Browsing by Author "Assefa Fassil (Professor)"

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    Diversity, Symbiotic Effectiveness and Plant Growth Promoting Characteristics of Rhizobia and Rhizospheric Bacteria on Growth and Production of White Lupin (Lupinus Albus L.) Under Greenhouse and Field Conditions In North Western Ethiopia
    (Addis Ababa University, 2018-04-03) Sintie Esubalew; Assefa Fassil (Professor)
    White Lupin (Lupinus albus L.) is a traditional legume crop in Ethiopia that can play complementary or alternative roles as sources of organic fertilizers to replenish nitrogen depleted soils in farming systems as it forms symbiotic associations with soil bacteria that have atmospheric nitrogen fixation and other plant growth promoting abilities. This study was aimed to investigate the diversity and symbiotic effectiveness of rhizobia and rhizobacteria on growth and production of white lupin (Lupinus albus L.) from major producing areas of the country. A total of 100 bacterial isolates were obtained from the root nodules of White lupin and characterized based on their phenotypic, biochemical and plant growth promoting characteristics using various standard methods. Accordingly, 39 isolates were confirmed as rhizobia on the basis of presumptive and authentication tests. These rhizobial isolates showed wide diversity in their symbiotic and cultural characteristics as well as heterotrophy, such as C and N-substrates utilization, tolerance to metal toxicity, antibiotics, pH, salt and temperature. Moreover, some of the isolates showed interesting PGP traits (IAA, siderophore and HCN production, P solubilization and antagonistic activity against Fusarium oxysporum) which can make them prior candidates for the production of inoculants for enhancing White lupin production in the country. Furthermore, the total of 136 phosphate solubilizing bacterial isolates were obtained from rhizospheric soils of White lupin producing area of the country and 40 effective phosphate solubilizing bacterial isolates were selected on the basis of their solubilization index (SI). The phosphate solubilizing bacterial isolates showed variations in their cultural characteristics, doubling times, Gram reactions and C and Nsources utilization patterns. Quantitative estimation of phosphate solubilizing efficiency of the phosphate solubilizing rhizospheric bacterial isolates on different inorganic phosphate sources showed that some of the isolates have better solubilization potential of tricalcium, aluminum and iron phosphates in Pikosvikya broth upon 5 days of incubation. In addition, they showed good ii tricalcium phosphate solubilization potential under different stress conditions, such as various salt concentrations, pH and temperature ranges and most of them showed multiple plant growth promoting characteristics. Furthermore, three rhizobial and two phosphate solubilizing rhizospheric bacterial isolates that showed better relative symbiotic effectiveness, plant growth promoting features, stress tolerance and heterotrophic competence were selected to study the effect of single and co-inoculation on the growth and nodulation of the White lupin under greenhouse and field conditions. Accordingly, co-inoculated white lupin plants resulted in higher nodule number, nodule and shoot dry matter accumulation, percent nitrogen and improved relative symbiotic effectiveness than single rhizobial inoculation, and N-fertilized plants on sand and soil cultures under greenhouse conditions. Similarly, inoculation of White lupin plants either with the selected rhizobial isolates alone or co-inoculation of rhizobial and phosphate solubilizing rhizospheric bacterial isolates showed improved performance on the nodulation, growth and yield of the crop compared to the negative control under field conditions. In general, the present study showed that Ethiopian soils contain symbiotically effective White lupin rhizobia and efficient phosphate solubilizing rhizospheric bacteria with multiple plant growth promoting traits to enhance growth and production of the crop under natural conditions. However, repeated tests should be carried out on different field conditions in order to use these rhizobial and phosphate solubilizing bacterial isolates for inoculant production.
