Helicoverpa armigera NPV

ge elic leo -Cu , Kaix revisi e 27 distinction for NPVs is their recognition as either single NPVs (MNPVs) depending on the number of nucleocapsids al., 1983). So far, the genomic nucleotide sequences of 25 baculo- viruses have been completely sequenced. These include 15 nica (Ac) MNPV Virology 333 (2005) 1 $ The GenBank accession number of the HaSNPV-C1 genomic The Baculoviridae is a family of invertebrate viruses with large, circular, and double-stranded DNA genomes ranging in size from 81.7 (NeleNPV) to 178.7 kb (XecnGV). They are pathogenic to arthropods, mainly insects of the orders Lepidoptera, Hymenoptera, and Diptera (Adams and McClintock, 1991). The family is subdivided into two genera, Nucleopolyhedrovirus (NPV) and Granulovirus (GV), based on the morphology of occlusion bodies (OBs). The NPVs are designated as viruses forming polyhedral OBs, each of which contains many virions, whereas the GVs typically produce ovoid OBs with a single virion (Blissard et al., 2000). A further phenotypic packaged into each virion (Blissard et al., 2000). This, however, is not correlated to genetic relatedness and appears to have no phylogenetic trait (Murphy et al., 1995). NPVs are pathogenic to a number of lepidopteran insects and are attractive biological agents for the control of agriculturally important insect pests. The current interest in the molecular biology of these viruses is fostered by their potential as modified virus pesticides with increased toxicity (Stewart et al., 1991) and as gene therapy vectors in medical science (Huser and Hofmann, 2003; Tani et al., 2003), also by their successful use as vectors for the expression of foreign proteins in the baculovirus-insect system (Smith et The complete nucleotide sequence of Helicoverpa armigera single nucleocapsid nucleopolyhedrovirus isolate C1 (HearSNPV-C1) was determined and analyzed by comparing with the genome of HearSNPV-G4 isolate. C1 and G4 isolates occurred in the same host species and geographic location but showed different virulence. The HearSNPV-C1 genome consisted of 130,759 bp and 137 putative open reading frames larger than 150 nucleotides were identified. The two genomes shared 98.1% nucleotide sequence identity, with a total number of 555 bp substitutions, 1354 bp deletions, and 710 bp insertions in HearSNPV-C1. Comparison of ORFs and homologous repeat (hr) regions of the two genomes showed that there were four highly variable regions hr1, hr4, hr5, and bro-b, all in repeat regions. These results suggest that baculovirus strain heterogeneity may be often caused by SNPs and changes in the hrs and bro genes. D 2005 Elsevier Inc. All rights reserved. Keywords: Baculovirus; Helicoverpa armigera; Nucleocapsid nucleopolyhedrovirus; Complete genome; Isolate comparison