Acta Agriculturae Zhejiangensis ›› 2025, Vol. 37 ›› Issue (3): 548-558.DOI: 10.3969/j.issn.1004-1524.20240356
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QIAN Mingzhu1,2(), WANG Shenfeng1, CHANG Xiaoran2, HU Junying2, ZHU Hongbiao3, WANG Xinping2,*(
)
Received:
2024-04-25
Online:
2025-03-25
Published:
2025-04-02
CLC Number:
QIAN Mingzhu, WANG Shenfeng, CHANG Xiaoran, HU Junying, ZHU Hongbiao, WANG Xinping. Characterization and analysis of the complete genome sequence of two strains of bovine enterovirus in Henan Province of China[J]. Acta Agriculturae Zhejiangensis, 2025, 37(3): 548-558.
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URL: http://www.zjnyxb.cn/EN/10.3969/j.issn.1004-1524.20240356
片段 Fragment | 引物序列 Primer sequences(5'-3') | |
---|---|---|
F1 | S | TTTAAAACAGCCTGGGGGTT |
AS | TGGGATTAGCAGCATTCACG | |
F2 | S | ATGGGCGGTCAGTTCAGCAG |
AS | GTACGAGGTGAGGCTAGCCCTGT | |
F3 | S | GGGCCTTTTGGACAACAACAAGG |
AS | TTGAACGAGACCTCGAGATCT | |
F4 | S | GGCCCCGTAACCTACAAAGGTA |
AS | GAATCGAATACATACCATTCGAC | |
3' | S | TTGTGCGCGATCAGCGCTACC |
Table 1 Primers for amplifying the full-length genome sequence of HeN-B78 strain
片段 Fragment | 引物序列 Primer sequences(5'-3') | |
---|---|---|
F1 | S | TTTAAAACAGCCTGGGGGTT |
AS | TGGGATTAGCAGCATTCACG | |
F2 | S | ATGGGCGGTCAGTTCAGCAG |
AS | GTACGAGGTGAGGCTAGCCCTGT | |
F3 | S | GGGCCTTTTGGACAACAACAAGG |
AS | TTGAACGAGACCTCGAGATCT | |
F4 | S | GGCCCCGTAACCTACAAAGGTA |
AS | GAATCGAATACATACCATTCGAC | |
3' | S | TTGTGCGCGATCAGCGCTACC |
片段 Fragment | 引物序列 Primer sequences(5'-3') | |
---|---|---|
F1 | S | TTTTAAAACCAGCCTGGGGGTT |
AS | TACGCGTTACGACGTAGCAAC | |
F2 | S | ATGGGTGCGCAAGTAAGCAA |
AS | CTGGAGGGCTGCGGTCTGAGTCA | |
F3 | S | GGGGATGTTAAGGACTCAATC |
AS | TTGCAGTCCAGCAAAGAGCTTG | |
F4 | S | GGACCCTACTCAGGGATCCACAC |
AS | GAAGGAGTCGTACCAGGAGCG | |
3' | S | GCAAAACTGCTCGCGGACGC |
Table 2 Primers for amplifying the full-length genome sequence of HeN-YR91 strain
片段 Fragment | 引物序列 Primer sequences(5'-3') | |
---|---|---|
F1 | S | TTTTAAAACCAGCCTGGGGGTT |
AS | TACGCGTTACGACGTAGCAAC | |
F2 | S | ATGGGTGCGCAAGTAAGCAA |
AS | CTGGAGGGCTGCGGTCTGAGTCA | |
F3 | S | GGGGATGTTAAGGACTCAATC |
AS | TTGCAGTCCAGCAAAGAGCTTG | |
F4 | S | GGACCCTACTCAGGGATCCACAC |
AS | GAAGGAGTCGTACCAGGAGCG | |
3' | S | GCAAAACTGCTCGCGGACGC |
序号 Numbers | 毒株名称 Strains | GenBank登录号 GenBank accession number | 年份 Year | 来源国家 Country |
---|---|---|---|---|
1 | K2577 | AF123432 | 1999 | 澳大利亚Australia |
2 | SL305 | AF123433 | 1999 | 澳大利亚Australia |
3 | VG-5-27 | NC_001859 | 2000 | 美国USA |
4 | JL-DH13 | MN598020 | 2012 | 中国China |
