Patil Basavaprabhu L, Gopalkrishna Amulya M, G M Sandeep Kumar, R Umamaheswari
ICAR-Indian Institute of Horticultural Research, Hessaraghatta Lake Post, Bengaluru 560089, Karnataka, India.
J Appl Microbiol. 2025 Jan 6;136(1). doi: 10.1093/jambio/lxaf010.
Bacillus subtilis is usually found in soil, and their biocontrol and plant growth-promoting capabilities are being explored more recently than ever. However, knowledge about metabolite production and genome composition of endophytic B. subtilis from seeds is limited. In the present study, B. subtilis EVCu15 strain isolated from the seeds of Vasconcellea cundinamarcensis (mountain papaya) was subjected to whole genome sequencing and detailed molecular and functional characterization.
Whole genome sequencing and sequence analysis of the endophytic bacterium from mountain papaya seed revealed that the bacterium was B. subtilis, strain EVCu15. The genomic sequence had more than 98% nucleotide similarity with two published whole genome sequences of B. subtilis strains. Some of the important secondary metabolite gene clusters involved in production of bioactive compounds such as surfactin, fengycin, plipastatin, bacillibactin, bacillaene, subtilomycin, subtilosin A, and bacilysin were identified from the whole genome sequence analysis. Genes encoding several plant growth-promoting metabolites, mostly involved in the nutrient metabolism, were identified in the bacterial genome. These included factors coding for nitrogen, phosphorus, iron, sulfur, potassium, and trehalose metabolism. Genes involved in auxin, riboflavin, acetoin biosynthesis, ACC deaminase activity, and xylan degradation were also identified. Proteomic analysis confirmed the biosynthesis and release of several bioactive secondary metabolites in the endophytic B. subtilis strain EVCu15. Liquid chromatography-mass spectrometry-based profiling for hormones and vitamins identified extracellular secretion of several important plant growth-promoting compounds such as IAA, salicylic acid, zeatin, vitamin D1, D2, E, K1, and pyridoxine. The in vitro and in vivo studies with the endophytic B. subtilis against various plant pathogenic fungi showed moderate to high levels of resistance. The B. subtilis EVCu15 compared to B. amyloliquefaciens showed better control over the root-knot nematode Meloidogyne incognita, in terms of egg hatching inhibition and the mortality of J2 juveniles.
Overall, this study underscores the biocontrol and plant growth-promoting potential of B. subtilis EVCu15, an endophyte isolated from mountain papaya seeds. Genomic analysis revealed a significant proportion of genes linked to biocontrol and plant growth promotion, corroborating its efficacy against M. incognita and various plant pathogens in vitro and in greenhouse studies. Furthermore, the bacterium's ability to produce diverse bioactive compounds, including proteins, hormones, and vitamins, was confirmed, highlighting its complex interactions within the plant system.
枯草芽孢杆菌通常存在于土壤中，其生物防治和促进植物生长的能力近来得到了前所未有的深入研究。然而，关于种子内生枯草芽孢杆菌的代谢产物产生和基因组组成的知识却很有限。在本研究中，对从番木瓜（山地番木瓜）种子中分离出的枯草芽孢杆菌EVCu15菌株进行了全基因组测序以及详细的分子和功能表征。
对来自山地番木瓜种子的内生细菌进行全基因组测序和序列分析，结果表明该细菌为枯草芽孢杆菌EVCu15菌株。该基因组序列与已发表的两株枯草芽孢杆菌全基因组序列具有超过98%的核苷酸相似性。从全基因组序列分析中鉴定出了一些参与生物活性化合物如表面活性素、丰原素、脂肽素、杆菌铁载体、芽孢杆菌烯、枯草菌素、枯草杆菌素A和杆菌溶素产生的重要次生代谢物基因簇。在细菌基因组中鉴定出了几个编码促进植物生长的代谢产物的基因，这些基因大多参与营养代谢。其中包括编码氮、磷、铁、硫、钾和海藻糖代谢的因子。还鉴定出了参与生长素、核黄素、3-羟基丁酮生物合成、ACC脱氨酶活性和木聚糖降解的基因。蛋白质组学分析证实了内生枯草芽孢杆菌EVCu15菌株中几种生物活性次生代谢物的生物合成和释放。基于液相色谱-质谱联用的激素和维生素分析确定了几种重要的促进植物生长的化合物如吲哚-3-乙酸、水杨酸、玉米素、维生素D1、D2、E、K1和吡哆醇的细胞外分泌。对内生枯草芽孢杆菌针对各种植物病原真菌的体外和体内研究表明其具有中度到高度抗性。与解淀粉芽孢杆菌相比，枯草芽孢杆菌EVCu15在抑制卵孵化和二龄幼虫死亡率方面对南方根结线虫表现出更好的防治效果。
总体而言，本研究强调了从山地番木瓜种子中分离出的内生枯草芽孢杆菌EVCu15的生物防治和促进植物生长的潜力。基因组分析揭示了很大比例的与生物防治和促进植物生长相关的基因，证实了其在体外和温室研究中对南方根结线虫和各种植物病原体的防治效果。此外，该细菌产生多种生物活性化合物（包括蛋白质、激素和维生素）的能力也得到了证实，突出了其在植物系统内的复杂相互作用。
