Microbial, Cellular and Molecular Biology

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    Diversity and Phytobeneficial Properties of Indigenous Root Nodule Bacteria and Arbuscular Mycorrhizal Fungi Associated with Erythrina Species and their Effect on Growth of the Host Plant under Greenhouse Conditions
    (Addis Ababa University, 2021-04-09) Berza, Belay; Assefa, Fassil (Professor)
    Erythrina is a leguminous tree used in agro-forestry practices in the southern and southwestern Ethiopia. The legume-rhizobium symbiosis provides N to plants, while Legume-AMF symbiosis enhances P availability. Despite the crucial agro-forestry attributes, there were scarcities of information on species diversity of arbuscular mycorrhizal fungi (AMF) associated to E. brucei, genetic and functional diversity, and eco-physiological stress tolerance traits of bacteria isolated from the root nodules of E. abyssinica. The objectives of this study were to; 1) determine AMF species diversity and richness associated with Erythina brucei; 2) the genetic and functional diversity and eco-physiological stress tolerance traits of bacteria isolated from the root nodules of E.abyssinica ;3) evaluate inorganic phosphate solubilizing efficiency and multiple phytobeneficial properties of root nodule bacteria associated with E. abyssinica; 4) evaluate the symbiotic effectiveness of Bradyrhizobium species, and evaluate the effects of multiple inoculation of the consortia of microorganisms on Erythrina abyssinica growth, nodulation, and shoot TN and P contents in greenhouse condition. Soil samples were collected from different E.spp growing areas involved various land use types and trap cultures were established. AMF spore extraction and species identification was done from soils obtained from the rhizosphere of E. brucei. The root nodule bacteria of E. abyssinica were obtained by plant infection method. The genetic diversities were studied using Amplified ribosomal DNA restriction analysis (ARDRA). The nifH was gene screened and the taxonomic position was determined using 16S rRNA sequence analysis. Plant growth promoting traits were also evaluated and selected microbial inputs were studied for their effects on E. abyssinica growth, nodulation, nitrogen fixation and uptake P in the greenhouse condition. Eleven AMF genera and 33 species were recovered. The ARDRA grouped the bacterial isolates in to twelve clusters at 70 % similarity level. NifH gene was amplified in 15 (50%) of the root nodule bacteria the 16S rRNA sequence analysis grouped the bacterial in to nine genera (species): Bradyrhizobium (n=3), Paenibacillus (n=2), Bacillus (n=2), Staphylococcus (n=2), Enterobacter (n=2), Achromobacter (n=1), Acinetobacter (n=3), Gluconobacter (n=4) and Stenotrophomonas (n=2). Achrmobacter, Acinetobacter, Bacillus, Gluconobacter, Paenibacillus, Staphylococcus and Stenotrophomonas are the first reports from E. brucei root nodules. In addition, Bradyrhizobium cajani and Bradyrhizobium cytisi are the first reports from E. abyssinica root nodules and from Ethiopia as well. The 61.9%, 76.2%, 47.6%, 33.3% and 19.04% of the isolates were inorganic phosphate solubilizers, IAA, NH3, HCN and chitinase producers respectively. The maximium shoot length and dry weight improvements (140%) and (267%) were recorded by multiple inoculations involved B. shewense (AU27) + Glomus sp.1(AMF1) + Acaulospora sp.1 (AMF2) + A. soli (AU4). Similarily, the maximum shoot total nitrogen improvement (260%) was recorded in treatment received B. shewense (AU27) + A. soli (AU4) inoculation; whereas the highest shoot P improvement (1200%) was recorded by B. shewense (AU27) + Glomus sp.1 (AMF1) inoculation. AMF species richness was affected by land use types and geographic locations. Phylogenetically and functionally diverse bacteria were inhabited in the root nodules of E. brucei. Multiple inoculations enhanced E. abyssinica growth and development, biomass production, nitrogen fixation and multiple inoculations also improved phosphorus uptake by the host plant. Synergistic interaction was observed among the microsymbionts and macrosymbiont. More studies are required to explore the rhizosphere of E. abyssinica and the performance of the microbial inputs which exhibited potential performance in greenhouse needs evaluation in the field condition.
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    Genetic and Physiological Diversities of Bacteria from the Rhizosphere of Woody Plants Grown on Soil from Girar Jarso District and their Effect on the Growth and Establishment of Tree Seedlings in North Shewa Zone Ethiopia
    (Addis Ababa University, 2020-04-19) Getahun, Alemayehu; Muleta, Diriba (PhD); Assefa, Fassil (Professor)
    Land degradation (LD) is one of the major problems the planet earth has been facing. It has severely affected 1.9 billion hectares and decreased ecosystem services by 60%. In Eastern Africa, particularly in the northern highlands of Ethiopia, 1 billion tons of topsoil are lost annually. Thus, the land becomes bare, nutrient-depleted, water-stressed and abandoned by the community. Hence, it needs urgent rehabilitation with the application of eco-friendly microbes and organic amendments (OAs) through the exploitation of different types of trees/shrubs. Therefore, the overall objective of the study was to rehabilitate degraded habitat through the application of OAs and phytobeneficial bacteria for the establishment of multipurpose trees. Soil samples were collected from nine random corners at depth of 30 cm for soil physicochemical analysis before and after OAs. Six different treatments (biochar, compost, manure, mixed, bacterial inoculation (BI), and control) were considered at a 1:1 ratio of OAs per pot. Application BI was done for field trials during transplanting and quarterly for a year. Six plots measuring 41 m x 4 m were established in completely randomized block design and assigned at the random block for the field trial. Following OAs, microbial counts were done for one year every month. Bacteria were isolated from the rhizosphere of Acacia and Juniperus. The primary selection of isolates was based on drought stress (DS) tolerance and phosphate solubilization, other stress, and plant growth-promoting traits. The potential isolates were subjected to carbohydrate and amino acid utilization tests, BOX PCR, and 16S rRNA profiling. DS tolerant, phosphate solubilizer strains with multiple plant growth-promoting traits were chosen for Acacia seeds germination and field application alone and in combination with OAs. Plant growth parameters and their survival rate at each amended plot were assessed. There is a significant increase in soil pH (5.69-8.13), CEC (43.78-49.98 cmolc/kg), OM (2.43-3.91%), total nitrogen (0.13-0.76%), and available P (18.9-26.31 ppm) following OAs compared the control. Combined treatment had the largest effect on cover crops biomass with 3.43 g, 4.54 g, 0.7 g, 2.07g in alfalfa, grass pea, and control respectively p ≤ 0.05. The C and N utilization revealed metabolic versatility of the strains (14.29 to 100%). Ochrobactrum, Pseudomonas, and Klebsiella spp expressed remarkable metabolic versatilities. BOX-PCR showed greater genetic diversity and confirmed by Simpson’s Index of Diversity (0.883) took the leading position with Bacillus species. The 16S rRNA genes sequence showed 21.92% Firmicutes and 78.08% Proteobacteria with Pseudomonas 23% and Ochrobactrum 21% dominant species. Out of 73 isolates, 10 (14%) were highly tolerant of 40% polyethylene glycol. All the isolates can grow in wider ranges of pH (5-9), temperature (15-45°C). The inoculated bacterial strains significantly p ≤ 0.05 increased root, shoot length, and dry biomass of acacia. According to solubilization index (SI) 45% isolates were classified as high and medium phosphate solubilizers with 195 to 373 μg/mL. The maximum P and IAA were produced by Pseudomonas FB-49 (373 and 659.07μg/mL), respectively. The highest (100%) seed germination caused by Pseudomonas BS-26 and Pseudomonas FB-49. There is a significant difference in microbial counts following OAs compared to the control. The greatest counts in bacteria, actinomycetes, and fungal (21.66, 2.29, 0.82 x 105 CFU g-1) of soil, respectively in the combined amended plot. There was a significant increase in stem height, girths, and branch numbers in amended plots relative to the control. The survival rate was observed in apple (80%) followed by acacia (66%) and prunus (51%). The survival is in the order of BI x OAs > BI > biochar > compost > manure > control amended soil. This study concluded that degraded land could be rehabilitated with cheap OAs, potential bacterial strains, and bring multipurpose tree establishment with greater survival rate and best performance.
