Partial biological characteristics and genomic analysis of Vibrio cholerae typing phage VP2
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摘要:
目的 对霍乱弧菌分型噬菌体VP2基因组、蛋白质组和增殖特征等开展分析,获取VP2的生物学特征。 方法 电镜观察噬菌体VP2的形态,测定最佳感染复数和一步生长曲线;通过在线RAST(https://rast.nmpdr.org/)对噬菌体VP2基因组进行编码基因预测及功能注释;提取VP2成熟颗粒中的蛋白质进行质谱分析;使用pyani(https://github.com/widdowquinn/pyani)进行噬菌体全基因组平均核苷酸一致性(ANI)聚类分析。 结果 噬菌体VP2为典型的20面体短尾噬菌体,最佳感染复数为1∶100,一步生长曲线结果显示其潜伏期约为60 min,60~120 min为爆发期,120 min后为稳定期。 预测存在49个开放阅读框,VP2成熟颗粒蛋白质谱分析显示有34个蛋白与预测的基因相对应,ANI聚类分析结果显示VP2与弧菌噬菌体CJY的ANI最高。 结论 明确了分型噬菌体VP2的形态和基因组特征,预测的功能蛋白及结构蛋白的鉴定为后续研究同源噬菌体及其与霍乱弧菌的相互作用奠定了基础。 Abstract:Objective This study aimed to further understand the genetic information of VP2 by analyzing the whole genome of Vibrio cholerae typing phage VP2, and analyzing the protein of mature particles in combination with biological characteristics. Methods The morphology of phage VP2 was observed under electron microscope, and some of its biological characteristics were determined. Online RAST (https://rast.nmpdr.org/) was used to predict and annotate the encoding genes of the phage VP2 genome, and the proteins in the mature VP2 particles were analyzed by mass spectrometry. Phage genome average nucleotide consistency (ANI, business, nucleotide identity) was analysis by pyani (https://github.com/widdowquinn/pyani). Results Phage VP2 was a typical 20-hedron short-tailed phage, the optimal MOI is 0.01. One step growth curve showed that incubation period is about 60 minutes, 60 minutes to 120 minutes for the outbreak period, after 120 minutes for the stable period. Forty-nine open reading frames (ORFs) were predicted. Protein spectrum analysis of mature VP2 particles showed that 34 proteins corresponded to the predicted genes. ANI cluster analysis showed that ANI was the highest in VP2 and vibro phage CJY. Conclusion The morphological and genomic characteristics of typing phage VP2 were identified, which laid a foundation for the further study of homologous phage and its interaction with Vibrio cholerae. -
Key words:
- Typing phage VP2 /
- Vibrio cholerae /
- Biological characteristics /
- Genomic analysis
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表 1 VP2噬菌体与宿主菌16017的最佳感染复数
Table 1. The optimal MOI between VP2 phage and bacteria 16017
噬菌体滴度 宿主菌浓度 MOI(噬菌体∶菌) 结果 106 106 1∶1 2.00×106 105 106 1∶10 5.20×107 104 106 1∶100 6.80×107 表 2 VP2颗粒全蛋白质谱与VP2全基因组测序预测蛋白[14]比对结果
Table 2. Comparison between the results of VP2 granule protein spectrum and the predicted proteins of VP2 whole genome sequencing[14]
VP2ORF 起始基因位点 终止基因位点 氨基酸长度(aa) 功 能 蛋白覆盖度(%) 1 409 630 222 hypothetical protein − 2 647 1147 501 terminase small subunit 36 3 1134 2840 1707 terminase 15 4 2851 4494 1644 Head-tail connecting protein 78 5 4494 4721 228 hypothetical protein 96 6 4734 4991 258 hypothetical protein − 7 4996 5838 843 hypothetical protein 58 8 6006 6974 969 structural protein 84 9 7041 7310 270 hypothetical protein 97 10 7325 8173 849 hypothetical protein 77 11 8170 8535 366 hypothetical protein 59 12 8537 9010 474 LysM peptidoglycan-binding domain-containing protein 27 13 9003 9305 303 Phage protein 93 14 9329 11554 2226 PE family protein/tail protein 76 15 11563 13347 1785 structural protein 76 16 13353 13757 405 Phage protein 98 17 13754 15838 2085 Phage protein 93 18 15838 17010 1173 hypothetical protein 78 19 17012 19360 2349 structural protein 79 20 19364 20437 1074 Phage tail fibers 54 21 20439 21821 366 Outer capsid protein 53 22 22220 21846 474 hypothetical protein − 23 22677 22240 303 hypothetical protein 45 24 22828 22667 2226 hypothetical protein − 25 23022 22825 1785 metal dependent phosphohydrolase 65 26 23336 23019 405 Hydrolase 91 27 23598 23338 2085 hypothetical protein − 28 24631 23600 1032 Adenylosuccinate synthetase 71 29 26961 24673 2289 integrase 54 30 28819 26951 1869 DNA polymerase I 17 31 29337 28861 477 Single stranded DNA-binding protein,phage-associated 15 32 30212 29403 810 hypothetical protein 29 33 30800 30333 468 hypothetical protein 17 34 32332 30863 1470 superfamily II DNA/RNA helicase 29 35 32750 32397 354 hypothetical protein 17 36 33442 32750 693 hypothetical protein 20 37 33832 33575 258 hypothetical protein 18 38 34446 34036 411 structural protein − 39 34774 34448 327 hypothetical protein − 40 35624 34998 627 hypothetical protein 14 41 35828 35667 162 hypothetical protein − 42 36418 35933 486 hypothetical protein − 43 36859 36428 432 Phage protein 38 44 37407 37036 372 hypothetical protein − 45 37631 37407 225 hypothetical protein − 46 37999 37709 291 hypothetical protein − 47 38345 38106 240 hypothetical protein − 48 38946 38356 591 hypothetical protein − 49 39817 39548 270 hypothetical protein − -
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