Introduction nucleocapsid NPVs (SNPVs) or multiple nucleocapsid Comparison of the complete and G4 isolates of the H nucleocapsid nuc Chuan-Xi Zhang*, Xiu Institute of Applied Entomology, Zhejiang University Received 14 November 2004; returned to author for Available onlin Abstract 0042-6822/$ - see front matter D 2005 Elsevier Inc. All rights reserved. doi:10.1016/j.virol.2004.12.028 sequence reporte * Corresponding author. Fax: +86 571 86971697. E-mail address: chxzhang@zju.edu.cn (C.-X. Zhang). nome sequence between C1 overpa armigera single polyhedrovirus $ i Ma, Zhong-Jian Guo uan Road 268, Hangzhou, Zhejiang 310029, China on 30 November 2004; accepted 22 December 2004 January 2005 90–199 www.elsevier.com/locate/yviro lepidopteran NPVs: Autographa califor d in this paper is AF303045. (Ayres et al., 1994), Bombyx mori (Bm) NPV (Gomi et al., 1999), Orgyia pseudotsugata (Op) MNPV (Ahrens et al., box followed by a 20–25 bp downstream CAC/GT motif) within 180 bp of the initiation codon, 53 (38.7%) contain a rology 1997), Lymantria dispar (Ld) MNPV (Kuzio et al., 1999), Spodoptera exigua (Se) MNPV (Ijkel et al., 1999), Helicoverpa armigera (Hear) SNPV-G4 (Chen et al., 2001), Helicoverpa zea (Heze) SNPV (Chen et al., 2002), Adoxophyes honmai (Adho) NPV (Nakai et al., 2003), Spodoptera litura (Spli) MNPV (Pang et al., 2001), Epiphyas postvittana (Eppo) NPV (Hyink et al., 2002), Mamestra configurata (Maco) NPV-90/2 (Li et al., 2002a, 2002b), MacoNPV-96B (Li et al., 2002a, 2002b), Rachi- plusia ou (Raou) MNPV (Harrison and Bonning, 2003), Choristoneura fumiferana (Cf) MNPV (GenBank NC_004778), and Choristoneura fumiferana defective (CfDEF) NPV (GenBank NC_005137). The genomes of these NPVs range in size from 113.2 kb (AdhoNPV) to 161.0 kb (LdMNPV). In contrast to NPVs, only seven of GVs, Xestia cnigrum (Xecn) GV (Hayakawa et al., 1999), Plutella xylostalla (Plxy) GV (Hashimoto et al., 2000), Cydia pomonella (Cypo) GV (Luque et al., 2001), Phthorimaea operculella (Phop) GV (NC_004062), Adox- ophyes orana (Ador) GV (Wormleaton et al., 2003), Cryptophlebia leucotreta (Crle) GV (Lange and Jehle, 2003), and Agrotis segetum (Agse) GV (NC_005839) have been determined, with the genomes ranging in size from 99.7 kb (AdorGV) to 178.7 kb (XecnGV). A baculovirus from dipteran insect host, Culex nigripalpus (Cuni) bacu- lovirus (Afonso et al., 2001) and two NPVs from hymenopteran hosts, Neodiprion lecontei (Nele) NPV (Lauzon et al., 2004) and Neodiprion sertifer (Nese) NPV (Garcia-Maruniak et al., 2004), have also been determined, with genome sizes of 108.3, 86.5, and 81.8 kb, respectively. All these genomic sequences give us a better understanding of the distinctive features, evolution, and extent of diversity of baculoviruses. However, data concerning the strain polymorphism