Cell and Molecular Biology

Characterization of maize rhizospheric microflora and evaluation of their antagonistic potential against Ustilago maydis

Wed, 01 Jan 2025 00:00:00 GMT

Characterization of maize rhizospheric microflora and evaluation of their antagonistic potential against Ustilago maydis OSENDI, Rosemary Opwondi Maize common smut, attributed to Ustilago maydis, poses a significant risk to maize farming across Kenya. Despite the known potential of rhizospheric microorganisms in biological disease control, limited information exists on their population, characterization, and efficacy against U. maydis in local maize cultivars. Additionally, data on their in vivo effect on disease incidence and severity remain largely unexplored. This study was designed to determine the bacterial and fungal populations within the maize rhizosphere, characterize them using morphological and biochemical characters, and evaluate the impact of selected microbial isolates on disease incidence and severity of common smut in maize. The study was conducted under field, laboratory, and greenhouse conditions at Maseno University. Two maize varieties, DK 8033 and Duma 43 were sown at the university farm in a completely randomized block design. Rhizospheric soil samples were collected at 30 and 60 days after sowing with a soil auger and trowel, into aseptic containers, then conveyed to the botany lab for analysis. Triplicate soil samples were processed via serial dilution followed by plating to recover colonies, which were counted and characterized morphologically and biochemically. Dual culture assays were conducted in vitro to evaluate antagonism against U. maydis. Four promising antagonists: Serratia spp., Bacillus spp., Aspergillus spp., and unidentified fungal isolate MF14, were selected for in vivo greenhouse trials in three replicates, with ten treatments arranged in a completely randomized layout. Treatments included maize cultivars with microbial isolates alongside pathogen inoculation. Controls comprised maize inoculated with the pathogen and distilled water. Comparisons among treatment means used Fisher’s Least Significant Difference, at P ≤ 0.05. There were 25 bacterial and 26 fungal isolates obtained and characterized. Results revealed significantly higher bacterial counts (134.69 × 10⁸ cfu /g) than fungal counts (34.58 × 10⁸ cfu /g), both peaking at 60 days. There were 15 bacterial genera, including Bacillus, Pseudomonas, and Serratia, and six fungal genera (Penicillium, Trichoderma, Fusarium, Alternaria, Aspergillus, Curvularia). In vitro, MF14 isolate showed the highest inhibition zone (22.00 mm), followed by Serratia spp. (19.00 mm), Bacillus spp. (16.00 mm), and Aspergillus spp. (15.00 mm). Greenhouse trials showed that the unidentified fungal isolate MF14 reduced disease incidence and severity to 49.3%, outperforming other treatments. Serratia spp and Aspergillus spp. showed moderate suppression, while Bacillus spp. showed minimal but superior protection to control. This study highlights indigenous rhizospheric microorganisms with potent antagonistic activity, particularly MF14 isolate. Findings provide foundational data for developing microbial-based biocontrol strategies, offering eco-friendly biocontrol options for maize farmers in Kenya upon further research. Master's Thesis

Evaluation of phsysicochemical, bacterial and fungal parameters of river Kibos water in Kisumu county, Kenya

