Biotechnology

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Alkaline Protease Production by an Alkaliphilic Bacterial Isolate Under Solid State Fermentation
(Addis Ababa University, 2009-08) Haile, Gizachew; Gessesse, Amare(PhD)
A total of 240 alkaliphilic microorganisms isolated from samples collected from alkaline soda lakes of Ethiopia were screened for the production of alkaline proteases. Of these, 30% were protease positive indicating the abundance of protease producing microorganisms in these habitats. This again is a reflection of the abundance of protein substrates in the form of bird`s feather and left over from dead cells of spirulina and other microorganisms. Out of the 80 protease positive isolates, 20 (25%) grew well and produce appreciable level of enzyme activity when grown in solid state culture. Of these, one isolate designated as C45 was selected for further study. The protease produced by isolate C45 was characterized to determine its potential industrial application. The enzyme was active in the pH range of 6.5-11.5, with optimum activity at pH 8-9; and stable at alkaline pH. The optimum temperature for activity was 40°C and 50°C in absence and presence of 5mM of Ca+2, respectively. The enzyme displayed appreciable activity and stability at low temperature. These properties suggest that protease C45 could find potential application for dehairing and detergent at moderate temperature. When protease C45 was added to raw hide enabled dehairing, suggesting the potential usefulness of the enzyme in the leather industry. The commercial application of enzymes greatly depends on the cost of the enzyme which again is determined by the production cost of the enzyme. Currently most commercially available enzymes are produced through capital intensive submerged fermentation (SmF). An alternative method for the growth of microorgianims which is currently receiving significant attention is solid state fermentation (SSF). In this study, isolate C45 was grown under solid state fermentation using wheat bran as the growth substrate. Maximum protease secretion was achieved at inoculum size of 20% (v/w), bran to moistening agent ratio of 1:2 when incubated at 30°C for 144 hr. Addition of inorganic nitrogen sources and organic carbon sources as a supplement of SSF medium repressed protease induction. These results indicate that the microbial isolate shows a good potential for production of low cost alkaline protease by using inexpensive substrate such as wheat bran alone and/or low cost complex nitrogen source such as Millettia ferruginea (Berbra) seed flour as supplement in SSF. Key words: Alkaline protease, isolate C45, solid state fermentation (SSF).
Antagonistic and Antibiotic Effects of Endophytic and Rhizobacteria of Sugarcane Plant at Wonji Sugar Estate, Ethiopia
(Addis Ababa University, 2013-06-06) Tilahun Sefinew; Lakew Mekuria (PhD); Assefa Fassil (PhD)
The thesis targeted to identify the endophytic, (leaf and root) and rhizobacteria of three sugarcane sample plants taken from different sites of Wonji sugar estates plantation. Accordingly, leaf and root samples cleared of surface bacteria by chemical sterilization, crushed using sterile mortar and pestle and the extract plated on nutrient agar media. The soil samples were also sieved and serially diluted from 10-1 -10-6 and 100µl of the solution spread plated on nutrient agar media. A total of 40 representative isolates were identified based on standard morphological characterization. These isolates were sub cultured for purity and preserved on nutrient agar slant for future use in different testes: Antagonism, Antibiotic effect and development of serological methods of screening of the isolates. The identification of the isolates was done in two steps. First, grouping of the isolates by morphological, physiological and Biochemical tests and second, sorting of the groups that had more than one genera in to the specific genus was done by Busnson’s method. The result showed 11 genera, the dominant of which were Shigella, Bacillus and Klebsiella. Antagonstic interactions between all the isolates revealed 22 of them were susceptible to 18 isolates that had inhibitory effects. The test for antibiotic effect of isolates against four human pathogenic bacteria: E. coli, Salmonella thyphimurium, Staphylococcus aureus and Streptococcus pneumonae showed 18 isolates with anti-biotic effects ranging from 10-17mm clear zone diameter. This was less sensitive compared to the positive control antibiotics, chloranphenicol and ampicillin. The serological method development for screening of the isolates was started by SDS-PAGE fractionation of the proteins and induction of antibodies in swiss albino mice shall be tested as soon as the conjugated antibodies arrive. The potential use of the identified isolates; in sustainable agriculture, environmental protection and animal and human benefits are discussed in detail.
Antimicrobial Properties of Endophytic and Rhizospheric Fungi Associated With Some Medicinal Plants
(Addis Ababa University, 2020-09-09) Shemsedin, Fertuna; Muleta, Diriba (PhD)
Nowadays the development of multidrug resistant human pathogenic microorganisms and the emergence of new diseases are the most challenging problems in public health care on a global scale. To overcome this problem it needs intensive searching for new sources of effective antimicrobial agent producing organisms. Therefore, the main objective of this study was to isolate and identify the antimicrobial properties of entophytic and rhizospheric fungi associated with some medicinal plants. A total of 150 plant parts and 50 soil samples were collected from five medicinal plants around Bale Zone, west Arsi Zone and Chancho Oromia Special Zone, Oromia Regional State of Ethiopia. All collected samples were processed following standard protocols. In vitro antimicrobial activities were tested against common resistant pathogenic organisms (Escherichia coli, Staphylococcus aureus, Enterococcus faecalis, Pseudomonas aeroginosa and Candida albicans). A total of 582 (316 endophytes and 266 rhizospheric) fungal isolates were obtained from the collected medicinal plants and soil samples. Accordingly, 78 (19.89%) isolates displayed antimicrobial activities against at least one target microorganism by fungal agar plug method. The ethyl acetate extracts of the crude metabolites of 18 isolates, showed antagonistic activity against at least one tested organisms with higher inhibition zone. Ethyl acetate extracts of isolate 30CRS showed highly significant (p≤0.001) inhibition zone against E. coli (30.33+0.57 mm), E. faecalis (25.33 + 0.28 mm) and S. aureus (19.16+ 0.28 mm) than positive control chloramphenicol whereas fungal isolate 37BRaL showed significantly (p≤0.001) higher inhibition zone against S. aures (19.16+ 0.28 mm) and C. albicans (26.83 + 0.76 mm). The mean MIC, 3.125 - 50mg/ml for gram positive bacteria, 6.25 - 50 mg/ml for gram negative bacteria and 12.5 – 50 mg/ml for yeast test organism. MBC 6.25-50mg/ml and MFC ranged from 12.5-50 mg/ml. The phytochemical screening of the fungal metabolite revealed the presence of flavonoids, alkaloids, glycosides, terpenoids, steroid, saponin, phenol, and tannin. A total of five potential fungi were examined by morphological characterization and Biolog identification, from this, isolates 30CRS and 37BRaL were identified as P. simplicissimum and T. flavus var flavus, also characterized for different Biolog carbon source utilization test using Biolog microbial identification system.