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    Evaluation of Phyto Beneficial Traits of Indigenous Phosphate Solubilizing Bacteria and Fungi as Microbial Inoculants for Enhancing Growth and Production of Coffee (Coffea Arabica) Under Greenhouse and Field Conditions in Jimma South West of Ethiopia
    (Addis Ababa University, 2021-12-17) Abafita, Reshid; Assefa, Fassil (PhD); Muleta, Diriba (PhD)
    Exploitation of phosphate solubilizing bacteria (PSB) and fungi as microbial inoculants is known to promote plant growth through the supply of plant nutrients and supression of pathogens. In view of this, the present investigation was planned to assess the phytobeneficial traits of phosphate solublizing bacterial and fungal isolates recovered from coffee (Coffee arabica) and vermicompost to determine their potential in growth promotion of coffee seedlings under low input agriculture.. The microbes were isolated and purified following standard methods. The selected isolates were investigated for their plant growth promoting properties, eco-physiological tolerance under laboratory conditions, and further tested under greenhouse and nursery experiments. The greenhouse and nursery experiments were conducted with completely randomized design (CRD) in three (3) replications per treatments. Thus, a total of 154 bacteria and 72 fungi isolates were recovered from which twelve potent bacterial and nine fungal isolates were selected and investigated for their plant growth promoting properties. Among the twelve bacterial isolates, three of them were tentatively identified to the genera of Pseudomonas (RCHVCB1) and Bacillus (RScB1.19 and RMaB2.11), and showed significant potential to solubilize Ca3 (PO4)2 and posessed several phytobeneficial traits, viz, indole acetic acid, NH3, HCN productions and N-fixing ability. They also exhibited remarkable tolerance to ecophysiological factors such as heavy metal, acidity and salinity, and inherent antibiotic resistance (IAR). Similarly, three fungal isolates with superior phosphate solubilizeation ability were characterized and identified as genera of Penicillium (RSCF1.19) and Aspergillus (RCHVCF2 and RLVCF2). During co-culture, RSCF1.19 (Penicillium sp.) slightly inhibited the test pathogen, Fusarium xyloriodes. The bacterial (RCHVCB1, RScB1.19, RMaB2.11) and fungal isolates (RSCF1.19, RCHVCF2, RLVCF2) enhanced rate of coffee seed germination under laboratory conditions and promoted coffee seedlings growth under glasshouse conditions. The results of inoculated seeds showed significant (p≤0.05) differences in germination rate and vigor index compared to the control. Isolates RScB1.19, RMaB2.11+RSCF1.19 and RMaB2.11 + RLVCF2 showed high germination rate (20.59%) over the control (13.33%). Moreover, a single inoculation of RLVCF2, RSCF1.19 and co-inoculation of RMaB2.11 with RLVCF2 also showed significant (p≤0.05) mean root length (1.31 cm) and mean shoot length (1.48 cm) over the control. Under greenhouse conditions, single inoculation of RSCF1.19+phosphate and dual inoculation of RSCF1.19 and RCHVCB1 in the presence of inorganic phosphate fertilizer led to significantly higher plant height, root length, stem girth, leaf number, leaf area, fresh and dry weights. Due to high pH value of the potting medium (vermicompost alkaline pH-pH>7.5), all the treatments combined with vermicompost showed suppressive effect and no any seedlings were emerged at all. Under nursery conditions, co-inoculation of RSCF1.19 with three bacterial isolates (RCHVCB1, RScB1.19, and RMaB2.11) in combination with inorganic phosphate led to significantly increase the tested growth parameters. Similar increase in growth attributes was observed in both single and dual inoculations due to vermicompost used compared with both positive and negative controls. Higher NPK-uptake was observed in a combination of bioinoculants and inorganic phosphate fertilizer compared to the positive and negative control. In general, inoculation of RSCF1.19 and RLVCF2 isolates to coffee 74110 variety combined with inorganic phosphate fertilizer resulted in good vigor and healthier coffee seedlings (RSCF1.19, 34.42%) and (RLVCF2, 37.09%) when compared to control (28.49%). Therefore, both RSCF1.19 and RLVCF2 fungal isolates could be used as bioinoculants after field trials in coffee 74110 variety productions.
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    Detection of Major Enteropathogens Associated with Calf Diarrhea in Dairy Farms of Afar Region Ethiopia
    (Addis Ababa University, 2021-10-21) Abera, Eskedar; Desalegn, Asnake (PhD); Kebede, Nigatu (Professor)
    Diarrhea is one of the most common causes of sickness and mortality among newborn calves. It has serious financial and animal welfare indications in both dairy and beef herds. Diarrhea in a newborn calf is one of the most common diseases in young animals, causing huge economic and productivity losses to livestock industry worldwide. Dairy farming is a growing cattle production system in Ethiopia. However, morbidity and mortality of calves are among the factors that have been hindering success of dairy industry. Multiple enteric pathogens such as virus, bacteria and parasite are the most common causes of calf diarrhea. However, various facets of diarrheal disease caused by rotavirus, coronavirus, E. coli K99 and Cryptosporidium parvum infections in calves in Afar region are inadequately understood. A Crossectional study was carried out from February 2020 to April 2021 with the aim of prevalence and associated risk factors of rotavirus, coronavirus, E. coli K99 and Cryptosporidium parvum infections in calves from selected districts of Afar national regional state, in north east Ethiopia. A total of 176 fecal samples of 0 up to 6 months old diarrheic calves were collected by purposive sampling. Samples were tested for those four pathogens using a commercially available enzyme-linked immunosorbent assay (ELISA). In addition, different factors were measured the level of association between variables. Out of the 176 samples tested 59 were positive for at least one pathogen by ELISA. Positivity rates for each pathogen were, coronavirus 45 (25.5%), Cryptosporidium parvum 29 (16.5%), rotavirus 18 (10.2%) and K99 enterotoxigenic E.coli 11(6.25%). From positive viral samples of ELISA test 38 were propagated in Madin Darby bovine kidney cells. After 3 following passage, progressive cytopathic effect (CPE) i.e. rounding, detachment as well as demolition of mono-layer cell of all samples was observed. The study sit, time of first colostrum feeding, and duration of colostrum feeding were significantly associated with the occurrence of rotavirus, coronavirus, Cryptosporidium parvum and E. coli and this are indicative of the need for strict prevention and control mechanisms such as practicing early colostrum feeding in newborn calves.