5 | HeN-A2 | MN598008 | 2018 | 中国China |
6 | HeN-A3 | MN598009 | 2018 | 中国China |
7 | HeN-A12 | MN598010 | 2018 | 中国China |
8 | HeN-B81 | MN598014 | 2018 | 中国China |
9 | HeN-B83 | MN598015 | 2018 | 中国China |
10 | HeN-B84 | MN598016 | 2018 | 中国China |
11 | HeN-C4 | MN598017 | 2018 | 中国China |
12 | JL-40 | MN598019 | 2018 | 中国China |
13 | JL-JY42 | MN598032 | 2018 | 中国China |
14 | NX-b-DR47 | MN598037 | 2018 | 中国China |
15 | NX-FY40 | MN598039 | 2018 | 中国China |
16 | E1_NSWL13 | OP020153 | 2022 | 澳大利亚Australia |
17 | SD-S67 | MK639928.1 | 2018 | 中国China |
18 | W17_02174-75 | MW959114.1 | 2017 | 新西兰New Zealand |
19 | W17_02174-23 | MW959113.1 | 2017 | 新西兰New Zealand |
20 | W17_00094-318 | MW959112.1 | 2017 | 新西兰New Zealand |
21 | W16_00634-15 | MW959109.1 | 2016 | 新西兰New Zealand |
22 | W16_00544-Red6 | MW959108.1 | 2016 | 新西兰New Zealand |
23 | W16_00544-CVGW336 | MW959107.1 | 2016 | 新西兰New Zealand |
24 | W16_00059-139 | MW959105.1 | 2015 | 新西兰New Zealand |
25 | W16_00059-89 | MW959104.1 | 2015 | 新西兰New Zealand |
26 | HeN-B62 | MN598011.1 | 2018 | 中国China |
27 | NGR_2018 | MN650196.1 | 2018 | 尼日利亚Nigeria |
28 | IL-alpaca | KC748420.1 | 2007 | 美国USA |
29 | IS2-Bos_taurus-JPN-1990 | LC150010.1 | 1990 | 日本Japan |
30 | Ho12-Bos_taurus-JPN-2014 | LC150008.1 | 2014 | 日本Japan |
31 | BEV-261 | NC_021220.1 | 1958 | 美国USA |
32 | W1 | AY462106.1 | 2002 | 新西兰New Zealand |
33 | BJ001 | HQ663846.1 | 2009 | 中国China |
34 | BJ50 | HQ917061.1 | 2008 | 中国China |
35 | BHM26 | HQ917060.1 | 2008 | 中国China |
36 | 3A | AY508697.1 | 2004 | 美国USA |
37 | PS87 | AY508696.1 | 1962 | 美国USA |
Table 3 BEV reference strains information
序号 Numbers | 毒株名称 Strains | GenBank登录号 GenBank accession number | 年份 Year | 来源国家 Country |
---|---|---|---|---|
1 | K2577 | AF123432 | 1999 | 澳大利亚Australia |
2 | SL305 | AF123433 | 1999 | 澳大利亚Australia |
3 | VG-5-27 | NC_001859 | 2000 | 美国USA |
4 | JL-DH13 | MN598020 | 2012 | 中国China |
5 | HeN-A2 | MN598008 | 2018 | 中国China |
6 | HeN-A3 | MN598009 | 2018 | 中国China |
7 | HeN-A12 | MN598010 | 2018 | 中国China |
8 | HeN-B81 | MN598014 | 2018 | 中国China |
9 | HeN-B83 | MN598015 | 2018 | 中国China |
10 | HeN-B84 | MN598016 | 2018 | 中国China |
11 | HeN-C4 | MN598017 | 2018 | 中国China |
12 | JL-40 | MN598019 | 2018 | 中国China |
13 | JL-JY42 | MN598032 | 2018 | 中国China |
14 | NX-b-DR47 | MN598037 | 2018 | 中国China |
15 | NX-FY40 | MN598039 | 2018 | 中国China |
16 | E1_NSWL13 | OP020153 | 2022 | 澳大利亚Australia |
17 | SD-S67 | MK639928.1 | 2018 | 中国China |
18 | W17_02174-75 | MW959114.1 | 2017 | 新西兰New Zealand |
19 | W17_02174-23 | MW959113.1 | 2017 | 新西兰New Zealand |
