Location: Agricultural Water Efficiency and Salinity Research Unit
Project Number: 2036-13210-013-012-R
Project Type: Reimbursable Cooperative Agreement
Start Date: Oct 1, 2023
End Date: Jul 31, 2026
Objective:
1. Comparative metagenomic investigation of microbial communities in the rhizosphere and endosphere of native plants -Ceanothus velutinus (Snowbrush), Cercocarpus ledifolius var. intricatus (Mountain Mahogany), and halotolerant bacteria from salt lake.
2. Isolation of the bacteria from the rhizosphere and endosphere, purification, and characterization of bacterial isolates for plant growth promoting activities.
3. Testing plant growth-promoting ability of these bacterial isolates under normal, drought, and salt stress on Arabidopsis thaliana and Medicago truncatula.
4. Identification of the plant growth-promoting bacteria by whole genome sequencing.
5. Validation of identified plant growth promoting bacteria in the greenhouse on maize, wheat, onion, watermelon and alfalfa under normal, drought, and salt stress.
6. Validation of bacteria tested positive in step (v) in field conditions, and releasing the plant growth promoting microbes as biofertilizers.
Approach:
The proposed project will investigate the rhizosphere and endosphere microbiome of the native plants of the Intermountain West Region of the United States; the study will then isolate, purify, characterize these microbes, and test them for plant growth promotion under abiotic stresses on grain crops, vegetables, and forage crop alfalfa. The final goal is to develop biofertilizers/ biostimulants to promote the growth and development of various crops under drought and salt stress.
i) Sample collection for bacterial isolation:
Rhizosphere soil and roots samples from Ceanothus velutinus (snowbrush ceanothus) will be collected from the Tony Grove region and Cercocarpus ledifolius var. intricatus will be collected from the wind caves, both in Logan, Utah. Halotolerant bacteria will be collected from the salt lake by University of Utah. Microbial DNA will be isolated from the rhizosphere soil and roots, the 16S ribosomal ribonucleic acid (rRNA) gene will be amplified and the samples will sequenced on the MiSeq using a 2×250 paired-end sequencing chip size (Illumina). The sequenced data will be analyzed using various packages available in R ver 4.0 (Team, 2021).
(ii) Isolation of the bacteria from the rhizosphere and endosphere:
Serial dilutions of the collected samples will be spread plated onto the five media compositions. The plates will be incubated at 28 degrees C for 3-5 days. Once growth appears, single colonies will be selected based on their different visual characteristics such as color, texture, transparency, size, consistency, and any other distinct morphological trait. The isolated bacterial colonies will be amplified for the whole length of the 16S rRNA and sequenced.
iii) Testing plant growth-promoting ability of these bacterial isolates under normal, drought, and salt stress on Arabidopsis thaliana and Medicago truncatula.
Drought stress will be imposed slowly by withdrawing water for nine days after 2nd bacterial treatment. The survival rate and biomass will be measured after one week of rewatering. Salt stress will be created by treating plants with 90 mm of sodium chlroide (NaCl) after 2nd bacterial treatment Survival rate and biomass will be measured at the end of the experiment. The plant growth-promoting bacteria will be characterized by whole genome sequencing.
iv) Validate identified plant growth promoting bacteria in the greenhouse on maize, wheat, onion, watermelon and alfalfa under normal, drought, and salt stress.
Identified PGPB containing will be screened on maize, wheat, onion, watermelon, and alfalfa under normal, drought, and salt stress in the greenhouse by bacterial seed coating and the pouring bacterial suspension after one week of germination. The bacterial colonization in the rhizosphere and roots will be monitored after regular intervals by collecting samples, serial dilution, and observing under a fluorescent microscope.
(v) Validation of selected bacteria in field conditions, and releasing the plant growth promoting microbes as biofertilizers.
The shortlisted PGPB from step (v) will be tested in fields condition for plant growth and development with and without abiotic stress.