Isolation, Identification and Whole-genome Sequence Analysis of Phytophthora nicotianae Antagonistic Bacteria XC-29

doi:10.19586/j.2095-2341.2024.0086

Current Biotechnology ›› 2024, Vol. 14 ›› Issue (6): 1004-1015.DOI: 10.19586/j.2095-2341.2024.0086

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Isolation, Identification and Whole-genome Sequence Analysis of Phytophthora nicotianae Antagonistic Bacteria XC-29

Sujie WANG¹(), Mengli GU¹, Jiemeng TAO¹^,², Zhijun TONG³, Junjia GUO¹, Jingjing JIN¹^,², Mengxiao XU¹, Lijun MENG¹, Jianfeng ZHANG¹^,², Peijian CAO¹^,², Peng LU¹^,²()

^1.Zhengzhou Tobacco Research Institute of China National Tobacco Corporation，Zhengzhou 450001，China
^2.Beijing Life Science Academy，Beijing 102200，China
^3.Yunan Academy of Tobacco Agricultural Sciences，Kunming 650106，China

Received:2024-04-17 Accepted:2024-06-21 Online:2024-11-25 Published:2024-12-27
Contact: Peng LU

烟草疫霉拮抗菌XC-29的分离鉴定及其全基因组序列分析

汪苏洁¹(), 顾梦丽¹, 陶界锰¹^,², 童治军³, 郭俊佳¹, 金静静¹^,², 徐梦晓¹, 孟利军¹, 张剑锋¹^,², 曹培健¹^,², 卢鹏¹^,²()

^1.中国烟草总公司郑州烟草研究院，郑州 450001
^2.北京生命科技研究院，北京 102200
^3.云南省烟草农业科学研究院，昆明 650106

通讯作者: 卢鹏
作者简介:汪苏洁 E-mail: wangsujie951230@163.com；
基金资助:
河南省科技攻关项目(212102110257);中国烟草总公司重点研发项目〔110202102050;110202201001(JY-01）〕;郑州烟草研究院省席科学家项目(902023CK0890)

Abstract

Abstract:

Phytophthora nicotianae is a soil-borne oomycete that can cause black shank disease of tobacco， which is very harmful to tobacco production. A strain XC-29 with strong biocontrol function against P.nicotianae was screened， and its antagonistic metabolites and genes were mined by analyzing its genomic information. The antibacterial activity and control effect of biocontrol bacterium XC-29 were verified by the plate confrontation method and pot test. The strain was identified by morphological observation and 16S rRNA amplicon sequencing. The antagonistic mechanism of strains was verified the disease prevention effect of the strain. The strains were identified by exploration by whole genome and transcriptomic sequencing. The results showed that the antagonistic strain XC-29 isolated from rhizosphere soil was identified as Bacillus safensis. Its whole genome sequencing predicted 136 carbohydrate-active enzymes and 11 gene clusters encoding the synthesis of secondary metabolites， of which six were identified as the synthesis clusters of antimicrobial substances， encoding lichenysin， zwittermicin， schizokinen， fengycin， bacilysin and bacilibactin. The results of transcriptome sequencing further confirmed that the strain genes could encode corresponding antibacterial substances. In conclusion， Bacillus safensis XC-29 has a strong antagonistic effect on Phytophthora nicotianae. The whole genome and transcriptome analysis preliminarily revealed the disease inhibition mechanism of Bacillus safensis XC-29， which provides a theoretical basis for further exploration of the antibacterial mechanism and biological prevention and control of antagonistic bacteria.

Key words: tobacco black shank, Bacillus safensis, whole genome sequencing, secondary metabolites

摘要：

烟草疫霉（Phytophthora nicotianae）是一种可引发烟草黑胫病的土传卵菌，对烟草生产造成极大危害。为了应对这一问题，筛选出一株对烟草疫霉菌具有较强生防功能的菌株XC-29，通过解析基因组信息，挖掘其拮抗代谢产物及拮抗基因。采用平板对峙法和盆栽试验鉴定生防菌XC-29的抑菌活性和防治效果；通过形态观察和16S rRNA扩增子测序技术准确鉴定了XC-29菌株；利用全基因组测序、转录组测序探究菌株拮抗机制。结果表明，根际土分离的拮抗菌株XC-29鉴定为沙福芽孢杆菌（Bacillus safensis），其全基因组测序预测出136种碳水化合物活性酶和11个编码次生代谢产物合成相关的基因簇，其中6个基因簇已被确认为抗菌物质合成簇，分别编码地衣素、双效菌素、schizokinen、丰原素、杆菌溶素和嗜铁素；转录组测序结果进一步确定菌株基因具有编码相应抑菌物质的能力。综上，沙福芽孢杆菌XC-29对烟草疫霉菌具有较强的拮抗作用，全基因组和转录组分析初步揭示了沙福芽孢杆菌XC-29的抑病机理，为进一步探究拮抗菌的抑菌机制和生物防控提供理论依据。

关键词: 烟草黑胫病, 沙福芽孢杆菌, 全基因组测序, 次级代谢产物

CLC Number:

Sujie WANG, Mengli GU, Jiemeng TAO, Zhijun TONG, Junjia GUO, Jingjing JIN, Mengxiao XU, Lijun MENG, Jianfeng ZHANG, Peijian CAO, Peng LU. Isolation, Identification and Whole-genome Sequence Analysis of Phytophthora nicotianae Antagonistic Bacteria XC-29[J]. Current Biotechnology, 2024, 14(6): 1004-1015.