Antimicrobial Resistance Profile of Escherichia Coli Isolated from Hospital Sewage and Polluted River: the Cases of Adama Hospital Medical College in Adama and Yerer River in Dukem
(Addis Ababa University, 2024-11) Kajelcha Fikadu; Alemayehu Godana
The antimicrobial resistance profile of Escherichia coli is dramatically increasing across the world, particularly in low- and middle-income countries where untreated wastewater can disseminate resistant bacteria into the environment. Untreated wastewater discharged from hospitals and industries is the most known vehicle for the emergence and spread of antimicrobial resistance genes. The study aimed to evaluate the antimicrobial resistance profile of E. coli isolates from hospital sewage and polluted river samples in Adama and Dukem, respectively. The hospital sewage and polluted water samples were collected from Adama Hospital Medical College and two rivers in Dukem. The samples were transported to Addis Ababa University, Institute of Biotechnology Laboratory. For the purpose of isolating E. coli, the samples were cultivated on Eosin Methylene Blue agar, MacKonkey agar, and Nutrient agar media. Biochemical tests, including Gram staining, IMViC, TSI, and SCA, were used for the identification of E. coli isolates. The researchers evaluated the antimicrobial susceptibility patterns of the E. coli isolates using the Kirby-Bauer’s disc diffusion method. The identified E. coli isolates were then cultured on Mueller Hinton agar media at 37°C for 24 hours, after which the inhibition zone was measured by a digital ruler. Polymerase chain reaction (PCR) was used to confirm the presence of resistance-associated genes in the E. coli isolates. A total of 75 presumed E. coli isolates were obtained via culture. Among these isolates, 50 isolates were subjected to AST. The isolates from Yerer River before the industrial waste entry site (YRBI) did not show multiple antimicrobial resistance (MAR), while the isolates from Xadacha River (XR) and Yerer River After Industrial waste entry site (YRAI) showed resistance profiles of 66.67% and 90%, respectively. This study also revealed that 86.67% of hospital sewage isolates and 65% of river water isolates showed MAR. PCR amplification confirmed the presence of tetA and blaTEM genes in 83.33% and 57.14% of the AMR isolates, respectively. In conclusion, the study showed that untreated hospital sewage and pollute river water are considered major reservoirs for the emergence of antimicrobial-resistant E. coli among humans, animals, and the environment. The study advocates for improved wastewater treatment, stringent regulations on antimicrobial use, and regular monitoring to curb the emergence and spread of antimicrobial resistance.
Assessing the Prevalence and Causes of Sport Injuries in Selected Ethiopian Premier League Football Clubs.
(Addis Ababa University, 2018-07-02) Lemma, Admasu; Taddese, Aschenaki (PhD)
The purpose of this study was to assess and evaluate the prevalence and causes of sport injures in selected Ethiopian premier league football clubs. For the implementation of the Study, a descriptive survey design was employed. The study subjects were selected from the availability samples. In this attempt, data were collected through questionnaires, structured interviews and observation check list. Consequently, the study demonstrated that the selected Ethiopian men’s premier players ,coaches physiotherapists and technical staffs contributed for the study to be conducted by giving their own views regarding the raised issues were selected on the base of their own voluntary cooperation. The total number of players is 90 of these the researcher selected 3 of the clubs (i.e. 10× 3=30 One club has an average of 10 players). Of this total population, the researcher has taken 3 male Ethiopian male premier football clubs from this 30 male players selected from 3 Ethiopian male premier league football clubs by using Simple random sampling technique was used to select trainees. And, 6 club coaches, 3 physiotherapists and 3 club technical staffs also purposeful sampling technique was applied to gather data. They all, owing to their limited and manageable size, have been taken as a sample study subjects. The study also paid a thorough consideration to the players, coaches’ physiotherapists and for the club technical staffs should contribute for the preventing prevalence and causes of sport injuries as a whole. Moreover, the players awareness about sport injuries ,the coaching system ,and roles of administrative bodies to work cooperatively, the facilities provided, the roles of coaches, lack of un proper using of protective sport equipment’s including shin guards, warm up programs, attention to environmental conditions, first aid and rehabilitation, all other related issues are taken as focal point of the study to achieve these objectives and to detect the factors that hindered the concerned bodies the researcher included male players, coaches, physiotherapists and technical staffs and encouraged them all to involve themselves in the issues raised in terms of the prevalence and causes of sport injuries Their genuine responses are collected and used as analytical framework for the effective implementation of the study.
