MST-Department of Biochemistry and Biotechnology
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Item Antimicrobial Activity, Phytochemical Screening, and Lignocellulolytic Enzyme Quantification of Microbial Endophytes from Aloe Secundiflora and Azadirachta Indica(Kenyatta University, 2024-11) Mwendwa, Peter K.Azadirachta indicahas been used in treatment of diseases such as skin inflammations, Chikungunya, Measles and fungal infections. Aloe secundiflora has been shown to inhibit Staphylococcus aureusand Cryptococcus neoformans. Medicinal plants have been shown to contain microbialendophytes that live and feed within the plantsand play a role in protection against pathogens, either by triggering their defense mechanisms or directly acting on the pathogens. The current antimicrobial resistance has led to the need to explore alternative sources for new generation antibiotics. These alternative sources include endophytes of Azadirachta indicaand Aloe secundiflora which have the potential to produce new generation antibiotics. The current industrial catalysts are expensive, toxic and difficult to remove from reaction mixtures. Endophytes are potential sources of the desired enzymes. These endophytes have not been comprehensively researched. The objectives of this study wereto isolate and identify endophytes from young and mature leaves of Az. indica and A. secundiflora in Kitui and Kiambu Counties, determine the antimicrobial activity of the endophytes against selected pathogens, screen for phytochemicals produced by the endophytes and quantify the cellulase enzymes produced by the endophytes. The endophytes were isolated on sterile growth media and identified by gene sequencingand using the NCBI genomesequence database. The agar diffusion and dual culture techniques were used to determine antimicrobial activity for bacteria and fungi respectively. For phytochemical screening, the endophytic isolates were transferred from solid culture media to broth media for fermentation and the phytochemicals extracted by solvent extraction method, using ethyl acetate. For enzyme production, the endophytes were subjected to SSF and their enzymes extracted and quantified. Bacterial isolates obtained belonged to Bacillus sp., Alcaligenes faecalis, Providencia vermicola, and Priestia megaterium. Fungal isolates belonged to Colletotrichum sp., Trichoderma lixii, Galactomyces candidum, Penicillium sp., Fusarium luffae, Cladosporium cladosporioides and Candida sp. Endophytic isolates with antimicrobial activity were Bacillus subtilis, Priestia megaterium, Bacillus aerophilus and Penicillium sp. The isolates were positive for terpenoids, saponins, alkaloids and flavonoids. All fungal isolates produced cellulase enzymes across all assays (p ≤ 0.05). This study showed that Azadirachta indica and Aloe secundiflora contain fungal and bacterial endophytes which are potential sources forproducing bioactive compounds and cellulose enzymes. Thus the recommendation is that the endophytes of these plants can be used in developing new antibiotics and more research need to be done to quantify the bioactive compounds present in the isolates.Item Stability and Immune Responses of a Newly Formulated Thermo-Tolerant Peste Des Petits Ruminants Virus Vaccine(Kenyatta University, 2024-09) Kipkoritoo R. ErickPeste des Petits ruminants (PPR) is an acute and contagious viral disease of small ruminants (goats and sheep) with a registered high morbidity and mortality rate levels in naive populations. The disease is largely endemic in most Asian and African countries and is known to cause annual losses of approximately USD 1.45-2.1 billion affecting 330 million individuals of the world’s poorest 76 countries. Peste des petits ruminants virus is a highly infectious transboundary disease with no treatment and can only be sustainably controlled through vaccination. Small ruminants often play a fundamental role in the livelihoods of pastoralists as they act as a source of food and are sold off quite often to meet daily financial needs. Small ruminants require fewer resources, reproduce faster, and are more resistant to environmental shocks and hence are more relied on than large ruminants. The current conventional PPRV vaccines are thermo-labile and require to be kept at 2°C - 8°C for them to remain viable during storage and transportation. This present study was aimed at evaluating the stability and immune response of a newly formulated thermo-tolerant PPRV vaccine. The study prepared PPRV vaccine cultures using live attenuated PPR vaccine (N75/1), Sucrose, and Trehalose sugars augmented with proteins (lactalbumin hydrolysate (LAH)) and adopted the Xerovac lyophilization technique. Comparative analysis of the thermotolerance of both vaccines was assessed using the Accelerated Stability Tests (ASTs) at different temperatures (25°C, 37°C and 40°C). The ability of the vaccine to elicit PPRV neutralizing antibodies was determined in rabbits. Serum samples were collected weekly from day 0 to day 28 and on day 42 and 56 and IgG antibodies were quantified by competitive enzyme linked immunosorbent assay (c- ELISA). Data were analyzed using the GraphPad Prism software and utilized Spearman-Karber, regression analysis, t-test, and One Way Analysis of Variance (ANOVA) statistics. A P values ≤ 0.05 was considered statistically significant. The vaccine shelf life was determined as the time the vaccine retained its minimum infectivity dose at a specified temperature excursion. Results indicated that, the vaccines produced using Trehalose + LAH stabilizers were the most stable across all three temperature ranges. The shelf life of the trehalose-based vaccine was 21 days at 25°C, 19 days at 37°C, and 5 days at 40°C. There was a significant difference (p=0.0001) in vaccine degradation at 25°C and at 37°C. Comparative thermo- degradation analysis between the newly formulated thermo-tolerant and the conventional thermo-labile PPRV vaccines showed a significant difference (p=.0001) in thermo-tolerance across the two temperatures (25°C and 37°C). There was also a significant difference (p=0.116) in vaccine degradation at 40°C. Vaccinated rabbits showed protective antibodies for the 56 days of observation period. The study concludes that the use of trehalose and sucrose can provide successful thermo- stabilization of