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    Genetic Characterization and Estimation of Genotype by Environment Interaction of Ethiopian Sesame (Sesamum Indicum L.) Germplasm
    (Addis Ababa University, 2021-07-02) Tesfaye, Tewodros; Tesfaye, Kassahun (PhD)
    Sesame is one of the major oil crops that has great economic importance for the country. In Ethiopia, sesame is among the foremost important oil crops both in terms of area coverage and total national annual production. However the crop suffers from low productivity due to biotic and abiotic stresses. Therefore, the present study conducted in different sets to generate information that can be used to design the future breeding program of sesame in the country. The first set of experiments was to study the morphological and molecular genetic diversity of the sesame germplasm collected from Ethiopia and other countries (Asian and other African countries). The same genotypes planted at three locations for phenotyping and genotyping using the two high throughput diversity array technology (DArT) markers (silicoDArT and SNP). Further to understand the impact of different putative genes Genome-wide association study of yield-related traits using 2997 SNPs in two environments was performed. The second set of experiments was conducted in 19 environments to assess the performance and stability of sesame varieties, and to characterize sesame growing environments in Ethiopia. Based on morphological characterization, genotypes showed wide variability for most morphological traits, except for plant growth type, leaf glands, anther filament color, and anther connective tip gland. High heritability combined with high genetic advance was recorded for plant height, primary branch, days to flower initiation, days to 50% flowering, pod bearing zone, seed yield per plant, and bacterial blight reaction indicating the potential of improving the population through a direct selection for these traits. Grain yield showed a significant and positive genotypic correlation with plant height, the number of capsules per plant, and pod bearing zone, the magnitudes of the positive genetic correlation suggest that the selection by those characters produces a significant increase in grain yield. Genetic divergence using Mahalanobis D2 statistics was computed, and the genotype lines were grouped into six different clusters. Clustering was not associated with the geographical distribution; instead, genotypes were grouped mainly based on morphological differences. The maximum inter-cluster distance was observed between clusters IV and VI (D2 =342.56, followed by clusters I, and VI (D2 =217.9783). Maximum genetic recombination and variation in the subsequent generation are expected from crosses that involve parents from the clusters characterized by maximum distances. The genetic diversity analysis showed that the average nucleotide diversity of the panel was 0.14. Considering the genotypes based on their geographical origin, Africa collections (0.21) as a whole without Ethiopian collection was more diverse than Asia and when further portioned Africa, North Africa (0.23) collection was more diverse than others, but at the continent level, Asia (0.17) was more diverse than Africa (0.14). The genetic distance among the sesame populations ranged from 0.015 to 0.394, with an average of 0.165. The structure analysis divided the panel into four hypothetical ancestral populations and 21 genotypes were clustered as an admixture. Under Genome-wide association study (GWAS) a total of 21 significant SNPs with 7 yield-related traits in two environments were identified and, these explaining the phenotypic variation ranged from 7.02 (DF) to 16.11% (CAPL), with an average of 9.76%, suggesting a moderate contribution to the traits. All significant loci found in LG 2, 6, and 11 associated with capsule length except one associated with the physiological period. The significant loci found in LG 3, 7, and 8 associated with a physiological period (Days to flower initiation, Days to 50% flowering, and Days to physiological maturity) except one associated with capsule length. Dissecting genetic control of flowering time and maturity is importance to foster sesame breeding and to develop new varieties able to adapt to changing climatic conditions. Indeed, flowering time and maturity strongly affect yield and plant adaptation ability. Since several favorable alleles detected in this study have not yet been intensively selected, our GWAS results will assist in incorporating further useful alleles into the elite sesame germplasm for a seed yield increase in the future. Based on the genotype x environment interaction study the test locations were divided into six groups. Humera, Banat, and Tach Armacho were highly discriminating and representative in the first, the second, and the third group respectively, and were identified as a core test site in that group. While Alemaya, Worer, and Mender67 were identified as the only test site in groups four, five, and six. The core testing sites identified would be used to facilitate the identification of superior sesame varieties and to reduce testing costs in the country. Environment Tach Armacho in 2017/18 and 2018/19 were close to the ideal environment. The GGE biplot analysis identified genotype G2 (setit-1) as the “ideal” genotype and among the highest mean seed yield. Setit-1 considered the most stable across variable environments.
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    Estimation of Breeding Parameters within and between Malt Barley (Hordeum Vulgare L) Crosses using Phenotypic Traits and Kasp SNP Markers
    (Addis Ababa University, 2021-12-31) Tadesse, Endeshaw; Tesfaye, Kassahun (PhD)
    The genetic variation in a breeding program is created by crossing genetically divergent parents. The resulting genetic variation between and within crosses is the determining factor for the offspring’s performance, which is defined as the level of compliance of the offspring with preset breeding goals. Estimation of breeding parameters plays a key role in crop improvement. The aim of this study was to (i) estimate and compare mid-parent value and the cross mean (ii) estimate and compare the variance between means of crosses (σ²c) and segregation variance of recombinant inbred line within crosses (σ²g) (iii) estimate correlation among the traits measured and their heritability (iv) estimate and compare Rogers Distances (RD) of parental lines and their correlation to the segregation variance within crosses, and (v) estimate usefulness of crosses. 900, F4:5 recombinant inbred lines randomly derived from 30 crosses were evaluated at two locations in modified split plot p-rep design whereas the parental where genotyped using SNP-KASP markers. The variance component analysis revealed significant (P <0.05) genetic variation among parents, crosses and RIL for almost all traits. Based on generation mean analysis, the means of recombinant inbred line did not deviate significantly from the means of parental lines. The range of σ²g for the individual crosses was very high for days to heading, days to maturity, plant height, thousand kernel weight and grain protein concentration and for these traits the respective standard deviation were high. Heritability for parents ranged from 49.50 % (malt extract) - 93.60 % (plant height), heritability for crosses ranged from 29.52 % (grain protein concentration) - 87.0% (days to maturity), whereas heritability for RIL was lowest with 27.40% (beta-glucan) - 73.60% for thousand kernel weight. Significant (P<0.01) genotypic correlations with high impact for practical breeding were found between malting traits. Significant (P<0.01) regression of cross mean on mid-parent value were obtained for all traits demonstrating that cross means can accurately be predicted from mid parent values and selection among crosses at an early stage of line development is highly effective. Further, based on the usefulness criteria, 16 outperforming crosses were identified that surpass Planet as the actual leading malt variety. Diversity analysis revealed an average RD between lines was 0.46 with a range of 0.32- 0.64 and homogeneity within the 17 parental lines varied from 71-100%. The dendrogram grouped the lines into three clusters. Importantly, the high malting European lines were grouped together with some of the ICARDA and Ethiopian lines Correlations of RD to σ²g ranged from -0.19 to 0.34 and were not significantly deviating from zero for all traits except DH, hence, RD proved to be not predictive for σ²g. In this study the two variances, σ²c and σ²g, proved to be significantly deviating from zero for almost all traits; hence breeder can exploit both of them for selection. Generally, differences between generation means (Parental lines vs. RIL) were small across all crosses. But when grouping into crosses with and without Planet as a parental line, deviation between parent and RIL were larger possibly due to epistastic effects. Hence breeder should estimate breeding values rather than genetic values of parental lines. The use of mid-parent value and usefulness for cross selection were found to be promising. The national barley breeding program should exploit this result by starting with a large number of initial crosses and reducing them to the most promising crosses. Some heterogeneity was observed within the lines. When originating from technical mixture this should be considered as a wake-up call for the breeder to closely follow maintenance breeding. RD based on KASP markers was not predictive for σ²g; hence further research will be required by improving accuracy of σ²g estimate and linkage disequilibrium between markers and quantitative trait loci (QTL). Over all, the large diversity revealed among lines indicates the potential that the genotypes have to improve the productivity and quality of the crop and the uses of molecular markers can benefit the breeding program in order to have an effective breeding program by selecting diverse parental genotypes with complementary gene action.