20 | W17_00094-318 | MW959112.1 | 2017 | 新西兰New Zealand |
21 | W16_00634-15 | MW959109.1 | 2016 | 新西兰New Zealand |
22 | W16_00544-Red6 | MW959108.1 | 2016 | 新西兰New Zealand |
23 | W16_00544-CVGW336 | MW959107.1 | 2016 | 新西兰New Zealand |
24 | W16_00059-139 | MW959105.1 | 2015 | 新西兰New Zealand |
25 | W16_00059-89 | MW959104.1 | 2015 | 新西兰New Zealand |
26 | HeN-B62 | MN598011.1 | 2018 | 中国China |
27 | NGR_2018 | MN650196.1 | 2018 | 尼日利亚Nigeria |
28 | IL-alpaca | KC748420.1 | 2007 | 美国USA |
29 | IS2-Bos_taurus-JPN-1990 | LC150010.1 | 1990 | 日本Japan |
30 | Ho12-Bos_taurus-JPN-2014 | LC150008.1 | 2014 | 日本Japan |
31 | BEV-261 | NC_021220.1 | 1958 | 美国USA |
32 | W1 | AY462106.1 | 2002 | 新西兰New Zealand |
33 | BJ001 | HQ663846.1 | 2009 | 中国China |
34 | BJ50 | HQ917061.1 | 2008 | 中国China |
35 | BHM26 | HQ917060.1 | 2008 | 中国China |
36 | 3A | AY508697.1 | 2004 | 美国USA |
37 | PS87 | AY508696.1 | 1962 | 美国USA |
Fig.1 3'RACE PCR amplification results of HeN-B78 strain and HeN-YR91 strain M, DL5000 DNA marker; 1, 3' UTR of HeN-B78 strain amplified gene fragment; 2, Negative control; 3, 3' UTR of HeN-YR91 strain amplified gene fragment; 4, Negative control.
Fig.2 RT-PCR identification of BEV HeN-B78 strain by 4 pairs of primers M, DL5000 DNA marker; 1-4, The target gene fragment amplified by primers F1-F4 for HeN-B78; 5, Negative control.
Fig.3 RT-PCR identification of BEV HeN-YR91 strain isolate by 4 primers M, DL5000 DNA marker; 1-4, The target gene fragment amplified by primers F1-F4 for HeN-YR91; 5, Negative control.
功能区 Function regions | HeN-B78 | HeN-YR91 | ||||
---|---|---|---|---|---|---|
位置 Positions | 基因大小 Gene size/nt | 氨基酸数量 Amino acid number | 位置 Positions | 基因大小 Gene size/nt | 氨基酸数量 Amino acid number | |
5'UTR | 1~812 | 812 | — | 1~825 | 825 | — |
VP4 | 813~1 025 | 213 | 71 | 826~1 032 | 207 | 69 |
VP2 | 1 026~1 763 | 738 | 246 | 1 033~1 770 | 738 | 246 |
VP3 | 1 764~2 489 | 726 | 242 | 1 771~2 499 | 729 | 243 |
VP1 | 2 490~3 329 | 840 | 280 | 2 500~3 324 | 825 | 275 |
2A | 3 330~3779 | 450 | 150 | 3 325~3 774 | 450 | 150 |
2B | 3 780~4 076 | 297 | 99 | 3 775~4 071 | 297 | 99 |
2C | 4 077~5 066 | 990 | 330 | 4 072~5 058 | 987 | 329 |
3A | 5 067~5 333 | 267 | 89 | 5 059~5 325 | 267 | 89 |
3B | 5 334~5 402 | 69 | 23 | 5 326~5 394 | 69 | 23 |
3C | 5 403~5 951 | 549 | 183 | 5 395~5 943 | 549 | 183 |
3D | 5 952~7 334 | 1 383 | 461 | 5 944~7 326 | 1 383 | 461 |
P1 | 813~3 329 | 2 517 | 839 | 826~3 324 | 2 499 | 833 |
P2 | 3 330~5 066 | 1 737 | 579 | 3 325~5 058 | 1 734 | 578 |
P3 | 5 067~7 334 | 2 268 | 756 | 5 059~7 326 | 2 268 | 756 |
3'UTR | 7 335~7 453 | 118 | 7 327~7 460 | 133 |