at the complete genomic sequence level are limited. H. armigera is one of the most serious pests in China. As an economically polyphagous pest, it has caused considerable economic loss to many vegetable and field crops such as cotton, corn, baccy, tomato, and wheat. As an important pathogen to H. armigera, HearSNPV was the first commercial baculovirus pesticide used to control the H. armigera in China, and also has been extensively used for the control of the pests in cotton and vegetable crops (Zhang, 1994). HearSNPV strains with different virulence or molecular characteristics have been isolated (Jia et al., 2003; Sun and Zhang, 1994). Bioassay showed that the virulence of HearSNPV isolate C1 was higher than that of HearSNPV isolate G4. The median lethal doses (LD50) for C1 and G4 against the third instar H. armigera were 568 (95% confidence interval 424–740) and 1584 (95% confidence interval 1065–2221) PIBs/larva, respectively. The genomic sequence of HearSNPV isolate G4 has already been determined (Chen et al., 2001). During the present study the genome of HearSNPV-C1 was com- C.-X. Zhang et al. / Vi pletely sequenced and compared with the genome of HearSNPV-G4 to further understand the strain heteroge- late promoter motif ((A/T/G) TAAG) within 120 bp of the initiation codon, and 17 (12.4%) have both an early and late promoter motif, which may allow transcription of these neity and the possible reasons for variation of the virulence in this virus and to provide clues for baculovirus evolution. Results and discussion Analysis of the HearSNPV-C1 genome The complete nucleotide sequence of the HearSNPV-C1 genome has been determined. The sequence data were assembled into a contiguous sequence of 130,759 bp (Tables 1), which was in good agreement with a previous estimate of 130 kb based on restriction enzyme analysis and physical mapping of DNA fragments (Zhang and Wu, 2001). According to the adopted convention (Hayakawa et al., 1999; Ijkel et al., 1999; Vlak and Smith, 1982), polyhedrin was designated as the first gene (ORF1) and the adenine residue at the translation-initiation codon of the polyhedrin gene was designated as the start point of the circular HearSNPV map. Using computer-assisted analysis and the criteria of selecting ORFs starting with methionine-initiated codons (ATG) and at least 50 aa having minimal overlap with other ORFs, 137 putative ORFs and five homologous repeat (hr) regions were identified for further detailed analysis in the HearSNPV-C1 genome. The location, orientation, and size of the predicted ORFs are shown in Table 1. The number of 137 ORFs is proportional to that of other completely sequenced baculoviruses ranging from NeleNPV (89) to MacoNPV-90/2 (169), especially similar to that of HezeSNPV (139), EppoNPV (135), SeMNPV (139), SpliMNPV (141), and BmNPV (143). The HearSNPV-C1 ORFs have average length of 843 bp with ORF84 (Heli- case) being the largest (3762 bp) and ORF40 being the smallest (153 bp). The 137 predicted ORFs encoded 38,362 aa. The total coding sequence and the intergenic regions were 114,394 and 10,041 bp and represent 87.5% versus 7.7% of the genome, respectively. The five hrs were distributed along the genome with sizes varying from 297 to 2253 bp and the total sequence was 6324 bp accounting for 4.8% of the genome. Twenty-three ORFs overlapped with adjacent ORFs in lengths ranging from 4 to 161 bp, totaling 1103 bp. Of the 137 HearSNPV-C1 ORFs identified, 135 (99.3%) had homologues in HezeSNPV, a possible variant of HearSNPV isolated from Helicoverpa zea (Chen et al., 2002). Of the 137 ORFs identified in HearSNPV-C1, only 27 (19.7%) possess a consensus early promoter motif (TATA 333 (2005) 190–199 191 genes during both early and late stages of infection, as has been reported for Spli19 (p10) and Spli57 (fp) (Kool and Table 1 Comparison of ORFs between HearSNPV isolate C1 and G4 ORF Name Position Length (aa) Predicted C1 compared with G4 MW G4ORF Nucleotide Amino acid residue (kDa) Substitution Insertion Deletion Substitution Insertion Deletion G4 length Identity % Note 1 polyhedrin 1 N 741 246 28.9 1 5[5] 3 0 5 1 0 245 97.6 d/i 2 p78/83 738 b 1982 414 46.1 2 14[8] 3 0 6 1 0 413 98.3 d/i 3 pk 1997 N 2800 267 31.6 3 2[1] 0 0 1 0 0 267 99.6 S 4 hoar 2923 b 5190 755 85.5 4 53 [15](3) 33 36 34 11 12 756 94.7 d/i 5b 5447 b 5638 63 7.4 2(2) 1 4 – – – – – – 6 5733 N 6590 285 34.4 6 10 [9] (1) 0 0(1) 2 0 0 285 98.6 S 7 6790 b 6966 58 6.7 7 0 6 0 0 2 0 56 94.8 d/i 8 ie-0 6954 N 7811 285 33.2 8 2 [2] 0 0 0 0 0 285 100 I 9 p49 7828 N 9234 468 55.2 9 5[5] (1) 0 0 1 0 0 468 99.8 S 10 odv-e18 9245 N 9490 81 8.8 10 2[2] 0 0 0 0 0 81 100 I 11 odv-ec27 9505 N 10359 284 33.3 11 9[9] 0 0 0 0 0 284 100 I 12 10404 N 10682 92 10.8 12 1[1] 0 0 0 0 0 92 100 I 13 ep23 10709 b 11314 201 22.6 13 8[5] 0 6 3 0 2 203 97.5 T 14 ie-1 11356 N 13341 661 76.5 14 18[14] 18 0 4 6 0 655 98.5 d/i 15 odv-e56 13395 b 14459 354 38.8 15 8[8] 0 0 0 0 0 354 100 I 16 me-53 14620 N 15699 359 42.7 16–17 0 0 1 0 0 0 284 100a E 18 15702 N 15869 55 6.4 18 0 0 0 0 0 0 55 100 I 19 15922 b 16203 93 11.1 19 0 0 0 0 0 0 93 100 I 20 p74 16224 N 18290 688 78.4 20 3 0 0 2 0 0 688 99.7 S 21 p10 18344 b 18607 87 9.3 21 0 0 0 0 0 0 87 100 I 22 p26 18690 b 19493 267 30.5 22 0 0 0 0 0 0 267 100 I 23 19607 N 19810 68 8.3 23 0 0 0 0 0 0 67 100 I 24 lef6 19886 b 20449 187 22.2 24 0 0 0 0 0 0 187 100 I 25 dbp 20463 b 21434 323 37.6 25 0 0 0 0 0 0 323 100 I 26 21578 N 22054 158 18.1 26 0 0 1 0 0 0 133 100a E repeat hr1 22129–24097 hr1 8 0 106 27 24225 b 24992 255 29.5 27 1b 0 0 1 0 0 255 99.6 S 28 ubiquitin 24832 N 25083 83 