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    Evaluation of Rhizobacteria and Trichoderma Isolates From the Rhizosphere of Faba Bean (Vicia Faba L.) for their Potential In the Management of Chocolate Spot (Botrytis Fabae Sard.) Disease and Host Plant Growth-Promoting Properties
    (Addis Ababa University, 2021-02-02) Firdu Zewdineh; Assefa Fassil (Professor); Alemu Tesfaye (PhD)
    Faba bean (Vicia faba L.) is one of the legume grains used as a source of food for human. However, the productivity of the crop is less due to chocolate spot (Botrytis fabae Sard) in Ethiopia. The disease can be managed through the applications of fungicides such as MORE 720 WP and ORZEB 80 WP. Alternatively, an eco-friendly approach is initiated using Trichoderma and bacterial species to manage the disease. Therefore, in this study, Trichoderma and bacterial species were evaluated against Botrytis fabae under in vitro and in vivo conditions. The data were analyzed by SPSS version 24. The bacterial strains inhibited B. fabae (6-68%) upon 3-9 days in the dual culture method. The highest inhibition was recorded by B. tequilensis AAUB100 that displayed 80% and Serratia nematodiphila AAUB146b with 79% inhibition at 20% (v/v) of the culture filtrate. A 200-760 μg mL-1 of solubilized phosphate was also released upon 3-6 days by the isolates. Trichoderma harzianum AAUT14 was the most effective antagonist with more than 88% of B. fabae inhibition in the dual culture upon 9 days. The isolates of Trichoderma also showed inorganic phosphate solubilization that ranged from 135-509 μg mL-1 upon 3-9 days. Under the greenhouse study conducted using a completely randomized design, the disease incidence reduced by 36-50% on Ashebeka variety compared to the control (T1) in which T. harzianum AAUT14 and B. subtilis AAUB95 (T4) displayed the highest reduction (50%) of disease incidence upon 60 days. The disease severity reduction was much pronounced upon 80 days ranging from 41-55% compared to the disease severity, which did not show significant difference (p>0.05) between T4 and T5 (MORE 720 WP) upon 60 days. A 3.8-34% of disease incidence and 15-26 disease severity reduction was also observed upon 60 days on Hachalu variety. An area under disease progress curve (AUDPC) of 405.97% and 415.33% was displayed by T5 and T6 (ORZEB 80 WP), respectively on Ashebeka variety. Similarly, 377.98% by T5 and 412.48% by T6 of AUDPC was illustrated in Hachalu variety showing no significance difference (p>0.05) with T4. Under field conditions conducted using a completely randomized block design, the combination of Trichoderma harzianum AAUT14+B.subtilis AAUB95 (T4) reduced the disease incidence and severity showing no significance difference (p>0.05) with T5 upon 70 and 90 days. Nevertheless, in Hachalu variety (Trial 2), the treatment reduced the disease incidence from 23-51% and disease severity 37-54% upon 90 days. In addition, the AUDPC ranged from 1586.1-2250.0% in trial 1 and 1382.0-2454.5% in trial 2. The grain yield was also increased by 34% and 38% in trial 1 and 2, respectively over the controls by T4. In conclusion, the mixture of T. harzianum AAUT14 and B. subtilis AAUB95 or in some cases T. harzianum AAUT14 performed best in controlling chocolate spot and growth promotion of faba bean under greenhouse and field conditions.
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    Pilot Scale Windrow Composting of Coffee Husks and Flower Residues: Biochemical and Microbiological Determination of the Composts
    (Addis Ababa Universty, 2015-04) Shemekite Fekadu; Assefa Fassil (Professor)
    Two hundred forty thousand (240,000) tons of coffee waste and about 4000 tons of flower residue wastes are generated annually from the age-old coffee processing industries and the booming cut flower industries in Ethiopia, respectively. Most of these wastes are released to the environment without treatment, and their direct release pollutes the environment, mainly inhibiting plant growth. This necessitates the transformation of these wastes into beneficial products with a dual purpose of production of bio fertilizer and protection of the environment. To this end, a study was made to evaluate the process of composting of coffee husks and flower residues separately in order to determine their compost quality. Each material was arranged in to three different piles in a trapezoidal Windrow composting for 90 days and more, and monitored for their physico-chemical, microbiological and phyto-toxicological properties during the whole process. Coffee husk (CH) amended with cow dung plus house hold compost (CHCD: Pile 1) and with fruit/vegetable wastes plus house hold compost (CHFVW: Pile 2) were arranged and compared against the control (CH: Pile 3) without any amendment. As a result, the two amended treatments showed similar pattern of temperature profile with the first mesophilic stage (18-42oC) in the first 2 days followed by the thermophilic stage (45- 70oC) up to 45 days, and declined to the second mesophilic stage afterwards and remained so until the end of the experiment. In all cases, there was a steady decline of moisture content (MC %), reduction