桃杏果实内生细菌多样性分析及软腐病原菌的分离与验证

为了解桃和杏采后果实内生细菌群落组成,并对潜在病原细菌进行筛选与验证,为其贮藏保鲜、软腐病害防治等相关研究奠定基础,利用高通量测序技术,对新疆地产油桃和库车小白杏采后果实内生细菌群落组成进行分析；同时,利用微生物传统分离培养,采用16S rDNA序列分析对潜在病原细菌进行分子鉴定,并利用离体接种法和再分离法,对相关菌株的致病能力进行验证。研究结果表明,油桃和库车小白杏内生细菌共包括128个操作分类单元,涉及9个门117个属,其中变形杆菌门(Proteobacteria)为绝对优势菌门,其次为厚壁菌门(Firmicutes)和拟杆菌门(Bacteroidetes)；在属水平上,油桃以泛菌属(Pantoea)、芽孢杆菌属(Bacillus)、假单胞菌属(Pseudomonas)、拟杆菌属(Bacteroides)等菌属为优势菌群；库车小白杏以葡糖杆菌属(Gluconobacter)、泛菌属(Pantoea)、克里斯滕森菌(Christensenella)、普氏菌属(Prevotella)等菌属为优势菌群。通过分离筛选获得了1株潜在采后致腐病原细菌XAAS-P1,经分子鉴定初步确定其归属于泛菌属(Pantoea)。XAAS-P1对采后油桃和库车小白杏均具有较强的致腐能力,并呈现出与自然软腐一致的病状。接种4d后,XAAS-P1致腐率可达100%,与阴性对照组相比,接种组样品的软腐现象提前了1d。对接种组病灶中的微生物进行再分离、鉴定,结果显示,菌株XAAS-P1是病灶组织中的绝对优势菌群,证明了菌株XAAS-P1是导致采后油桃和库车小白杏软腐变质的主要病原细菌。研究表明,桃和杏果实中存在丰富多样的内生细菌,其内生泛菌XAAS-P1可导致采后油桃和库车小白杏的软腐变质。

Diversity of Endophytic Bacteria in Nectarine and Apricot and Isolation, Identification of Soft Rot Pathogen

In order to reveal the main composition of endophytic bacterial communities in nectarine and Kuqa apricot, and screen and verify the potential pathogenic bacteria, then provide the foundation for the research of storage, freshness, and prevention of soft rot disease, the endophytic bacterial communities in postharvest nectarine and Kuqa apricot in Xinjiang were analyzed by high-throughput sequencing. The pathogenic bacteria were screened by traditional isolation and culture methods, then identified based on 16S rDNA sequences analysis. The pathogenicity of strains was tested and identified by in vitro inoculation method and re-isolation method. The results showed that endophytic bacterial communities in nectarine and Kuqa apricot contained 128 operational taxonomic units (OTUs), involving 117 genera of 9 phylums. Proteobacteria was absolute dominant phyla followed by Firmicutes and Bacteroides. At the genus level, Pantoea, Bacillus, Pseudomonas and Bacteroides were the dominant microflora in nectarine, while Gluconobacter, Pantoea, Christensenella and Prevotella were the dominant population in Kuqa apricot. A potential pathogenic bacteria named XAAS-P1 related to soft rot disease was obtained, which was classified in Pantoea by phylogenetic tree construction of 16S rDNA sequence. It exerted the ability to cause the postharvest decay of nectarine and Kuqa apricot, which had consistent symptoms with natural soft rot. After 4 days inoculation, the incidence of decay was 100%, and the decay appeared 1 days earlier compared with the negative control group. Results of re-isolation and identification showed that strain XAAS-P1 was the absolutely dominant bacteria in lesion, which proved that strain XAAS-P1 was the pathogenic bacteria caused the postharvest decay of nectarine and Kuqa apricot. The study indicated that diversity of endophytic bacteria in nectarine and Kuqa apricot was rich, and the endophytic bacterium, Pantoea sp. XAAS-P1 could lead to the postharvest decay of nectarine and Kuqa apricot.