Wed, 12 Nov 2025 00:00:00 GMT

Evaluation of phsysicochemical, bacterial and fungal parameters of river Kibos water in Kisumu county, Kenya OLUOCH, Nashon Okoth Wastewater discharges in river water is a major source of fecal microorganisms and poor water quality. Most important bacterial diseases are transmitted through water. River Kibos water quality is constantly changing due to waste water discharge from industries and the surrounding agricultural farms and settlements. Toxic wastes, chemicals and pathogens in the river water pose health risks to people using river water for drinking and recreation activities. Information on the effects of the season, water level, degree of eutrophication on physicochemical and microbial parameters of river Kibos is limited. The main objective of this study was to evaluate the physicochemical, bacterial and fungal parameters of River Kibos water. Specific objectives were to determine physicochemical parameters (pH, DO, temperature, turbidity, TDS, Nitrates, phosphates, BOD, ORP, Specific conductivity, COD and salinity), to characterise the fungi and bacteria of River Kibos water, to determine the fungal and bacterial population, and to determine the relationship between physicochemical and microbial parameters. Water samples (72) were collected for a period of 2 months three times per month from the river at four sampling sites randomly selected for wet (April to May 2019) and dry (February to March-2019) season in triplicates: The sites were Katieno I, Katieno II, Kotunga and Kibos Prison Bridge. The water was drawn slightly below the surface near the river edge. The water samples were preserved in an ice box at 4˚C to avoid destabilisation and fixed using acidic Lugols solution during transportation to the laboratory for analysis(You need to explain a bit how the samples were handled before the bacteria culturing)Bacteria were cultured using Nutrient Agar while fungi were cultured using Potatoe Dextrose Agar. Identification of bacteria and fungi was done using the standard microbiological techniques. Microbial count was determined by colony counter. Physicochemical parameters (temperature, DO, ORP, salinity, turbidity, specific conductivity, pH) were determined insitu by multiparameter instrument while nitrates, Phosphates, COD and BOD), were determined at the Maseno University microbiology laboratory. Data on physicochemical and microbial parameters was subjected to analysis of variance. Means were separated and compared using Pearson’s LSD at p≤0.05. Physicochemical parameters for dry season were: pH 7.4633, turbidity 249.92NTU, COD 0.83mg/l, NO - 3 3.623mg/l and PO - 4 2.4958.Wet seasons were; pH 11.0017, turbidity 341.58 NTU, COD 0.68 mg/l, NO - 3 3.89 mg/l and PO 4- 3.6625mg/l.Nine bacteria; Klebsiella spp, E.coli, Shigella spp, Salmonella typhi, Pseudomonas aeriginosa, Enterobacter cloacae, Shigella flexneri, Proteus spp and Staphylococcus spp. And Fungi ;Alternaria tenuissima, Aspergillus tubingensis, Penicillium citrinum and Penicillium crustosum. Bacterial and fungal counts in wet season was 50.25 and 141 cfu/ml respectively while for wet season were 12.5 and 51.33 cfu/ml respectively. The parameters were above the WHO and NEMA limits. The findings indicate that the water is polluted and hence a potential threat to human health. The water should be treated before human consumption. Master's Thesis

Phytochemical composition and bioactivity of extracts of Adenia lobata, Ipomoea aquatica, and Rubia cordifolia against Candida albicans, Pseudomonas aeruginosa, and Staphylococcus aureus

Mon, 10 Nov 2025 00:00:00 GMT

Phytochemical composition and bioactivity of extracts of Adenia lobata, Ipomoea aquatica, and Rubia cordifolia against Candida albicans, Pseudomonas aeruginosa, and Staphylococcus aureus JUMA, Meshack Mayianda Emergence and spread of antimicrobial resistance (AMR), is of great concern to the global health, necessitating the search for new therapeutic agents. Candida albicans, Pseudomonas aeruginosa and Staphylococcus aureus are among the serious AMR pathogens worldwide. Furthermore, AMR infections are associated with higher levels of oxidative damage in the human cells which is linked to various diseases, including cancer and cardiovascular diseases. Medicinal plants are of significant importance worldwide, serving as alternatives to modern medicine. Plant extracts contain phytochemicals, such as phenolic compounds, which exhibit bioactivities including antioxidant and antimicrobial activities. Antioxidants prevent oxidative stress in human cells, aiding in the management of oxidative stress-related diseases whereas antimicrobials kill or inhibit the growth of microorganisms, helping in managing infectious diseases. Despite several benefits of medicinal plants in herbal medicine, there is insufficient scientific data to support the use of the herbal drugs. Though a lot of studies have dealt with the studies for antioxidant and antimicrobial drugs from plants, there is less information on antioxidant and antimicrobial activity especially from Adenia lobata, Ipomoea aquatica, and Rubia cordifolia against Candida albicans, Pseudomonas aeruginosa, and Staphylococcus aureus. This study was conducted at Maseno University and Kenya Marine and Fisheries Research Institute. The objectives of the study were to determine (i) the phytochemicals present in leaf, stem, and root extracts of the selected climbers; (ii) determine the antioxidant activity of the extracts and (iii) determine the effect of the extracts on the growth of the specified test organisms. A. lobata, I. aquatica, and R. cordifolia samples were collected using targeted random sampling method from Kisii (0°54'38.7"S 34°48'28.95"E), Lake Victoria shorelines-Kisumu City (0°7'33"S 34°44'33.6"E), and Maseno University farm (0°0'32.6"S 34°36'28.6"E) respectively. The samples were separated into leaf, stem and root parts which were air-dried in the shade, ground into powder and then soaked in methanol. Thereafter, filtration was done and the extracts concentrated. Phytochemical analysis was conducted to test the presence of saponins, tannins, terpenoids, steroids, flavonoids, alkaloids, cardiac glycosides, and anthraquinones. Thin layer chromatography (TLC), was conducted as a semi-quantitative phytochemical analysis. The antioxidant activity was assessed using 2, 2-diphenylpicryl-1-hydrazyl free radical scavenging method with ascorbic acid as reference standard. Microbial growth was assessed using the disc diffusion method at 25, 50, 75, and 100 mg/ml concentrations. Minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and minimum fungicidal concentration (MFC) were also determined. The antioxidant and antimicrobial assays were done in triplicates in a completely randomized design. Data collected was subjected to analysis of variance (ANOVA) followed by post hoc tests. Means were separated and compared using the least significant difference (LSD) at P≤0.05. Phytochemical analysis revealed presence of alkaloids, tannins, terpenoids, steroids, glycosides, saponins, flavonoids, and anthraquinones in the extracts. TLC analysis indicated R. cordifolia leaf extract had the highest number of phytochemical spots (14), while A. lobata and I. aquatica root extracts had the lowest spots (2). A. lobata root extract showed relatively high antioxidant activity with IC50 value of (116.25 μg/ml), and I. aquatica roots had lowest antioxidant activity of (355.16 μg/ml). The extracts significantly inhibited the growth of the three test organisms. The largest zones of inhibition were observed in 100 mg/ml R. cordifolia leaf extract against P. aeruginosa (21.00 mm), and S. aureus (15.57 mm), and 100 mg/ml in R. cordifolia root extract against C. albicans (11.67 mm). The extracts exhibited significant bacteriostatic and fungistatic activity. MIC values for R. cordifolia extracts against C. albicans was 5 mg/ml for both leaf and root extracts and 10 mg/ml for the stem whereas for both P. aeruginosa and S. aureus were all at 10 mg/ml. The study has demonstrated that A. lobata roots is a potential source of antioxidants. R. cordifolia has been shown to be active against C. albicans, P. aeruginosa and S. aureus. Bioactive activity in plants is primarily linked to phytochemicals. The study has revealed that the extracts from these climbing plants can be used as potential antioxidant and antimicrobial agents for the development of new drugs. With this evidence of bioactive secondary metabolites, there is need for further studies to isolate and characterize phytochemicals associated with these plants. Master's Thesis

