oligonucleotide / Sequences / Reference
S segment / NCRS
255 R
342 F
393 R
883 F
883 R
1027 F
1102 R
1280 R
1581 F / 5´- TAG TAG TAG ACT CCT TGA A -3´
5´- TGG ACA CAG CAT CTG CCA -3´
5´- TAT GGT AAT GTC CTT GAT GT-3´
5´- CTC CAA TTG TAT ACC AAT CT-3´
5´- CAT GTG GCT GAC ATT GAT A - 3´
5´- TAT CAA TGT CAG CCA CAT G -3´
5´- ATG GCA GAG TTA GGT GCA -3´
5´- GCC ATD ATD GTR TTY CTC AT-3´
5´- GAT CAA TGA GTG CTT GTG C -3´
5´- CTG CTG CTT GTT ATT GTT ACC -3´ / This paper
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(20)
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(20)
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M segment / Han_Uni_M
257 R
M896 F
948 R
M1539 R
PUM1F(2371)
PUM1R(2863)
3303 F
5NCR
1232 R
1516 F
2401 R
2401 F
3148 F
3385 F / 5´-TAG TAG TAK RCW CCG CA -3´
5´- CTG TGA TGA TGT GCT AGT ATG -3´
5´- CHC ATG GHG AAG AYC ATG A -3´
5´- CTC ATT GCA CTT ATG GCA -3 ´
5´- ABC CAG CCA AAG CAR AAT GT-3´
5´- ACC TGA TTG TCC AGG GGT AGG -3´
5´- GGG TTC TGT AAC ATT GAA TGA TTG -3´
5´- GAT GGY GCW CCW SAR TGT GG -3´
5´- TAG TAG TAG GCT CCG CA -3´
5´- CAG AAT ATG CCT CAC AAT C -3´
5´- GGC TAC AAT GCT ACT CTT AT -3´
5´- GTA TAC ACC ACA TGC TGT AC -3´
5´- TAC AGC ATG TGG TGT ATA C -3´
5´- ATA CAG TTC ATA TCG TTG G -3´
5´- GAT GGT AGT GGC TGT ATT AGT -3´ / This paper
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L segment / 5NCRL
285 R
980 F
980 R
1243 F
1513 F
1513 R*
2245 F
2245 R
2935 F
2961 R
3367 F
4216 F*
4480 F
4480 R
5245 R
5292 F
5893 F
5893 R
5566 R*
6412 F / 5´-TAG TAG TAG ACT CCG AGA TA -3´
5´-TGA TCT CAG TAT CTG ACC ACT A -3´
5´- TCT GAY CAG TAT GAT ACA CG - 3´
5´- CGT GTA TCA TAC TGR TCA GA -3´
5´- GAC AGT ATT GAT GTA GCA AG -3´
5´- ATT GCA CAT GCA GGG CTT A -3´
5´- TAA GCC CTG CAT GTG CAA T - 3´
5´- GCT TGC ATG GCA ATC TTA CA -3´
5´-TGT AAG ATT GCC ATG CAA G -3´
5´- ATG TAT GTA AGT GCA GAT GC -3´
5´- AAT TAT CAC CAG GTG ACC A -3´
5´- ATG ACG GAA CAG ACT GGT T -3´
5´- ACT GCA ATG ACA ATG CAA TC -3´
5´- CAT AGA CCT AAG GTG GCA -3´
5´- GTG CCA CCT TAG GTC TAT -3´
5´- GAT CTC TAA CCT TGC ACA AT -3´
5´- CCT AGA GAC ATC AAT AAC TC -3´
5´- GGC TAT ACA ACA GTT AGT AC -3´
5´- GTA CTA ACT GTT GTA TAG CC -3´
5´- CTT CAT CAA GCT GTG CCA T -3´
5´- AGA AGA TGT GCT ACC CGA TT -3´ / This paper
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(22, modified HanL F1)
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(22, modified HanL F1)
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Supplementary Table 1: Oligonucleotides used for RT-PCR amplification and sequencing
*Theprimer sequences differedfrom those obtainedfor strain Astrup07_1219.