in total organic carbon (TOC%) and a rise in total Nitrogen (TN%) because of the loss of carbon in the form of CO2 thereby decreasing the C:N ratio of Pile 1, Pile 2, and Pile 3 to 11, 13, and 18, respectively. The highest bacterial count of 9.78logMPN g-1dw (Pile1) and 9.45logMPN g-1 dw (Pile 2) was seen at the start of the process; while the highest actinobacterial counts of 9.78logMPNdw (Pile 1 and Pile 2) and fungal counts of 9.78log MPN g-1 dw (Pile 1) and 9.45 MPN g-1 dw (Pile 3) were recorded at the end of the composting process, respectively. This change in number indicated a clear microbial succession along the process. The effect of the different microbial activities was reflected in the better maturation of Pile 1 with a Germination Index value >80% and C/N ratio of 11 at the end of composting compared to the control that required much more days for the material to become free from phytotoxicity. This transformation could be associated with higher volatile solid reduction (75% in Pile 1 and 65% in Pile 2), substantiating the importance of chemical parameters in coffee husk compost. The same feed stock was also composted with cow dung (Pile 1), with fruit/vegetable wastes (Pile 2) and coffee husk alone (Pile 3) again to study their microbial diversity and enzyme activities; and samples were collected on days 0, 32 and 90. Similar changes in TOC (%) and in TN(%) were found resulting in lower C/N ratios of 11 and 13 for Pile 1 and Pile 2, respectively due to changes in temperature profile, pH and water content of the mixture. This change which directed the succession of different microbial groups was accompanied by the release of compost relevant enzymes: hydrolases, phosphatases, peptidases and proteases in Pile 1 and Pile 2 at the start; and esterase at the end of the process. Furthermore, denaturing gradient gel electrophoresis (DGGE) analysis of coffee husk composting indicated distinctive community shifts during the composting process. The DGGE revealed that bacterial and fungal communities of samples from day 0 were clustered separately from communities of samples from day 32 and 90, indicating a change in the bacterial and fungal community composition. This result attributed that microbial communities at the start were responsible in the degradation of labile organic substrates while those communities at the end involved in the stabilization of the process. Young compost of Pile 1 and 2 were clustered separately from Pile 3 as the two cosubstrates introduced different and diverse microbial communities into the composting process. On the contrary, microbial community in the matured compost was grouped together showing the end compost were homogenous with well-defined microbial communities. Principal Component Analysis (PCA) of compost communities from day 0 and day 90 composts analyzed by COMPOCHIP microarray explained 62.5% of the variations. As a result, probes KO443 and 444 (Stenotrophomonas maltophilia), KO609, KO610 and KO614 (Brevundimonas/Caulobacter), KO500 (Derxia gummosa) KO612, 615, 616 and 617 (Flavobacterium/Flexibacter), KO541 (Pseudomonas putida), KO252 (Acinetobacter) and KO342 (Actinomyces sp.) were found to be more influential in discriminating the samples into different composting phases. Besides, Brevundimonas, Caulobacter, Chryseobacterium Sphingobacterium were dominant during the first mesophilic phase in Piles 1 and 2. These species have broader degradation activity of complex biopolymers. On the other hand, Flavobacteria/Flexibacter was detected significantly in all samples, suggesting their importance in the composting process. In general, microbial diversity and numbers were lower in the mature composts as indicated by DELTA 495a, Alpha proteobacteria and Low G+C, Xylella/Xanthomonas/Stenotrophomonas (KO241), Azotobacter beijerinckii (KO277) and the Actinomyces (KO342). Especially, the presence of Actinomycetes was an indicator of mature compost. In flower residue, the major feed stock (flower residue) was blended with cow dung (Pile 1), with activated EM (Effective Microorganisms) and molasses (Pile 2) and compared against the control (Pile 3) without any amendment for the same period of composting and the same composting system. Changes in NH4-N, NO3-N, NH4:NO3 ratio, available P and K and concentration of micronutrients (Cu, Zn, Fe, Mn) were included in the chemical analysis in addition to TOC%, TN% and C/N ratio to better understand the efficiency of chemical parameters in maturity indices. Consequently, the reduction in TOC (%) and the steady increase in TN(%) resulted in 10%, 16% and 24% of C/N ratios at the end of the process for Pile 1, Pile 2 and Pile 3, respectively. The use of inorganic N forms (NH4-N, NO3-N and NH4-N to NO3-N ratios) as indicators of maturity showed significant differences among the three piles. While NH4-N reduced below 400 mg/kg in all piles which was the maximum threshold level for matured compost, NO3-N showed an increasing trend, resulting in lower NH4-N to NO3- N ratio (0.12) of Pile 1, compared to Pile 2 and Pile 3 which could not reduce below 0.38 and 0.58, respectively. The analysis of available P and K also showed an