Table 4 Nucleotide and amino acid numbers in coding regions of BEV HeN-B78 and HeN-YR91
功能区 Function regions | HeN-B78 | HeN-YR91 | ||||
---|---|---|---|---|---|---|
位置 Positions | 基因大小 Gene size/nt | 氨基酸数量 Amino acid number | 位置 Positions | 基因大小 Gene size/nt | 氨基酸数量 Amino acid number | |
5'UTR | 1~812 | 812 | — | 1~825 | 825 | — |
VP4 | 813~1 025 | 213 | 71 | 826~1 032 | 207 | 69 |
VP2 | 1 026~1 763 | 738 | 246 | 1 033~1 770 | 738 | 246 |
VP3 | 1 764~2 489 | 726 | 242 | 1 771~2 499 | 729 | 243 |
VP1 | 2 490~3 329 | 840 | 280 | 2 500~3 324 | 825 | 275 |
2A | 3 330~3779 | 450 | 150 | 3 325~3 774 | 450 | 150 |
2B | 3 780~4 076 | 297 | 99 | 3 775~4 071 | 297 | 99 |
2C | 4 077~5 066 | 990 | 330 | 4 072~5 058 | 987 | 329 |
3A | 5 067~5 333 | 267 | 89 | 5 059~5 325 | 267 | 89 |
3B | 5 334~5 402 | 69 | 23 | 5 326~5 394 | 69 | 23 |
3C | 5 403~5 951 | 549 | 183 | 5 395~5 943 | 549 | 183 |
3D | 5 952~7 334 | 1 383 | 461 | 5 944~7 326 | 1 383 | 461 |
P1 | 813~3 329 | 2 517 | 839 | 826~3 324 | 2 499 | 833 |
P2 | 3 330~5 066 | 1 737 | 579 | 3 325~5 058 | 1 734 | 578 |
P3 | 5 067~7 334 | 2 268 | 756 | 5 059~7 326 | 2 268 | 756 |
3'UTR | 7 335~7 453 | 118 | 7 327~7 460 | 133 |
基因名称 Function regions | 比对类型 Comparison type | HeN-B78 | HeN-YR91 | ||
---|---|---|---|---|---|
同E种参考毒株 Reference strains of EV-E | 同F种参考毒株 Reference strains of EV-F | 同E种参考毒株 Reference strains of EV-E | 同F种参考毒株 Reference strains of EV-F | ||
Genome | 核苷酸Nucleotide | 75.9~88.0 | 67.1~68.1 | 67.2~68.5 | 74.6~82.4 |
氨基酸Amino acid | — | — | — | — | |
VP1 | 核苷酸Nucleotide | 71.3~77.7 | 51.9~56.9 | 54.1~58.2 | 61.5~80.7 |
氨基酸Amino acid | 76.4~97.5 | 54.6~58.3 | 54.0~57.0 | 67.2~90.2 | |
VP2 | 核苷酸Nucleotide | 74.7~79.7 | 65.0~67.6 | 65.3~68.8 | 71.0~81.2 |
氨基酸Amino acid | 93.0~98.8 | 74.6~77.5 | 76.6~78.9 | 84.6~97.2 | |
VP3 | 核苷酸Nucleotide | 74.5~78.7 | 59.0~64.6 | 60.7~63.4 | 70.2~83.0 |
氨基酸Amino acid | 89.3~95.5 | 62.0~66.1 | 61.2~64.0 | 78.2~90.5 | |
VP4 | 核苷酸Nucleotide | 69.6~81.2 | 67.1~72.5 | 66.7~71.0 | 78.3~86.0 |
氨基酸Amino acid | 87.0~92.8 | 76.8~79.7 | 71.0~79.7 | 94.2~100.0 | |
2A | 核苷酸Nucleotide | 72.0~95.1 | 66.9~72.0 | 65.3~70.4 | 66.9~72.0 |
氨基酸Amino acid | 84.7~99.3 | 74.0~79.3 | 72.7~79.3 | 78.7~98.0 | |
2B | 核苷酸Nucleotide | 73.4~96.0 | 59.6~66.0 | 63.6~67.0 | 71.0~83.8 |
氨基酸Amino acid | 90.9~99.0 | 67.7~72.7 | 68.7~72.7 | 84.8~100.0 | |
2C | 核苷酸Nucleotide | 77.1~83.8 | 67.7~71.5 | 67.0~71.6 | 78.0~86.2 |
氨基酸Amino acid | 95.2~99.4 | 78.4~80.2 | 78.1~79.3 | 93.6~99.4 | |
3A | 核苷酸Nucleotide | 74.9~80.5 | 61.0~67.0 | 58.4~64.8 | 74.5~82.8 |