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    Characterization of Ethiopian Durum Wheat Landraces and Cultivars for Processing Quality Using Phenotypic Traits and Simple Sequence Repeat (SSR) Markers
    (Addis Ababa University, 2021-09-02) Dagnaw, Temesgen; Tesfaye, Kassahun (PhD); Haileselassie, Teklehaimanot (PhD); Geleta, Mulatu (PhD)
    Durum wheat (Triticum turgidum L. ssp. durum Desf.) is getting attention in terms of production and area coverage at the global level. Its unique characteristics to produce pasta and related end-products increased the preference for durum wheat by millers and processors. Ethiopia is considered as a center of diversity for durum wheat. However, in Ethiopia, durum wheat is facing serious genetic erosion due to the expansion of teff and bread wheat, and the cultivation of exotic durum wheat materials. The genetic resource of the Ethiopian durum wheat genotypes has been limitedly explored for its processing quality attributes at the level of both phenotypic and molecular diversity. Therefore, the present study was aimed to assess the genetic diversity and evaluate the processing quality attributes of Ethiopian durum wheat genotypes using phenotypic traits and SSR markers. The field experiment was conducted at two locations (Sinana in Bale zone and Chefe Donsa in East Shewa zone) to assess the phenotypic diversity. The phenotypic characterization of genotypes showed a wide range of variability for most quantitative and qualitative traits. The combined analysis of variance (ANOVA) showed highly significant (P<0.01) variations among genotypes for the majority of the traits studied. Gluten content (GL) and grain yield (GY) showed high and intermediate heritability, respectively, combined with moderate genetic advance, and grain protein content (GPC) showed intermediate heritability combined with low genetic advance. Both, GPC and GL showed a significant and negative correlation with GY. Cluster analysis based on quantitative traits grouped the genotypes into 5 major clusters. The first four principal components (PCs) explained 64% of the total variation. Based on qualitative traits, high genetic variation was observed among genotypes. Correspondence analysis discriminated cultivars from other populations of landraces. Finally, 104 best-qualified genotypes for processing quality traits were selected and assessed using quality traits associated 14 SSR markers, which had high mean polymorphic information content (PIC) of 0.56 and gene diversity (0.61). High levels of genetic diversity was obtained among all populations (I = 0.86; He = 0.46). Analysis of molecular variance (AMOVA) revealed high genetic variation within populations (88.35%) and among populations (11.65%). The Neighbor-joining (NJ) clustering and principal coordinate analysis (PCoA) grouped the genotypes into three major clusters. In addition, Bayesian model-based population structure analysis revealed two major genetic groups. Overall, this study revealed high genetic diversity among Ethiopian durum wheat landraces and cultivars, and these genotypes can be used to identify the best genotype/s for processing quality attributes and subsequent use in the Ethiopian durum wheat improvement programs.
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    Analysis of Five Year District Health Office Data for Intestinal Parasitosis in Aleta-Chuko Sidama Ethiopia
    (Addis Ababa University, 2021-09-15) Degife, Teshale; Mamo, Hassen (PhD)
    Intestinal parasitic infections (IPIs) are widely distributed, particularly in low-income countries. The objective of this study was to retrospectively assess the prevalence of IPIs among preschool (1-4 year) and school-age (5-18 year) children in Aleta-Chuko District (ACD), Sidama Region, South Ethiopia, in the past five-year (September 2015 - August 2019). The study was retrospective study. It used secondary data from disease registers of ACD health office (2015 - 2019) to assess the prevalence of IPIs among preschool and school-age children.Overall, 199,241 children visited district health centers during this period and 140,706 (65,292 males, 75,414 females) were school-age children. Of these, 50,057(25.1%) were positive for IPIs with slightly more females (13.2%) than males (11.9%). The number of children diagnosed for IPIs had increased continuously although there was some decline in 2016 and 2017. Six different types of intestinal parasites were identified. Ascaris lumbricoides was the most prevalent (41.5%) followed by Entamoeba histolytica/dispar/moshkovskii (14.3%). The least identified parasite was Schistosoma mansoni. Intestinal parasites remain major health problem among preschool and school-aged children in ACD and emphasis on control measures including periodic deworming is needed. Comprehensive community-based studies are required for better understanding of the impact of IPIs on children in ACD.
