Rega HIV-1 Subtying Tool V3 Methods, Reference Sequences and CRFs

The subtyping process consists of four steps:
  • The initial step involves construction of a master alignment for the query sequence and the reference strains of subtypes and CRFs.
  • The second step involves the construction of a phylogenetic tree using HKY distance method in PAUP* software. The alignment contains the sequence under query and group M pure reference sequences for subtypes A-D, F-H, J and K.
  • The next step, involves construction of a second tree using the sequence under query, HIV-1 pure subtypes, and CRFs sequences.
  • In the third step for sequences > 800 bps, the pure subtype alignment is divided into small segments and is analysed for recombination using bootscanning using a sliding window of 400 bps moving in steps of 40 bps.
  • In addition, if recombination and clustering with CRF is detected for > 800 bps, the pure subtype and the detected CRF reference sequences alignment is divided into small segments and is analysed for recombination using bootscanning using a sliding window of 400 bps moving in steps of 40 bps.
  • For sequences < 800 bps the alignment is examined to determine whether it contains sufficient phylogenetic signals for subtype determination using Treepuzzle software.
1. Reference Alignment:
Subtypes or CRFs Reference Sequences
Subtype A1 A1_UA_01_01UADN139_DQ823357
A1_RU_00_RU00051_EF545108
A1_UG_UG031_AB098330
Subtype A2 A2_CD9797CDKTB48
A2_CY9494CY01741
Subtype B B_US_1986_5019_86_AY835780
B_US_90_US2_AY173953
B_BR_03_BREPM1028_EF637053
Subtype C C_ZA_98_TV001_AY162223
C_IN_x_VB39_EF694033
C_IL_99_99ET1_AY255823
Subtype D D_UG_99_99UGG35093_AF484495
D_UG_98_98UG57143_AF484514
D_ZA_84_R2_AY773338
Subtype F1 F1_ES_x_X1670_DQ979024
F1_BR_89_BZ126_AY173957
Subtype F2 F2_CM_95_MP255_AJ249236
F2_CM_97_CM53657_AF377956
Subtype G G_NG_01_01NGPL0669_DQ168576
G_ES_05_ES_EU786670
G_GH_2003_GHNJ175_AB231893
Subtype H H_CF_90_056_AF005496
H_BE_93_VI997_AF190128
H_BE_93_VI991_AF190127
Subtype J J_SE_93_SE7887_AF082394
J_SE_1994_SE7022_AF082395_1
J_CD_97_J_97DC_KTB147_EF614151
Subtype K K_CD_97_EQTB11C_AJ249235
K_CM_96_MP535_AJ249239
CRF01_AE 01_AECF19090CF11697
01_AETH90CM240
CRF02_AG 02_AGCM9797CMP807
02_AGFR91DJ264
CRF03_AB 03_ABRU97KAL153
03_ABRU98RU98001
CRF04_CPX 04_CPXG1R9197PVCH
04_CPXGR9797PVMY
CRF05_DF 05_DFBEVI1310
05_DFBE93VI961
CRF06_CPX 06_CPXAU96BFP90
06_CPXML9595ML84
CRF07_BC 07_BCC1N9797CN001
07_BCCN9898CN009
CRF08_BC 08_BCCN19797CNGX7F
08_BCCN9898CN006
CRF09_CPX 09_cpxCI0000IC_10092AJ866553
09_cpxGH9696GH2911AY093605
CRF10_CD 10_CDTZ19696TZBF071
10_CDTZ9696TZBF110
CRF11_CPX 11_CPXF1R99MP1298
11_CPXGRGR17
CRF12_BF 12_BFAR97A32989
12_BFU1Y99URTR23
CRF13_CPX 13_CPXC1M961849
13_CPXCM964164
CRF14_BG 14_BGE1S00X477
14_BGES99X397
CRF18_CPX 18_cpxCM97CM53379AF377959
18_cpxCU99CU14AY586541
CRF19_CPX 19_cpxCU99CU29AY588971
19_cpxCU99CU7AY894994
CRF20_BG 20_BGCU99Cu103AY586545
20_BGES99R77AY586544
CRF24_BG 24_BGCU03CB378AY900574
24_BGCU03CB471AY900575
CRF25_CPX 25_cpxCM0606CM_BA_040EU693240
25_cpxSA03J11451EU697908
CRF27_CPX 27_cpxCD9797CDKTB49AJ404325
27_cpxFR0404CD_FR_KZSAM851091
CRF29_BF 29_BFBR01BREPM16704DQ085876
29_BFBR99BREPM11948DQ085871
CRF31_BC 31_BCBR02110PAEF091932
31_BCBR0404BR142AY727527
CRF35_AD 35_ADAF0505AF026EF158043
35_ADAF0505AF094EF158040
CRF37_CPX 37_cpxCM0000CMNYU926EF116594
37_cpxCM97CM53392AF377957
CRF39_BF 39_BFBR0303BRRJ103EU735534
39_BFBR0303BRRJ327EU735536
CRF40_BF 40_BFBR0404BRRJ115EU735538
40_BFBR0404BRSQ46EU735540
CRF42_BF 42_BF_LU_04_EU170142
42_BF_LU_03_EU170151
CRF43_02G 43_02GSA03J11223EU697904
43_02GSA03J11243EU697907
CRF47_BF 47BFES2008P1942GQ372987
47BFES2008X24572FJ670529
Information on the selection of CRFs for usage in the subytping tool is here 2. Program software used in the tool:
This application is built and maintained by Tulio de Oliveira, Pieter Libin, Koen Deforche, Sharon Cassol and Anne-Mieke Vandamme.