increasing trend in all piles, but the concentration of available P (0.7-1%) at the end of the process was sufficient for soil nutrient supplementation. On the contrary, available K and all micronutrients were below the sufficiency level for plant production. In general, the effect of the different microbial activities was reflected in the better maturation of Pile 1, showing a low C/N ratio (10-11) and high GI% (80-85%) at the end of the composting; and significantly correlated with temperature, water content, pH over the 90 days of composting of both coffee husk and flower residue composts. Comparatively, Pile 2 and control of both the composting processes required more days for maturity in this respect. In conclusion, a polyphasic approach using physico-chemical and biological parameters including temperature, C/N ratio, GI%, NH4:NO3 ratio, enzyme assay together with molecular tools are very essential to evaluate maturity and stability of coffee husk and flower residue composts. Bulking agents that have much amendment potential like cow dung must be encouraged in large scale composting industries to tackle the current environmental challenges; and further studies on composting of various sources of solid organic wastes with a different composting system can offer greater insight on the quality, safety and age of composting. The nutritional and economic feasibility of these compost products must be addressed in order to fully realize their benefits and to understand the implication of the end products on plant production. Key words/phrases: Coffee husk compost; COMPOCHIP microarray; Enzyme activity; Organic matter degradation; PCR-DGGE; Germination index
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    Plant Growth Promoting Properties of Rhizobactel'ia Isolated from Chickpea and Lentil Producing Areas of Ethiopia: Implication for Productivity in Low-inputs Agricultural system
    (Addis Ababa Universty, 2013-06) Jida Mulissa; Assefa Fassil (Professor)
    Chickpea (Cieer arielinulIl L.) and lentil (Lens euinaris Medik.) are among the major sources of dietary protein for the majority of population in Ethiopia. They also restore and maintain soil fertility through their symbiotic nitrogen-fixation in association with rhizobia. This study was intended to investigate plant growth promoting propelties of chickpea and lentil nodulating rhizobia and phosphate solubilizing bacterial isolates fi'om major producing areas of the country. Accordingly, a total of 157 bacterial isolates were obtained and characterized using different standard methods. Phenotypic and plant gro\\1h promoting characteristics of 66 rhizobia I isolates recovered fi'omroot nodules of chickpea and lentil grown in soils collected fi'Dln producing areas of the country were investigated. These rhizobial isolates showed wide diversity in their different C and N-sources utilization pattern and tolerance to salinity, high temperatures, acid and alkaline pH, heavy metals and antibiotics. Symbiotic and morphological characterization also showed a wide diversity among these rhizobial isolates. Moreover, some of these rhizobial isolates exhibited plant groMh promoting characteristics such as phosphate solubilization, IAA production and antagonistic activity against FusariulIl o.\)'sporulll. In addition, a total of 91 phosphate solubilizing bacterial isolates were obtained fi'om chickpea and lentil rhizosphere soil samples collected fi'om different producing areas of the country. These isolates were identified using the API biochemical test kits and partial 16S rDNA sequences analysis. The result showed the presence of genera such as Aeinelobacler, Aerolllonas, Agl'ObacleriulIl, Bacillus, Brevibacillus, BUl'khoideria, Chl)iSeOmOnas, Empedobaclel', Enlel'Obaclel', Pseudomonlls, Raislonia, Sphingomonas and Slenoll'Ophomonas. Phosphate solubilizing efficiency of the isolates was assessed qualitatively using solubilization index and quantitatively. Quantitative estimation of the phosphate solubilization efficiency of different insoluble phosphorus sources showed that most isolates have good solubilization ability of tricalcium phosphate, rock phosphate and bone meal. Furthermore, some of these isolates showed multiple plant growth promoting characteristics; production of indole acetic acid and fluorescent siderophore, and inhibition of FusariulII oxysporulII growth under ill vitro conditions. The effects of inoculation of isolates with multiple plant growth promoting properties on chickpea and lentil growth in pot culture showed significant improvement in number of nodules, shoot dry matter, shoot nitrogen and phosphorus contents. Generally, the present study indicated that Ethiopian soils contain symbiotically effective chickpea and lentil rhizobia which are endowed with different plant growth promoting attributes. Moreover, chickpea and lentil rhizosphere harbor phosphate solubilizing bacteria which are diverse in their taxonomy and phosphate solubilizing efficiency. Some of these isolates are of particular interest for further research under different tield conditions. Key words: IAA, MesorhizohiulII species, PGPR, PSB, RhizobiulII species