9.2 28 1[1] 0 0 0 0 0 83 100 I 29 25147 N 25653 168 20.4 29 0 0 0 0 0 0 168 100 I 30 e125 25673 N 26251 192 22.6 30 1 6 0 1 2 0 190 98.4 d/i 31 39K/pp31 26310 b 27248 312 35.3 31 1 3 0 1 1 0 311 99.3 d/i 32 lef11 27214 b 27597 127 14.6 32 0 0 0 0 0 0 127 100 I 33 bv-e31 27566 b 28282 238 28.4 33 0 0 0 0 0 0 238 100 I 34 28513 N 29592 359 41.2 34 1 0 0 1 0 0 359 99.7 S 35 p47 29667 b 30905 412 48.1 35 0 1 0 0 0 0 333 100a E 36 lef12 30978 N 31649 223 25.8 36 0 0 0 0 0 0 223 100 I 37 31735 N 31977 80 9.5 37 0 0 0 0 0 0 80 100 I 38 lef8 31974 b 34679 901 104.9 38 2[2] 0 0 0 0 0 901 100 I 39 34732 N 35316 194 22.5 39 0 0 0 0 0 0 194 100 I 40 35457 N 35609 50 6.3 40 0 0 0 0 0 0 50 100 I 41 chitinase 35617 b 37329 570 65.5 41 0 0 0 0 0 0 570 100 I 42 37408 b 37950 180 21.3 42 0 0 0 0 0 0 180 100 I 43 38067 N 38477 136 16.4 43 1(1) 0 0 1 0 0 136 99.3 S C .-X . Z h a n g et a l. / V iro lo g y 3 3 3 (2 0 0 5 ) 1 9 0 – 1 9 9 1 9 2 44 38484 b 39620 378 42.8 44 3[3] 0 0 0 0 0 378 100 I 45 39628 b 39855 75 9.1 45 0 0 0 0 0 0 75 100 I 46 lef10 39815 N 40030 71 7.7 46 1b 0 0 1 0 0 71 98.6 S 47 vp1054 39903 N 40958 351 41.7 47 4[1] 0 0 3 0 0 351 99.1 S 48 41078 N 41284 68 8.0 48 1 0 0 0 0 0 68 100 I 49 41285 N 41479 64 7.4 49 0(1) 0 0 0 0 0 64 100 I 50 41765 N 42280 171 20.7 50 3[2] 0 0 1 0 0 171 99.4 S 51 8.2kDa 42331 b 42810 159 18.9 51 3 0 3 3 0 1 160 97.5 d/I 52 42822 b 43088 88 10.2 52 0 0 0(1) 0 0 0 88 100 I 53 fp 25K 43300 b 43953 217 25.4 53 0 0 0 0 0 0 217 100 I 54 44125 N 44310 61 7.3 54 0 0 0(2) 0 0 0 61 100 I 55 lef9 44420 N 45979 519 59.6 55 1 0 0 1 0 0 519 99.8 S 56 cathepsin 46063 b 47160 366 42.4 56 0 0 0 0 0 0 365 100 I 57 47201 b 47788 195 21.3 57 0 0 0 0 0 0 195 100 I 58 gp37 47859 b 48698 279 32.1 58 0 0 0 0 0 0 279 100 I repeat hr2 48813–49719 hr2 2(1) 1 0 59 bro-a 49850 N 50584 244 28.3 59 1(14) 0 0 1 0 0 244 99.6 S 60 bro-b 50708 N 51781 357 40.2 60 70 [13] (1) 21 531 34 7 177 527 59.6 d/i repeat hr3 51949–52245 hr3 3(2) 0 0(1) 61 he65 52536 N 53246 236 27.5 61 0 0 0(1) 0 0 0 236 100 I 62 iap-2 53322 b 54074 250 29.2 62 7[7] 0 0 0 0 0 250 100 I 63 54122 b 54946 274 31.6 63 0 0 0 0 0 0 274 100 I 64 54915 b 55316 133 15.6 64 0 0 0 0 0 0 133 100 I 65 lef3 55336 N 56475 379 44.0 65 7[5] 0 0 2 0 0 379 99.5 S 66 93 kDa 56584 b 58940 785 88.9 66 2[2] 0 0 0 0 0 785 100 I 67 DNA pol 58971 N 62033 1020 119.2 67 11[11] 0 0 0 0 0 1020 100 I 68 30.5 kDa 62114 b 62572 152 17.6 68 0 0 0 0 0 0 152 100 I 69 62634 b 63017 127 14.9 69 0 0 0 0 0 0 127 100 I 70 63023 b 63280 85 10.0 70 0 0 0 0 0 0 85 100 I 71 vlf-1 63321 b 64562 413 48.0 71 0 3 0 0 1 0 412 99.8 d/i 72 64575 b 64907 110 12.7 72 0 0 0 0 0 0 110 100 I 73 gp41 64976 b 65944 322 36.6 73 2[2] 0 0 0 0 0 322 100 I 74 65874 b 66599 241 27.7 74 1[1] 0 0 0 0 0 241 100 I 75 66472 b 67149 225 24.9 75 0 0 0 0 0 0 