Molecular determination of Plasmodium falciparum parasites with histidine-rich protein 2/3 gene deletions in a holoendemic area, Siaya county, western Kenya

Sun, 01 Jan 2023 00:00:00 GMT

Molecular determination of Plasmodium falciparum parasites with histidine-rich protein 2/3 gene deletions in a holoendemic area, Siaya county, western Kenya ACHIENG’, Sharley Wasena Malaria remains endemic in western Kenya despite the various control interventions. Accurate diagnosis is key to the treatment and control of malaria. As such, the World Health Organization (WHO) recommends parasite-based confirmation of malaria prior to treatment. Plasmodium falciparum Histidine Rich Protein 2 (PfHRP2) based malaria Rapid Diagnostic tests (mRDTs) kits are commonly used throughout western Kenya as an alternative to microscopy in malaria diagnosis. However, the emergence of P. falciparum isolates with HRP2/3 deletions has threatened the sensitivity and performance of the mRDT kits. In western Kenya, a high transmission holoendemic area, polyclonal P. falciparum infection could be present but masked by the wild type gene. This may lead to both underestimation of deletion prevalence and increase in the spread of parasites with deletion. False negative mRDTs pose a public health threat towards malaria treatment and elimination progress. This is because the proportion of malaria infected patients having these gene deletions will go undetected by the PfHRP2-mRDTs, and therefore remain untreated. Siaya County borders western Uganda, an area where massive reports of HRP2/3 gene deletions have been reported and hence the need to conduct HRP2 surveillance in the study area. The study therefore investigated PfHRP2/3 deletions in a paediatric cohort from Siaya county, in western Kenya. Specifically, the study determined the prevalence of PfHRP2/3 deletion and the relative strain abundance of deleted strains in polyclonal infections, compared P. falciparum parasite densities estimated by qPCR and microscopy and evaluated the performance of the mRDT and microscopy techniques that routinely used for malaria diagnosis in Siaya County, western Kenya. The study being retrospective in nature, archived RBC pellets extracted from EDTA blood of children (n=219) who were previously enrolled in the study was used for DNA extraction. In order to achieve all the objectives, the study utilized one-step multiplex quantitative polymerase chain reaction (qPCR). The multiplex qPCR was designed using three differently labelled TaqMan assays detecting the PfHRP2 (PF3D7_0831800) and PfHRP3 (PF3D7_1372200) genes. Overall, PfHRP2/3 deletions were detected in 12 (5.6%) parasite isolates. The PfHRP2 monoclonal deleted strains were present in 2 isolates (1%) isolates while no parasite isolates harbored PfHRP3 single deletion. Further, 9 (4.1%) isolates had deleted PfHRP2 and 1 (0.5%) had PfHRP3 deleted but were masked in polyclonal infection. The average relative abundance of PfHRP2 deleted parasites was 9.6% while wild type was 90.4% in polyclonal infections. The multiplex qPCR demonstrated a higher ability in detecting malaria parasites compared to microscopy with median (IQR) of 8.28E4 (2.75E6) and 6.24E3 (2.45E4) respectively (P≤0.001). Further, there was a positive correlation between parasite detection by qPCR and microscopy (r=0.59, P≤0.001). On evaluation of the performance of clinical diagnostic techniques used in Siaya County Referral Hospital (SCRH), microscopy demonstrated a higher diagnostic sensitivity of 97.6% (95%CI, 89.8-105.0) and specificity of 26.0% (95%CI, 22.4-29.6) as compared to mRDT. Cohen Kappa’s test revealed a fair agreement between microscopy and qPCR, (k=0.30, P≤0.001). The study provides evidence of PfHRP2 deleted strains including those that are masked in polyclonal infections and their relative abundance in Siaya County. Further the study highlights microscopy as a more sensitive and specific technique in detecting malaria infection as compared to mRDT test. Master's Thesis

