生防副地衣芽孢杆菌ZYGT1811的GFP标记及其在小麦植株和根际土的定殖动态

doi:10.16409/j.cnki.2095-039x.2025.02.069

摘要/Abstract

摘要： 为明确副地衣芽孢杆菌Bacillus paralicheniformis ZYGT1811对小麦茎基腐病的防治机制，本研究采用电击转化法，获得有绿色荧光的ZYGT1811-GFP标记菌株，通过测定生长曲线、拮抗活性、游动性以及生物膜形成能力，发现外源基因的导入对该原始菌株无明显影响，采用灌根法明确了菌株ZYGT1811-GFP在小麦植株和根际土的定殖状况。结果表明，灌根6 d后，ZYGT1811-GFP在小麦根际土、根、茎、叶中定殖量达到最大，分别为3.24× 10⁶、1.76× 10⁶、9.00× 10⁵、6.47× 10⁴ cfu/g，第21 d定殖量有所下降，但仍维持一定定殖量，各部位的菌群密度仍表现为根际土＞根＞茎＞叶。本研究证实菌株ZYGT1811具有稳定的定殖能力，可为揭示副地衣芽孢杆菌防治小麦茎基腐病的机制提供理论依据。

关键词: 小麦茎基腐病, GFP, 生物防治, 副地衣芽孢杆菌, 定殖

Abstract: This study employed electroporation to generate the ZYGT1811-GFP labeled strain exhibiting green fluorescence, aiming to elucidate regulatory mechanism of Bacillus paralicheniformis ZYGT1811 on wheat crown rot. Assessment of growth curve, antagonistic activity, motility, and biofilm formation revealed that the incorporation of foreign genes did not significantly impact the original strain. Colonization of strain ZYGT1811-GFP in wheat plants and rhizosphere soil was elucidated using root irrigation. The findings indicated that the colonization of ZYGT1811-GFP in the wheat rhizosphere soil, roots, stems, and leaves peaked at 6 days after root irrigation, with population densities 3.24× 10⁶, 1.76× 10⁶, 9.00× 10⁵, and 6.47× 10⁴ cfu/g, respectively. Colonization diminished on the 21st day, although persisted at a certain level, with population density ranking as follows: rhizosphere soil>root>stem>leave. The above results indicate that ZYGT1811 possesses sustainable colonization capability, which offers a theoretical foundation for elucidating the control mechanism of Bacillus paralicheniformis against wheat crown rot.

Key words: wheat crown rot, green fluorescent protein (GFP), biocontrol, Bacillus paralicheniformis, colonization

中图分类号:

S476

王艺璇, 柳敬龙, 李惠霞, 费芍丹, 孙佳聪, 任兴平, 田晓明, 刘永刚, 张海英. 生防副地衣芽孢杆菌ZYGT1811的GFP标记及其在小麦植株和根际土的定殖动态[J]. 中国生物防治学报, 2025, 41(6): 1403-1411.

WANG Yixuan, LIU Jinglong, LI Huixia, FEI Shaodan, SUN Jiacong, REN Xingping, TIAN Xiaoming, LIU Yonggang, ZHANG Haiying. GFP labeling of Bacillus paralicheniformis ZYGT1811 and Its Colonization Dynamics in Wheat Plants and Rhizosphere Soil[J]. Chinese Journal of Biological Control, 2025, 41(6): 1403-1411.