vaccines while eliminating use of expensive cold chain systems during storage, transport, and field application. It is recommended that sugar-stabilized PPR vaccines can be used to facilitate successful mass vaccinations in the tropical regions as part of the global PPR eradication.Item Production of Cellulolytic Enzymes by Chaetomium Globosum, Hypocrea Lexii and Gibberella Intermedia for Biomass Saccharification and Ethanol Production(Kenyatta University, 2024-10) Munyasi, KelvinThe world is staring at an energy crisis because of increase in population and utilization of non-renewable energy resources; petroleum and coal. Availability of these nonrenewable resources is abundant presently but will be exhausted in a limited period of time. Renewable energy resource provides an opportunity to serve as a substitute to the ever diminishing non-renewable energy resources, in this regard lignocellulosic plant biomass as an abundant renewable resource generated from plants and algae can be utilized. Exploration of plant biomass as a renewable energy resource not only allows for the conservation of non-renewable energy resources, but also for environmental and biodiversity protection. This study was aimed at isolation and identification of cellulase producing fungi from a decaying tree trunk, determining the effect of incubation time, moisture content level and initial medium pH levels on cellulase production using untreated maize cobs, rice husks and sugarcane bagasse as substrates under solid state fermentation thereafter, saccharifized untreated maize cobs, eucalyptus and sugarcane bagasse for ethanol production. Samples of wood were collected from Ngong forest, Nairobi County (1°19'13"S 36°44'54"E) and screened for isolation of cellulase producing fungi on carboxymethylcellulose agar stained in 1% Congo red. The isolated fungi were further grown to obtain pure cultures on Potato dextrose agar before DNA extraction and amplification by Polymerase Chain Reaction (PCR) was carried out to identify the isolated fungi. The PCR products were sent to Macrogen, Netherlands where sequencing was done by Sanger dideoxy sequencing technique. The isolated fungi were cultured on the untreated substrates in glass flasks for cellulase enzyme production in solid state fermentation. The crude cellulase obtained after fermentation was used to carry out cellulase assays; filter paper assay, exoglucanase assay and endoglucanase assay. Molecular data analysis was carried out using the National Center for Biotechnology Information (NCBI) and Basic Local Alignment Search Tool (BLAST) algorithm and Molecular Evolutionary Genetics Analysis (MEGA X version 10) software to identify the isolated fungi, while enzyme activity analysis was done using one-way Analysis of Variance (ANOVA) on R software at P≤0.05 significance level and the significant differences were determined by Tukeys Post Hoc test. The isolated fungi were identified as Chaetomium globosum, Hypocrea lexii and Gibberella intermedia. For effect of time of incubation on cellulase production on the three untreated substrates, the three fungi expressed high enzyme production on different days within the incubation period, highest cellulase enzyme activity was recorded at moisture content ratio of 1:2 and initial medium pH of 5. The three untreated plant biomass; maize cobs, eucalyptus and sugarcane bagasse were hydrolyzed with the crude cellulases produced from the three fungi, the saccharification studies showed that 7% (v/v) enzyme concentration, 12% (v/w) substrate concentration and a hydrolysis time of 72 hours were optimal for maximum yield of reducing sugars by the Dinitro salicylic acid method. The total reducing sugar produced maximum bioethanol at 72hours when Saccharomyces cerevisiae was used as a fermentation agent. Consequently, bioethanol was successfully produced from the cellulose of the three substrates using cellulases produced by the three fungi. Therefore, cellulase enzymes can be produced from microorganism found locally and by using readily available agricultural waste.Item Isolation and Characterization of Therapeutic Lytic Phages against Multi-Drug Resistant Enterobacter Cloacae in Kenya(Kenyatta University, 2023-06) Mutai, Ivy JepkuruiNosocomial infections due to Enterobacter cloacae affect 7% of patients in high-income nations, whereas they affect 15% of patients in low- and middle-income countries. This has been attributed to the ability of E. cloacae to confer resistance on carbapenems. With the scarce information on surveillance data and mortality rates of E. cloacae in Kenya, a recent report described E. cloacae as the most predominant multidrug-resistant organism with a resistance rate of 42.9% in critically ill patients in national referral hospitals and 12.3% in neonates from a rural hospital. E. cloacae is a unit of the ESKAPE family. This is composed of key resistant bacterial pathogens of Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and other Enterobacter species. With a 12% mutation rate, which is twice as high as those reported in other Gram-negative bacteria, E. cloacae has been referred to as one of the greatest mutators in a clinical environment. This has been attributed to its complexity and exponential evolution of phenotypic and genotypic characteristics. Following this rise in antimicrobial resistance, the use of bacteria-eating viruses, termed as bacteriophages (phages) has been recommended. This study determined the antibacterial profile of E. cloacae for phage isolation, evaluated the existence and therapeutic potential of lytic phages against the MDR E. cloacae, evaluated the phage's stability in different physicochemical properties, and determined their genetic characteristics. The phages were screened from environmental wastewater sourced from the informal settlements of Korogocho, Huruma, Kibera, Dandora, Kariobangi, and Mathare in Nairobi County. The identification and antimicrobial profiling of the bacteria was done through Gram staining, biochemical tests, and confirmation using the automated Vitek II Machine. Phages were screened from the individual samples using enrichment and spot test assays. The isolated phages were assessed for cross-reactivity on a panel of thirty clinical isolates using spot tests as part of a host range analysis. From host range analysis, the