Prevalence and antibiotic susceptibility patterns of cotrimoxazole resistant bacterial uropathogens from Hiv patients attending Maseno mission hospital, western kenya

Sun, 01 Jan 2023 00:00:00 GMT

Prevalence and antibiotic susceptibility patterns of cotrimoxazole resistant bacterial uropathogens from Hiv patients attending Maseno mission hospital, western kenya MUSASIA, Philip Sakwa Urinary tract infections (UTI) are inflammatory disorders within the urinary system due to presence of pathogenic microorganisms. Taking advantage of lowered immunity in Human Immuno-deficiency Virus (HIV) patients, bacterial UTI globally contribute up to 60% of opportunistic infections (OI). Previous HIV related (OI) studies concentrated on malaria and sexually transmitted diseases ignoring UTI. Owing to its wide interceptive range, cotrimoxazole was recommended for prophylaxis by world health organization and has been standard against opportunistic infections. However, with prolonged use, bacterial resistance is being reported. Uncertainty of UTI management following rising reports of bacterial resistance to commonly used drugs in AIDS era is real. This hospital based descriptive study investigated prevalence and antibiotic susceptibility patterns of cotrimoxazole resistant bacterial uropathogens from HIV patients attending Maseno mission hospital in western Kenya where treatment is blindly based on chance and history during clinic visits without laboratory testing. Specifically described bacterial UTI prevalence, determined their response to cotrimoxazole and established antibiotic susceptibility patterns. At interval of six, 354 participants were systematically selected from population of 2000. Mid-stream urine was cultured on cysteine lactose electrolyte deficiency agar and samples obtaining >105 colony forming units determined by colony counters were biochemically characterized before being subjected to cotrimoxazole broth dilution sensitivity testing. Isolates not responding to cotrimoxazole were subjected to disc diffusion susceptibility testing in accordance with Clinical Laboratory Standards Institute’s guidelines. Tables, graphs and charts were used to present generated data which was analyzed alongside structured questionnaire captured demographic information. Prevalence of UTI among HIV patients was 117(33%) where 75.4% originated from females and 25.6% males with Escherichia coli (54.7%) being most encountered. Chi square analysis of bacterial responses to cotrimoxazole evaluated at 95 % confidence level was statistically significant at P<0.05 (x)2 (5)=14.3, p< 0.005 with an average susceptibility of 46.6%. Significantly, study results will serve as reference point for future researchers and medics, formed policy reevaluation for prophylaxis program and enhanced better OI management by establishing susceptibility patterns and suitable antibiograms. Study recommended for constant monitoring profile aetiological bacteria, periodic surveillance of for prophylactic drug and practicing scientifically determined treatment recommending most susceptible gentamicin (80.3%) empirical antibiotic for region with 53.4% resistance Master's Thesis

Sero-prevalence of Brucellosis and associated risk factors to Brucella pathogen in Tiaty, Baringo County, Kenya