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    Characterization of Ethiopian Chickpea (Cicer Arietinum L) Genotypic Diversity Population Structure Cold Tolerance and Assessment of Change of Variability Over time
    (Addis Ababa University, 2021-12-29) Admas, Sintayehu; Shiferaw, Eleni (PhD); Tesfaye, Kassahun (Professor); Hailesilassie, Teklehaimanot (Professor)
    Plant genetic resources are valuable source of desirable agronomic traits that can be used in crop improvement. These resources are constantly threatened by both natural and human-caused risks. Chickpea (Cicer arietinum L.) germplasm is one of the crops affected by these factors. Regular chickpea diversity monitoring in farmers' fields, as well as characterization and evaluation of Ethiopian chickpea germplasm in ex-situ, are essential in order to build viable conservation strategies and undertake breeding initiatives. However, in Ethiopia chickpea, the abovementioned information is limited. As a result, this research began with the general objective of determining the diversity of Ethiopian chickpea germplasm and its potential for frost resistance, as well as determining the degree to which chickpea on-farm diversity has changed over time. In the study, molecular analysis was conducted to determine the genetic diversity, population structure, and relationships of 152 chickpea genotypes using simple sequence repeats (SSR) markers. There were 133 alleles observed using 23 polymorphic SSR markers, with a mean of 5.8 alleles per locus. Pairwise population Nei's genetic distance, heterozygosity (0.4), Shannon's information index (1.2), Gene Diversity (0.67), polymorphic information content (0.63), and percent polymorphism (99.5%) were among the genetic-based statistics used in the study. These studies demonstrated that there was high genetic variation within and among chickpea genotypes. The 152 genotypes were divided into two groups on UPGMA tree developed based on Nei's genetic distances. The exotic genotypes were all grouped together in one cluster, showing that they are distinct from Ethiopian genotypes. However, seed exchange caused inconsistencies in the patterns of grouping Ethiopian chickpea genotypes by geographic location. Two separate populations were discovered using model-based population structure clustering. These findings can be used to create efficient conservation approaches and breeding initiatives to enhance the genetic diversity of chickpeas. To assess the potential of Ethiopian chickpea for frost resistance, a frost screening study was carried out in a natural field and in controlled conditions. Six hundred seventy-three genotypes were characterized for two years (2018/2019 to 2019/2020) using an augmented design at Bakelo, Debre Birhan, Ethiopia, a place as national frost screening site. A significant (p<0.01) variability amongst genotypes was recorded for all agronomic traits considered. In terms of agronomic performance, 94 (12.6%) of the genotypes examined outperformed the frost susceptible genotypes. Stem/leaf pigmented genotypes had a better response to frost stress than non-pigmented genotypes. The majority of black seeded chickpeas were adapted well to frost stress, when compared to brown and white seeded genotypes. According to the freezing tolerance rate (FTR) and plant survival rate (SR), 83 (12.3%) and 85 (12.6%) genotypes were identified as frost tolerant. Grain yield had a significant (p<0.01) correlation with FTR, SR, seed shriveling score, stem/leaf pigmentation, and seed color. Based on the field experiment genotypes performances, 72 genotypes were selected and evaluated further for cold tolerance variability at seedling stage under controlled environment using a complete randomized design with two replications. The analysis of variance revealed a significant (P<0.01) difference among genotypes for plant height, number of foliage, number of primary branches and fresh weight. Cold tolerant genotypes had a higher growth rate than susceptible genotypes, which had a slower growth rate or completely failed. According to SR, 31 (43.1%) genotypes scored above 0.8 values. Based on FTR, 37 (51.4%) and 31 (43.1%) genotypes were rated at a score of 1 to 3 in freezing test 1 (T1) and freezing test 2 (T2), respectively indicating better frost tolerance than others. Fresh biomass yields had a high negative correlation (p<0.01) with SR, but a substantial positive correlation (p<0.01) with FTR value. At seedling stage, based on the combined results of FTR and SR scores, 26 genotypes were shown to be cold tolerant genotypes at temperatures as low as -5oC. Therefore, the field and controlled environment experiments results revealed that Ethiopian chickpea landraces have a good genetic potential for cold tolerance traits for use in breeding programs. In addition, a diversity monitoring research was conducted to look into the differences in phenotypic diversity between the present (2017) and prior chickpea collections (1979-1983). The experiment was conducted using simple lattice design with two replications at Debre Zeit Agricultural Research Center for two consecutive years (2018/2019 to 2019/2020). The result showed that chickpea has been under genetic erosion ranging from 30.4% to 100% during the last 35 years. During this time, estimations of Shannon-Weaver diversity index for black seeded and ivory white seeded chickpea kinds have declined from 1.99 to 0.69 and 1.33 to 0.0, respectively. In previous collections, 25.8% of black coated genotypes and 6.5% of white coated genotypes were found, but these chickpea varieties are now rare (black 6.5%) or unobtainable (white 0.0%). Cluster analysis revealed that genotypes clustered according to the time of collections regardless of their geographic origins. The genotypes of chickpeas in the current and prior collections were different. Landraces of black and white seeded chickpeas were vulnerable to genetic erosion and chickpea genotypes at farmer’s field are found to be losing their variability. Black and white seeded chickpea are missing at farmers’ fields and are under greater threat of extinction. So, implementation of immediate restoration of chickpea landraces is required to recover and maintain the lost chickpea landraces in the studied region and we strongly recommended revision of the current conservation strategies.
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    The Relative Importance of Three Cabbage Aphid Species (Brevicoryne Brassicae, Lipaphis Erysimi; Myzus Persicae) and Their Parasitoid Complex in Kenya.
    (Addis Ababa, 2000) Gacii, Peter; Seyoum, Emiru(PhD)
    Studies on the relative importance of the three known species of cabbage aphids (Brevicoryne brassicae (Linneaus), Myzus persicae (Sulz) and Upaphis erysimi (Kaltenbach)) and their parasitoid complex in Kenya were carried out. The studies involved farmers' fields' surveys and interviews and also field experiments conducted at the Jomo Kenyatta University of Agriculture and technology (JKUAT) Research and Production Department land and in a farmer's field at Limuru. Gloria F1 hybrid was sown at JKUAT while Copenhagen Market variety was sown at Limuru. The field experiments were conducted in a Completely Randomized Block Design (CRBD) involving three treatments and a control, with three repl icates each. The treatments comprised of applications of: - Lambda-Cyhalothrin (Karate; Bacillus thuringiensis Berliner (Bt.) var. Kurstaki toxin (Dipel 2x) that selectively kills Diamondback moth (DBM); A combination of the two pesticides (Karate and Bt.) and Untreated controls. Parameters included; relat ive abundance of the three species of aphids; their corresponding damage to cabbage; the number of mummies per sample; percent parasitism and the proportions of the primary- and hyper-parasitoids hatched. The parasitoid complex was constructed from the identified parasitoids. The results showed aphid counts to be higher in non karate-treated plots than in the karate-treated ones. More hyperparasitoids notably Alloxysta sp and Pachyneuron sp. than the only primary parasitoid (Diaeratiella rapae) emerged from non karate-treated plots. This might suggest that karate might have the advantage of repelling hyperparasitoids or inhibiting their emergence. Yield showed some negative correlation with aphid infestations. A combined treatment gave the best overall yield at both sites.
