Legume-rhizobia interactions in a complex microbiome

Abstract

Biological nitrogen fixation is important for agriculture, carbon sequestration, and ecosystem restoration. This is primarily conducted by rhizobia (nitrogen fixing bacteria) in association with legume plants. Most research in improving rhizobial strains involves single strain experiments. However, improved metagenomics methods have demonstrated considerable differences between single strain inoculations and strain behaviour when exposed to a complex microbiome. To identify some these differences, this experiment applies two treatment factors in a controlled environment of containers with autoclaved sand. All main experimental containers were inoculated with several strains of Bradyrhizobia japonicum. The first treatment factor was the planting of surface-sterilised seedlings of host plant Acacia acuminata; the second treatment factor was inoculation with an external soil microbiome. Several negative controls without planting or inoculation were also present. A novel method of whole genome metagenomic sequencing to observe known strain abundance, without amplification or culturing, was developed. Using this method, abundance patterns of these B. japonicum strains were compared between initial inoculation and the end of a growth period of several weeks. Analysis reveals a single strain as the preferred nodulation strain within this experiment, but also shows that all strains inoculated continued to persist in the substrate at detectable levels. The use of long reads with the MinION DNA sequencer also allowed the potential of identification of horizontal gene transfer events. None were detected in an initial screen, but a framework for further inspection of this dataset for such events is described.