Genetic manipulation of Rhizobium for tolerance to heat, acid and salt stress in soils impacted by climatic variability

Abstract

Use of Rhizobium inoculants is a proven technology to increase legume crop yields, improve soil fertility through symbiotic biological nitrogen fixation (BNF). In Sub-Saharan Africa, BNF is challenged by increasing temperatures due to climatic variability. One possible solution is use of genetically modified heat tolerant strains since use of field selected tolerant strains has been unsuccessful. This research aims to understand genetic changes associated with heat stress in Rhizobium in order to enhance stress tolerance through genetic manipulation. Specific objectives are 1) to characterize selected soybean Rhizobium strains by 16S rRNA sequencing and identification of differentially expressed genes from heat tolerant strains; 2) to construct plasmids for introduction and expression of the identified genes into a susceptible Rhizobium. 3) to evaluate stress tolerance of transformed Rhizobium and impact of expressing these genes in Rhizobium on improvement of symbiotic growth parameters in soybeans.
Characterization identified USDA193 and HH103 as heat tolerant and susceptible strains respectively while 22 Malawi isolates as non-soybean Rhizobium. Proteomic study identified 3628 proteins, 8 differentially expressed at 20oC and 40oC by four-fold were selected, only hsp20, shsp, IbpA and 3 uncharacterised proteins (UC1, UC2 and UC3) were amplified from USDA193 for construction of pLMB51plasmids for HH103 transformation. All transformants show phenotypic gene expression in normal liquid media. Under stress, the empty plasmid control, UC1, hsp20, UC2, and shsp transformants were acid and salt tolerant while hsp20, UC2 and shsp were heat tolerant. Non-stressed plants (20oC) showed no differences in nodule number (NN) and root dry weight (RDM), but nodule score (NS) except hsp20 and shoot dry mass (SDM) (except UC1 and UC2) were lower than the wild type. Heat stressed plants (32oC) show no differences in NN, NS, SDM and RDM but visual observations on plant vigor predict that hsp20, UC2, and shsp can maintain plant growth.

Date of Award	23 Jan 2020
Original language	English
Awarding Institution	University of Bristol
Supervisor	Gary D Foster (Supervisor) & Andy M Bailey (Supervisor)