一株分离自烟草根际的生防菌株鉴定及其生防潜力研究

doi:10.13560/j.cnki.biotech.bull.1985.2025-1076

摘要/Abstract

摘要：

目的确定生防菌株YC25的分类地位，评估其抑菌能力，深入解析其基因组特征与次级代谢产物潜力，并阐明其对烟草植株的诱导抗病作用机制，为开发高效、环保的烟草土传病害生防制剂提供理论依据和技术支撑。方法采用形态学、生理生化和分子生物学方法对菌株YC25进行鉴定；通过对峙试验评估其抑菌能力，并进行全基因组测序和次级代谢产物预测；利用转录组学分析YC25处理对烟草植株基因表达的影响。结果菌株YC25被鉴定为斯氏芽胞杆菌（Bacillus spizizenii），对9种植物病原真菌具有广谱拮抗活性。田间试验表明，菌株YC25与甲霜恶霉灵联用可有效防治烟草根腐病，同时减少甲霜恶霉灵的用量。全基因组测序显示，YC25基因组总长4 120 588 bp，平均GC含量43.85%，编码基因4 045个。预测到13个次级代谢产物合成基因簇，编码surfactin、bacillaene、fengycin、bacillibactin、subtilosin和bacilysin等抑菌活性物质。转录组分析表明，施用YC25后，烟草中共诱导205个差异表达基因（DEGs），其中119个上调，86个下调。多个参与植物抗逆响应和免疫反应基因上调表达，包括FLS2、CaM、CYP84A、ERF1、ERF038、bHLH041、bHLH92和PR-1等。结论 YC25表现出抑制病原菌和诱导植物抗病性的综合能力，具有进一步开发为生物农药的应用潜力。

关键词: 斯氏芽胞杆菌, 抑菌能力, 基因组分析, 转录组分析

Abstract:

Objective The purpose of this study is to determine the taxonomic status of biocontrol strain YC25, evaluate its antibacterial ability, deeply analyze its genomic characteristics and potential of secondary metabolites, and elucidate its mechanism of induced disease resistance to tobacco (Nicotiana tabacum L.) plants, so as to provide theoretical basis and technical support for the development of efficient and environmentally friendly biocontrol agents for tobacco soil-borne diseases. Method The strain YC25 was identified by morphological, physiological, biochemical and molecular biological methods. The antibacterial ability was evaluated by confrontation test, and whole genome sequencing and secondary metabolite prediction were performed. The effect of YC25 treatment on gene expression in tobacco plants was analyzed by transcriptomics. Result The strain YC25 was identified as Bacillus spizizenii and showed broad-spectrum antagonistic activity against 9 plant pathogenic fungi. Field experiments showed that the combination of strain YC25 and metalaxyl effectively controlled tobacco root rot and reduced the amount of metalaxyl. The whole genome sequencing showed that the total length of the genome was 4 120 588 bp, the average GC content was 43.85%, and the coding genes were 4 045. A total of 13 secondary metabolite synthesis gene clusters were predicted, encoding antimicrobial active substances such as Surfactin, Bacillaene, Fengycin, Bacillibactin, Subtilosin and Bacilysin. Transcriptome analysis showed that a total of 205 differentially expressed genes (DEGs) were induced in tobacco after YC25 application, of which 119 were up-regulated and 86 were down-regulated. Several genes involved in plant stress response and immune response were up-regulated, including FLS2, CaM, CYP84A, ERF1, ERF038, bHLH041, bHLH92 and PR-1. Conclusion YC25 demonstrates the comprehensive ability to inhibit pathogens and induce plant resistance to disease, and thus has the potential to be further developed as a biological pesticide.

Key words: Bacillus spizizenii, antimicrobial activity, genome analysis, transcriptome analysis

危潇, 黎妍妍, 袁勤峰, 姚经武, 曹春霞, 黄大野. 一株分离自烟草根际的生防菌株鉴定及其生防潜力研究[J]. 生物技术通报, 2026, 42(7): 279-291.

WEI Xiao, LI Yan-yan, YUAN Qin-feng, YAO Jing-wu, CAO Chun-xia, HUANG Da-ye. Identification of a Biocontrol Strain Isolated from Tobacco Rhizosphere and Its Biocontrol Potential Analysis[J]. Biotechnology Bulletin, 2026, 42(7): 279-291.