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    Genetic Diversity of Faba Bean ( Vicia Faba L). Landrace Accessions in Ethiopia
    (Addis Ababa University, 2021-10-08) Neda, Bullo; Feyissa, Tileye (PhD); Dagne, Kifle (PhD)
    Faba bean is one of the most important legume species cultivated in Ethiopia for both human and animal protein source. Morphological and molecular characterization of faba bean accessions is important for future collection, conservation, and crop improvement programs. However, the genetic diversity of the Ethiopian faba bean accessions has not been comprehensively characterized. The present study was conducted to determine the genetic diversity of faba bean using morphological and ISSR markers. A total of 96 accessions including 90 landraces and six released varieties of faba bean were analyzed. Analysis of variance showed significant to very high significant differences for most of the traits. In this study phenotypic coefficient of variation, genotypic coefficient of variation, broad-sense heritability, and genetic advance as percent of mean revealed medium to high values for most traits. Traits that had a positive direct effect and positive correlation with grain yield could be used as a reliable indicator in indirect selection for higher grain yield. The D2 analysis showed a highly significant difference (P<0.01) among all intra and inter-cluster distances. Therefore, this study indicated that there is an opportunity to improve this crop through simple selection and hybridization in these accessions. Using ISSR molecular markers, the polymorphic information content, marker index, resolving power, and effective multiplex ratio showed average values of 0.32, 5.87, 7.14, and 18.34 per primer, showing high polymorphism values. The marker showed average gene diversity of 0.26, ranging from 0.15 to 0.36. The amount of polymorphism among the landrace accessions ranged from 44.91% for the Sidama population to 72.46% for the East Hararghe population, with a mean of 57.27%. AMOVA indicated 99% and 1% variation within and among populations, respectively. The current study exhibited little to moderate population differentiation and high gene flow. Both distance-based and model-based cluster analysis distinguished seven distinct XVII groups showing the conformity of results obtained from the study. This study exhibited high genetic diversity of collections from most of the administrative zones.
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    Marker Assisted Introgression of Striga Resistant and Drought Tolerant Quantitative Trait Loci into Popular Ethiopian Sorghum Cultivars
    (Addis Ababa University, 2021-03-30) Abebe, Teklay; Belay, Gurja (PhD); Tadesse, Taye (PhD); Keneni, Gemechu (PhD)
    Sorghum (Sorghum bicolor (L.) Moench) is the fifth important dryland cereal crop worldwide, and a dietary staple food for more than half a billion poor and food-insecure people dwelling in the semi-arid tropics of Africa and Asia. Sorghum is one of the strategic crops in Ethiopia, which provides more than one third of the cereal diet and is almost entirely grown by subsistence farmers to meet needs for food, feed, income, beverage, house and fuel purposes. Despite the multiple uses, its productivity is constrained at least partly by the intense Striga infestation and frequent drought, which causes up to 100% crop loss. The possible approaches to overcome the two prevailing constraints include genetic manipulation of the crop and the growing environment. The use of host-plant resistance offers a potentially cost-effective and environmentally sound complementary management strategy for incorporation into integrated management of Striga and drought. The studies were, therefore, designed to introgress Striga resistance and drought tolerance genes/quantitative trait loci (QTLs) into popular and farmer-preferred cultivars through marker assisted backcrossing (MABC). One-hundred-eighteen and 61 backcrossed lines were developed for Striga and drought, respectively. The 118 backcrossed lines and 15 parental lines were evaluated for Striga resistance based on low Striga seed germination stimulant activity in the agar gel assay (AGA) and under field condition. The AGA study was conducted at Holetta national agricultural biotechnology research center (NABRC) arranged in completely randomized design with four replications, while the field evaluation was conducted at two selected Striga hotspots and two non-infested locations layout in -lattice design with triplicates. The 61 drought tolerant converted and nine parental lines were evaluated under two-contrasting (optimum vis-à-vis stress) environments in -lattice design with three replications. The well-watered experiment was irrigated throughout the season, while the water-limited trial was irrigated well during the early growth stages with irrigation withheld at anthesis in order to allow moisture stress during grain-filling stage. The results from the AGA indicated that genotypes significantly differed (P<0.01) for their low production of Striga germination stimulant activity (lgs). The mean maximum germination distance (MGD) and germination rate (GR) ranged from 0.0 mm to 29.45 mm and 0.0% to 72.4% in the agar, respectively. Of the 118 backcrossed lines, 22.9% showed less than 10 mm of MGD and GR of <30%, revealing provision of low germination stimulant/strigolactones production (lgs). There were significant positive (r = 0.4-0.81) correlations showing the roles of these parameters as selection criteria in breeding for resistance. The existence of higher heritability (h2b = 77-83%) and genetic advance (GA = 62-93%) for the germination parameters indicated possibilities for improving resistance against Striga through selection. The results from the field evaluation also revealed that genotypes significantly differed (P<0.01) for both agronomic traits and reaction to Striga. About 21.2% of the backcrossed lines were promising with relatively good level of resistance to Striga and yield ranging from 773.3 to 1220 kg ha-1. Grain yield showed highly significant negative associations with all the studied Striga characters indicating severe damage. The heritability and genetic advance ranged from 37.7 to 88.7% and 7.72 to 52.9%, respectively, in which their values are greater for agronomic than Striga resistance traits. The aspects of interaction with heterogeneity of natural Striga infestation, pathogenicity, and concomitant environmental effects may have increased genotype by environment interactions resulted relatively low heritability. Genotypes that carry different QTLs showed different capacity to produce Striga germination stimulants in the AGA and agronomic performances under natural Striga infestation. The combined effect of both QTL (lgs2_SBI-05_60404021 and lgs_3_60629027) at a time showed lower Striga germination stimulant activity and better field resistance indicating existence of possible cumulative effects. The pre-attachment traits (MGD and GR) based on the AGA and emerged Striga under field condition showed significant positive relationships. This indicated that the AGA proved to be a useful, precise and fast indirect selection method to screen for sorghum lines with the low production of Striga germination stimulants resistance mechanism. The analysis of variance (ANOVA) for the stay-green (drought tolerant) experiment revealed that the developed backcrossed lines significantly differed for most of the agronomic and physiological characters. The mean grain yield of the genotypes under full-water (2069.2 to 5704.2 kg ha-1) and water-limited (923 to 4585 kg ha-1) conditions were 3633 and 1991 kg ha-1, respectively. Out of the 61 backcrossed lines, only 9.8% showed superior performance in yield ranging from 2831-4585 kg ha-1, under water-limited condition indicating the potential to withstand drought after anthesis. They were also characterized by high chlorophyll content, leaf area and greenness at physiological maturity. Relatively high heritability (35.4-79.5%; 35.1-76.3%) and genetic advance (1.7-35.5%; 1.4-42.7%) was obtained for most agronomic and physiological characters under full-irrigation and water-limited regimes, respectively, indicating that selection for such characters could be fairly easy. Grain yield showed significant positive association with chlorophyll content (r = 0.24 & 0.45), green leaves (r = 0.31 & 0.29) and leaf area (r = 0.29 & 0.27) both at booting and maturity under the former and later moisture regimes, respectively. This indicated that the presence of greater green leaf area with high chlorophyll content at booting and maturity contributed higher photosynthesis, better availability of food reserves for grain-filling and enhanced yield. Of the 13 groups of QTLs, the cumulative effect of stg1+stg2+stg3a+stg4, stg1+stg2+stg3a+stg3b, stg1+stg3a+stg3b+stg4, and stg2+stg3b+stg4 showed overall superior performance in most of the agronomic and physiological characters under both moisture regimes. Thus, the study provided useful evidence that the MABC transfer of lgs and drought tolerance genes from donor into popular cultivars have the potential to enhance tolerance/resistance to Striga and drought in sorghum, respectively, and could be used as noble tools supporting effective future sorghum breeding.
