宁杞7号枸杞根腐病发生的微生物学机制

doi:10.13560/j.cnki.biotech.bull.1985.2022-0363

摘要/Abstract

摘要：

从根表微生物群落结构及病原菌两个角度探究宁杞7号枸杞根腐病发生的微生物学机制。基于高通量测序分析微生物群落组成及多样性；采用组织分离法获取腐根真菌，结合形态学与分子生物学特征鉴定真菌的分类，依据柯赫法则进行致病性验证。患病株根表真菌群落结构已严重失衡，不仅丰富度和多样性显著低于健康株，而且优势类群的变化很大。患病株根表的优势属为镰刀菌属和粘帚霉属，相对丰度分别为38.81%和31.24%；而健康株根表的优势属为被孢霉属（14.09%）和镰刀菌属（10.38%），患病株根表潜在的病原菌丰度显著高于健康株。从腐根样品中共分离到33株真菌，分属于镰刀菌属、青霉属、俄氏孔菌属和粘帚霉属4个属，其中分离频率最高的是镰刀菌属，达到 75.76%。镰刀菌属的2个种：腐皮镰刀菌（Fusarium solani）和尖孢镰刀菌（F. oxysporum）均可致宁杞7号枸杞患根腐病。本研究明确了宁杞7号枸杞根腐病的发生不仅与病原菌关系密切，而且与根表微生物的群落结构密切相关。

关键词: 枸杞, 根腐病, 病原菌, 鉴定, 微生物群落结构

Abstract:

The purpose of this study is to explore the microbiological mechanism of the root rot of Lycium barbarum Ningqi-7 based on the analysis of microbial composition on rhizoplane and pathogenic microbes. High-throughput sequencing was used to analyze the microbial community composition and diversity. The root rot fungi were isolated with the tissue separation method. The combination of morphology and molecular characteristics was used to identify the fungi. The pathogenicity was identified according to Koch’s rule. The structure of fungal community on the rhizoplane of the diseased plant had been seriously imbalanced，with the richness and diversity being significantly lower than those of the healthy plants. The dominant groups of rhizoplane microorganisms were also quite different between the root rot diseased plants and the healthy plants. The dominant genera on the rhizoplane of the diseased plants were Fusarium and Clonostachys，with relative abundance being 38.81% and 31.24%，respectively；while the dominant genera on the rhizoplane of the healthy plants were Mortierella（14.09%）and Fusarium（10.38%）. The abundance of potential pathogens on the rhizoplane of the diseased plants was significantly higher than that of the healthy plants. Total 33 fungi were isolated from the root rot of the diseased plants，and they were classified into four genera：Fusarium，Penicillium，Earliella，and Clonostachys，among which，the highest frequency of isolation was Fusarium，reaching 75.76%. Two species of Fusarium，F. solani and F. oxysporum，caused the root rot of Lycium barbarum Ningqi-7. This study clarified that the occurrence of root rot of L. barbarum Ningqi-7 was not only closely related to pathogenic fungi，but also related to the community structure of rhizoplane microorganisms.

Key words: Lycium barbarum, root rot, pathogenic microbe, identification, microbial community structure

高惠惠, 贾晨波, 韩琴, 苏建宇, 徐春燕. 宁杞7号枸杞根腐病发生的微生物学机制[J]. 生物技术通报, 2022, 38(12): 244-251.

GAO Hui-hui, JIA Chen-bo, HAN Qin, SU Jian-yu, XU Chun-yan. Microbiological Mechanism of Root Rot of Lycium barbarum Ningqi-7[J]. Biotechnology Bulletin, 2022, 38(12): 244-251.

图/表 8

图1 健康株与患病株根表真菌的OTU韦恩图

Fig. 1 Venn diagram of rhizoplane fungi in the healthy and diseased plants based on OTU analysis

表1 根表真菌的Alpha多样性

Table 1 Alpha diversity of rhizoplane fungi of the healthy and diseased plants

样品Sample	Sobs	Simpson	Shannon	ACE	Coverage
患病株Diseased	195.00±35.12b	0.32±0.18a	1.88±0.60a	340.94±39.89b	99.9%
健康株Healthy	461.67±14.10a	0.11±0.01a	3.17±0.12a	572.73±10.77a	99.8%

图2 健康株和患病株门水平（A）和属水平（B）上根表真菌的组成与相对丰度

Fig. 2 Composition and relative abundance of rhizoplane fungi at the phylum（A）and genus（B）levels of healthy and diseased plants

图3 代表性菌株的菌落形态及显微形态

Fig. 3 Colony morphologies and microscopic morphologies of representative strains

图4 基于ITS基因序列构建的系统发育树

Fig. 4 Phylogenetic tree based on ITS gene sequences

图5 基于TEF基因序列构建的镰刀菌的系统发育树

Fig. 5 Phylogenetic tree of Fusarium based on TEF gene sequences

图6 接种3种菌株25 d后枸杞根腐病的发病症状

Fig. 6 Symptoms of L. barbarum root rot 25 d after inocul-ation with 3 strains

图7 接种3种菌株25 d后发病枸杞的腐根真菌再分离

Fig. 7 Re-isolation of root-rot fungi from L. barbarum 25 d after inoculation with 3 strains

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