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Supplementary Table 2: Lengths of the 5´ non-coding region (NCR), coding sequence (CDS) and 3´-NCR of the novel Puumala virus (PUUV) strain Astrup, PUUV strains Sotkamo and Pieksamaki from Finland, PUUV strain Umea from Sweden, PUUV strains Kazan, Samara, DTK/Ufa-97 and CG1820 from Russia and outgroup viruses Hantaan Virus (HTNV) and Tula virus (TULV)
Strain* / S segment / M segment / L segment5´-NCR / CDS / 3´-NCR / 5´-NCR / CDS / 3´-NCR / 5´-NCR / CDS / 3´-NCR
Astrup / 42 / 1302 / 484 / 40 / 3447 / 193 / 36** / 6471 / 43
Sotkamo / 42 / 1302 / 486 / 40 / 3447 / 195 / 36 / 6471 / 43
Pieksamaki / 42 / 1302 / 483 / 40 / 3447 / 195 / 36 / 6471 / 43
Umea / 42 / 1302 / 485 / 40 / 3447 / 195 / 36 / 6447 / 43
Kazan / 42 / 1302 / 482 / 40 / 3447 / 195 / 36 / 6471 / 43
Samara 49 / 42 / 1302 / 484 / 40 / 3447 / 195 / 36 / 6471 / 43
Samara 94 / 42 / 1302 / 484 / 40 / 3447 / 195 / 36 / 6471 / 43
DTK/Ufa-97 / 42 / 1302 / 485 / 40 / 3447 / 195 / 36 / 6471 / 43
CG1820 / 42 / 1302 / 440 / 40 / 3447 / 195 / 36 / 6471 / 43
TULV / 42 / 1290 / 499 / 55 / 3426 / 213 / 36 / 6462 / 43
HTNV / 36 / 1290 / 370 / 40 / 3408 / 168 / 37 / 6456 / 40
*Accession numbers of the S, M, and L segment sequences of PUUV strains: Astrup (KJ994776, KJ994777, KJ994778), Sotkamo(NC_005224, NC_005223, NC_005225), Pieksamaki (NJ831943, JN831944,JN831945),Umea (AY526219, AY526218,AY526217),Kazan (Z84204, Z84205, AF405801), Samara 49(AB433843, AB433850, AB574183), Samara 94 (AB433845, AB433852, AB574184), DTK/Ufa-97 (AB297665, AB297666, AB297667),CG1820 (M32750, M29979, M63194), TULV strain Moravia (NC_005227,NC_005228,NC_005226), and HTNVstrain 76-118 (NC_005218,M14627,NC_005222).
**The proximal sequence TAGTAG of the 5´-NCR of the L segment could not be determined and stems therefore from the used primer.
Supplementary Table 3: Nucleotide (above the diagonal) and amino acid sequence divergence(below the diagonal) of the coding sequences of the S (A), M (B) and Lsegments (C) and the encoded proteins between Puumala virus (PUUV) strain Astrup to PUUV strains from other regions in Europe and Tula virus(TULV) as an outgroup (in %).