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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, 2020-07-09) 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 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 fertility; and provide that further field trials recommended over several seasons and sites in Ethiopia.
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    Identification and Characterization of Raw Starch Degrading Β-Amylase from Endophytic Bacillus Species
    (Addis Ababa University, 2008-08-06) Gashaw, Metages; Gessesse, Amare (PhD)
    Starch is a major reserve carbohydrate of all higher plants. In some cases it accounts 70% of the wet weight of plant material. It occurs in the form of water insoluble granules. The size and shape of the granules are often characteristic of the plant species from which they are extracted. When heated in water the hydrogen bonds holding the granules together begin to weaken and this permits them to swell and gelatinize. Ultimately they form paste or dispersion, depending on the concentration of polysaccharide (Prasanna, 2005). Starches are produced commercially from the seeds of plants, such as corn, wheat, sorghum or rice; from the tubers and roots of plants such as cassava, potato, arrowroot and the pith of sago palm. The major commercial source of starch is corn from which it is extracted by a wet milling process (Crab, et al., 1997). Starch is a heterogeneous polysaccharide composed of two high molecular weight entities called amylose and amylopectin which are differ in structures and physical properties. The insoluble amylose, composed of linear chains of α-1, 4 linked D-glucose residues can be separated from soluble amylopectin fraction by addition of a polar solvent, e.g. n-butanol. Even if amylose has a degree of polymerization of several thousands of glucose units, because of the molecular shape and structure, it is not stable in aqueous solution and retrogrades (precipitates spontaneously) this is because linear chains align themselves by hydrogen bonding and thus forms aggregates. Due to its molecular shape amylose has also considerable viscosity in alkaline solutions and forms complex with iodine to form intense blue color and this forms the basis of a method for quantitative determination of amylose. Amylopectin accounts for 75 to 85% of most starches. It has molecular weight of 107 – 108 and has a branched structure composed of chains of about 20 – 25 α-1,4 linked D-glucose residues. Amylopectin which is branched by α-1,6 linkages may contain 4 to 5% α-1,6-D-glucosidic bonds. In aqueous solutions, amylopectins are relatively stable due to branched molecules and are not able to form compact aggregates (Fogarty and Kelly, 1990).
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    Phytochemical Screening, Acute Toxicity and Anti-Rabies Activities of Extracts of Selected Ethiopian Traditional Medicinal Plants
    (Addis Ababa University, 2019-09-20) Tesera, Yeweynshet; Desalegn, Asnake (PhD)
    Despite the existence of safe and effective vaccines, rabies disease still causes an estimated 59,000 human deaths a year in the endemic areas in Asia and Africa. In most developing countries people believe to cure rabies with different traditional and religious treatment rather than seeking effective post exposure prophylaxis. The objective of this study was, therefore, to investigate the phytochemical constituents, acute toxicity and antirabies activity of crude extracts of the leaves of Justicia schimperiana and Ricinus communis and the stem bark of Croton macrostachyus. Extraction was done by maceration technique. Standard procedures were used to test the presence of various phytochemicals. For the determination of acute toxicity and antirabies activities, Organization for Economic Corporation and Development (OECD) Guideline No.423 was used. Different concentrations of extracts (0.4, 0.8, 1.6, 3.2, 6.4 and 12.8 mg/ml) were tested for their cytotoxic effect on vero cells through 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-Diphenyltetrazolium Bromide (MTT) assay. The mean cytotoxic concentration (CC50) was estimated and the antirabies assay was carried out using the minimal cytotoxic concentration of extracts. The phytochemical screening result has revealed the presence of alkaloids, flavonoids, phenols, steroids, tannins and terpenoids in all plant extracts screened. The toxicity evaluation of the extracts revealed that they are slightly toxic. The antirabies assay result showed that all plant extracts had a moderate to good antirabies potential. The 80 % methanol extracts exhibited higher antirabies activity compared to the other extracts under investigation. The present study concluded that these medicinal plants have possessed different phytochemicals that helps in the anti-rabies properties of the studied plants commonly used in Ethiopia. Further study on the mechanism of those phytochemicals must be elucidated for the potential as antirabies agent. Evaluation of those medicinal plants for their long-term toxicity would also be very important.
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    Genomic Diversity, Eco-Physiological Competence and Symbio-Agronomic Characteristics of Mesorhizobium Species Nodulating Chickpea (Cicer Arietinum L.) from Ethiopia
    (Addis Ababa University, 2021-01-01) Mohammed Zehara; Assefa Fassil (PhD); Cook Douglas (Professor); Fikre Asnake (PhD)
    Chickpea is among widely cultivated legumes for human consumption and grown as a rotation crop for the enhancement of soil fertility in succeeding cereal crops. The crop has the ability to fix atmospheric nitrogen in association with symbiotic nitrogen fixing bacteria in the genus Mesorhizobium. Thus, in order to fully realize the potential of nitrogen fixation in the crop and improved soil fertility, it is important to understand symbiont diversity, their eco-physiological competence among strains and to identify elite Mesorhizobium strains for their symbio-agronomic effectiveness. To this end, 138 farmer fields throughout major chickpea production regions were sampled for collection of root nodules, soils and recording of their farming practices. Pure rhizobial isolates (80) were subjected to Illumina whole genome sequencing and characterized using average nucleotide identity analyses. 64 strains occur within the Mesorhizobium genus that were assigned to eleven distinct Mesorhizobium genospecies and 3% of the surveyed diversity (genus) previously observed on chickpea, corresponding to genospecies 7A which is M. ciceri. The largest group of 26 strains belonged to M. genospecies 2A, which is conspecific M. spp. ORS3359, corresponding to unnamed symbiont isolated from Acacia seyal in Senegal. Genospecies 2A is related to but distinct from M. plurifarium. Among the diverse clade, 20 representative isolates were tested in-vitro for their eco-physiological competency and symbiotic efficiency in a greenhouse using Natoli and Arerti chickpea varieties. Most isolates were tolerant to high salinity 35%, high temperature 20%, acidity 25%, antibiotics resistance 67% and heavy metal resistance 83% as well as substrate (Carbon, Nitrogen) utilization and inorganic phosphate solubilization. The strains showed difference in nodule number, nodule dry weight and shoot dry weight of which 85% of the Mesorhizobium strains were either effective or highly effective on Natoli and Arerti varieties. Ten of the best performing symbiont strains were tested at three experimental locations for three consecutive years in split plot design using Natoli and Arerti chickpea varieties. Results from field evaluation indicate that the number of nodule and nodule dry weight showed two-fold difference between the low nodulating and high nodulating strains. Similarly, these strains increased shoot dry weight of Natoli and Arerti varieties (21-40%), above ground biomass accumulation (13-22%) and seed yield (19-31%) compared to uninoculated control irrespective of planting years at three sites. The robust strains comprising of M. loti 45P4S, M. amorphae 80P4S2, M.sp. LSJC280B00 2P3S1-b and M plurifarium 43P2S1 revealed more pronounced response in both nodulation and yield enhancement. These four strains are recommended for future inoculant production after validation with further field trials at different soil conditions. This study demonstrated symbiont diversity, eco-physiological competence and symbio-agronomic potential of selected elite Mesorhizobium strains in soils of chickpea growing regions of Ethiopia.