参考文献

[1] Li H, Yuan H, Fu B, et al. First report of Fusarium pseudograminearum causing crown rot of wheat in Henan, China[J]. Plant Disease, 2012, 96(7): 1065-1065.
[2] 郭炜, 李雪萍, 漆永红, 等. 西北地区冬小麦茎基腐病原假禾谷镰刀菌的鉴定及种质资源抗性筛选[J]. 甘肃农业大学学报, 2018, 53(6): 164-170.
[3] 徐飞, 宋玉立, 周益林, 等. 2013-2016年河南省小麦茎基腐病的发生危害情况及特点[J]. 植物保护, 2016, 42(6): 126-132.
[4] 马璐璐, 闫翠梅, 冯彩莲, 等. 玉米秸秆还田对假禾谷镰刀菌及小麦茎基腐病化感效应的模拟研究[J]. 河北农业大学学报, 2019, 42(3): 38-44.
[5] Kazan K, Gardiner D M. Fusarium crown rot caused by Fusarium pseudograminearum in cereal crops: recent progress and future prospects[J]. Molecular Plant Pathology, 2018, 19(7): 1547-1562.
[6] 杨云, 贺小伦, 胡艳峰, 等. 黄淮麦区主推小麦品种对假禾谷镰刀菌所致茎基腐病的抗性[J]. 麦类作物学报, 2015, 35(3): 339-345.
[7] Chen Y, Wang J, Yang N, et al. Wheat microbiome bacteria can reduce virulence of a plant pathogenic fungus by altering histone acetylation[J]. Nature Communications, 2018, 9(1): 3429.
[8] 王超, 郭坚华, 席运官, 等. 拮抗细菌在植物病害生物防治中应用的研究进展[J]. 江苏农业科学, 2017, 45(18): 1-6.
[9] 胡江涛, 刘奇, 周英, 等. 芽孢杆菌hjt6的鉴定、生长特性及其对瓠瓜灰霉病菌的拮抗效果[J]. 微生物学通报, 2024, 51(3): 832-845.
[10] Shen L, Xu J Q, Fu L M, et al. Biocontrol of wheat crown rot using Bacillus halotolerans QTH8[J]. Pathogens, 2022, 11(5): 595-595.
[11] Xu W, Yang Q, Xie X, et al. Genomic and phenotypic insights into the potential of Bacillus subtilis YB-15 isolated from rhizosphere to biocontrol against crown rot and promote growth of wheat[J]. Biology, 2022, 11(5): 778.
[12] Dong Q, Liu Q, Goodwin P H, et al. Isolation and genome-based characterization of biocontrol potential of Bacillus siamensis YB-1631 against wheat crown rot caused by Fusarium pseudograminearum[J]. Journal of Fungi, 2023, 9(5): 547.
[13] 刘青祥, 徐文, 董迁迁, 等. 特基拉芽胞杆菌YB-1145的分离鉴定及其对小麦茎基腐病的生防效果评价[J]. 中国生物防治学报, 2024, 40(4): 884-893.
[14] 李健, 何鹏飞, 李咏梅, 等. 枯草芽孢杆菌L1-21在柑橘上的定殖能力及其防治绿霉病效果[J]. 华中农业大学学报, 2021, 40(5): 31-36.
[15] 董爱菊, 邱慧珍, 董莉, 等. 类芽孢杆菌QHZ11-gfp在马铃薯植株上的定殖特征及促生效果[J]. 微生物学报, 2021, 48(11): 4075-4086.
[16] 陈楠楠, 秦平伟, 尹珺伊, 等. 解淀粉芽孢杆菌抗菌机制研究进展[J]. 中国微生态学杂志, 2018, 30(12): 1464-1469.
[17] 薛丽香, 童坦君, 张宗玉. 报告基因的选择及其研究趋向[J]. 生理科学进展, 2002, 33(4): 364- 366.
[18] 汪恒英, 周守标, 常志州, 等. 绿色荧光蛋白(GFP)研究进展[J]. 生物技术, 2004, 14(3): 70-72.
[19] 张小蕊, 王睿, 李宇诗, 等. 解淀粉芽孢杆菌FS6绿色荧光蛋白标记菌株的获得及其生物学特性研究[J]. 吉林农业大学学报, 2025, 47(3): 437-444.
[20] 李蓉, 洪赫, 姜文筱, 等. 贝莱斯芽胞杆菌LQ-3防治小麦纹枯病的效果及根际定殖能力[J]. 麦类作物学报, 2024, 44(7): 947-954.
[21] Zhou L, Song C X, Muñoz C Y, et al. Bacillus cabrialesii BH5 protects tomato plants against Botrytis cinerea by production of specific antifungal compounds[J]. Frontiers in Microbiology, 2021, 12: 707609.
[22] 刘忠伟, 孙振才, 杨佑明, 等. 小麦苗期浸种和根灌浅黄隐球酵母菌的定殖特征及对茎基腐病的防治效果[J]. 中国农业大学学报, 2021, 26(2): 24-31.
[23] 杨洪凤, 余向阳, 薛雅蓉, 等. 内生解淀粉芽孢杆菌CC09在小麦根部定殖的电镜观察及防病效果[J]. 中国生物防治学报, 2014, 30(6): 839-844.
[24] 汪盼盼, 杨野, 刘迪秋, 等. 宏基因组学在植物病害研究中的应用[J]. 生物技术通报, 2020, 36(12): 146-154.
[25] Tokman M, Lee H J, Levine A Z, et al. Electric field-driven water dipoles: nanoscale architecture of electroporation[J]. PLoS ONE, 2017, 8(4): e61111.
[26] Aune T E V, Aachmann F L. Methodologies to increase the transformation efficiencies and the range of bacteria that can be transformed[J]. Applied Microbiology and Biotechnology, 2010, 85: 1301- 1313.
[27] 高竞, 方伟, 顾佳悦, 等. 荧光标记解淀粉芽孢杆菌WK 1在山核桃树体和土壤中的定殖规律[J]. 浙江农业学报, 2021, 33(1): 77-86.
[28] 何朋杰, 崔文艳, 何鹏飞, 等. 表面活性素促进枯草芽胞杆菌XF-1在大白菜叶际定殖能力研究[J]. 植物保护, 2021, 47(5): 28-34.
[29] 齐永志, 赵斌, 李海燕, 等. 多功能菌B1514在小麦根际的定殖及对纹枯病的防治作用[J]. 植物保护学报, 2014, 41(3): 320-326.
[30] Lugtenberg B J, Dekkers L, Bloemberg. Molecular determinants of rhizosphere colonization by Pseudomonas[J]. Annual Review of Phytopathology, 2001, 39: 461-490.
[31] Sasse J, Martinoia E, Northen T. Feed Your Friends: Do plant exudates shape the root microbiome?[J]. Trends in Plant Science, 2018, 23(1): 25-41.
[32] 李全芬, 张婷婷, 马磊. 羽毛针禾根部内生假单胞菌XG11在小麦上的定殖动态及促生效应[J]. 江苏农业科学, 2022, 50(3): 219-224.
[33] Knight N L, Macdonald B, Sutherland M W. Colonization of durum wheaT (Triticum turgidum L. var. durum) culms exhibitinG premature senescence (dead headS) associated with Fusarium pseudograminearum crown roT[J]. Plant Disease, 2017, 101(10): 1788-1794.