Assessment of Diversity, Morphological Variation and Description of Grasspea (Lathyrus Sativus) and Other Related Species
(Addis Ababa University, 2007-03) Tsegaye, Martha; Demissew, Sebsebe (Professor); Jorge, Alexandra (PhD)
Lathyrus sativus (grasspea) has been widely cultivated in South Asia and Ethiopia for over 2500 years and is used as a food and feed. It is rich in protein content, around 30 g/100 g edible seeds. Agronomically, the species is able to withstand both severe drought as well as water logging. Although seeds of grasspea are tasty and protein rich, excessive consumption of the seeds causes a motor neuron disease called neurolathyrism which is characterized by the paralysis of the lower limbs. The neurotoxic causal agent of this disease is believed to be a non protein aminoacid called Oxalyl Di aminopropionic Acid (ODAP). Morphological marker analysis and molecular analysis have been used widely to estimate genetic variability of populations. These methods have useful in addressing questions on population genetic structure and genetic conservation. Knowledge of genetic diversity of species is particularly important, since modern breeding practices have narrowed the genetic diversity of cultivated crops. In the case of grasspea, the problem of Lathyrism is leading to the banning of its production which in turn aggravates genetic erosion and loss of diversity of the crop. Fifty one grasspea accessions which were selected from the genebank collection of International Livestock Research Institute (ILRI) were evaluated and characterized for different qualitative and quantitative morphological characters. Cluster analysis was performed to estimate differences between accessions. Randomly Amplified Polymorphic DNA (RAPD) was also used to study the nature of variation. In addition to L. sativus, three other species of the genus (L.cicera, L. clymenum and L. ochrus) and seventeen unidentified populations of Lathyrus were also evaluated for morphological and biochemical characters and characterized accordingly. Cluster analysis of both the morphological and the RAPD data showed that all of the unidentified Lathyrus populations were found to be L. sativus. The result also showed that two of the accessions (5295 and 5296) represented by L. ochrus and one accession (5282) represented by L. cicera were found to be L. sativus. The results would suggest that germplasm evaluation is important for proper characterization of populations. Key words: Lathyrus sativus, grasspea, genetic diversity, morphological characters, RAPD, cluster analysis
Bioethanol Production Via Fermentation of Waste Substrates Using Yeast Isolates
(Addis Ababa University, 2019-05-05) Tsegu, Getu; Tesfaye, Anteneh (PhD)
Bioethanol is one biomass derived platform molecules which has a potential to be a sustainable feedstock for a variety of commodity chemicals. The aim of this study was to produce bioethanol from grain waste flour, tella spent, pineapple and papaya peels. Dilute sulfuric acid was used to hydrolyze polysaccharide in the raw materials. Starch content in grain waste flour and reducing sugar concentration in the hydrolysates of each raw material was determined. Indigenous yeast was isolated from areki difdif and tej samples. Indigenous yeast isolates with high fermentation efficiency and high ethanol producing ability were used to ferment hydrolysates in each of raw material to bioethanol. The results indicate that pretreatment of grain waste flour, papaya peels, pineapple peels and tella spent with 2% sulfuric acid at 110oC for 90 minutes yielded 29.7–30.8, 28.2–28.9, 16.2–16.6 and 10.3–10.5% of reducing sugar, respectively. The yeast isolate 1T-10 from tej samples was efficient in converting the D-glucose to ethanol (75.58%) and produced highest yield of ethanol (7.72%) from 20% D-glucose compared to other yeast isolates. Fermentations of grain waste flour with 1T-10 produced the highest yield of ethanol 1.380% (w/v) from pretreatment of substrates with 2% sulfuric acid at 168 hours of fermentation time compared to other combinations or 0% at (72, 120 and 168 hours), 1% at (72, 120 and 168 hours), 1.5% at (72, 120 and 168 hours) and 2% at (72 and 120 hours). Similarly, fermentation of pineapple peels, papaya peels and tella spent produced the highest yield of ethanol; 7.883%, 6.400% and 4.813% (w/v) with pretreatment of 2% sulfuric acid at 120, 72 and 72 hours of fermentation time, respectively compared to other combinations when fermented with the same yeast isolate (1T-10). It was concluded from the present study that it is possible to produce economically viable bioethanol by digesting waste and cheap resources like grain waste flour, tella spent, pineapple and papaya peels using indigenous yeast isolates.
Biological and Ecological Studies on Acacia Drepanolobium Harms Ex Sjöstedt in Borana Zone of Oromiya Regional State, Ethiopia
(Addis Ababa University, 2003-06) Maryo, Melesse; Nemomissa, Sileshi (PhD); Kelbessa, Ensermu (PhD)
The biological and ecological studies of A. drepanolobium: the floristic composition in A. drepanolobium wooded grassland, soil properties, seed production, seed dispersal, soil seed bank, percent seed germination at different treatments, capacity of coppicing , and its interactions (symbiosis) with ants, insects and microbes were investigated in four A. drepanolobium wooded grassland sites in Negele Borana , Oromiya Regional State, S. Ethiopia. The results indicated that in A. drepanolobium wooded grassland 114 plant species were identified. Of these 70.2 %, 23.7% and 6.1% were herbs, trees/shrubs and climbers respectively. More over, 33.3 % were forage species whereas 14.4% and 2.6% had socioeconomic and medicinal importance respectively. Asteraceae, Fabaceae and Poaceae have constituted 36 % of the total number of species. The number of species was found to be smaller than previous studies on non- A. drepanolobium wooded grassland of the study area. This may suggest the impact of bush encroachment by A. drepanolobium, which had a mean density of 1798 plants/ hectare with a large number of individuals at the younger stage. The soils studied had higher proportion of clay (> 30 %) with properties that favor the growth of most plant species. An average of 2417 ± 23 (X ± SE) seed production per plant was encountered, and only 1 ± 0.4 (X ± SE) trees bore seeds in average per plot. Seeds are mainly dispersed by wind. 267 seeds (8.3 ± 2.6 seeds /m 2) were found only at the litter layer and none in mineral soil layer. There was statistically significant difference in percent germination among treatments [F (5, 17), P < 0.05]. Fast rate and higher percent germination was achieved by scarification treatments whereas dry heat treatment (90 oC) and moist heat (98 oC for greater than 30 minutes) resulted in almost all mold outgrowths after a week’s period. Tukey’s HSD indicated that moist heat treatments didn’t improve the percentage germination. High percent germination of a control experiment within week’s time may suggest the absence of pronounced seed dormancy in the study species. There is no statistically significant difference among stumping treatments both in number and in height of coppice but the coppice number and height increased down to a tree height (soil surface). Four A. drepanolobium occupant ant species (3 Crematogaster and 1 Tetraponera species) were identified. However, a black cotton soil habitat hosted only 2 Crematogaster species. Although each tree was occupied by a single ant species, Crematogaster mimosae occupied the