I N44K N58Q / G9C Q13R S16F C24Y F25L A27V A29V D34N S35N L39I N44K
S66T A92G V94A F101Y V102Y I111V T121I F127L A137S / N58Q S66T A92G V94A F101Y V102Y C109S I111V T121I F127L
R151G I161V R164G V185A I189L / A137S I161V R164G V185A I189L
BD-8 / G9C S16F L17S C24F F25L A29V N33S D34N S35N L39I / G9C Q13R S16F L17S C24F F25L A27V A29V N33S D34N S35N
N44K N58Q K59R S66T A92G V94A F101Y V102Y T121I / L39I N44K N58Q K59R S66T A92G V94A F101Y V102Y C109S
F127L A137S I161V R164G V185A I189L R191K / T121I F127L A137S G151R I161V R164G V185A I189L R191K