氨基酸Amino acid | 85.4~98.9 | 64.0~69.7 | 66.3~69.7 | 89.95~97.8 | |
3B | 核苷酸Nucleotide | 72.5~85.5 | 60.9~73.9 | 59.4~68.1 | 75.4~85.5 |
氨基酸Amino acid | 91.3~100.0 | 69.6~78.3 | 69.6~78.3 | 91.3~100.0 | |
3C | 核苷酸Nucleotide | 73.4~83.8 | 64.1~68.9 | 65.4~69.6 | 72.9~82.9 |
氨基酸Amino acid | 89.6~97.3 | 72.1~74.9 | 72.1~74.3 | 85.8~97.8 | |
3D | 核苷酸Nucleotide | 78.6~86.9 | 70.9~73.3 | 70.7~73.7 | 79.2~86.2 |
氨基酸Amino acid | 92.8~98.5 | 82.9~84.6 | 82.9~83.7 | 93.3~98.3 |
Table 5 Results of homology analysis of isolated strains with reference strains %
基因名称 Function regions | 比对类型 Comparison type | HeN-B78 | HeN-YR91 | ||
---|---|---|---|---|---|
同E种参考毒株 Reference strains of EV-E | 同F种参考毒株 Reference strains of EV-F | 同E种参考毒株 Reference strains of EV-E | 同F种参考毒株 Reference strains of EV-F | ||
Genome | 核苷酸Nucleotide | 75.9~88.0 | 67.1~68.1 | 67.2~68.5 | 74.6~82.4 |
氨基酸Amino acid | — | — | — | — | |
VP1 | 核苷酸Nucleotide | 71.3~77.7 | 51.9~56.9 | 54.1~58.2 | 61.5~80.7 |
氨基酸Amino acid | 76.4~97.5 | 54.6~58.3 | 54.0~57.0 | 67.2~90.2 | |
VP2 | 核苷酸Nucleotide | 74.7~79.7 | 65.0~67.6 | 65.3~68.8 | 71.0~81.2 |
氨基酸Amino acid | 93.0~98.8 | 74.6~77.5 | 76.6~78.9 | 84.6~97.2 | |
VP3 | 核苷酸Nucleotide | 74.5~78.7 | 59.0~64.6 | 60.7~63.4 | 70.2~83.0 |
氨基酸Amino acid | 89.3~95.5 | 62.0~66.1 | 61.2~64.0 | 78.2~90.5 | |
VP4 | 核苷酸Nucleotide | 69.6~81.2 | 67.1~72.5 | 66.7~71.0 | 78.3~86.0 |
氨基酸Amino acid | 87.0~92.8 | 76.8~79.7 | 71.0~79.7 | 94.2~100.0 | |
2A | 核苷酸Nucleotide | 72.0~95.1 | 66.9~72.0 | 65.3~70.4 | 66.9~72.0 |
氨基酸Amino acid | 84.7~99.3 | 74.0~79.3 | 72.7~79.3 | 78.7~98.0 | |
2B | 核苷酸Nucleotide | 73.4~96.0 | 59.6~66.0 | 63.6~67.0 | 71.0~83.8 |
氨基酸Amino acid | 90.9~99.0 | 67.7~72.7 | 68.7~72.7 | 84.8~100.0 | |
2C | 核苷酸Nucleotide | 77.1~83.8 | 67.7~71.5 | 67.0~71.6 | 78.0~86.2 |
氨基酸Amino acid | 95.2~99.4 | 78.4~80.2 | 78.1~79.3 | 93.6~99.4 | |
3A | 核苷酸Nucleotide | 74.9~80.5 | 61.0~67.0 | 58.4~64.8 | 74.5~82.8 |
氨基酸Amino acid | 85.4~98.9 | 64.0~69.7 | 66.3~69.7 | 89.95~97.8 | |
3B | 核苷酸Nucleotide | 72.5~85.5 | 60.9~73.9 | 59.4~68.1 | 75.4~85.5 |
氨基酸Amino acid | 91.3~100.0 | 69.6~78.3 | 69.6~78.3 | 91.3~100.0 | |
3C | 核苷酸Nucleotide | 73.4~83.8 | 64.1~68.9 | 65.4~69.6 | 72.9~82.9 |
氨基酸Amino acid | 89.6~97.3 | 72.1~74.9 | 72.1~74.3 | 85.8~97.8 | |
3D | 核苷酸Nucleotide | 78.6~86.9 | 70.9~73.3 | 70.7~73.7 | 79.2~86.2 |
氨基酸Amino acid | 92.8~98.5 | 82.9~84.6 | 82.9~83.7 | 93.3~98.3 |
Fig.4 Genetic evolution analysis of BEV ●The isolates in this test; a, Phylogenetic tree analysis of the whole genome; b-l, Phylogenetic tree analysis of VP1-VP4, 2A-2C, 3A-3D.
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