most potent phage from each source was selected for physicochemical properties studies and whole genome sequencing (WGS) using Oxford Nanopore Technology (ONT). According to antimicrobial tests, the isolate was resistant to six classes of antimicrobial agents that include: beta-lactams (penicillin, cephalosporins, monobactams, and carbapenems), chloramphenicol, tetracyclines, glycylcyclines, quinolones, and sulphonamides. The only antibacterial agents that worked were tigecycline (glycylcycline) and meropenem (carbapenem). Twenty-nine lytic phages were screened with 4 from Dandora, 5 from Huruma, 5 from Kibera, 5 from Kariobangi, 7 from Korogocho, and 3 from Mathare. The chosen phages remained steady from 4 -50ºC with a 5.1% (4/6) drop and a 1.8% (2/6) increase in phage titre at 50ºC respectively. After two weeks of incubation at 4°C, the phages were effective, with peak activity at 37°C (human body temperature) and lowest activity at -20°C. There was no phage activity at pH 2 but was active from pH 5-11 with optimal activity at pH 7.5. Furthermore, they were efficient with Ca2+ ion intensities of 0.002M and 0.015M. All phages' effectiveness decreased with time after UV exposure. All the phages (n=29) showed cross-reactivity against isolates of E. cloacae (n=30) during host range evaluation, with the highest potency being 67% and the lowest being 27%. In WGS, the phages were virulent, and none harboured AMR genes, virulence factors, toxins, plasmids, chromosomes, transfer (tRNA), ribosomal (rRNA), and lysogenic genes. This study revealed that Kenya has therapeutic super lytic phages that can be used to eliminate multidrug-resistant E. cloacae infections. This study recommends the need to explore on phage dosage and in vitro studies for safety and side effects.Item Antinociceptive and Anti-Inflammatory Effects of Leaves, Wood, Stem and Root Barks Extracts of Croton Megalocarpus (Hutch)(Kenyatta University, 2024-11) Kiarie Peter KanogaThe use of herbal medications has risen as a remedy for various ailments as they are low cost and are readily available to a high number of people across the world living below the poverty line. Croton megalocarpus has been used by some communities to manage ailments like malaria, stomachache, pneumonia, and wound clotting. The Current study blended leaves, stems and root bark extracts. This study aimed to determine the plant parts responsible for antinociceptive and anti-inflammatory effects among leaves, wood, stem and root barks of C. megalocarpus in mice. The samples were collected, ground to powder, and then extracted using methanol. White albino young mice were in eight clusters of 5 mice each for each set of experiments. These groups were standard control, negative control, positive control and five experimental groups. These experimental groups were in five sets: I, II, III, IV and V. Set I to V animals were treated with leaves, stem bark, root bark, woody part of stem and root methanolic extracts of C. megalocarpus, respectively. The experimental groups received 50, 100 and 200 mg/kg bw extract doses. The antinociceptive activity was evaluated by means of a formalin test, while an anti-inflammatory assay was undertaken using formalin-induced paw oedema. The formalin was infused through subcutaneous injection of the sub-plantar region of the hind paw. The qualitative analysis was undertaken to identify the secondary metabolites in all C. megalocarpus extracts. Chronic toxicity was conducted using mice administered with high doses of the extracts for 30 days. The blood was collected for biochemical and haematological tests. Tissues were harvested, and their weights were taken. The data collected was expressed as means and SEM. It was then analyzed using one-way ANOVA and Scheffes post hoc tests. All extracts of the various plant parts showed significant reduction (p < 0.05) in the period spent in pain behaviour. In this study, extracts from various parts of the plant demonstrated substantial inflammation reduction (p < 0.05) in mice except for the woody part of the root. Studies has shown that various parts of C. megalocarpus have medicinal values, and the plant parts have antinociceptive and anti-inflammatory effects. Methanolic extracts of C, megalocarpus, contain toxic properties and phytochemicals. Therefore, there is a great need to look at ways the secondary metabolites may be harnessed to develop novel drugs of medicinal value.Item Prevalence and Distribution of Foot-and-Mouth Disease Virus Serotypes and Serotype O Subtypes in Kenya in 2019 and 2020(Kenyatta University, 2024-10) Mumo, Josiah JudithLivestock is a source of livelihood, contributing to the reduction of poverty in many parts of the globe. In Africa, livestock is an asset, a source of food and income. In Kenya, approximately 12% of the country’s Gross Domestic Product (GDP) is contributed by livestock production and similarly, 42% of the agricultural GDP. Additionally, livestock offers employment to 50% of the agricultural labour force. Animal diseases, however, pose a great challenge in the industry among them being FMD (Foot-and-Mouth Disease), which is a transboundary viral disease typical among the cloven-hoofed animals. This disease has a huge economic impact on farmers as it affects many animals and disrupts the trade in animals as well as their products. It is also known to be endemic in most parts of the world including Africa, exists in seven serotypes (A, O, SAT 1, SAT 2, SAT 3, C, and Asia 1) and numerous subtypes (topotypes) and the antisera from one serotype cannot neutralize another serotype. Moreover, the movement of infected animals is closely linked with the spread of the disease, and quarantine measures and vaccination are key in controlling its spread, especially in endemic areas. In Kenya, the disease occurrence and severity have continued to increase despite the implemented control measures. A cross sectional study was done to establish the disease prevalence, distribution, circulating serotype O topotypes, and their genetic divergence from the current vaccine strain. A total of 267 bovine epithelial samples were analysed, sourced from cattle presenting with clinical disease in the years 2019 and 2020. FMDV (Foot-and-Mouth Disease Virus) antigen detection and serotyping ELISA was performed and those positive for serotype O