Sun, 01 Jan 2023 00:00:00 GMT

Sero-prevalence of Brucellosis and associated risk factors to Brucella pathogen in Tiaty, Baringo County, Kenya OUMA, Dickens Oloo Brucellosis is a global zoonotic disease caused by Brucella pathogen, which affects man and animals. The disease has been reported across the world including Kenya. Information regarding the sero-prevalence and risk factors of brucellosis in camels in the pastoral Tiaty is still scanty despite several reported loss of camels due to zoonotic diseases. The main objective of this study was to investigate sero-prevalence of brucellosis, and the Risk factors associated with Brucella pathogen infections in Tiaty Sub-county, Baringo County. Thus a cross-sectional study was conducted in the study area whereby a total of 105 sera samples were collected from camels i five study locations, dominated by camel farmers using a multi stage sampling technique. The samples were tested to detect antibodies against Brucella by competitive immunosorbent assay (cELISA) as per the manufacturer’s instructions. In addition, data, on location of camels, history of retained placenta or abortion, gender, and age was gathered using a questionnaire. DNA extraction and purification was done on the positive samples using the Norgen bacterial genomic DNA isolation kit, then the quality and quantity of DNA were determined according to the manufacturer’s instructions. The data on risk factors was analyzed using chi-squared test (X2) at 95% confidence interval (CI) to investigate the relationship between brucellosis and the risk factors. The overall sero-prevalence of 20.0% was reported in camels. The percentage sero-prevalence per study location indicated that Ribikwo had the highest seroprevalence of 38.1% while Silale recorded the least 14.3%. Chemolingot, Lolyamorok and Kollowa locations recorded 28.6%, 19.0% and 0.0% respectively. The proportions of seropositivity in the study locations were significantly higher which revealed a significant association between sero-positivity of camel brucellosis with the location, (p ═ 0.037). It was revealed that brucellosis was associated with age and gender of the camels, further logistic regression analysis on gender revealed that there was 4 times more likelihood of females being seropositive as compared to males (OR = 4.329, 95% CI = 0.971-19.307, P-value 0.050). Further, logistic regression analysis on age revealed that there was 5.8 times more likelihood of seropositivity of brucellosis occurring in camels < 2 years old compared to those aged 2-3 years and those over 3 years old (OR = 5.845, 95% CI = 1.340-25.489, P-value 0.019). History of abortion was found to have no significant association with camel brucellosis which implies that abortions in camels is mainly not linked to brucellosis. Further logistic regression analysis on the age of the camels found out that there is 0.5 times more likelihood of brucellosis occurring in camels with history of abortion (OR = 0.522, 95% CI = 0.118-2.305, P-value 0.391). The high sero-prevalence reported in this study incriminates cattle, sheep and goats as the source of infection since they are reared in close association with camels. The determinant of brucellosis seropositivity were gender and age of camels. Therefore, brucellosis control programs targeting multiple species cattle, goats’ sheep, and camel should be formulated to curb spread of the disease. Master's Thesis

Association between hiv-1 subtypes and drug resistance to first line therapy among individuals with advanced disease in Homa bay county, Kenya

Sun, 01 Jan 2023 00:00:00 GMT

Association between hiv-1 subtypes and drug resistance to first line therapy among individuals with advanced disease in Homa bay county, Kenya NYAKUNDI, Nelson Mokay Human Immunodeficiency Virus-1 remains a public health threat globally, and although antiretroviral therapy has greatly improved the lives of people living with HIV, challenges persist due to high HIV genetic diversity and drug resistance. Analysis of HIV-1 subtypes over time demonstrates high subtype diversity and dynamic changes with variations in drug resistance among different HIV-1 subtypes. Homa Bay County carries one of the highest HIV-1 burdens worldwide, yet up-to-date information on circulating subtypes and subtype-specific drug resistance is lacking. The general objective of this study was to determine the association between HIV-1 subtypes and drug resistance to first-line ART among individuals with advanced disease in Homa Bay County. The study specifically sought to characterize HIV-1 subtypes and recombinants, to determine HIV-1 subtype-specific drug-resistance mutations, and to determine HIV-1 subtype-specific polymorphisms associated with drug resistance among HIV-1 patients on first-line ART in Homa Bay County, Kenya. A facility-based, cross-sectional survey was conducted, enrolling individuals aged 15 years and above, with advanced HIV-1 disease, and who had been on first-line ART for at least 6 months. The sample size determined using Cochran's formula was 70 participants. Plasma samples were analyzed using CAP/CTM real-time PCR for HIV-1 viral load and genotyping was performed on dried blood spots for samples with a viral load ≥1000 copies/mL. A genetic analysis of a 1,084-bp fragment of the HIV-1 pol gene encoding amino acids 6-99 of the protease (PR) and 1-251 of the reverse transcriptase (RT) region from 65 participants was performed using an in-house assay. Drug resistance was determined using the Stanford University HIV Database. Subtypes were identified using phylogenetic analysis and REGA subtyping tool. Fisher’s exact test was used to assess the association between subtypes and drug-resistance mutations. Samples with a significant association were subjected to logistic regression analysis, controlling for regimen. The findings revealed that subtype A1 was most prevalent in 46 patients (70.8%), with lower prevalence rates for subtype D (n=9, 13.8%), recombinants (n=6, 9.2%) and A2 (n=4, 6.2%). No statistically significant association was observed between subtypes and nucleoside reverse transcriptase inhibitor (NRTI) mutations or polymorphisms. However, a notable finding was the lower likelihood of observing NNRTI mutations K101E/H and Y181C/I/V, associated with high-level resistance to Nevirapine (NVP) and Efavirenz (EFV), in subtype A1 than in other subtypes (OR=0.14, 95% CI=0.03-0.60 and OR=0.21, 95% CI=0.06-0.72, respectively). This analysis found that HIV-1 subtypes are associated with resistance mutations, with K101 and Y181 less likely in subtype A. This suggests that areas with a high prevalence of subtype A1 may experience reduced compromised efficacy of NVP and EFV as first-line ART options. This implies that ART choice may need to be tailored to the HIV-1 subtypes. However, further investigations with large sample sizes and longitudinal designs are warranted to confirm this and assess the clinical impact of these subtype-specific differences to inform ART regimen policies effectively. Master's Thesis

