ITS鉴定真菌病原:一例司法鉴定研究实例

本研究选择一例疑似空气污染物伤害松树的司法鉴定案作为研究实例,在现场勘察和初步形态鉴定的基础上,应用ITS序列分析鉴定了对疑似空气污染物伤害松树的病原做了分子鉴定。我们对委托鉴定的疑似空气污染物伤害松树的地及周边地区进行了勘查,现场勘查结果并不支持空气污染物伤害松树的看法。为明确松树枯死的原因,对取样病枝上的疑似病原物进行了显微镜观察和培养性状观察,根据显微镜下孢子的形态和病原菌的培养性状,初步判断导致马尾松枯死的病原可能为拟盘多毛孢属(Pestalotipsis)真菌。为进一步确定病原种类,我们应用分子生物学技术对危害马尾松的病原物进行了ITS分子鉴定。测定了疑似真菌的ITS序列,并进行了比对和系统发育分析。研究结果表明,导致松树枯死的病原菌系韦司梅拟盘多毛孢(Pestalotipsis vismiae)。在本研究案例中,利用真菌ITS序列的系统发育分析,快速、准确地鉴定了真正导致松树枯死的病原,为今后类似司法鉴定案件审理提供了证据鉴定的新手段。

Identification of Fungus Pathogen by ITS Sequencing:A Judicial Case Study

In this study we chose one judicial case of suspected injury to the pine by air pollutants as a case study,we made an on-site investigation and preliminary morphological identification prior to molecular identifying the pathogen that tentatively caused harm to the pine by air pollutants.The results seemed to be unsupported the charge of the injured pines by air pollutants based on the on-site survey in the pine field and the surrounding areas.In order to figure out reasons for the dead pine trees,we sampled the tentative disease pathogens from the branches of the pines to be observed under the microscope as well as to be incubated for observation of cultured characters.Based on the microscopic morphology of spores and cultured characters of the pathogens,we did preliminary determine the curse leading to dead pines that might be the Pestalotiopsis fungus.To further confirm the pathogenic species,we employed the techniques of molecular biology,the ITS molecular identification,to determine the pathogen by sequencing,aligning and phylogenetic analyzing.The results showed that the pathogen resulting in dead pines should be the fungus Pestalotiopsis vismiae.In this case study,we conducted the phylogenetic analysis of fungal ITS sequences to have rapidly and accurately identified the actual pathogen leading to dead pine trees,which would be the case to provide a new means for identification of forensic evidence in future.