The role of seed microbiota in germination

Microorganisms contribute to plant growth and a plants seed offers the possibility to design a plant that starts with an optimized microbial community. However, currently we do not have sufficient knowledge of how to achieve this, or even how the microbial community contributes to seed germination. The plant microbial community includes bacterial endophytes that live within plants, including seeds. To understand more about seed-microbe interactions, an emerging and poorly explored research topic, a collaboration between seed physiologists from the University or Innsbruck and molecular microbiologists from the Austrian Institute of Technology has been established to test the following hypotheses: 1) Metabolic changes in the seed during germination, rather than re- hydration alone, activate proliferation of bacterial endophytes; 2) Seed microbiota contribute to seed vigour and seedling establishment; 3) The diversity of the seed microbiome becomes imbalanced during seed ageing, increasing seedling infection, which can be ameliorated by inoculation with endophytes antagonistic to pathogens 4) Cultivar-dependent effects of germination-promoting microbiota are due to a lack in representation of functional endophytes in the natural seed microbial community. To test these hypotheses we will use soybean (Glycine max L.) because of its economic importance, the available literature, sequenced genome and our own research experience. Community profiling of the microbiota will be conducted by identification of their DNA in seed extracts. The influence of seed endophytes on seed germination will be studied after removal of the pre-existing seed microbial community and re-inoculation of seeds. The influence of altering the microbial community in terms of seed vigour will be tested under a range of environmental conditions. Furthermore, germination-influencing endophytes will be visually tagged with fluorescent proteins to allow their detection with microscopy at the seed tissue and cellular level. The changes in seed metabolites during germination and in response to ageing will be monitored by gas or liquid chromatographic techniques coupled to mass spectrometry (i.e., GC-MS and LC-MS, respectively). Overall, we hope to comprehend which specific endophytes proliferate in seeds, which metabolites they are responding too, how they affect seed germination, compete alongside soil-borne microbes and how these endophytes are affected during seed ageing. New knowledge on beneficial plant- microbe interactions may be applied to enhance agricultural productivity.

Bacteria are some of the smallest living organisms, much smaller than our own cells. However, what they do not have in size they can make up for in number! There is a growing awareness of the importance that the microbiome (the sum of the microbes, including bacteria and fungi) has for the health and fitness of much larger multicellular organisms, such as ourselves, but also for example plants. The research of this project investigate potential roles of native microbiota in various aspects of seed germination of Austrian-grown soybean (Glycine max L.). To achieve this, an interdisciplinary collaboration between seed scientists at the University of Innsbruck and microbiologists at the Austrian Institute of Technology (AIT) was established. By using amplicon sequencing we were able to identify the bacteria living within (so-called endopyhtes) and on (so-called epiphytes) of soybeans and how the community changed during seed germination. Our results showed that water uptake by the seed led to a redistribution of the epiphytes so that later during germination they became endophytes. Furthermore, the breakdown of energy reserves of the seed and related metabolism was essential for increase in abundance, so-called richness, of the microbiota, especially those that could grow rapidly and take advantage of the available nutrients. These results can be found free of charge (open access) in the journal Phytobiomes (https://doi.org/10.1094/PBIOMES-03-23-0019-MF). Moreover, our results have relevance to agriculture since it shows that an application of bacteria to seed coats, e.g. in pellets, would be a suitable way of enabling growth-promoting bacterial strains to be taken up during germination. We also investigated if native seed bacteria could have beneficial roles for seed germination. For this, over 200 bacterial isolates were isolated from soybean, leading to 47 unique taxa. We found that by mimicking the nutrient availability of the host (in this case using soy milk or ground seed as a growth media) we could isolate the highest bacterial diversity (https://doi.org/10.1128/spectrum.00172-22). We then tested the ability for these bacteria to influence the germination of low-vigour seeds which are susceptible to fungal growth. Interestingly, bacteria that promoted the germination of low vigour seed were also antagonistic to fungal growth. We further investigated the seed-bacteria relationship in an attempt to understand why these bacteria promoted seed germination. We hope to extend our research further by conducting field trials in a future academic-industry collaboration, as well as deepen our understanding of the specific relationships that plants can make with certain bacteria.