Evaluation of anti-leishmanial activities of Olea europaea, Kigelia africana, Terminalia mollis, Croton macrostachyus, and Bridella micrantha extracts

Sun, 01 Jan 2023 00:00:00 GMT

Evaluation of anti-leishmanial activities of Olea europaea, Kigelia africana, Terminalia mollis, Croton macrostachyus, and Bridella micrantha extracts WAFULA, Dennis Mukhwana Leishmaniasis pose a significant medical concern worldwide which if left untreated, can be fatal. Although expensive and toxic, pentavalent antimonial drugs are the first-line treatment for leishmaniasis raising the need to find an alternative. Plants extracts contain several bioactive compounds that may offer alternative therapeutic signifcance to already developed antileismanial drugs. Based on folkloric information, a number of plants such as Olea europaea, Kigelia africana, Terminalia mollis, Croton macrostachyus, and Bridella micrantha are used to cure Leishmaniasis in the endemic regions in Baringo County. However, no studies have been done to find out the phytochemical compounds, as well as the anti-leishmanial properties. The aim of this study was to investigate the anti-leishmanial activity of Olea europaea, Kigelia africana, Terminalia mollis, Croton macrostachyus, and Bridella micrantha extracts. The plants were collected from Marigat in Baringo County (Kenya), air-dried, ground into fine particles, and extracted at the KEMRI Center for Traditional Medicine and Drug Research. Completely randomized design (CRD) was used in the in vivo studies. In vitro investigations were carried on L. major promastigotes to determine inhibitory concentrations (IC50) and Minimum Inhibition Concentrations (MIC) and toxicity on Vero cells. A total of 35 mice, 6 in each experimental group were used. For in vivo bioassays, the mice were injected intradermally on the left posterior footpad with 1×106 stationary phase flagellate forms of cultured L. major, and then housed for 4 weeks to allow disease manifestation. Differences between treatment groups exposed to different drugs were examined using logistic regression for each parameter studied. Phytochemical components of bark of the plants contain glycoside, terpenoids, tannins, flavonoids, steroids, and saponins. T. mollis contaned higher concentration of tannins, phenols, alkaloids, and anthraquinone. The crude extracts from the plants significantly inhibited promastogote and amastigote growth (P ˂ 0.05) after 24 hours of exposure, with the standard drug (Amphotericin B) being the most effective, while T. mollis was highly potent on amastigote among the plant extracts, then, C. macrostachyus, while O. europeae was least effective. Among the plant extracts, T. mollis had the highest efficacy (IC50 = 85.5 mg/l) in promastigote, while the least effective was O. europaea. In amastigotes, T. mollis exhibited the highest anti-amastigote activity (IC50 = 96.5 mg/l). The Leishman Donovan Units (LDU) of the L. major infection was lowest (0.12 × 106) in L. major-infected mice treated with T. mollis, while that of O. europeae was highest (4.12 × 106). After four weeks, mice administered with T. mollis demonstrated the greatest decrease in lesion diameter (0.68 ± 0.07 mm). Therapy with T. mollis, resulted in a considerably greater lesion reduction in mice. After 4 weeks of treatment, mice administered with O. europeae had the smallest reduction in lesion diameters of all the drugs examined. After 24 hours of treatment, the different drugs significantly influenced mammalian cell survival (P ˂ 0.05), with percentage cell viability of herbal medications, B. microstachyus, and O. africana causing the most toxic effects. It is concluded that among the extracts from the back of the five test plants, T. mollis at concentraton 85.5 to 96 mg/l was the most effective against Leishmania infection. Over all, the results obtained from the crude extracts screening, suggest that these may be promising sources for the development of new drugs for controlling leishmaniasis. The study recommends the use of T. mollis as the most effective plant extract in management of leishmaniases. PhD Theses

