100 years of plant breeding - what have we done to the seed microbiome?

人类驯化植物已有几千年的历史，公元前9000年左右，谷物首次在中东新月沃土被驯化。植物传统上被认为是一个单一的实体，基于表型的选择导致植物基因组的修饰，最近，对基因组的理解被用来加速关键表型的选择。然而，随着下一代测序技术的出现，人们对植物微生物组及其宏基因组在植物生长中的作用有了更大的认识。种子生物库对于保护由于栖息地丧失和环境变化而可能丧失的野生植物多样性至关重要，但我们对长期储存对微生物组的影响一无所知。我们将验证驯化、育种和储存改变植物微生物组组成的假设，从而影响植物在产量、恢复力和质量方面的性能。种子微生物组是本研究的理想目标，因为它代表了植物招募的群落，并可能适应内生生存。我们将分析具有不同历史和农业用途的3种作物的细菌微生物组：1)燕麦（燕麦）：在公元前第一个千年期间作为人类食物的粮食作物而驯化。2)黑麦草（Lolium perenne）：一种用于家畜（肉和奶）生产的草料，已经培育了大约100年。芒草：一种高大的草，最近从野外被选中作为生物能源和工业产品的原料，以及营养食品（如益生元和甜味剂），代表了一种正在进行的新型驯化。非洲大学的种子生物库收藏了这三种物种的种子，包括野生亲缘种和育种系，分别可以追溯到大约40年前、100年前和15年前。我们将使用这个资源：1。A.测量收集（Y1）中不同年龄的三个物种的种子萌发和植物适合度，并研究它们与细菌组成（通过快速和具有成本效益的16S rDNA分析）和收集时提供的种子气候数据（Y1/2）的关系。B.比较燕麦和黑麦各代的微生物组（Y1），并将存在的群体与这些品系的广泛性状数据相关联，包括谷物和饲料质量（Y1/2）。C.对于芒草，i)将野生采集种子的微生物组组成与采集来源进行比较，ii)将这些与最近产生的杂交种进行比较，以确定种群跨代保守的程度（Y1/2）。2. 将最近一批种子置于不同条件下模拟长期储存（Y1/2），并确定这些处理在最佳条件和非生物胁迫（Y2/3/4）下对种子萌发、微生物组组成以及随后植株性能的影响。3. 从不同年龄和来源的燕麦、黑麦草和芒草中分离细菌内生菌（Y1），在最优条件和非生物胁迫下进行植物生长促进实验，鉴定有益菌株（Y2/3/4）。我们将比较历史上驯化的粮食作物、最近培育的饲料作物和第一次进行选择的野生物种的微生物组。我们将把收集地点的气候和其他环境数据与种子微生物组联系起来，并通过几代人进行追踪。最终，我们将确定选择和育种是否无意中导致了植物微生物组多样性的减少，从而降低了植物的潜在益处，或者它们是否导致了优化的宏基因组。这将对微生物组对植物适应性的影响提供新的理解，对未来的种子储存和育种策略以及提高英国农业对未来气候的可持续性和恢复力具有重要意义。

Humans have domesticated plants for millennia, with cereals first domesticated in the Fertile Crescent of the Middle East around 9000 BCE. The plant has traditionally been considered as a single entity, with selection based on phenotype resulting in modification of the plant genome, and more latterly, understanding of the genome being used to accelerate selection of key phenotypes. However, with the advent of next generation sequencing technology, there has been greater appreciation of the role of the plant microbiome and its metagenome in plant performance. Seed biobanks are critically important for preserving plant diversity which may be lost in the wild due to habitat loss and the changing environment, but we know nothing about the effect of long-term storage on the microbiome. We will test the hypothesis that domestication, breeding and storage alter the composition of the plant microbiome, with impacts on plant performance in terms of yield, resilience and quality. The seed microbiome is the ideal target for this study as this represents the community recruited by the plant and potentially adapted to endophytic existence. We will analyse the bacterial microbiomes of 3 crops with contrasting histories and agricultural uses: 1) Avena sativa (oats): domesticated as a grain crop for human food during the first millennium BC. 2) Lolium perenne: a forage grass for livestock (meat and milk) production that has been bred for ~100 years. 3) Miscanthus: a tall grass recently selected from the wild as a feedstock for bioenergy and industrial products, as well as nutraceuticals (e.g. prebiotics and sweeteners) and represents a novel domestication in progress. The seed biobank at AU holds collections, including wild relatives and breeding lines, of all three species, going back approximately 40 years, 100 years, and 15 years respectively. We will use this resource to: 1. A. Measure seed germination and plant fitness of the three species of different ages in the collection (Y1) and investigate their relationship to both the bacterial composition present (via rapid and cost-effective 16S rDNA profiling) and the climatic data provided for the seed at collection (Y1/2). B. Compare the microbiome of oats and Lolium through the generations of breeding lines (Y1), and correlate the populations present with the extensive trait data available for these lines, including grain and forage quality respectively (Y1/2). C. For Miscanthus, i) compare microbiome composition of wild collected seed with origin of collection and ii) compare these with recently generated hybrids to determine the extent to which the populations are conserved across generations (Y1/2). 2. Subject a recent batch of seed to various conditions to simulate long-term storage (Y1/2), and determine the effect of these treatments on seed germination, microbiome composition, and on subsequent plant performance under optimal conditions and abiotic stresses (Y2/3/4). 3. Isolate bacterial endophytes from oats, Lolium and Miscanthus of different ages and origins (Y1) and conduct plant growth promotion experiments, under optimal conditions and abiotic stresses, to identify beneficial strains (Y2/3/4). We will compare the microbiomes of a historically domesticated grain crop, a more-recently bred forage crop, and a wild species undergoing selection for the first time. We will correlate climatic and other environmental data from the collection site with the seed microbiome, and trace this through generations. Ultimately we will determine whether selection and breeding have inadvertently resulted in reduced diversity within the plant microbiome with reductions to the potential plant benefits, or whether they have resulted in an optimised metagenome. This will provide novel understanding about the impact of the microbiome on plant fitness, with implications for future seed storage and breeding strategies, and for improving sustainability and resilience of UK agriculture to future climates.