Browsing by Author "Muleta Atsede"

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    Genomic Diversity, Plant Growth Promoting Properties and Symbiotic Effectiveness of Mesorhizobum Spp. Nodulating Chickpea (Cicer Arietinum L.) on Acidic Soils of Ethiopia: Implication for Inoculant Development to Enhance Production Under Low Inputs Agriculture
    (Addis Ababa University, 2020-07-07) Muleta Atsede; Assefa Fassil (PhD)
    Nitrogen and phosphorus deficiencies are the two most limiting factors in crop production. Although chickpea fixes nitrogen in association with diverse Mesorhizobium species, effectiveness in the amount of nitrogen fixed depends upon the inherent character and other factors of the endosymbionts. The nitrogen-fixing process requires more phosphrous and phosphorus deficiency aggravated under acidic soils because of sorption. Thus, selection of acid tolerant rhizobia with effective nitrogen fixation and phosphate solubilizing activity is essential in order to improve phosphorus and nitrogen fertility to enhance the growth and production of the crop in acidic soil. This study was initiated to collect chickpea rhizobia from acidic soils of central, western, southern and northern parts of Ethiopia and determine their genomic diversity, and further screen for low pH tolerance, solubilization of insoluble phosphate on tri-calcium phosphate medium and their potential to ecological adaptations and symbiotic effectiveness with host varieties on sand culture at low pH and acidic soil culture under greenhouse conditions. The genomic diversity was studied using illumina sequencing a whole-genome sequence (WGS) approach. The results showed that 81 Mesorhizobium strains were obtained and estimated with average genome sizes 7Mbp, average depth of 143 X coverage. Annotation of the assembled genome predicted an average of 6725 protein-coding genes (CDSs). Genome completeness showed that 81 strains are near complete (>99% complete). Concatenation of 400 universal phylophlan marker genes, present in all genomes (strains) allowed ii detailed phylogenetic analysis, from which eight well-supported phylogenetic groups were identified. Phylogenetic reconstruction based on the symbiosis-related (nod C and nif H) genes were generally different to those shown by the phylophlan marker genes. The pattern of genomic diversity indicates the existence of multiple, broadly distributed phylotypes, with no relationship between geographic and genetic distance (r=0.10, p<0.01). The result indicated that 62 strains grew well at low pH 5; and 47 (76%) of them were phosphate solubilizers. The strains showed wide diversity in their substrate utilization and tolerance to salinity, high temperatures, Mn 2+ and Al3+ toxicity, heavy metals and antibiotics. Symbiotic characterization also showed a wide diversity among these mesorhizobial strains. Moreover, most of the phosphate solublizing strains had multiple growth promoting characteristics (production of indole acetic acid, hydrogen cyanide, siderophore, ACC diaminase and inhibition of Fusarium oxysporum f.sp.ciceris growth) under in vitro conditions. Under field conditions, the mesorhizobial inoculants (a.117L2, a.71 and a.15star) showed significant (P<0.01) improvement in biological nitrogen fixation, growth, yield, nitrogen and phosphorus uptakes of chickpea compared to the commercially available local reference strain Cp41. The indigenous mesorhizobial strain a.117L2 was superior inoculant for almost all the tested parameters. Generally, the present study indicated that Ethiopian acidic soils contain symbiotically effective, phosphate solublizing, chickpea nodulating Mesorhizobium spp. endowed with different plant growth promoting attributes which are diverse in their genomic and taxonomic identities. Therefore, there is a potential advantage using these mesorhizobial inoculants to enhance chickpea production in acidic soils by improving phosphorus and nitrogen iii fertility; and provide that further field trials recommended over several seasons and sites in Ethiopia.
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    Isolation and Screening of Antibiotic Producing Actinomycetes from Soil
    (Addis Ababa University, 2011-10) Muleta Atsede; Assefa Fassil (PhD)
    Antibiotics are secondary metabolites produced by microorganisms. Actinomycetes represent the top producer of antibiotics that are widely distributed in the different types of soils. The study was designed to isolate and screen antibiotic producing actinomycetes on samples obtained from soil and rhizosphere of plants. A total of 30 actinomycetes were isolated from Debrezit, Holeta and Arat Kilo campus. Out of the 30 actinomycetes, 18 (60%) showed antimicrobial activities of one of the tested microorganisms of which 9 isolates were selected for their wide spectrum of antibiotic activities. In this study, isolate AAUBA5 and AAUBA30 were found to inhibit all of the test organism, except E.coli ATCC 25922 and the latter was found to be the most resistant that was only inhibited by 3/18 ( 17 %) of the isolates. The isolates were identified and characterized by cultural, morphological, physiological and biochemical characteristics and found to be classified under the genus Streptomycetes. In order to isolate the active compounds of the extract, fermentation was carried out on nine isolates and the former was extracted by using ethyl acetate. The antimicrobial activity of the active compound of the isolates was studied using Disc Diffusion Assay method and found to inhibit the test microorganisms that could not during the preliminary screening. Finally, isolates AAUBA13 was selected for further study based on spectrum activity and inhibition zone diameter in the antimicrobial bioassay. Antimicrobial compound obtained from AAUBA13 isolate was separated and purified with thin layer chromatography (TLC), column chromatography and preparative chromatography. The purified compound found to be active against gram positive bacteria (S.aureus clinical isolate, S.pneumoinae ATCC49619 and S.aureus ATCC25923), gram negative bacteria (E.coli ATCC25922, S.typhi ATCC 6539 Shigella boydii clinical isolates and P.aeruginosa ATCC2585) and effective against Candida albicans ATCC62376. The data, in general, showed that the antimicrobial compounds obtained from AAUBA13 demonstrate broad spectrum and a remarkable antimicrobial activity against bacterial and Candida albicans ATCC62376. It is also suggested that the other isolates should be further processed to fully realize their antibiotic property on different test microorganisms. Key words: Actinomycetes; Antibiotics; Antimicrobial bioassay; Antimicrobial compound; Disc Diffusion Assay