图/表 12

参考文献 46

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基因名称 Gene name	引物名称 Primer name	序列 Sequence（5'-3'）
GAPDH	N. tabacumGAPDH F	ATGAGAGAGTGCATATCGAT
GAPDH	N. tabacumGAPDH R	TTCACTGAAGAAGGTGTTGAA
PR-1	N. tabacum PR-1 F	ATTGGAGAGGACAACGTCCC
PR-1	N. tabacum PR-1 R	ACACACCTCCATAATACCACC
PR-3	N. tabacum PR-3 F	ACAGATTCTGCCGGACACAC
PR-3	N. tabacum PR-3 R	AAGGAAGGTCCCAAAAGGGT
NPR-1	N. tabacumNPR1 F	GGATGCGGATGACTTGTCTGA
NPR-1	N. tabacumNPR1 R	GGAACAAGATGACGAGAGGTG
PAL	N. tabacumPAL F	AAGCATCACCCCGGACAAAT
PAL	N. tabacumPAL R	GGTGATGTTCTGAGGGCGTA
GST	N. tabacumGST F	CCTCACCCCCTCCCTCATAA
GST	N. tabacumGST R	TCGCCTTGATCTGTTTGTAGACT
SOD	N. tabacumSOD F	TAGCACCACACGCACAGAAG
SOD	N. tabacumSOD R	TGAAGGCTTCTCTTCTGCCT
CAT	N. tabacumCAT F	TGAAGGCTTCTCTTCTGCCT
CAT	N. tabacumCAT R	TGGAGGCCTTGTCTGATCCT

基因名称 Gene name	引物名称 Primer name	序列 Sequence（5'-3'）
GAPDH	N. tabacumGAPDH F	ATGAGAGAGTGCATATCGAT
GAPDH	N. tabacumGAPDH R	TTCACTGAAGAAGGTGTTGAA
PR-1	N. tabacum PR-1 F	ATTGGAGAGGACAACGTCCC
PR-1	N. tabacum PR-1 R	ACACACCTCCATAATACCACC
PR-3	N. tabacum PR-3 F	ACAGATTCTGCCGGACACAC
PR-3	N. tabacum PR-3 R	AAGGAAGGTCCCAAAAGGGT
NPR-1	N. tabacumNPR1 F	GGATGCGGATGACTTGTCTGA
NPR-1	N. tabacumNPR1 R	GGAACAAGATGACGAGAGGTG
PAL	N. tabacumPAL F	AAGCATCACCCCGGACAAAT
PAL	N. tabacumPAL R	GGTGATGTTCTGAGGGCGTA
GST	N. tabacumGST F	CCTCACCCCCTCCCTCATAA
GST	N. tabacumGST R	TCGCCTTGATCTGTTTGTAGACT
SOD	N. tabacumSOD F	TAGCACCACACGCACAGAAG
SOD	N. tabacumSOD R	TGAAGGCTTCTCTTCTGCCT
CAT	N. tabacumCAT F	TGAAGGCTTCTCTTCTGCCT
CAT	N. tabacumCAT R	TGGAGGCCTTGTCTGATCCT

生理生化指标 Physiological and biochemical index	结果 Result	生理生化指标 Physiological and biochemical index	结果 Result
淀粉水解 Starch hydrolysis	+	L-苹果酸	+
接触酶	+	好氧生长	+
氧化酶	+	D-乳糖	-
蔗糖Sucrose	+	D-岩藻糖	-
D-甘露醇	+	L-鼠李糖	-
柠檬酸	+	pH 5.0	+
D-山梨糖醇	+	8% NaCl	+

生理生化指标 Physiological and biochemical index	结果 Result	生理生化指标 Physiological and biochemical index	结果 Result
淀粉水解 Starch hydrolysis	+	L-苹果酸	+
接触酶	+	好氧生长	+
氧化酶	+	D-乳糖	-
蔗糖Sucrose	+	D-岩藻糖	-
D-甘露醇	+	L-鼠李糖	-
柠檬酸	+	pH 5.0	+
D-山梨糖醇	+	8% NaCl	+

编号 Number	病原菌名称 Name of pathogens	抑制率 Inhibition rate （%）
A	尖孢镰孢菌 Fusarium oxysporum	67.45±1.47d
B	核盘菌 Sclerotinia sclerotiorum	72.55±1.47bcd
C	灰葡萄孢菌 Botrytis cinerea	83.92±2.93a
D	烟草炭疽菌 Colletotrichum micotianae	69.02±2.42cd
E	茶拟盘多毛孢菌 Pestalotiopsis theae	74.90±2.00bc
F	烟草疫霉菌 Phytophthora nicotianae	70.20±0.55bcd
G	互隔链格孢菌 Alternaria alternata	76.08±1.47b
H	立枯丝核菌 Rhizoctonia solani	74.90±1.47bc
I	多主棒孢菌 Corynespora cassiicola	72.94±0.96bcd