225 100 I 76 vp91capsid 67079 N 69529 816 93.5 76 0 0 0 0 0 0 816 100 I 77 cg30 69657 b 70508 283 32.3 77 1[1] 0 0 0 0 0 283 100 I 78 p39 70597 b 71478 293 33.4 78 0 0 0 0 0 0 293 100 I 79 lef4 71477 N 72862 461 50.0 79 1 0 0 1 0 0 461 99.8 S 80 p33 72915 b 73679 254 30.8 80 0 0 0 0 0 0 254 100 I 81 73681 N 74169 162 19.1 81 0 0 0 0 0 0 162 100 I 82 odv-e25 74215 N 74907 230 25.9 82 0 0 0 0 0 0 230 100 I 83 74939 b 75436 165 18.8 83 3 0 0 3 0 0 165 98.2 S 84 helicase 75455 b 79216 1253 146.0 84 8[3] 0 0 4 0 0 1253 99.7 S 85 79173 N 79694 173 19.8 85 1 0 0 1 0 0 173 99.4 S 86 79753 b 80718 321 37.9 86 0 0 0 0 0 0 321 100 I 87 lef5 80614 N 81561 315 37.0 87 0 0 0 0 0 0 315 100 I 88 p6.9 81555 b 81884 109 11.5 88 0 0 0 0 0 0 109 100 I 89 81949 b 83058 369 42.6 89 1 0 0 0 0 0 369 100 I 90 13.1kDa 83104 N 83472 122 13.8 90 2 0 0 0 0 0 122 100 I (continued on next page) C .-X . Z h a n g et a l. / V iro lo g y 3 3 3 (2 0 0 5 ) 1 9 0 – 1 9 9 1 9 3 ble 1 (continued) RF Name Position Length (aa) Predicted C1 compared with G4 MW G4ORF Nucleotide Amino acid residue (kDa) Substitution Insertion Deletion Substitution Insertion Deletion G4 length Identity % Note 83472 b 84605 377 44.0 91 6[5] (1) 0(1) 0 1 0 0 377 99.7 S vp80/p87 84701 N 86518 605 69.7 92 13[3] 0 0 10 0 0 605 98.3 S 86515 N 86691 58 6.9 93 0 0 0 0 0 0 58 100 I odv-ec43 86706 N 87791 361 41.5 94 0 0 0 0 0 0 361 100 I 87837 N 88121 94 11.0 95 0(1) 0 0 0 0 0 94 100 I odv-e66 88188 b 90206 672 76.0 96 8[6] 0 0 2 0 0 672 99.7 S p13+ 90227 b 91057 276 32.5 97 0 0 0 0 0 0 276 100 I peat hr4 91222–93474 hr4 4 550 0 a 93485 b 93652 55 6.4 0 0 0 0 0 0 55 100 I 93913 N 94512 199 22.4 98 0 0 0 0 0 0 199 100 I 94516 N 94872 118 14.4 99 0 0 0 0 0 0 118 100 I 0 94968 N 96500 510 58.1 100 0 0 0 0 0 0 510 100 I 1 96579 N 97340 253 29.0 101 0 0 0 0 0 0 253 100 I 2 97355 N 97687 110 12.8 102 0 0 0 0 0 0 110 100 I 3 iap-3 97745 b 98551 268 31.5 103 0 0 0 0 0 0 268 100 I 4 98548 b 98703 51 5.9 104 0 0 0 0 0 0 100 100 I 5 bro-c 98814 b 100319 501 58.3 105 4[2] 0 0 2 0 0 501 99.6 S 6 100487 N 100966 159 16.8 106 2 0 0 1 0 0 159 99.4 S 7 sod 100973 N 102346 457 51.2 107 5[2] 0 0 2 0 0 457 99.6 S 8 102399 b 102977 192 22.7 108 1 0(3) 0 1 0 0 192 99.5 S 9 103149 N 103505 118 13.6 109 2[2] 0 0 0 0 0 118 100 I 0 103516 N 103782 88 10.1 110 0 0 0 0 0 0 88 100 I 1 pif 103850 N 105436 528 60.3 111 2[2] 0 0 0 0 0 528 100 I 2 105433 N 105669 78 9.1 112 0 0 0 0 0 0 78 100 I 3 fgf 105692 b 106597 301 34.3 113 7[5] (7) 0 0 2 0 0 301 99.3 S 4 alk-exo 106724 b 108010 428 49.4 114 2[2] 0 0 0 0 0 428 100 I 5 108030 b 108419 129 15.2 115 3[1] (2) 0 0(1) 2 0 0 129 98.4 S peat hr5 108500–109398 hr5 37(2) 43 648 5a 109493 b 110419 308 37.1 6[5] 0 0(2) 1 0 0 308 99.7 S 6 110618 N 110833 71 8.2 116 2 0 0 2 0 0 71 97.2 S 7 lef2 110951 b 111679 242 27.9 