云南草莓属植物的微生物组生物地理特征研究

The research project is aiming to reveal the ecological basic issues and biogeologic features of Fragaria spp. plant microbiomes/phytomicrobiomes (a.k.a. microbiota) in the different climatic and geographical regions of Yunnan (seven regions: north tropical climate region, subtropical climate region, south temperate climate region and north temperate climate region of Yunnan, respectively), i.e. the correlationships between the spatio-temporal dynamics of Fragaria spp. phytomicrobiomes and those of biotic and abiotic factors in their habitats, and biogeologic features of phytomicrobiomes in Yunnan. The project is designed: to carry out researches on the spatio-temporal dynamics of Fragaria spp. plant microbiomes in two seasons (Summer and winter) by applying culture-independent methods of molecular microbial ecology, including high throughput sequencing methods of the next generation sequencing platforms (Illumina MiSeq System or Roche 454 Pyrosequencing platform) (with universal prokaryotes-specific and fungi-specific primer sets or barcodes of the ribosomal small subunit-SSU/internal transcribed spacer region-ITS region/large subunit-LSU rRNA genes of prokaryotes and fungi, respectively, in Fragaria spp. plant microbiomes/phytomicrobiomes), as well as those of microbial community structures and community functions by using fluorescent dyes, such as Calcofluor White and DAPI, or FISH, or SEM/TEM and Biolog EcoPlate, respectively; meanwhile, to determine and analyize the spatio-temporal dynamics of biotic and abiotic factors of the habitats (climate factors, geographic information, vegetations, physical and chemical parameters of soils and plant samples); furthermore, to reveal the correlationships between the spatio-temporal dynamics of Fragaria spp. phytomicrobiomes and those of biotic and abiotic factors of the habitats by using comprehensive analysis of ecological factors, i.e., to settle the ecological fundamental issues of the Fragaria spp. plant holobionts (microecosystems); finally, to plot out the biogeologic features of Fragaria spp. phytomicrobiomes in Yunnan, based on the research results of this study and the biogeographic informations of the Fragaria spp. habitats. The project is expected both to meet the needs and to be the theoretical basis of further scientific researches, conservation and sustainable exploitation and utilization of microbial resources of Fragaria spp. and their phytomicrobiomes such as plant probiotics (as biofertilizers or biocontrol agents), as well as for ecological cultivation and sustainable agriculture of strawberry cultivars in Yunnan.

在前期引种云南草莓属野生植物种质资源(Fragaria spp.;4-6个种)及栽培种(F. ananassa)的基础上，针对其植物微生物组(phytomicrobime)的生物地理特征(biogeographic features)，提出计划：拟采用宏基因组扩增子高通量测序技术，研究云南不同地理与气候区域草莓属各野生种及栽培种根际、内际(endosphere)、茎叶际(phyllosphere)的内生菌(endophytes)与附生菌(epiphytes)，以及生境非根际土壤(bulk soil)的微生物组时空动态；同时分析其生境的生物与非生物因子时空动态(如地理与气候因子等)；通过生态因子综合分析，揭示各植物种微生物组与其生境的生物与非生物因子时空动态的相关性(生态学基本问题)，并揭示其生物地理特征；从而为资源(如植物益生菌,plant probiotics)的可持续利用奠定理论基础。

以往对草莓属植物微生物的研究，绝大多数都是采用经典纯培养分析方法，且只针对草莓植物某个器官或部分的微生物开展研究，而针对草莓属植物所有器官及其微生物组的所有微生物类群，以及考虑和分析包括草莓属植物地理分布与气候因素在内影响微生物组群落动态的生物与非生物因子的系统研究，显得十分有限与不足，有待深入、系统开展相关工作。为此，开展本研究计划：首先对云南野生草莓属植物种质资源的分布进行调查和物种分布预测，采集云南野生草莓属植物种质资源并建立种质资源苗圃，构建野生草莓属植物DNA条形码，以及分析其遗传多样性在不同地理区域、不同气候区和植物区系单元的地理分布格局；其次，采用宏基因组高通量扩增子测序技术，对不同地理区域和不同气候区云南野生草莓六个不同部位（非根际、根际、根部表面、根部内生、茎叶表面和茎叶内生）相关微生物群落的多样性、组成与结构进行研究，并结合相关的生物与非生物因子进行分析，其结果揭示了草莓属植物微生物组（原核生物和真菌）特征及其与环境因子的相关性；此外，阐述了草莓属植物微生物组地理分布特征，分析了形成地理分布格局的驱动因子，并探讨了草莓属植物遗传多样性地理分布格局与植物微生物组地理分布格局之间的关系；同时，分析了不同野生草莓种微生物组的差异，以及野生草莓与栽培草莓植物微生物组的异同，从而为野生草莓种质资源及其微生物组菌种资源的研究、保护、开发与可持续利用奠基理论基础。