Relationship between the concentration of microcystin in serum and primary liver cancer in patients attending JOOTRH, Kisumu, Kenya

Sat, 01 Jan 2022 00:00:00 GMT

Relationship between the concentration of microcystin in serum and primary liver cancer in patients attending JOOTRH, Kisumu, Kenya CHEMUTAI, Evaline Microcystins (MCs) are toxins released by unicellular micro-organism known as cyanobacteria. These unicellular micro-organisms flourish in aquatic habitats with warm temperatures and nutrients rich water bodies. Microcystins are known to cause Liver cancer via inhibition of serine-threonine protein phosphatases (PP1 & 2A). Previous studies in China and Serbia have shown an association between MCs and occurrence of primary liver cancer (PLC). However, in Kenya, there is no data that links MCs and occurrence of PLC in humans despite there being an existence of harmful cyanobacteria within Nyanza gulf. This cross- sectional study was therefore carried out to investigate the association of serum microcystins to occurrence of PLC. The specific objectives were to determine the levels of MCs and nutrients within the Nyanza gulf; to determine the levels of serum serine-threonine protein phosphatases (PP 1 & 2A) in PLC patients and controls and to determine the correlation between mcyE and mlrA genes. A total of 40 study participants were enrolled; 20 PLC patients confirmed from JaramogiOgingaOdinga Teaching and Referral Hospital were recruited and 20 controls obtained from the source population giving rise to PLC cases. Their demographic information was obtained through questionnaire. Blood samples were collected and analyzed for total serum-serine-threonine PP1 and 2A levels using enzyme-linked immunosorbent assay. Water samples were collected from three sites along Nyanza gulf; Dunga, Homabay, Mbita; and filtration was done using 0.45 μmmembrane filter using vacuum pump to concentrate phytoplanktons. MC-toxin levels in water were measured using MC-ELISA Kit, whereas Nutrients analysis was done using photometric methods. Total RNA was extracted and quantification of mlrA and mcyE gene expression was done using RT-PCR software. MCs levels were detected using ELISA reader, Nutrients levels were quantified using spectrophotometer; whereas total serine-threonine protein phosphatase quantification were done using ELISA reader and analyzed using independent t-test, while correlation of gene expression analysis was done using Pearson correlation. RESULTS: MCs toxins were present in all samples sites, and their levels exceeded recommended WHO guidelines of 1.0µg/l for quality water safety. Highest MCs levels were detected in Homa/Bay B site (21.4 µg/L) which corresponded to high nitrogen and phosphorus levels within the same station (498.64 µg/L: 136.145 µg/L) respectively. This was followed by Mbita A (13.1 µg/L; TP = 46.14 µg/L; TN = 327.32 µg/L), Homa/Bay A (7.9 µg/L; TP = 100 µg/L; TN = 395.21 µg/L), as well as Dunga B (7.1 µg/L; TP = 84.71 µg/L; TN = 308.37 µg/L) and Mbita B (5.9µg/L; TP = 59 µg/L; TN = 330.47 µg/L). However, Dunga site A had the low level of toxin detection of about (2.2 µg/L, TP = 71.14 µg/L; TN = 247.58 µg/L) despite having high levels of nitrogen. Higher levels of serine-threonine protein phophatase activities were observed in liver cancer patients (103.9 ± 17.83 µg/ml) than in controls (25.91 ± 3.342 µg/ml). The variation was statistically significant (P < 0.05; t38 = 4.298;). Water sample analysis provided evidence of expression of MC-synthesizing gene (mcyE) and MC-biodegrading gene (mlrA). Pearson correlation revealed an insignificant moderate positive relationship between mcyE and mlrA genes, (r = 0.625, P (two-tailed) = 0.053). In conclusion, Nyanza gulf presented a high levels of MCs beyond recommended WHO values (1 µg/L) hence posing a great health risk to riparian communities. This could be attributed to variations in the levels of serine-threonine liver enzyme in PLC patients in the study, due to consumption of MC-contaminated water. Therefore, it is necessary to strengthen the protection and monitoring of drinking water source for effective control of water pollution and safeguarding of human health. Masters Thesis

HEMATOLOGICAL AND BIOCHEMICAL PARAMETERS AS ALTERNATIVE MARKERS OF STOCKING DENSITY- INDUCED STRESS IN CULTURED NILE TILAPIA, (Oreochromis niloticus Linnaeus, 1758) UNDER CULTURE CONDITIONS.

