Document Type

Dissertation - Open Access

Award Date

2024

Degree Name

Doctor of Philosophy (PhD)

Department / School

Agronomy, Horticulture, and Plant Science

First Advisor

Senthil Subramanian

Abstract

Nitrogen is a crucial nutrient for the growth and development of plants. Legume crops like soybeans develop symbiotic associations with soil bacteria from the genus, Bradyrhizobium to obtain fixed nitrogen. In this mutualistic association, rhizobia are released into root nodules that provide a compatible environment for rhizobia to grow and fix nitrogen (Symbiotic Nitrogen Fixation (SNF)). A caveat in such beneficial associations is the competition between different rhizobium strains in soil for infection and nodule occupancy. The elite inoculant strains encounter competition by indigenous rhizobial population in the soil to infect and nodulate a susceptible legume host, because of which they often fail to infect and occupy root nodules, affecting the overall nitrogen fixation efficiency in legume hosts. In response to this, host plants might employ sanction mechanisms to favor nodulation by more beneficial rhizobium strains over less beneficial ones. The goal of this study was to evaluate the host plant responses towards rhizobia competition for nodulation at the phenotypic and molecular level using several rhizobium strain combinations with varying nitrogen fixation capacities/efficiencies. For this purpose, we utilized a split-root system approach, which allows studying plant responses by simultaneously inoculating separate root halves of the same host plant with different strains. Our initial experiments with high-capacity nitrogen fixing and non-fixing B.diazoefficiens strains (USDA 110 and USDA 126 respectively) at endpoint revealed that host plants can favor USDA 110 while imposing sanctions on USDA 126 during nodulation. Here we performed parallel inoculations of these strains at three-time points to determine when the sanctions are initiated by the host. To further determine the gene expression patterns in these root halves over time, we conducted a time-course transcriptomic analysis and identified differential expression of several nodulation, phytohormone, and clock-related genes/pathways. Subsequently, we conducted timecourse experiments using combinations of intermediate-capacity fixer (USDA 140) which indicated that soybeans may not show preference for nodulation between high-capacity strain, USDA 110, and 140 or with USDA 140 and the non-fixing strain, USDA 126. Instead, we observed differences only in the nodule bacteroid density from USDA 126 and 140 combinations which suggested that host sanctions might be implemented against the non-fixer post nodulation. Additionally, we evaluated host responses with parallel inoculations of two Bradyrhizobium elkanii strains, one with a high nitrogen-fixation capacity (USDA 83) and the other with a low nitrogen-fixing capacity (USDA 26). Results from this study showed no difference in overall nodule numbers between the strains at 21 days post-inoculation. We further tested host responses to different Bradyrhizobium species (diazoefficiens vs. elkanii strains) wherein we observed that soybeans sanctioned USDA 126 when in competition with USDA 26 at the nodulation level. However, with USDA 110 vs. USDA 26 combination, we neither observed differences in nodule numbers nor bacteroid densities, suggesting that the host plants do not select against the lowcapacity strain, USDA 26. Previously, transcriptomic analysis of USDA 110 and 126 root halves at endpoint suggested that the Autoregulation of Nodulation (AON) pathway might be involved in such sanctioning. Here, we validated this using nod 1-3 mutant plants defective in AON pathway. Overall, our study provides evidence for potential host sanctioning mechanisms that soybeans might employ against poor/low-capacity nonnitrogen fixers at different levels and stages depending on the strain combinations.

Publisher

South Dakota State University

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Rights Statement

In Copyright