A
Strain / 1 / 2 / 3 / 4 / 5 / 6 / 7 / 8 / 9 / 101 / Astrup 07_1219_KJ994776 / 15.3 / 16.1 / 15.7 / 16.1 / 16.1 / 16.6 / 16.6 / 16.6 / 26.9
2 / Sotkamo_NC_005224 / 2.1 / 7.3 / 14.6 / 14.6 / 14.3 / 14.7 / 14.5 / 14.5 / 26.9
3 / Pieksamaki_JN831943 / 1.9 / 0.5 / 15 / 13.6 / 13.6 / 14 / 14.4 / 14.4 / 27.8
4 / Umea_AY526219 / 2.1 / 3.5 / 3.1 / 15.6 / 15.5 / 15.9 / 16.2 / 16.2 / 26.7
5 / Samara49_AB433843 / 2.6 / 3.3 / 2.8 / 3.7 / 3 / 5 / 5.8 / 5.8 / 27.5
6 / Samara94_AB433845 / 2.6 / 3.3 / 2.8 / 3.7 / 0 / 4.9 / 6.2 / 6.2 / 27.2
7 / Kazan_Z84204 / 2.8 / 3.3 / 2.8 / 3.7 / 1 / 1 / 5.7 / 5.7 / 27.8
8 / DTK/Ufa-97_AB297665 / 3.1 / 3.7 / 3.3 / 4.2 / 1 / 1 / 1.4 / 0.4 / 27.2
9 / CG1820_M32750 / 3.1 / 3.7 / 3.3 / 4.2 / 1 / 1 / 1.4 / 1 / 26.9
10 / TULV_NC_005227 / 20.6 / 21.1 / 20.6 / 20.6 / 20.6 / 20.6 / 20.8 / 20.6 / 19.9
B
Strain / 1 / 2 / 3 / 4 / 5 / 6 / 7 / 8 / 9 / 101 / Astrup07_1219_KJ994777 / 19.6 / 19.1 / 19 / 19.3 / 19 / 18.7 / 19.6 / 19.9 / 27.6
2 / Sotkamo_ NC_005223 / 10.3 / 9.2 / 19.1 / 15.6 / 15.9 / 15.2 / 16.8 / 17.1 / 28.7
3 / Pieksamaki_JN831944 / 9.7 / 2.4 / 18.7 / 15.7 / 15.9 / 15.1 / 16.3 / 16.6 / 28.3
4 / Umea_AY526218 / 8.1 / 8.5 / 8.2 / 19.3 / 18.8 / 19.2 / 19.2 / 19.4 / 27.8
5 / Samara49_AB433850 / 9.3 / 5.1 / 4.7 / 8.6 / 6.5 / 6.2 / 14.6 / 14.9 / 28.3
6 / Samara94_AB433852 / 9.4 / 5.5 / 5 / 8.8 / 0.9 / 6.1 / 14.4 / 14.7 / 27.6
7 / Kazan_Z84205 / 9.1 / 4.8 / 4.4 / 8.4 / 0.7 / 1.1 / 13.8 / 14.1 / 27.9
8 / DTK/Ufa-97_AB297666 / 9.9 / 5.6 / 5.3 / 8.8 / 3 / 3.4 / 3 / 0.5 / 27.6
9 / CG1820_M29979 / 10.5 / 6.2 / 5.9 / 9.4 / 3.6 / 4 / 3.6 / 0.8 / 27.9
10 / TULV NC_005228 / 21 / 21.7 / 21 / 22.2 / 21 / 21 / 21.1 / 21.3 / 21.8
C
Strain / 1 / 2 / 3 / 4 / 5 / 6 / 7 / 8 / 9 / 101 / Astrup_07_1219_KJ994778 / 17.7 / 17.8 / 19 / 17.2 / 17.7 / 17.5 / 17.7 / 17.8 / 24.5
2 / Sotkamo_NC_005225 / 4.4 / 7.1 / 18 / 15.1 / 15.1 / 15.2 / 15.6 / 15.6 / 25.2
3 / Pieksamaki_JN831946 / 4.1 / 1.1 / 17.5 / 15.2 / 15.3 / 15.5 / 15.4 / 15.4 / 24.6
4 / Umea_AY526217 / 6.7 / 6.6 / 6.5 / 18.1 / 18.2 / 18.2 / 18.8 / 18.8 / 26.1
5 / Samara49_AB574183 / 3.9 / 2.7 / 2.5 / 5.9 / 3.3 / 6.3 / 12.7 / 12.7 / 25.1
6 / Samara94_AB574184 / 4 / 2.9 / 2.7 / 5.9 / 0.6 / 6.5 / 12.8 / 12.8 / 25.2
7 / Kazan_EF405801 / 3.9 / 2.6 / 2.4 / 5.8 / 0.4 / 0.5 / 13 / 13.1 / 25.3
8 / DTK/Ufa-97_AB297667 / 3.9 / 2.7 / 2.6 / 6.1 / 1.2 / 1.3 / 1 / 0.3 / 25.3
9 / CG1820_M63194 / 4 / 2.8 / 2.6 / 6.1 / 1.3 / 1.4 / 1.1 / 0.3 / 25.3
10 / TULV NC_005226 / 14.5 / 15 / 15 / 16.9 / 14.6 / 14.6 / 14.5 / 14.6 / 14.6
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