antigen were further analyzed for genetic diversity. Viral RNA was extracted from the confirmed serotype O samples using PureLink® Viral RNA/DNA Mini Kit from Invitrogen according to the manufacturer’s instructions. The partial FMDV VP 1 gene was amplified through RT (Reverse Transcription) PCR (Polymerase Chain Reaction) and directly sequenced. The generated sequences were edited using BioEdit version 7.2.5 and phylogenetically analysed using MEGA X version 10.2.6 by neighbor-joining at 1000 bootstrap replicates before viewing them through Tree view. The antigen detection and serotyping gave an overall virus prevalence of 65.9% (176/267). From the same results, serotype SAT 1 had the highest prevalence of 45.5%(80/176) followed by serotype O at 37.5% (66/176). Serotype SAT 2 had a prevalence of 14.2% (25/176) and serotype A had the least of 2.8% (5/176). The phylogenetic analysis on FMDV serotype O revealed that all the sequences analysed were of the EA (East African) 2 topotype. The genetic analysis of these sequences showed a divergence from the current vaccine strain by the replacement of some amino acids on the gene. This study, therefore, confirms that the 2019 and 2020 outbreaks were predominantly caused by serotype SAT 1 with some serotype O viruses causing these outbreaks likely to have mutated and not being covered by the current vaccine for serotype O. This information is useful to disease control policymakers, implementors and farmers in effecting disease control at their specific levels and recommends continual monitoring of the FMDV serotypes and topotypes that are in circulation, and regularly matching the vaccine strain with the circulating viruses to ensure effective vaccination measures.Item Distribution and Diversity of Fungi and Their Biocontrol Potential in Managing Coffee Berry Disease in Kirinyaga County Kenya(Kenyatta University, 2024-11) Msenya, Happiness NyambugeCoffee is among the major traded goods across the world after oil. Its production in Kenya faces constraints majorly coffee berry disease (CBD) which is caused by the fungus Colletotrichum kahawae. Diversity of biological species confers essential benefits in creating sustainability which contributes to increase in diversity thereby providing resistance to disturbance and stress, and change in conditions in the soil. Understanding the diversity of microorganisms in berries and soils from coffee farms could be of importance in determining which of these micro-organisms could control the disease. The significance of this study was to isolate and identify fungi from different zones in coffee farms in Kirinyaga County Kenya with an aim of further studying their biocontrol potential to Colletotrichum kahawae. One hundred (100) coffee farms were sampled. The species of fungal were isolated from coffee berries; soil and from Upper Midland 1, 2 and 3. The berries of coffee were washed and allowed to sporulate for 24hrs. Lesions were excised, suspended in distilled water and serial dilutions made. This was plated in potato dextrose agar (PDA) and incubated at room temperature for ten days. Fungi was isolated from soil by inoculating 1ml from serial dilutions 10-3 on PDA and incubating at room temperature for ten days. Recovered colonies were sub cultured individually by inoculating the spores scooped from the culture into a separate plate with PDA. Identification of fungi at the genus level was carried out by using macroscopic and microscopic examination. Five isolates were tested for antagonism against CBD by co culturing 3cm diameter discs of the test isolate and the pathogen on PDA media. The degree of antagonism was determined by measuring and comparing the radial growth of pathogen with the bio agent against the control. Fungi were identified as Colletotrichum, Phoma, Trichoderma, Epichoccum, Aspergillus, Alternaria, Fusarium, Rhizopus, Penicillium, and Cladosporium. The results indicated that Colletotrichum and Cladosporium were the most abundant in the berries at 76% for both species. Fusarium and Cladosporium were the most abundant in soil at 50% and 20% respectively. From the five isolates that were tested against C. kahawae, two isolates Penicillium and Fusarium proliferetum significantly inhibited growth of the test pathogen at 55% and 60% respectively. The remaining isolates Aspergillus fumigatus, Chaetomium perithecia and Fusarium Ceraneasum showed inhibition of growth at 40%, 18.18 %, and 45.45% respectively. Statistical analysis (p<0.05) indicated significance difference of distance of growth between the potential isolates with a higher growth inhibition to the test pathogen against control. The test results indicated the potential of utilizing saprophytic coffee surface micro flora and rhizosphere fungi in bio control of the coffee berry disease upon further verification of the species of fusarium not being a disease causing agentItem Assessment of Physico-Chemical Water Parameters, Habitat Characteristics, and Bacterial Influence on Anopheles Funestus Mosquito Larvae in Fihoni, Kwale County(Kenyatta University, 2024-08) Okumu, Clifton OmondiAnopheles funestus is a primary malaria vector yet remains understudied in East Africa. Resistance to available control measures has led to increased prevalence of the vector. Studies have revealed that chemical cues mediating oviposition attraction may be associated with bacterial communities. However, the bacteria responsible have not been profiled and characterized. This study, therefore, provides missing information on bacterial species associated with oviposition attraction in Anopheles funestus. This study aimed to isolate and identify bacteria isolates from the larval and non-larval sites of A. funestus, characterize the larval habitats using physiochemical parameters, and then conduct a bioassay-guided determination of oviposition attraction by A. funestus. Larval habitats were characterized using an estimation method on habitat parameters like size, distance from homestead, and water depth, among others. Physiochemical parameters of the larval water were determined using multi-parameter equipment and other standard laboratory methods. The experiment used two larval water samples (obtained from mixing samples from the four different transects in Fihoni) categorized as either preferred or non-preferred. Bacteria were isolated from the preferred and non-preferred larval water and grown on TSA agar. Isolates were then characterized using morphological, biochemical, and molecular techniques. 