largest proportion (85%) of seed bearing trees. The mutualistic association of Crematogaster nigriceps is doubtful because this species sterilizes flower buds and new shoots. Two seed feeding bruchid beetles (Callosobruchus maculatus and Acanthoscelides obtectus) were identified and found to reduce the reproductive vigor of the study species by mass predation of its seeds. A. drepanolobium was found to be nodulated by slow growing rhizobia called Bradyrhizobium species. There was no statistically significant difference in nodulation status between two soils (t at 24 df = -1.22 and P=0.268). The mean nodule number and weight were 4.93 ± 0.6 and 0.00381 ± 0.0008 (X ± SE) in clay soils respectively. Reduction in nodulation may be due to 1) richness of clay soil in mineral elements including nitrogen 2) absence of adequate aeration 3) missing of some nutrients elements such as molybdenum and 4) Slowness of the fixer species. From the socio-economic view point, 50% of informants declared the importance of A. drepanolobium, and the rest expressed their hatred for its bush encroachment impacts on their surrounding. However, from the present study , it can be suggested that sterilization of flower buds and young shoots by Crematogaster nigriceps ants, low soil seed bank, seed predation by bruchid beetles and low recruitment being the limiting factors, further expansion of bush encroachment by A. drepanolobium can be managed through integrated bush management systems such as reducing cultivation of dry season grazing areas, encouraging traditional rangeland management systems and applying proper land use policy, reducing excess livestock, stumping late in rainy seasons and periodic burning though complete recovery of the previous range condition is a difficult task. Key words: A. drepanolobium, bush encroachment, seed characteristics, Crematogaster, bruchid beetles.
Characterization of Diarrheagenic Escherichia Coli Strains Isolated From Dairy Calves in Fiche, Debretsige and Muketuri Towns; North Shoa, Ethiopia
(Addis Ababa University, 2018-06-03) Endale, Abinet; Sisay, Tesfaye (PhD)
Diarrheagenic E. coli (DEC) strains are associated with several outbreaks and sporadic cases worldwide. Understanding the nature of the organisms is important in order to tailor preventive and control strategies before any harm emanates. Thus, the distribution of these organisms among humans as well as animals should be studied, which was the major objective of the current study. In this study a total of 73 E. coli samples isolated from feces of dairy calves found in 56 farms located in Fiche, Debretsige and Muketuri areas were characterized. Isolates were assessed and characterized based on their virulence gene and plasmid content. Moreover, the study tried to show the relationship between virulence gene, plasmid content antibiotic resistance traits. The study identified the occurrence of virulence genes in 69.86% of isolates, carrying at least one of the six VGs screened. Among the VGs eae was most frequent, observed in 58.90% of isolates, followed by stx1 34.25 %, stx2 24.66%, ehlyA 23.29%, bfpA 12.33% and aatA 5.48% observed in the total sample isolates. A total of eleven distinct virulence profiles were identified based on combination of virulence genes and isolates were placed into five different pathotypes with a frequency of EHEC 30%, tEPEC 12%, aEPEC 16% and 6% of STEC and EAEC each. Plasmid analysis on the other hand revealed the occurrence of ten different kinds of plasmids ranging in size from 2.6 + 0.14 Kbp to 98.2 + 4.17 Kbp and exhibited 13 different kinds of distribution profile among isolates. In an effort made to observe any possible association among virulence factors, plasmid content and antibiotic resistance traits, a significant one to one correlation was observed. a Pearson product moment correlation in a range of r = 0.17 to r = 0.56. Accounting to the total dataset, isolates showed significant segregation in to the sample site, indicating spatial clustering of isolates based on overall pathogenic characteristics. The demonstration of pathogenic potential in substantial amount of E. coli isolates from dairy farms from all the three sites indicate the level of risk posed in humans as well as animals. Thus, identifying potential sources and route of transmission of DEC is vitally important in order to establish control and prevention strategies for DEC infection. Moreover, nationwide screening for virulence factors and antibiotic resistance genes and associated plasmids is recommended in order to prevent the spread of these pathogens among various sources.
Characterization of Wild Indigenous Yeasts from Molasses and other Sugary Substrates and their Potential for Bioethanol Production
(Addis Ababa University, 2021-05-09) Degu, Sisay; Muleta, Diriba (PhD); Tesfaye, Anteneh (PhD)
The increasing demand of energy has been supplied through the combustion of petroleum throughout the world. Due to escalating cost of petroleum, its contribution to global warming and non-renewable nature of this oil, there is collectively a need for renewable and ecofriendly energy sources such as bioethanol. Thus, the objective of this study was aimed at isolating, characterizing and evaluating the potent wild yeasts under different stress conditions from locally available resources for bioethanol production to minimize the utility of fossil fuel. In this study, a total of 35 samples of sugary substrates were collected from Metehara Sugar Factory for wild yeasts isolation following the standard protocols. A total of 305 yeast isolates were retrieved and screened using physiological and osmotic stress tolerance tests. Fermentative and potent wild yeasts were identified to species level using morphological and biolog-based biochemical methods. Out of 305 yeast isolates, 20 (6.56%) and 7 (2.29%) of them were found to be tolerant to 18 and 20% of ethanol, respectively. Out of these 20 ethanol tolerant yeast isolates, 17 tolerated the temperature of 45°C for 48 hrs. From 17 ethanol-thermotolerant isolates, 5 (29.41%), 5 (29.41%) and 7 (41.18%) were found producing gas from glucose at 24, 48 and 72 hrs, respectively. Out of the 17 ethanol-thermotolerant yeast isolates, 12 yeast isolates were able to tolerate 35% of glucose. Out of these 12 ethanol-thermo-and-sugar tolerant yeasts, the 7 were found tolerant to pH 2. From the 7 acidic tolerant yeast isolates, 5 yeast isolates were shown tolerant to 7% of NaCl and identified as K. lodderae, P. guilliemodii B, S. boulardii, Z. rouxii and T. globosa, respectively. During fermentation, K. lodderae, P. guilliemodii B, S. boulardii, Z. rouxii and T. globosa were able to produce bioethanol with the values of (% v/v) 12.62, 11.61, 10.58, 10.82 and 10.44, respectively at 48 hrs and 12.56, 12.52, 12.08, 11.11 and 11.48, respectively at 72 hrs. The initial inoculum cell density was increased from (1.83 to 3.08, 0.59 to 1.81, 0.54 to 1.36, 0.48 to 1.69 and 0.45 to 1.56) x108 cells/ml for K. lodderae, P. guilliemodii B, S. boulardii, Z. rouxii and T. globosa, respectively 0 hr to72 hrs. The fermentation efficiency (%) for K. lodderae, P. guilliemodii B, S. boulardii, T. globosa and Z. rouxii was shown as 98.0, 97.8, 93.1, 90.8 and 88.1, respectively. Thus, on the basis of highest stress tolerant and higher fermentation efficiency features, K. lodderae and P. guilliemodii B wild yeasts were considered to be the best bioethanol producers. Molecular characterization and optimization of fermentation parameters is recommended to utilized these potent yeasts.