Sat, 01 Jan 2022 00:00:00 GMT

HEMATOLOGICAL AND BIOCHEMICAL PARAMETERS AS ALTERNATIVE MARKERS OF STOCKING DENSITY- INDUCED STRESS IN CULTURED NILE TILAPIA, (Oreochromis niloticus Linnaeus, 1758) UNDER CULTURE CONDITIONS. ONYANGO, ELIJA ODHIAMBO Fish and fisheries play a significant role in meeting nutritional food security, more so of the poor communities in the low and middle income countries (LMIC).Oreochromis niloticusisone the most valuable warm water fishes used in aquaculture systems. Various stressors trigger specific stress responses in O. niloticus. High stocking density (HSD) causes persistent stress response, which brings about inhibited fish breeding, growth, and decreased immune capacity thus decreased yield. Studies have been done in other regions of the world to evaluate the various hematological and biochemical parameters in O. niloticus as markers of stress, thus this study evaluated four specific parameters and to customize them for use by O. niloticus farmers in the Counties boardering Lake Victoria in Kenya. Two specific objectives were evaluated in this study namely; to determine the relative hematological parameter; erythrocyte count in cultured O. niloticus, subjected to stocking density induced stress and to determine the relative biochemical parameters; plasma cortisol, whole blood glucose, and blood electrolytes concentrations in cultured O. niloticus, subjected to stocking density induced stress. Stocking rates for the aquaria was done by matching all the fish for body weight/standard mass, Mean± SD (15 ± 1 g) thus, 150 g and 300 g of body weight/ 0.05 m3aquarium for low stocking density (LSD) and HSD respectively both in triplicates. The fish were cultured for 21 days following 3 days of acclimatization to the laboratory conditions after being obtained from the University`s fish rearing facility by seining technique. The fish were fed on a carbohydrate based feed of chick mash at 18 % protein supplemented with crushed Rastrineobola argentea to 25 % protein at a feed portion of 10 g/kg of life body weight. Blood samples were drawn using cardiac puncture technique from fish anaestethised with 2 – phenoxyethanol at 0.30 ml.l-1 from each aquarium (n = 5) using a large sieve. Erythrocyte counts (n = 18) were assayed for using hemocytometer. Plasma cortisol concentration (ng/ml) (n =18) levels were assayed for by use of Enzyme-Linked Immunosorbent Assay. Whole blood glucose concentration levels (n = 30) were determined using a hand-held One Touch Ultra glucose meter (MD-300) and test strips. Electrolyte concentrations (n =18) were assayed using Sherwood Flame Photometer 410. Whole blood glucose analysis revealed statistical (P< 0.05) difference in the means in HSD and LSD O. niloticus groups. Mean plasma glucose concentration was statistically significantly (P ≤ 0.01) higher for HSD than LSD O. niloticus groups at mean ± SD, 96.84 ± 5.28mgd-1 and 76.80 ± 5.92 mgd-1respectively. One-way Analysis of Variance (ANOVA) was done on the data collected and comparison of significant differences in means done between LSD and HSD at 0.01%. Plasma cortisol levels revealed statistically (P ≤ 0.01) significant values of HSD at mean ± SD, 6.35 ± 0.89ng/ml and 4.49 ± 1.08 ng/ml respectively. One way ANOVA analysis of the true electrolyte means for the LSD and HSDO. niloticus groups revealed significantly (P ˂ 0.05) higher mean at mean ± SD (1.63 ± 0.18 mmol/l) HSD and (1.11 ± 0.08 mmol/l)LSD for Na+, (0.73 ± 0.03 mmol/l)HSD and (0.42 ± 0.02 mmol/l)LSD for K+ with no statistical (P˂ 0.05) difference at mean ± SD (0.13 ± 0.00)HSD and (0.13 ± 0.00)LSD for Ca++.One-Way ANOVA analysis revealed significant (P ˂ 0.05) difference in the erythrocyte count means in LSD and HSD O. niloticus groups at mean ± SD, 7.01 ± 0.77 x 106 mm-3 and 3.36 ± 0.63 x 106 mm-3 respectively. Overally, the findings of this study demonstrate that high HSD increase erythrocyte count, plasma cortisol, whole blood glucose, and Na+ and K+ concentration in O. niloticus fish indicating a marked increase in stress levels. Elevated erythrocyte count, plasma cortisol, whole blood glucose, and Na+ and K+ concentrations can be used as alternative biomarkers for acute stress in O. niloticus produced under aquaculture systems. The findings of this study can help improve aquaculture practices on management of chronic stress in O. niloticus and related Cichlids under industrial aquaculture production through timely diagnosis and deployment of appropriate mitigation measures.