16S rRNA gene was used in amplification and Sanger sequencing. Bacterial primer pairs (27F and 1492R) were used in sequencing. Data on the number of A. funestus and non-A. funestus larvae were compared using the Mann-Whitney U test in Scientific Analytical System (SAS) version 9.4, and results were presented in table format. Data on counts of oviposition due to inoculation of substrate (Media) by different bacteria isolates was analyzed by Kruskal Wallis test in SAS version 9.4. Data on the association between habitat parameters and the number of larvae was analyzed using chi-square (α = 0.05) and Pearson correlation coefficient in SAS version 9.4. Significance medians were compared using Steel-Dwass-Critchlow-Fligner in SAS version 9.4. Phylogenetic analysis was conducted on MEGA X version 11.0, and alignment was done using the MUSCLE program in MEGA X version 11.0. Conductivity, total dissolved solids, pH, sulfate, conductivity, and Biological Oxygen Demand had an effect (p<0.0001) on the presence of Anopheles and non-Anopheles funestus larvae at the oviposition sites. Phylogenetic analysis shows that the bacterial isolates had a sequence identity between 82.05% and 96.56%. They included Staphylococcus saprophyticus, Enterobacter sp strain EDB2, Uncultured actinobacterium clone SGR 192, Enterobacter sp MLB, and Citrobacter werkmanii which attracted A. funestus to their oviposition cups. The vector's attraction rate to the preferred oviposition cup was stronger (median=52.00) with crude bacterial isolates than with pure isolates (median=38.00). Bacteria (Enterobacter sp strain EDB2 and Enterobacter sp MLB) synergistically increased oviposition attraction. The rate of attraction was substantially increased from (median= 23.00 for CFI3 [Pluralibacter sp] and median=35.00 for CPIp [Bacillus sp]) when isolates were pure to (median =123.00 for mixture M) when isolates were mixed. No oviposition (median=0.00) was seen in cups amended with Pluralibacter gergoviae, Citrobacter sp, and Enterobacter sp MLB07, among others, when used individually. There are no sequence differences between CPI7 [Enterobacter sp. strain EDB2], CNPI3 [Pluralibacter gergoviae], and CPI5 [Pluralibacter gergoviae]. CFI4 [Uncultured actinobacterium] was most attractive (median=58.00), followed by CPI11 [Enterobacter sp] (median=57.00). The present study shows the ability of some bacterial isolates from A. funestus larval sites to influence oviposition site preference. Information on bacteria isolates with positive oviposition response adds knowledge crucial for researchers and policymakers dealing with malaria control. For example, government agencies concerned with public health could use the oviposition attraction of mosquitoes by bacteria to reduce human mosquito contact and the spread of malaria.Item Phytochemical Composition, Antidiarrheal Activity and Antibacterial Effect of Aqueous Leaf Extract of Plectranthus Barbatus (Andrews)(Kenyatta University, 2024-05) Ajwang, Emmah ClariceDiarrhea is defined as the movement of unformed or watery stool more than three times a day. Globally, diarrheal infections remain a public health problem, especially in children. With nearly 1.7 billion episodes and 1.3 million fatalities reported annually. Developing countries bear 78% of diarrhea burden worldwide. A range of viral, bacterial, and parasitic species induce diarrhea in humans, including rotavirus and Escherichia coli. The mainstays of pharmacological therapy for infectious diarrhea include probiotics, antibacterials, antiviral drugs, and intestinal adsorbents. However, these clinical treatments are not devoid of shortcomings, including prohibitive costs, drug-drug interactions, and adverse effects such as lethargy, constipation, respiratory depression, and coma. Medicinal plants, including Plectranthus barbatus have folkloric remedies against diarrhea. However, there is a paucity of knowledge to scientifically validate the efficacy of the leaves of P. barbatus on diarrheal infections. The study, therefore, was undertaken to ascertain the antidiarrheal efficacy, antibacterial activity, bioactive composition, and toxicity profiles of P. barbatus aqueous leaf extracts. Antidiarrheal activity and acute toxicity were carried out on mice. Using Swiss albino mice, castor oil-induced diarrhea, charcoal meal-based gastrointestinal motility, and castor oil-induced secretion models were employed to assess antidiarrheal activity. In all of the test models, animals were randomly assigned into six groups consisting of six animals in each. Group I received distilled water, group II received 10 ml/kgbw of the vehicle (distilled water), while group III was treated with standard drug (3 mg/kgbw loperamide) in the respective models, whereas groups IV to VI received 100, 200, and 400 mg/kgbw of the aqueous leaf extracts of Plectranthus barbatus. Antibacterial activity was carried out on selected bacterial pathogens. Quantitative phytocompound analysis was evaluated using liquid chromatography mass spectrometry. Data were analysed using one-way analysis of variance followed by Tukey’s test, and p< 0.05 was considered statistically significant at 95% confidence of interval.The study results indicated that P. barbatus extract has antidiarrheal activity. The plant extract prolonged the onset of diarrhea, caused a significant decline in the occurrence of wet feces and intestinal transit. Additionally, the extract elicited a reduction in the accumulation of intraluminal fluid, resulting in a decrease in distension, intestinal overload, and water content in the fecal drops. Loperamide showed a statistically similar antidiarrheal effect with the extract at a dosage of 200mg/kgbw suggesting a probable effective dosage of the extract. This study demonstrated that the aqueous leaf extracts of P. barbatus exhibited diarrheal inhibition activity. The percentage inhibition was dose dependent with 100, 200 and 400mg/kgbw showing 49.98±1.61, 66.12±2.17 and 75.80±2.16% inhibition of diarrheal output respectively (p<0.05). Further, P. barbatus demonstrated antibacterial activity against pathogens associated with diarrheal diseases. The extract had