Characterization of Wild Yeasts Isolated from Selected Fruits for their Bread Leavening Capacity
(Addis Ababa University, 2017-06) Lakew, Eshet; Muleta, Diriba (PhD); Tesfaye, Anteneh (PhD)
Leavening agents are important in raising flour dough. Biological leavening agents are microorganisms that have the ability to produce carbon dioxide from the utilization of Sugar and thereby ferment and raise the dough. The present study was carried out to characterize yeast isolates isolated from selected fruits and to assess their leavening potential of wheat dough under laboratory scale. The collected fruit samples were processed to isolate yeasts using Potato Dextrose Agar (PDA) amended with 0.1 g/L chloramphenicol. Initially, 88 yeasts were isolated from the fruits and were first tested for their carbohydrate fermentation in yeast extract peptone dextrose (YEPD) broth medium. Six yeast isolates with their sugar fermentative abilities were selected and tested for H2S production. Among them, AAUGr5, AAUOr7 and AAUPi3 found not produce undesirable H2S for bread baking quality on both Kligler Iron Agar (KIA) and Bismuth Sulfite Agar (BSA) media. The three yeast isolates were identified as Saccharomyces using colonial, morphological parameters and biochemical tests. The optimum growth pH and temperature values for the three selected yeast isolates were recorded as 5 and 30 oC, respectively, in YEPD medium. In addition, 30% (w/v) D-glucose and 5 %( w/v) NaCl concentrations showed optimum growth of the three selected yeast isolates in yeast extract peptone broth medium. In all the cases, the maximum biomass was achieved at 96 hrs of incubation and there was a rapid decrease in biomass for all the yeast isolates after 96 hrs of incubation. In terms of CO2 and biomass production as well as leavening potential, starter cultures which were formulated from the combination of the three yeast isolates (AAUGr5+AAUOr7+AAUPi3) showed better performance than starter cultures formulated from paired combination of the three isolates or each of the three isolates separately. However, isolate AAUGr5 was found to be satisfactorily potent for leavening action from the single isolates. The present study could therefore be important with respect to screening of wild yeast isolates that possess better bread leavening potential for extending the use of indigenous microbes as starter culture in bakery sector. Keywords/Phrases: biomass; carbon dioxide ;fruits, hydrogen sulphide; laboratory scale leavening; yeast isolation
Chloroquine Resistance Transporter (Pfcrt) and Multidrug Resistance 1 (Pfmdr1) Genes Mutation in Plasmodium Falciparum Population Under Varying Level of Endemicity with Plasmodium Vivax in Selected Parts of Ethiopia
(Addis Ababa University, 2023-02) Tajudin Abdurahman; Alemayehu Godana
The global controlling of Plasmodium falciparum infections faces significant challenges due to the spread of parasites resistant to antimalarial drugs. In Ethiopia, where both P. vivax and P. falciparum coexist, the treatment for uncomplicated falciparum malaria shifted from chloroquine (CQ) to sulfadoxine-pyrimethamine (SP) in 1998 and then to Coartem (artemether-lumefantrine (AL)) in 2004. AL has been the standard treatment for over two decades for P. falciparum, while P. vivax is still treated with CQ. The coexistence of these two species and the accessibility of CQ for P. vivax treatment raise questions about whether switching from CQ to AL for P. falciparum treatment might lead to the resurgence of CQ -susceptible P. falciparum strains due to reversal mutations or the efficacy of AL. The sudy aimed to assess the prevalence of pfcrt-76 and pfmdr1-86 gene mutations in the P. falciparum population under varying levels of endemicity with P. vivax in selected parts of Ethiopia. In this study the frequency of gene mutations of P. falciparum chloroquine resistance transporter 76 (pfcrt-76) and P. falciparum multidrug resistance 1-86 (pfmdr1-86) in P. falciparum collected from malaria-infected patients in Abobo, Dera, Fentale, and Metema districts, Ethiopia, were examined. Confirmed P. falciparum samples (n = 258) with microscope were collected through health facility-targeted cross-sectional surveys in areas with varying levels of P. falciparum and P.vivax prevalence. Genomic DNA was extracted from dried blood spots by using MagMAX DNA Multi-Sample Kit, operated by the Kingfisher Flex Automated Extractor. An 18S rRNA gene-based multiplex real-time PCR assay was employed to confirm P. falciparum mono-infection samples (n = 258). The study analyzed 258 P. falciparum-infected patients using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), examining Pfmdr1-86 and Pfcrt-76 gene mutations. Fisher's exact test determined marker distribution significance. Out of 250 genotyped for Pfcrt K76T, 68.8% had mutant 76T, and 31.2% were wild-type K76. For 257 genotyped for Pfmdr1-N86Y, 98.44% was N86 wild-type, and 1.56% was 86Y mutants. The mutant Pfcrt-76T was more prevalent in areas with higher P. vivax endemicity, including 93.33% (Fentale), 84.71% (Dera), 57.5% (Metema), and 43.75% (Abobo). Pfcrt-76 gene single nucleotide polymorphisms (SNPs) significantly correlated with P. vivax endemicity (P = 0.000). Pfmdr1-N86 was predominantly 100% except in Abobo, where 95.18% were N86, and 4.81% were 86Y. Pfmdr1 gene SNPs were significantly associated with P. vivax endemicity (P = 0.025). Despite CQ discontinuation for over two decades in Ethiopia, a substantial proportion of P. falciparum isolates still carry mutant 76T genotypes, indicating latent CQ pressure. The use of AL for uncomplicated P. falciparum malaria may lead to the return of Pfmdr1 N86 wild-type genes. Further molecular epidemiological investigations in varied endemic regions with different CQ usage histories are recommended to understand chloroquine (CQ) susceptibility recovery and AL therapy efficacy.