汪苏洁, 顾梦丽, 陶界锰, 童治军, 郭俊佳, 金静静, 徐梦晓, 孟利军, 张剑锋, 曹培健, 卢鹏. 烟草疫霉拮抗菌XC-29的分离鉴定及其全基因组序列分析[J]. 生物技术进展, 2024, 14(6): 1004-1015.

Figures/Tables 13

References 32

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类型	特征	数值
基因组	染色体个数	1
基因组	基因组序列总长度/bp	3 719 058
编码蛋白基因	基因数	3 870
	基因总长度/bp	3 310 209
	基因平均长度/bp	855
	基因序列中GC含量/%	42.24
基因间区	基因间区总长度/bp	408 849
基因间区	基因间区GC含量/%	37.87
非编码	tRNA拷贝数	81
	5S rRNA拷贝数	8
	16S rRNA拷贝数	8
	23S rRNA拷贝数	8
	sRNA拷贝数	7

类型	特征	数值
基因组	染色体个数	1
基因组	基因组序列总长度/bp	3 719 058
编码蛋白基因	基因数	3 870
	基因总长度/bp	3 310 209
	基因平均长度/bp	855
	基因序列中GC含量/%	42.24
基因间区	基因间区总长度/bp	408 849
基因间区	基因间区GC含量/%	37.87
非编码	tRNA拷贝数	81
	5S rRNA拷贝数	8
	16S rRNA拷贝数	8
	23S rRNA拷贝数	8
	sRNA拷贝数	7

类型	基因数	占总基因百分比
COG	2 896	74.83%
GO	2 708	69.97%
KEGG	3 652	94.37%
CAZy	136	3.51%

类型	基因数	占总基因百分比
COG	2 896	74.83%
GO	2 708	69.97%
KEGG	3 652	94.37%
CAZy	136	3.51%

通路编号	类目	基因数（>40）
Map01100	代谢途径	566
Map01110	次生代谢物的生物合成	276
Map01130	抗生素的生物合成	211
Map01120	不同环境下的微生物代谢	170
Map02010	ABC转运蛋白	138
Map01230	氨基酸的生物合成	130
Map02020	双组分调节系统	111
Map01200	碳代谢	98
Map02024	群体效应	70
Map00230	嘌呤代谢	59
Map03010	核糖体	52
Map00240	嘧啶代谢	45
Map00010	糖酵解/糖异生	44
Map00620	丙酮酸代谢	44
Map00270	半胱氨酸与蛋氨酸代谢	40

Isolation, Identification and Whole-genome Sequence Analysis of Phytophthora nicotianae Antagonistic Bacteria XC-29

烟草疫霉拮抗菌XC-29的分离鉴定及其全基因组序列分析

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Figures/Tables 13

References 32

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类型	家族成员	基因数
AA	AA1	1
CBM	CBM12、CBM13、CBM48、CBM50	35
CE	CE0、CE12、CE14、CE4、CE7、CE8、CE9	18
GH	GH0、GH1、GH3、GH4、GH5、GH9、GH10、GH11、GH13、GH16、GH18、GH23、GH28、GH30、GH32、GH33、GH38、GH42、GH43、GH48、GH51、GH53、GH73、GH101、GH105、GH126、GH170、GH171	55
GT	GT0、GT1、GT4、GT9、GT26、GT28、GT30、GT51、GT58	28
PL	PL1、PL9	2

基因簇	次级代谢产物类型	基因数量	相似基因簇	相似度
簇1	非核糖体肽合成酶	43	地衣素	BGC0000381（92%）
簇2	非核糖体肽合成酶、Ⅰ型聚酮类化合物	48	双效菌素	BGC0001059（18%）
簇3	包含Rev响应元件	24	铁载体二异羟肟酸酯	-
簇4	萜类化合物、镍-铁载体	27	-	BGC0002683（60%）
簇5	β-内酯	24	丰原素	BGC0001095（53%）
簇6	萜类化合物、镍-铁载体	21	-	-
簇7	Ⅲ型聚酮类化合物	47	-	-
簇8	核糖体合成和翻译后修饰肽类似物	15	-	-
簇9	β-内酯	33	-	-
簇10	其他	46	杆菌溶素	BGC0001184（85%）
簇11	非核糖体肽-载体、非核糖体肽合成酶	39	嗜铁素bacilibactin（E/F）	BGC0002695（80%）
			嗜铁素bacilibactin	BGC0000309（100%）
			嗜铁素paenibactin	BGC0000401（100%）
			嗜铁素bacilibactin	BGC0001185（100%）

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