Department of Biochemistry and Biotechnology

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Browsing Department of Biochemistry and Biotechnology by Author "Okanya, Patrick"

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    Development and evaluation of a loop-mediated isothermal amplification (LAMP) diagnostic test for detection of whipworm, Trichuris trichiura, in faecal samples
    (2023-03-06) Okanya, Patrick; Ngari, M.G.; Mwangi, I.N.
    Whipworm infection or trichuriasis caused by Trichuris trichiura is of major public health concern in sub-Saharan Africa, particularly among pre-school and school-going children. It is among the neglected tropical diseases targeted for elimination through mass drug administration (MDA). One of the outcomes of MDA is a rapid decline in levels of infection intensity, making it difficult to monitor effectiveness of control measures using the conventional Kato–Katz procedure, which relies on the microscopic detection of parasite ova in faecal samples. In the present study, a loop-mediated isothermal amplification (LAMP) test was developed for the detection of T. trichiura infection in faecal samples. LAMP technology offers greater sensitivity and specificity than the microscopy-based tests. A set of four specific primers targeting the internal transcribed spacer 2 region of the ribosomal DNA were designed using Primer Explorer software. DNA was extracted from faecal samples using the alkaline lysis method (HotSHOT) and the LAMP reaction performed at 63°C for 1 h. The amplicons were visualized by both gel electrophoresis and with the naked eye following staining with SYBR green dye. Sensitivity and specificity tests were determined using the standard Kato–Katz diagnostic procedure as a reference test. The developed LAMP assay reliably detected T. trichiura DNA in faecal samples, with a specificity and sensitivity of 88% and 77%, respectively. No cross-reactivity was observed with several common helminth parasites. The developed LAMP assay is an appropriate diagnostic method for the detection of T. trichiura DNA in human faecal samples due to its simplicity, low cost, high sensitivity and specificity.
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    Lanyamycin, a macrolide antibiotic from Sorangium cellulosum, strain Soce 481 (Myxobacteria)
    (2023-03-06) Okanya, Patrick; Mulwa, Lucky; Jansen, Rolf
    Lanyamycin (1/2), a secondary metabolite occurring as two epimers, was isolated from the myxobacterium Sorangium cellulosum, strain Soce 481. The structures of both epimers were elucidated from HRESIMS and 1D and 2D NMR data and the relative configuration of their macrolactone ring was assigned based on NOE and vicinal 1H NMR coupling constants and by calculation of a 3D model. Lanyamycin inhibited HCV infection into mammalian liver cells with an IC50 value of 11.8 µM, and exhibited a moderate cytotoxic activity against the mouse fibroblast cell line L929 and the human nasopharyngeal cell line KB3 with IC50 values of 3.1 and 1.5 μM, respectively, and also suppressed the growth of the Gram-positive bacterium Micrococcus luteus.
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    Rotavirus prevalence and seasonal distribution post vaccine introduction in Nairobi county Kenya
    (2023-03-06) Okanya, Patrick; Mbatia, Betty; Gikonyo, Joshua
    Rotaviruses are one of the leading etiological agents of gastroenteritis in young children, for which a monovalent G1P(8) vaccine has been provided for free in Kenyan since July 2014. The main objective was to estimate the post vaccine prevalence and seasonal distribution of rotavirus diarrhea in children less than 5 years in Nairobi County, Kenya. Rotavirus positive samples were collected from children below 5 years of age in two hospitals within Nairobi County where vaccination status was card-confirmed. The children were examined and the demographic and clinical profiles of the children were recorded. Fecal specimens were analyzed for rotavirus antigen using an ELISA kit, followed by characterization by PAGE. Out of the total 323 samples, 49 had detectable rotavirus infection, representing 15.2% prevalence. Age distribution of rotavirus prevalence was as follows: ≤ 6 months-8.5%, 7-12 months-27.4%, 13-24 months - 41.4%, 25-36 months - 16.4% while 36-65 months had 6.3%. Rotavirus diarrhea was more common in wet and cold months of the year, the highest prevalence being observed in August (24.5%), 12.3% in both July and March, while April scored a prevalence of 10.2%. Out of the 49 rotavirus positive children, 48 had vomiting and abdominal cramps while all had fever and watery stool. The prevalence of Rotaviral diarrhea in children less than 5 years in Nairobi County Kenya has greatly reduced following the vaccine introduction and is more common during the wet and cold seasons of the year.
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    Screening and Characterization of Hydrocarbonoclastic Bacteria Isolated from Oil-contaminated Soils from Auto Garages
    (2023-03-06) Okanya, Patrick; Mbatia, Betty Nyambura; Muge, Edward Kirwa
    Release of petroleum oil and its products into the environment is a worldwide concern. The present study focused on isolation, molecular identification, morphological and biochemical characterization of bacteria possessing hydrocarbon- degrading properties. The study also aimed at optimizing appropriate culture conditions for the isolates as well as screening for alkane hydroxylase enzyme. Out of twenty one microbes isolated, nine were selected based on their ability to utilize different hydrocarbons. The isolates were observed to mineralize heating oil, hexane, octane, toluene and diesel oil. PCR amplification of 16S rDNA gene revealed that the isolates belong to six different genera; Pseudomonas, Acinetobacter, Klebsiella, Enterobacter, Salmonella and Ochrobactrum. Based on their ability to degrade diesel oil, three isolates were selected and their growth conditions optimized. Optimum degradation was recorded at less than 1% substrate concentration, pH 7 and temperature range of between 30°C and 37°C and using yeast extract as nitrogen source. GC-MS analyses confirmed their diesel oil degrading properties. Alkane hydroxylase gene from one of the isolates (isolate 1C) was successfully amplified indicating its catabolic capabilities in degrading alkanes. Overall, the characterized isolates may constitute potential candidates for biotechnological application in environmental cleanup of petroleum contaminants.