Chromatic Polynomial
(Addis Ababa University, 2018-05-07) Tamiru, Abebu; Tsegaye, Yirgalem (PhD)
In this project, based on articles published by(Coudy Fouts), we see how Incidence Algebra in particular, M obius Inversion Theorem is used to compute the Chromatic Polynomials of graphs. This is one of the various applications of M obius Inversion Theorem.
Comparative in Silico Study of Lodging-Resistant Genes of Tef (Eragrostis Tef (Zucc.) Trotter) Against Wheat, Barley and Rice
(Addis Ababa University, 2024-07) Tigist Shewafera; Abiy Zegeye
Tef (Eragrostis tef), a cereal crop indigenous to Ethiopia, is essential in ensuring food and nutrition security. It is well-known for its gluten-free properties, rendering it suitable for individuals with celiac disease. However, tef is highly susceptible to lodging that negatively impacts its productivity. Efforts to develop varieties that are lodging-resistant have had limited success; particularly, because there have been a dearth of targeted genetic interventions. The aim of this study is to computationally predict and characterize six lodging-resistant genes (Rht1, BRI1, COMT1, CAD8C, CESA1 and CESA4) and the corresponding polypeptides of tef vis-à-vis three economically important cereal crops. The study began by selecting three economically important crops, namely: wheat, barley and rice and their six lodging-resistant genes. Using in silico probes, six homologous candidate lodging-resistant genes were retrieved from tef subgenomes. The candidate tef lodging-resistant nucleotide sequences were analyzed to computationally predict their gene structures and then comparatively analyzed against their homologues in the selected three crops. Putative polypeptide sequences of the six lodging-resistant genes, derived from their obtained predicted tef genes, were subjected to further amino acid level analysis to identify variations unique to tef, map the variations to specific functional or structural domains, predict the potential effects of the said variations and examine the possible impact of significant variations on the 3 dimensional structure of the proteins. Multiple sequence alignment of polypeptide sequences revealed distinct amino acid differences unique to tef in key functional and structural domains encoded by these lodging-resistant genes, with potential implications for tef's lodging susceptibility attributes. Variant effect predictions of counterfactual amino acid variations point to a subset of the unique tef variations having significant likelihood of causing damage in four of the six proteins: Rht1, BRI1, COMT1 and CESA1. Further examination of significant variation effects on the 3D structures of the four proteins revealed possible effects on the secondary structure of the polypeptides. Overall, the findings provide insights into several of tef's genetic variations related to lodging resistance features. Further experimental validation studies are needed to confirm these findings and elucidate the precise roles of these genes in tef's lodging susceptibility.
Deciphering the Conserved Cis-Regulatory Elements of Major Milk Protein Genes by Computational Analysis
(Addis Ababa University, 2024-04) Addis Tekaw; Abiy Zegeye
Milk genes are exclusively expressed in MECs during lactation, regulated by lactogenic hormones like prolactin that act through specific TFs. The recruitment of TF is determined by cis-regulatory motifs in their gene promoters; but there is limited evidence of shared motifs among the major milk gene across species. In this study, Jalview, Meme-Suite, TomTom, and GOMo software were utilized to construct phylogenetic trees, discover motifs, identify TFs, and determine GO terms, respectively, in the 2kb upstream putative promoter region of CSN1S1, CSN1S2, CSN2, CSN3, LALBA and BLG milk genes among twenty-three mammalian species. The analysis revealed three common TFBSs for STAT1, STAT5a, and STAT5b TFs in all milk genes across the species studied, except for BLG, within the region -390 to -80bp from the canonical TSS, with a few shared TFBSs located in the distal promoter region upstream of -600bp. STAT3 was also detected in CSN1S1, CSN2, CSN3, and LALBA genes, sharing binding sites with STAT1, STAT5a, and STAT5b. Furthermore, TFBSs for Sox9 and Sox6 TFs were found to be shared between the LALBA and BLG putative promoters within -400 to -100bp. Moreover, GO analysis linked GPCR with regulatory motifs in all six milk genes across 23 species, essential for enhancing STAT5 phosphorylation through Gαq activation in the JAK-STAT pathway. The totality of the result suggests that the relative abundance of STAT5 and STAT3 proteins may play a role in regulating the expression level of caseins and LALBA commensurate with the stage of lactation. Together, these results offer valuable insights into the potential of the milk genes’ promoter for constructing eukaryotic expression vectors and provide essential information for transgenic studies.
