一株李维斯链霉菌的鉴定及其抗病促生性能

Characterization of Streptomyces levis L2 with plant disease-inhibiting and growth-promoting effects

摘要:

【目的】 探究一株李维斯链霉菌的抗病促生性能，鉴定其抑菌活性成分。【方法】 从耐旱稗草根际分离到一株能抑制多种病原真菌及细菌的链霉菌，采用形态学观察及16S rRNA基因序列分析的方法对菌株进行鉴定；运用液相色谱和高分辨质谱技术鉴定活性物质成分；利用二代Illumina与三代Nanopore测序技术进行全基因组测序；antiSMASH分析该菌株中活性成分的生物合成基因簇。【结果】 李维斯链霉菌L2的菌体及发酵液能高效抑制革兰氏阳性菌及多种植物病原真菌的生长，能在铬天青平板(chrome azurol sulphonate, CAS)上产生较大的噬铁圈，还具有较好的产吲哚乙酸(indole-3-acetic acid, IAA)的能力。其抑菌活性成分的m/z为537.102 0[M+H]⁺ (calcd for C₂₇H₂₀O₁₂, 537.103 5, 2.2×10⁻⁶)，523.086 3 [M+H]⁺ (calcd for C₂₆H₁₈O₁₂, 523.087 8, 2.9×10⁻⁶)，分别与α-鲁霉素、β-鲁霉素、γ-鲁霉素的精确分子量相符，误差小于1/5 000 000，其高效液相色谱保留时间与鲁霉素标准品一致。该菌基因组总长8.8 Mb，预测编码有鲁霉素等32个次级代谢产物生物合成基因簇。【结论】 李维斯链霉菌L2具有较好的抗病促生性能，适宜进一步开发为生防菌剂，其抑菌活性成分为鲁霉素。

Abstract:

[Objective] To study the plant disease-inhibiting and growth-promoting effects and identify the antimicrobial components of Streptomyces levis L2. [Methods] Morphological features and the phylogenetic tree based on the 16S rRNA gene sequences were employed to identify the strain L2 isolated from the rhizosphere of drought-tolerant Echinochloa crusgalli. The antimicrobial components produced by the strain were identified by chromatography and high-resolution mass spectrometry. The whole genome of the strain was sequenced by Illumina in combination with Nanopore. antiSMASH was employed to search for the biosynthetic gene clusters. [Results] Streptomyces levis L2 and its fermentation broth inhibited the growth of Gram-positive bacteria and phytopathogenic fungi, and the strain produced a large transparent zone on the CAS (chrome azurol sulphonate) plate and could produce indole-3-acetic acid (IAA). The antimicrobial components of the strain showed the m/z of 537.102 0 [M+H]⁺ (calcd for C₂₇H₂₀O₁₂, 537.103 5, 2.2×10^–6) and 523.086 3 [M+H]⁺ (calcd for C₂₆H₁₈O₁₂, 523.087 8, 2.9×10^–6), which were consistent with the exact masses of α-rubromycins, β-rubromycins and γ-rubromycins, with the errors less than parts per 5 million. Their HPLC retention time were in agreement with that of standard rubromycins. The whole genome of L2 had a length of 8.8 Mb and carried 32 biosynthetic gene clusters for secondary metabolites including rubromycins. [Conclusion] S. levis L2 exhibited excellent plant disease-inhibiting and growth-promoting properties, thus could be further developed as biocontrol agents. It produces the antimicrobial components rubromycins and its biosynthetic gene clusters contained 6 more modification genes than the previously published gene clusters for rubromycins and several genes with unknown functions.