varying Mean Zones of Inhibition (MZI), 7.33±0.33 to 17.17±0.73mm, against the bacterial pathogens, with higher effects observed against P. aeruginosa and B. subtilis. Acute toxicity assays on mice showed that P. barbatus extract was non-toxic at the dosage level of 2000mg/kgbw. LC-MS analysis detected the presence of phytocompounds associated with antidiarrheal and antibacterial effects. Findings from this study offer scientific validation for the folkloric utilization of P. barbatus in the management of diarrhea. However, further studies should be conducted to explore the mechanistic approach to the reduction of diarrhea and the comprehensive chronic toxicological effects on biochemical and hematological parameters.Item Effects of Aqueous Extract of Spirulina Plantesis on Immunologic Dysfunction and Inflammation Associated with Aflatoxin B1-Induced Toxicity in Mice(Kenyatta University, 2024-06) Kipkoech, GilbertKenya is among African countries that face the burden of food contamination by aflatoxin. High levels of aflatoxins have been reported to kill 157 people in Kenya while also contributing to cancer burden. Aflatoxins are produced by Aspergillus flavus, Aspergillus parasiticus, Aspergillus and Aspergillus nomius. These toxins majorly affects cereal grains. This study aimed at bioprospecting Spirulina plantesis in ameliorating immune dysfunction and inflammation caused by aflatoxin B1 (AFB1). Spirulina plantesis have been consumed for decades as food supplement as it has been proven to have strong antioxidant and anti-inflammatory properties. Male BALB/c mice weighing 28 to 34 g were divided into six groups at random and given the following oral treatments: Group 1 was only provided with food and water during the treatment course. AFB1 was given orally to Group 2 at a dosage of 200µ g/kg body weight. AFB1 was administered orally to Group 3 an hour after receiving 1g/kg of activated charcoal. Groups 4, 5, and 6 each received 200µg/kg of AFB1 orally an hour after receiving Spirulina plantesis at doses of 50, 100, and 150mg/kg respectively. The mice were treated daily for 14 days. During the last day of the treatment schedule, mice were aseptically dissected and tissues isolated for immunological studies. The results indicate that intervention with spirulina at doses of 100mg/kg and 150mg/kg were enough to increase the body weight of mice significantly (p<0.05). It was also demonstrated the blood levels of interleukin 4 and interleukin 2 were not affected significantly when AFB1-induced mice are treated with spirulina extract (p>0.05). Interferon gamma (IFN- γ) and interleukin 2 (IL-2) blood levels were significantly lower in the group not treated with Aflatoxin b1 (p≤0.05). The findings also indicate that immunoglobulin A (IgA), immunoglobulin G (IgG), and immunoglobulin M (IgM) serum levels were unaffected by treatment with spirulina extract at various dosages (p>0.05). In addition, TNF and IFN-γ mRNA expressions were also highly up regulated, while interleukin 4 (IL 4) was down-regulated. The results further show that the over-production of TNF and IFN due to Aflatoxin B1 is correctable upon spirulina treatment (p<0.05). To sum up, the results suggest that spirulina treatment can be an innovative approach to correcting the aflatoxin B1-mediated immune aberration and inflammationItem Performance of Glucose Challenge Test in the Diagnosis of Gestational Diabetes Mellitus in Antenatal Mothers at Kenyatta National Hospital, Nairobi, Kenya(Kenyatta University, 2024-04) Ndege, Martin K.Gestational diabetes mellitus is a disorder that affects women during pregnancy. It is glucose intolerance that is detected in pregnant women but reverses to normal after delivery; thus, it requires re-classification post-partum. It impacts approximately 4% of all pregnancies, with a prevalence of 1–14%. The statistics are dependent on the screening method and population. Early detection of GDM is essential to reduce the risks associated with delivery as well as potential long-term health concerns, such as metabolic and cardiovascular problems, for both the mother and the child. Nevertheless, diagnosing GDM in Sub-Saharan Africa (SSA) still faces challenges due to logistical constraints and the financial burden, particularly for populations with limited resources. It is therefore imperative to develop a practical and affordable method of screening for GDM in these contexts. Currently, OGTT is the confirmatory method in use. The OGTT procedure is cumbersome and difficult, costly and time-consuming with up to four blood collections from the patients. It is lengthy and requires the patients to fast; it is not user-friendly for antenatal mothers. As a screening method, GCT confers several advantages such as it is a one-step method that follows simple steps that are easy to remember, it reduces delay in diagnosis which consequently reduces delays in care and management. The method does not require patients to fast, can be performed at any time of the day, is cheaper than OGTT and is less time-consuming hence it can be used for both diagnosis and prevalence surveillance studies. It is much friendlier to the users and the technical staff, rendering it a more financially feasible option for both healthcare systems and patients, particularly in resource-limited settings. However, the use of GCT for screening for GDM has not been evaluated in Kenya. Consequently, the purpose of this study was to ascertain the prevalence of gestational diabetes mellitus (GDM) and evaluate the effectiveness of the Glucose Challenge Test (GCT) in detecting GDM in the Kenyan population. This was done by comparing the performance of GCT to OGTT which is the reference method in the investigation of GDM in expectant mothers. GCT and OGTT were performed on 107 pregnant and 84 non-pregnant women totalling 191 participants. 