Detection of Selection Signatures Breed-Specific SNPs and Linkage Disequilibrium Analysis in Ethiopian Indigenous and European Dairy Cattle Breeds
(Addis Ababa University, 2021-11-20) Dejene, Genet; Sisay, Tesfaye (Professor)
Ethiopia ranks first in Africa with the largest number of cattle populations adapted to diverse environments. Detection of selection signatures and assessment of linkage disequilibrium enable us to asses genetic diversity and genomic region under positive selections. However, in Ethiopian indigenous cattle there is few studies regarding their origin, divergency and genetic adaptability. This study investigated detection of selection signatures, breed-specific SNPs and linkage disequilibrium in Ethiopian cattle populations and European dairy breeds. A total 135 animals representing four Ethiopian indigenous cattle populations: Arsi (n = 29), Begait (n = 40), Boran (n = 40) and Sheko (n = 26) were genotyped with 80K SNP chip. Two European dairy breeds (Holstein, n = 60 and Jersey, n = 38) were used for comparison. The mean of minor allele frequency (MAF) 0.32 ± 0.12, 0.32 ± 0.12, 0.31 ± 0.13, 0.30 ± 0.13,0.19 ± 0.17 and 0.18 ± 0.17 for Arsi, Begait, Boran, Sheko, Holstein and Jersey, respectively. The common variant MAF (≥0.10 and 0.5) distribution across Ethiopian cattle populations and European cattle breeds were 89% and 57%, respectively. Ethiopian cattle specific SNPs were located in genes(CSN2, ABCA7, PDE4B, ABCA1, JAK2, B4GALT4, FOXO3, GHR, ADCY8, ACACB) associated with milk production traits, fertility and growth traits (PGR, GHR, XKR4, ADCY5, POU2F1, IGF1, ABCC2, XKR4) thermo-tolerant, coat color (HSF1, HSPH1, HSPA4, KDR, RAD50, WNT1, KIT), feed intake (XKR4, ACCN1, ACAD11) and fat thickness (XKR4, IGFBP-3 and POU2F1).The top 1% fixation index (Fst) values representing positive selection harbored candidate genes (ABCG2, ABCA7, B4GALNT1, GHR, ITGAV) involved in milk traits such as, milk protein, milk fat and mastitis, milk production and (HSPH1, HSPA4, SOD1, MATR3, RAD50, KDR) for tropical adaptation. The estimated observed heterozygosity (Ho) and expected heterozygosity (He) for Ethiopian cattle populations were found to be, 0.40 and 0.39 respectively. These values were 0.24 and 0.25 for European dairy cattle breeds respectively. Principal component analysis (PCA)clearly separated Sheko from Ethiopian zebu populations with the value of 4.34%(PCA1) and 3.33% (PCA2). Similarly, PCA1 and PCA2 accounted for 65.97 % and 9.50 % of the variation and differentiated bos indicus from European taurine. Additionally, result of the phylogenetic tree analysis supporting PCA revealed that with the exception of Sheko, Ethiopian cattle population were closely clustered and two European dairy breeds (Holstein and Jersey) were located in a clade with Sheko. The overall mean of r2 values were 0.22, 0.23, 0.23, 0.22, 0.24, 0.16, 0.16 in Arsi, Begait, Boran, Sheko, Holstein and Jersey, respectively. In broad, this study revealed that there are genomic regions under strong divergent selection harboring genes involved in milk production traits and in adaptation to tropical environment.
Determinants of Leather Export From Ethiopia: Application of Vector Error Correction Model
(Addis Ababa University, 2018-06-06) Mamo, Getnet
Determinants of leather export from Ethiopia. Application of vector error correction model. Getnet Mamo Addis Ababa, 2018 Leather manufacturing is one of the oldest industry globally and particularly in Ethiopia which has remained and sprung forward as an economically important sector in terms of engaging citizens intensively and in export business. The study is aimed to use a multivariate time series model which explains the determinants of leather export from Ethiopia using vector auto-regression (VAR) and vector error correction (VEC) model. The data used are quarterly observations from September 2000 to august 2016. The variables are value of leather export, export price of leather, consumer price index (endogenous variables) And nominal exchange rate (exogenous variables). The series are seasonally adjusted after they were known to be seasonal through standard tests built in X-12 ARIMA program in E-Views 6 statistical software. Post seasonal adjustment tests also assured that all series are non-seasonal. Unit root tests of the series under study reveal that all the series are non-stationary at level and stationary after first difference. The result of Johansen test indicates the existence of two co-integration relation between the variables and there is long-term dynamics between value of leather export, nominal exchange rate, export price and consumer price index. The three information criteria AIC, SIC and HQ recommended one lag length. Johnsen co-integration test indicated two long term equilibrium relationship occurred between variables. This immediately implied the legitimacy of vector error correction model (VEC) model of order one to be fitted than a pure VAR (1) model for time series data. The final result shows that a Vector Error Correction (VEC) model of lag one with two co-integration equations best fits the data. Export price of leather has a negative effect on value of leather exports. A one percent increase in a unit price of leather export will cause 5.82219 percent decrease in value of leather export in the long run. In the short run Exchange Rate has a negative effect on exports of leather as expected. .
Determination of Viral Titer and Symptom Severity Variations in Maize Chlortic Mottle and Sugarcane Mosaic Virus Infected Maize Genotypes in Ethiopia
(Addis Ababa University, 2018-01-03) Tesfu, Kalkidan; Haileselase, Teklehaimanot (PhD)
The co-infection of Maize chlorotic mottle virus (MCMV) and Sugarcane mosaic virus (SCMV) can cause maize lethal necrosis. Though some studies were conducted to determine the incidence of MCMV and SCMV in Ethiopia, the role of their synergistic interaction in enhancing the severity of MLND in Ethiopian maize genotypes received little attention. This study was, therefore, designed to determine the role of MCMV and SCMV interaction in enhancing the severity of MLND in eight Ethiopian maize genotypes. Symptomatic leaves suspected with MCMV and SCMV infection were collected from different parts of Ethiopia. Eight maize genotypes (Melkassa 1, Melkassa 2, Melkassa 4, MH 140, MH 130,MHQ 138, Jibat and CML 445) were also obtained from Melkassa Agricultural Research Center (MARC). The collected leaves were checked for MCMV and/or SCMV infectionby DAS ELISA method. RNA was extracted from each leaf sample using HiPurA plant and fungal RNA Miniprep purification kit. The extracted RNA was then confirmed by RT PCR. Symptom severity was also determined by standard scores. The mean MCMV titer was significantly higher in five of the maize genotypes (Melkassa4, MH140, MH130, CML445 and Jibat) infected by both MCMV and SCMV than those infected by MCMV alone (P<0.05 for all). In contrast, there were no significant differences in mean MCMV viral titer between MCMV and MCMV-SCMV infected genotypes in Melkassa 1 (P=0.860), Melkassa 2 (P=0.572) and MHQ138 (P=0.734). Although higher mean SCMV titer was found in all genotypes infected by SCMV than those co-infected by SCMV and MCMV, the difference was not statistically significant (P>0.05 for all).As compared to the other genotypes lower score symptom severity was observed in Melkassa 1 and Melkassa 2. This study, therefore, revealed that on the basis of viral titer and symptom severity used to determine disease response, Melkassa 1 and Melkassa 2 have high resistance to MLND. Moreover, MH 138 was also found to have higher resistance to increment in viral titer than the other genotypes. These resistant genotypes might serve as markers for further investigation of resistant maize genotypes in Ethiopia.
