Optimizing the sporulation of endophytes Actinobacteria on broth and agar media and their effect on growth and nodulation of chickpea

Author: Sita Ram Sapkota

Sapkota, Sita Ram, 2020 Optimizing the sporulation of endophytes Actinobacteria on broth and agar media and their effect on growth and nodulation of chickpea, Flinders University, College of Medicine and Public Health

Terms of Use: This electronic version is (or will be) made publicly available by Flinders University in accordance with its open access policy for student theses. Copyright in this thesis remains with the author. You may use this material for uses permitted under the Copyright Act 1968. If you are the owner of any included third party copyright material and/or you believe that any material has been made available without permission of the copyright owner please contact copyright@flinders.edu.au with the details.


Chickpea is currently the second most important pulse crop produced in the world. It is a rich source of protein for human consumption and is cheaper compared to animal protein. Chickpea is a type of legume that can fix the atmospheric nitrogen via symbiosis with the rhizobial partner, Mesorhizobium. Recent research in our laboratory has found that the legume grain yield increased when the rhizobium inoculant was added together with selected endophytic actinobacteria in chickpea plants. The actinobacterium acts by colonizing plant roots and increases the nitrogen fixation capacity of the rhizobial partner. Some strains have the added benefit of controlling the losses caused by fungal root pathogens. The actinobacterial inoculants are added as spores to the seed before sowing. However, spores normally grow on solid surfaces, but they are also able to be formed in liquid submerged culture. In this study, endophytic actinobacterial strains CP21A2, CP56, CP84B, and CP200B isolated from chickpea were evaluated for the sporulation rate in solid and liquid media. These resultant spores were evaluated for their stability at different pH and temperature. The spores and rhizobia were then inoculated in chickpea seeds in order to determine whether the spores produced on agar and in liquid culture had a similar symbiotic relationship on the growth and development of chickpea plants.

Calcium carbonate in the liquid broth and MS medium in solid agar media can be used to increase the sporulation rate in the actinobacteria. Additionally, we found out that almost all spore strains were stable at 70 °C but temperatures greater than that were lethal to the spores obtained from both types of media. In addition, the tested spore strains were more sensitive and prone to lysis at alkaline pH rather than acidic. Furthermore, our study suggested that CP56 spores obtained from liquid media and CP84B from solid media were the best performers in promoting the overall growth of plants and nodules. However, further detailed investigations need to be carried out in order to determine their influence on the growth and development of the chickpea plants, which can be useful to increase the yield in the agricultural industry.

Keywords: Optimization, liquid, and solid media, Chickpea plant, stability, Endophytic actinobacteria

Subject: Medical Biotechnology thesis

Thesis type: Masters
Completed: 2020
School: College of Medicine and Public Health
Supervisor: Prof. Christopher Franco