河南黄河湿地放线菌多样性及植物病害生防放线菌的筛选

Diversity of actinobacteria in Henan Yellow River wetland and screening of cultivable actinobacteria with biological control on plant disease

摘要:

[目的]探究河南黄河湿地放线菌多样性，筛选对植物病原菌有拮抗活性的放线菌菌株。[方法]基于Illumina HiSeq技术的高通量测序分析了河南黄河湿地放线菌物种多样性及其分布特点；利用8种分离培养基对采集自三门峡黄河湿地、郑州黄河湿地和开封黄河湿地21份土壤样品中的放线菌进行了分离纯化，通过16S rRNA基因序列分析对分离株进行了初步的分类鉴定和系统学研究；以7种植物病原菌为靶标筛选有拮抗活性的分离株，并对活性菌株进行了聚酮合酶、非核糖体多肽合成酶和安莎类化合物等基因筛查。[结果]高通量测序结果表明，河南黄河湿地放线菌物种多样性丰富，且不同湿地区域以及同一湿地不同生境间放线菌多样性存在显著差异，放线菌优势属物种依次为Nocardioides、Streptomyces、CL500-29 marine group、Fodinicola、Mycobacterium和Micromonospora，此外，还具有大量的未知类群；通过分离培养共获得了261株纯培养菌株，分布于放线菌门中的7个目9个科9个属，包含97个可能的已知物种和8个潜在新物种，对植物病原菌有拮抗活性的放线菌86株，其中74株中至少含有一种活性代谢物质合成相关基因。[结论]河南黄河湿地放线菌物种多样性丰富，具有发掘新物种和新型植物病害生防资源的潜力。

Abstract:

[Objective] This study is aimed to analyze the diversity of actinobacteria in soils of the Henan Yellow River wetland and screen actinobacteria strains which have antagonistic activity against plant pathogens. [Methods] High-throughput sequencing based on Illumina HiSeq technology was used to analyze the actinobacterial diversity and distribution in the Henan Yellow River wetland. We used eight selective media to isolate actinobacteria from 21 soil samples collected from Sanmenxia, Zhengzhou and Kaifeng Yellow River wetland. The cultivable actinobacteria were identified and phylogeny analyzed based on the 16S rRNA gene sequences. The active isolates were screened using seven plant pathogens as targets, and 5 functional genes were assayed, including polyketide synthases gene (PKS-I, PKS-Ⅱ), nonribosomal peptide synthetases gene (NRPS), 3-amino-5-hydroxybenzoic acid synthases gene (AHBA) and 3-hydroxy-3-methylglutaryl coenzyme A gene (HMG CoA). [Results] The high-throughput sequencing results indicated that the actinobacteria were abundant in Henan Yellow River wetland and there were significant differences among different wetland areas and different habitats in the same wetland. Nocardioides was the most frequently isolated genus, followed by the genera Streptomyces, CL500-29 marine group, Fodinicola, Mycobacterium and Micromonospora. In addition, there are still a lot of unknown groups. We isolated a total of 261 dereplicated strains, belonging to 7 orders, 9 families and 9 genera of the phylum Actinobacteria. They were including 97 putative species and eight unknown species. Among them, 86 actinobacteria strains have antagonistic activity against plant pathogens; 74 strains contain at least one gene which involved active metabolites synthesis. [Conclusion] Actinobacteria species diversity was abundant in Henan Yellow River wetland and new species have potential as biocontrol agents against plant diseases.