Development and Evaluation of Multiplex Pcr for Simultaneous Detection of Major Bovine Mastitis Causing Pathogens
(Addis Ababa University, 2018-02-02) Assefa, Yared; Sisay, Tesfaye (PhD)
Mastitis is the most frequently occurring and economically one of the most important infectious disease in dairy cows. In most clinical laboratories, identification methods are based on microbiological culturing of milk and biochemical tests. However, there are many limitations associated with microbiological culture. Therefore, alternative, non-culture- based diagnostic methods are needed. This study was conducted with the objective of developing and assessing the use of multiplex PCR as a tool to identify bacterial species commonly associated with mastitis viz; Staphylococcus aureus, Escherchia coli, Streptococcus agalactiae and Streptococcus dysagalactiae. Conventional techniques based on biochemical and physiological characteristics of bacteria were utilized to identify the bacterial species. Then multiplex PCR assays were optimized using species specific primers for simultaneous detection of bacterial species. For rapid and accurate diagnosis, DNA was isolated from milk samples and subjected to multiplex PCR. Based on multiplex PCR results, S. aureus (60%) was found to be the predominant bacteria detected from milk followed by S. agalactiae (40 %) and E. coli (25%). The detection levels of S. aureus and S. agalactiae were higher when using mPCR compared to the culture-based method. However, S. dysagalactiae was not detected by multiplex PCR. Diagnostic sensitivity and specificity of the optimized multiplex PCR were calculated as 80% and 99.9% respectively. Furthermore, there was a substantial agreement between both diagnostic techniques (kappa result (K= 0.7)). Overall the results suggest that the multiplex PCR assay employed in this study could be used as an alternative routine diagnostic method for rapid, sensitive and specific simultaneous detection of major mastitis agents.
Distribution and Genetic Diversity of Maize (Zea Mays L.) Associated Dna and Rna Viruses in Ethiopia
(Addis Ababa University, 2019-10-10) Guadie Demsachew; Abraham Adane (Professor); Tesfaye Kassahun (Professor)
Maize (Zea mays L.) is leading cereal crop in terms of production in Ethiopia. However, achieving its yield potential is hindered by a number of biotic and abiotic factors. To enhance its productivity, these limiting factors should be addressed. Among the biotic factors, more than 50 viruses can infect and cause disease on maize. Of these, the rapidly emerged maize lethal necrotic disease (MLND) and maize streak disease (MSD) were reported to have high yield loss potential. This study was initiated to survey maize fields, study associated virus diseases and examine their diversity in Ethiopia. A total of 284 maize fields from five major maize growing regions were surveyed in four survey missions between 2015 and 2017, of which 846 leaf samples with virus like disease symptoms were collected from 191 fields. Ninety three fields with no observable symptoms were exempted from sampling. Up to 100% disease incidence was recorded in the Benishangul-Gumuz, Oromia and South Nations, Nationalities and People (SNNP) regions. In the first set of test experiment double or triple antibody sandwich enzyme-linked immuno-sorbent assay to test for eight common maize viruses confirmed the presence of Maize chlorotic mottle virus (MCMV), Sugar cane mosaic virus (SCMV) and Maize streak virus (MSV). MLND was the most important disease in SNNP and Oromia while MSD was predominant in Benishangul-Gumuz. Sequence analysis of coat protein genes of these three viruses showed little variability. In the test experiment for mastreviruses, three genetic groups, each representing distinct virus species were identified. The first group represented the A-strain of MSV. The second sequence group shared 96-98% identity with Maize streak reunion virus (MSRV) isolates, confirming the presence of MSRV also in continental East Africa for the first time. Sequence analysis of additional virus genomes (each 2846 nt) representing the third group revealed only a limited nucleotide identity of 70-71% with MSRV isolates belonging to a novel virus species tentatively named maize streak dwarfing virus (MSDV). PCR screening of 89 iv samples showing streak symptoms with designed general or specific mastrevirus primers showed that MSV is the most incident followed by MSRV and MSDV. Maize yellow mosaic virus (MaYMV) was also assessed from 47 leaf samples by RT-PCR using general polerovirus primer pairs embracing 325 nucleotides of the coat protein gene 34 samples were positive for this primer. Direct sequencing of the RT-PCR products confirmed ≥ 99% nt identity to each other and shared 98 to 99% identity with the reference sequence, KU248489. Full-genome sequence variability study of three MaYMV isolates using Illumina MiSeq sequencing revealed 99.6% nt identity to each other. One of the complete genome sequences, MF684369 shared 96.8% nt identity with KU248489. In conclusion, this study revealed the presence of six viruses in maize fields of Ethiopia, their distribution, incidence and provided sequence data for these viruses. Moreover, occurence of three different mastrevirus species on maize in Ethiopia, is unprecedented, and suggests that Ethiopia may be one of the potential hot spots for the diversity of maize mastreviruses.

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