23 pregnant women (21.5%) had a diagnosis of GDM at 95% CI. Compared to OGTT, GCT had sensitivity, specificity, PPV and NPV of 73.91%, 86.9%, 60.71% and 92.41% respectively using a diagnostic cut-off of 7.8 Mm. Of the 23 (21.5%) with GDM, 12 were followed up, and 4 (33%) transformed to type II diabetes while 8 (67%) reversed to normal glucose levels. Of the common adverse outcomes of GDM in antenatal mothers, only excessive amniotic fluid had a significant impact (ρ = 0.048). The study concluded that there is a similarity in the specificity and sensitivity of OGTT and GCT in detecting GDM. Further, a significant GDM prevalence exists in antenatal mothers in the population under study. The MoH should develop policy guidelines for screening, counselling, and treatment of GDM among ANC mothers. GDM mothers should be followed up with to determine their postpartum status as Type 2 Diabetes. Based on the high prevalence of GDM among ANC mothers, this research recommends that the MoH adopt GCT for GDM screening. Additionally, ANC mothers should be educated about the traditional adverse outcomes associated with GDM.Item In Vivo Ameliorative Effects of Vitamin E Supplements against Hydralazine Induced Systemic Lupus Erythematosus(Kenyatta University, 2024-03) Githaiga, Fiona MuthoniSystemic lupus erythematosus (SLE) is a chronic autoimmune disease where the immune system attacks body cells and tissues causing inflammation. The drug therapy in SLE states that organ threatening disease should be managed aggressively by high to low doses of corticosteroids often followed by institution of steroid sparing measures in the form of immunosuppressive. Long term use of these drugs have life threatening side effects. Toxic oxygen radicals such as hydrogen peroxide have been associated with the development of SLE. The aim of this project was to evaluate the potential of an anti-oxidant, Vitamin E to eliminate the hydrogen peroxide levels in hydralazine induced lupus, a disease model that closely resembles SLE. The experiment involved forty Bagg Albino (BALB) c mice aged three to four weeks comprising of twenty females and twenty males that were randomized into six different groups. Each group contained three males and three females. To mimic the pathogenesis of SLE, hydralazine hydrochloride 25mg/kg bdw was used to induce a lupus-like condition in mice from groups B to F. The mice from group A received 1.0ml of sterile water and served as the normal control. An anti-nuclear antibody test was carried out every seven days to monitor the development of autoantibodies, which are considered a hallmark of lupus. By the 35th day post hydralazine administration, ANA antibodies were confirmed in all treated groups. Drug treatments were initiated in all lupus-induced mice at the same time, daily for a period of another 35 days with the exception of group B that served as the negative control of lupus induced mice with no drug treatment. Group C received prednisolone 25 mg/kg bdw, Group D received methotrexate 25 mg/kg bdw, Group E received Vitamin E 25 mg/kg bdw, and Group F received Vitamin E 50 mg/kg bdw. Only the higher dose of Vitamin E significantly eliminated the levels of lymphocyte hydrogen peroxide associated with the pathogenesis of hydralazine induced lupus. Both doses of Vitamin E protected lupus-induced mice from alterations in all blood cells. Mice from the methotrexate treatment groups recorded significantly low levels of all blood cells. However, the prednisolone group recorded significantly low lymphocyte count with eosinophil and neutrophil counts significantly high. Notably, the prednisolone treated mice had significantly increased platelet count. Both doses of Vitamin E protected the mice from changes in liver biomarkers while prednisolone and methotrexate treatment groups showed significantly high levels of liver enzymes. Both doses of Vitamin E protected from alterations in the lipid profile. Prednisolone and methotrexate treatment groups had high ANA titer concentrations, Mice treated with Vitamin E had significantly low ANA titers. Vitamin E also was found to prevent injury to the kidney, liver, spleen, heart and brain. Prednisolone group had significantly elevated bodyweight compared to the normal control group. In conclusion, this study found Vitamin E supplementation was able to counter oxidative stress associated with the pathogenesis of lupus. Based on these findings, Vitamin E supplementation may be beneficial for improving the condition of SLE patients. Further research is recommended to explore other relevant free radicals in SLE and determine the optimal dosage of Vitamin E to effectively eliminate free radicals associated with SLE.Item The Association between Toll-Like Receptor 4 (-8984c/G) and 299 Asp/Gly and Severe Malaria Disease in Children below 3 Years in Siaya County Western Kenya(Kenyatta University, 2024-05) Omwandho, John Robert CharlesThe high paediatric mortality and morbidity linked to Malaria in Africa is majorly caused by the parasite: Plasmodium falciparum with many complications characterized by severe anaemia and parasitemia. Severe Malaria Anaemia (SMA; Hb<5.0g/L accompanied by parasitemia of any density) manifests in young children below three years old. Nonetheless, the absence of understanding regarding the molecular underpinnings of SMA continues to hinder progress in creating successful treatments. Genetic susceptibility factors provide a means to decipher the intricate molecular processes at play. Toll like Receptor signalling pathways play an essential immunological role in helping to clear the parasites. However, the contributions of genetic variations to SMA pathogenesis are partially understood. Therefore, the study aimed to unravel the associations between Toll-Like Receptor 4 (-8984C/G) and 299 Asp/Gly polymorphisms and vulnerability to SMA and parasitemia. Samples from children (n= 426) diagnosed with malaria at Siaya County Referral Hospital were analysed. DNA extraction and genotyping were performed on dry blood spots using Gentra Systems Isolation Kit and TaqMan® 5' allelic discrimination Assay-By-Design high-throughput technique respectively. The association between Toll-like receptor-4 (-8984C/G) and 299 Asp/Gly polymorphism and SMA was determined using bivariate logistic regression analysis while controlling for anaemia confounders; Bacteraemia (co-infections) and HIV-1, alpha-thalassemia (traits) and sickle cell. The association between parasitemia and carriage of respective genotypes and haplotypes was done using Kruskal Wallis test/ Mann U Whitney where applicable. Bivariate regression analysis revealed that the TLR-4 (-8984 C/G) genotypes (GG vs. GC, OR = 0.53, 95% CI = 0.07-3.91, P= 0.533, and GG vs. CC, OR = 1.02, 95% CI = 0.12–8.82, P= 0.988) and TLR-4 +299 Asp/Gly genotypes (Asp/Asp