117 4[4] 3b 0 0 1 0 241 99.6 d/i 7a 111543 b 111950 135 15.7 0 3 0 0 1 0 134 99.2 d/i 8 p24capsid 112041 N 112787 248 28.3 118 1 0 0 1 0 0 248 99.6 S 9 gp16 112849 N 113139 96 10.9 119 4[4] 6 0 0 2 0 94 97.9 d/i 0 calyx/pep 113191 N 114213 340 39.1 120 0 0 0 0 0 0 340 100 I 1 114292 N 114756 154 18.5 121 0(3) 0 0(1) 0 0 0 154 100 I 2 odv-c21 114886 N 115476 196 23.4 122 3[1] (1) 0 0 2 0 0 196 99.0 S 3 38.7kd 115520 b 116689 389 44.9 123 3[2] 0 1 1 0 0 385 98.7 E 4 lef1 116691 b 117428 245 29.0 124 2[1] 0 0 1 0 0 245 99.6 S 5 117403 b 117831 142 16.0 125 5[2] (3) 0(1) 6(1) 3 0 2 144 96.5 d/i 6 egt 117976 N 119523 515 58.9 126 6[2] 0 0 3 0 0 515 99.4 S 7 119723 N 120301 192 22.6 127 2[1] 0 0 1 0 0 192 99.5 S C .-X . Z h a n g et a l. / V iro lo g y 3 3 3 (2 0 0 5 ) 1 9 0 – 1 9 9 1 9 4 Ta O 91 92 93 94 95 96 97 re 97 98 99 10 10 10 10 10 10 10 10 10 10 11 11 11 11 11 11 re 11 11 11 11 11 11 12 12 12 12 12 12 12 12 1 2 8 1 2 0 2 5 2 N 1 2 1 0 5 2 2 6 6 3 0 .4 1 2 8 0 0 0 0 0 0 2 2 6 1 0 0 I 1 2 9 1 2 1 1 3 3 b 1 2 3 9 7 6 9 4 7 1 1 1 .4 1 2 9 7 [2 ] 0 0 5 0 0 9 4 7 9 9 .5 S 1 3 0 p ki p -1 1 2 4 3 4 1 N 1 2 4 8 5 0 1 6 9 2 0 .3 1 3 0 5 [1 ] (1 ) 0 0 4 0 0 1 6 9 9 7 .6 S 1 3 1 a ri f1 1 2 4 9 1 7 b 1 2 5 7 1 4 2 6 5 3 0 .3 1 3 1 6 [3 ] 0 (1 ) 0 3 0 0 2 6 5 9 8 .9 S 1 3 2 p if -2 1 2 5 9 7 2 N 1 2 7 1 2 3 3 8 3 4 4 .5 1 3 2 1 2 [8 ] 0 0 3 0 0 3 8 3 9 9 .2 S 1 3 3 f- p ro te in 1 2 7 1 6 4 b 1 2 9 1 9 7 6 7 7 7 8 .2 1 3 3 3 2 [2 6 ] (2 ) 0 0 6 0 0 6 7 7 9 9 .1 S 1 3 4 1 2 9 3 3 9 b 1 2 9 8 8 4 1 8 1 2 1 .9 1 3 4 1 0 0 1 0 0 1 8 1 9 9 .4 S 1 3 5 1 3 0 0 6 6 N 1 3 0 6 5 3 1 9 5 2 3 .5 1 3 5 1 0 [3 ] (1 ) 3 0 5 1 0 1 9 4 9 6 .9 d /i N o te . T : tr u n ca te d ; S : si m il ar ; I: co m p le te ly id en ti ca l; d /i : d el et io n o r in se rt io n ; E : ex te n d ed . F ig u re s in sq u ar e b ra ck et s in d ic at e si le n t m u ta ti o n s. F ig u re s in p ar en th es es in d ic at e th e n u cl eo ti d e ch an g es o cc u rr ed in in te rg en ic (I G ) re g io n s. a O n ly th e tr u n ca te d re g io n s w er e co m p ar ed fo r O R F 1 6 , 2 6 an d 3 5 . b In se rt io n s o r d el et io n s in th e o v er la p p ed re g io n s o f tw o n ei g h b o ri n g O R F s. C.-X. Zhang et al. rology / Vi Vlak, 1993; Pang et al., 2001). Thirty-nine ORFs lack any recognized consensus early or late promoter motif within 180 bp of the ATG. Comparison of ORFs between HearSNPV-C1 and HearSNPV-G4 The genome of HearSNPV-G4 was reported 131,403 bp long and contained 135 ORFs (Chen et al., 2001). All the 135 ORFs identified in G4 were also found in C1. For comparing and to avoid confusion of ORF names in related studies, we updated the C1 genomic sequence data in GenBank (accession number AF303045). The 133 homol- ogous ORFs in C1 were numbered similar to those of G4. ORF16/