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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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    Characterization of Striga Sp. Seed Bank Depleting Bacteria from Striga Infested Sorghum (Sorghum Bicolor (L.) Moench) Growing Areas of Northern and Eastern Ethiopia
    (Addis Ababa University, 2020-06-06) Yilma Getachew; Assefa Fassil (PhD)
    Striga hermonthica is a very devastative parasitic weed of (Sorghum bicolor (L.) Moench) which is the third important crop in Ethiopia. It is hemi-parasite living with the host plant by attaching small sucker root system to host plant. A study was undertaken to evaluate the effect of bacterial isolates obtained from soil samples collected from Striga infested sorghum growing fields in Amhara, Tigray and Oromia Regions to inhibit (deplete) the Striga seed bank. Surface sterilized and conditioned Striga seeds were buried in the soil by using teabag to determine germinated, viable and decayed seeds at different times after burial. A total of 44 isolates were isolated from germinated and decayed striga seeds and tested for their effect on germination and seed decay of Striga seeds. Six isolates (GS29, GS32, GS34, GS39, GS42 and GS45) were stimulated 10.33, 9.0, 9.67, 8.33, 5.33 and 8.33 Striga seeds germination from the total of 35 seeds respectively in the absence of host plant synthetic stimulant. On the other hand four isolates (SD3, SD9, SD36 and SD46) significantly decayed Striga seeds at P<0.05 (21, 20, 21.33 and 21 seeds respectively) in order to reduce Striga attack. Selected isolates were characterized by using biochemical tests, and three isolates were classified under the genus Pseudomonas while the other three isolates were grouped under the genus Klebsiella. The four isolates that showed seed decay were classified under the genus Bacillus.
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    Isolation and Characterization of Striga Hermonthica Seed Bank Depleting Bacteria from Striga Infested Sorghum Growing Areas of Northern and Eastern Ethiopia
    (Addis Ababa University, 2020-06-06) Yilma Getachew; Assefa Fassil (PhD)
    Striga hermonthica is a very devastative parasitic weed of (Sorghum bicolor (L.) Moench) which is the third important crop in Ethiopia. It is hemi-parasite living with the host plant by attaching small sucker root system to host plant. A study was undertaken to evaluate the effect of bacterial isolates obtained from soil samples collected from Striga infested sorghum growing fields in Amhara, Tigray and Oromia Regions to inhibit (deplete) the Striga seed bank. Surface sterilized and conditioned Striga seeds were buried in the soil by using teabag to determine germinated, viable and decayed seeds at different times after burial. A total of 44 isolates were isolated from germinated and decayed striga seeds and tested for their effect on germination and seed decay of Striga seeds. Six isolates (GS29, GS32, GS34, GS39, GS42 and GS45) were stimulated 10.33, 9.0, 9.67, 8.33, 5.33 and 8.33 Striga seeds germination from the total of 35 seeds respectively in the absence of host plant synthetic stimulant. On the other hand four isolates (SD3, SD9, SD36 and SD46) significantly decayed Striga seeds at P<0.05 (21, 20, 21.33 and 21 seeds respectively) in order to reduce Striga attack. Selected isolates were characterized by using biochemical tests, and three isolates were classified under the genus Pseudomonas while the other three isolates were grouped under the genus Klebsiella. The four isolates that showed seed decay were classified under the genus Bacillus.
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    Genetic Diversity Study of Kabuli and Desi Type Chickpea (Cicer Arietinum L.) Varieties Using Agro-Morphological Traits, Nutritional Composition and Issr Marker
    (Addis Ababa University, 2020-05-05) Ahmed, Ansha; Dagne, Kifle (PhD); Fiker, Asnake (PhD); Tessfaye, Kassahun (PhD)
    Genetic diversity study of Kabuli and Desi type chickpea (Cicer arietinum L.) varieties using Agro-Morphological Traits, Nutritional Composition and ISSR Marker Ansha Ahmed. MSc. Thesis Addis Ababa University, January 2020 Chickpea (Cicer arietinum L.) is one of the oldest and most widely consumed legumes in the world due to its relatively high protein content and wide adaptability as a food grain. In Ethiopia an average chickpea yield on farmer’s field is low. Even released varieties produce about 3.5t/ha in research area, although it’s potential yield is more than 5t/ha. In Ethiopia, no studies were conducted on chickpea varieties genetic diversity assessment based on biochemical (nutritional and anti-nutritional) analysis and this study is designed to fill this gap. The study was designed to reveal the genetic diversity existing among Ethiopian chickpea varieties using phenotypic traits, biochemical analysis and ISSR markers. Field experiment was conducted at two environments’ (irrigation, main seasons) for agro-morphological performance, while the molecular part was conducted at Addis Ababa University (ISSR) and biochemical experiment was conducted at Holeta Agricultural Research Center and Ethiopian public Health Institutions. A total of 28 chickpea varieties were used for biochemical and ISSR markers, whereas 7 varieties were used for phenotypic traits. A total of 10 phenotypic traits, 9 traits for biochemical 5 ISSR primers were used to assess genetic diversity and interrelationship among Ethiopian chickpea varieties. The genomic DNA extraction was done based on modified CTAB extraction methods. The combined analysis of variance over the two seasons showed highly significant (p<0.01) variations among the varieties for the majority of traits. Genotype by environment interaction showed highly significant difference (p< 0.01) among traits. UPGMA constructed based on phenotypic traits revealed four major clusters and the first three principal components (PCs) accounted for 93.7% of the entire diversity among the varieties for all the 10 t0raits and the first two PCA revealed 73% of the total variation. UPGMA constructed based on nutritional and anti-nutritional traits revealed three major clusters and the five principal components (PCs) accounted for 90% of the entire diversity among the varieties for all the 9 traits and the first three PCA revealed 67% of the total variation. Five ISSR primers amplified 54 bands, of which 47 loci were polymorphic. The highest gene diversity (0.38) and Shannon index (0.55) were recorded by primer 889, while the least gene diversity (0.22) was revealed by primer 824. Therefore, the observed genetic variation in the study indicated the opportunity of using these materials in future Chickpea breeding program via introgression with other germplasm resources for improvement of better genotypes.