vs. Gly/Asp, OR = 1.66, 95% CI = 0.42–6.50, P=0.471) were not associated with SMA. Further, haplotypes of TLR-4 -8984G and +299Gly (OR; 2.70, CI; 0.70-10.48, P= 0.151), TLR-4 -8984G and +299Asp (OR; 0.72, CI; 0.15-3.51, P= 0.682) and TLR-4 -8984C and +299Asp (OR; 1.29 CI; 0.56-3.63, P = 0.636) were also not associated with SMA. Moreover, there was no association between genotypes of TLR-4 (8489G /C) [GG (18530), GC (18303) and CC (14952) P= 0.858] or TLR-4 (Asp + 299 Gly) [Asp/Asp (18770), Asp/Gly (37714) and Gly/Gly(67324) P= 0.482] with parasitemia. Similarly, the haplotypes of TLR-4 (G-8984C) + TLR-4 (Asp+299Gly), [G/Asp (P=0.887), C/Asp (P=0.884) and C/Gly (P= 0.4032)] had no association with parasitemia. The results demonstrated that TLR-4 (8489G/C) and TLR-4 (Asp+299 Gly) genotypes and haplotypes are not associated with SMA or parasitemia in the population of interest. The investigation outcomes are important in providing insights on the effect of polymorphism in immune cells in designing novel treatments and vaccinations in the fight against malaria. The study recommends further longitudinal studies to fully understand the functions of the polymorphisms in SMA.Item Properties of Isolated Galaxies within a Redshift Range of 0.005 < Z < 0.080(Kenyatta University, 2024-05) Kalondu, Kinyumu MarcelinaIn the study of galaxy formation and evolution process, a number of processes and properties are known to influence the formation and evolution process such as protogalactic clouds, galaxy masses and galaxy environment. Despite studies done on different environments, the effect of environment on galaxy properties has not been fully quantified. In this study, a sample of isolated galaxies within a redshift of 0.005 < z < 0.080, from the Sloan Digital Sky Survey, data release sixteen, (SDSS-DR 16) was analysed. The aim of the study was to investigate the physical properties of single isolated galaxies in low-density environment, to understand the internal processes and mechanisms dominating galaxy evolution. The classification of galaxies into late-type (mostly spirals) and early-type (mostly ellipticals) in the sample was first done using the central concentration index tool. A Concentration Index, Cr , of 2.65 well classified the sample. The automated classification was later confirmed using visual classifications from the galaxy zoo, were the spiral galaxies dominated the sample of isolated galaxies with about 68%. Through the analysis of colour, magnitude and stellar mass relations, the isolated elliptical galaxies are redder, massive and more luminous, while spiral galaxies seem biased towards the blue, less massive and less luminous side. However, a significantly larger fraction of isolated red spiral galaxies with larger stellar masses and highly luminous was observed. A slightly different distribution by both the isolated blue elliptical and spiral galaxies, characterised by lower stellar masses at lower luminosity was also noted. With the tight correlation of the (u − r) and (r − z) colours, the galaxy sample was analysed as star-formation, quiescent with a fraction of recently quenched elliptical galaxies (RQE). Most of the elliptical galaxies were well selected as quiescent, with majority of the spirals being star forming. This was expected since most elliptical galaxies are thought to be older and have quenched their star-formation, while the spirals are younger and have star forming activities. 5% of the isolated elliptical galaxies seem to have recently quenched their star formation and are transiting to the red sequence, with 90% being blue. The red recently quenched elliptical galaxies are among the 20% low-mass red elliptical galaxies, confirming that most massive elliptical galaxies quenched their star formation long ago. The Hα luminosity star formation rates were derived and correlated with the galaxy morphology and stellar masses. From the analysis, spiral galaxies have higher star formation rates, indicating strong star formation activities, while elliptical galaxies dominate the passive region. An indicated fraction of passively evolving high-mass spiral galaxies and actively star-forming elliptical galaxies was observed. In the emission line diagnostics, 50% of the spiral galaxies had higher star-forming activities, while 54% of the elliptical galaxies dominated in the Active Galactic Nuclear (AGN) powered region. A number of galaxies, both spirals and ellipticals are in transition from star-forming to AGN and occupy the composite region. The star forming galaxies selected in the colour - colour analysis tightly follows the star formation sequence, with 47% having star forming emissions and only 18% have AGN emissions. Recommendations for future work includes, a detailed study on the effect of galaxy properties on the central concentration index morphological classification tool, and further investigation on the powering mechanism of isolated galaxies with other methods.Item Harnessing the Potential of Phosphate Solubilizing Bacteria for Improvement of Maize and Cowpea Production in Semi-Arid Agro-Ecosystems of Eastern Kenya(Kenyatta University, 2023-05) Kibet, Charles Kirui; Ezekiel Mugendi Njeru; Stephen RunoAbstractItem Evaluation of Sex Selection Activity and Safety of Vernonia Amygdalina (Del), Rubia Cordifolia (Linn), and Asparagus Racemosus (Willd), in Rats(Kenyatta University, 2023-01) Wambugu, Enoc Njoroge; Eliud NM Njagi; John K MwonjoriaAbstractItem Cognitive Enhancing Effects of Aqueous Extracts of Amaranthus dubius Mart. Ex Thell and Vigna unguiculata (L.) Walp. in Mice(Kenyatta University, 2023-02) Kipkemoi, Daisy Jepkosgei; Mathew Piero Ngugi; Antony IreriAbstractItem Antibacterial Antioxidant, Phytochemical Profile and Toxicological Efffects of Aqueous extract of Rhaphiolepis Bibas (Lour.)(Kenyatta University, 2023-05) Kariuki Ibrahim WaweruAbstractItem In Vivo Anxiolytic and Teratogenic Effects of the Aqueous Extracts of Carissa Spinarum and Azadirachta Indica(Kenyatta University, 2023-10) Wabai, Yvonne Wairimu; John K. Mwonjoria; Joseph J. N. NgeranwaAbstractsItem Phytochemical Composition, in Vitro Antmicrobial Activity, And Cytotoxicity of Essential Oils of Ocinum Gratissimum, Ocimum Kilimandscharicum and Lippia Javanica(Kenyatta University, 2023-04) Samoei, Chemutai Judith; Mathew Piero Ngugi; Christine BiiAbstract