Integrated modular microfluidic system for forensic Alu DNA typing

Driven by the numerous applications of genome-related research, fully integrated microfluidic systems have been developed that have advanced the capabilities of molecular and, in particular, genetic analyses. A brief overview on integrated microfluidic systems for DNA analysis is given in Chapter 1 followed by a report on micro-capillary electrophoresis (µCE) of Alu elements with laser-induced fluorescence (LIF) detection, in which the monomorphic Alu insertions on the X and Y chromosomes were utilized to detect male DNA in large female DNA background (Y: X = 1:19) without cell sorting prior to the determination. The polymorphic Alu loci with known restricted geographical distribution were used for ethnicity determination. A valveless integrated microsystem that consists of three modules is discussed as well: (1) A solid-phase extraction (SPE) module microfabricated on polycarbonate, for DNA extraction from whole cell lysates (extraction bed capacity ~209 ±35.6 ng/cm² of total DNA). (2) A continuous-flow polymerase chain reaction (CFPCR) module fabricated in polycarbonate (Tg ~150 ºC) in which selected gene fragments were amplified using biotin and fluorescently-labeled primers accomplished by continuously shuttling small packets of PCR reagents and template through isothermal zones. (3) µCE module fabricated in poly(methylmethacrylate), which utilized a bioaffinity selection and purification bed (2.9-µL) to preconcentrate and purify the PCR products generated from the CFPCR module prior to µCE. Biotin-labeled CFPCR products were hydrostatically pumped through the streptavidin-modified bed where they were extracted onto the surface of the poly(methylmethacrylate) micropillars (50-µm width; 100-µm height; total surface area of ~117 mm²). This SPE process demonstrated high selectivity for biotinylated amplicons and utilized the strong streptavidin/biotin interaction (Kd =10-15M) to generate high recoveries. The SPE selected CFPCR products were thermally denatured and single stranded DNA released for size-based separations and LIF detection. The multiplexed SPE-CFPCR-µCE yielded detectable fluorescence signal (S/N≥3; LOD ~75 cells) for Alu DNA amplicons for gender and ethnicity determinations with a separation efficiency of ~1.5 x105 plates/m. Compared to traditional cross-T injection procedures typically used for µCE, the affinity preconcentration and injection procedure generated signal enhancements of 17-40 fold, critical for CFPCR thermal cyclers due to Taylor dispersion associated with their operation

Design, synthesis, and structure-activity relationship studies of dual Plasmodium falciparum phosphatidylinositol 4-kinase and cGMP-dependent protein kinase inhibitors

Malaria is a life-threatening disease caused by protists in the genus Plasmodium and transmitted by the female Anopheles mosquito. Amongst five species which infect humans, Plasmodium falciparum (Pf) causes the severest form of the disease. Although significant efforts have been made to reduce the overall impact of malaria in endemic regions, the ever emergence and continuous spread of parasite resistance to available chemotherapeutics, threatens to undermine advances made thus far. In addition, the current portfolio of drugs is non-effective in addressing chemoprotection, transmission blockade and relapse in P. vivax and P. ovale species. Thus, drugs targeting multiple stages of the parasite life cycle and of low risk to resistance, are highly desirable to support malaria elimination and/or eradication efforts. Considering the success of human kinase inhibitors as anti-cancer drugs and the identification of Plasmodium kinases as promising targets for malaria chemotherapy, this study aimed to optimize anti-plasmodium phosphatidylinositol 4-kinase (PI4K) and the cGMP-dependent protein kinase (PKG) inhibitors, based on two distinct chemotypes. Plasmodium PI4K and PKG are validated targets, each with the potential to deliver pan-stage active compounds with potentially moderate to low risk of resistance. Part 1 of this study focused on the repositioning of the oncological clinical Phase-1 mammalian target of rapamycin (mTOR) inhibitor, MLN0128, as a dual Plasmodium PI4K/PKG inhibitor for malaria. MLN0128 was identified by GlaxoSmithKline (GSK) Cellzome facility as a Plasmodium multi-kinase inhibitor with potent PI4K and PKG inhibitory activity. In this study, an in silico-guided structural modification strategy was undertaken towards optimizing dual Plasmodium kinase inhibition and anti-plasmodium activity while also mitigating potency against its oncological human target, mTOR and off-target PI4KIIIb (Figure 1). Arising from this work, analogues equipotent against both the chloroquine sensitive (PfNF54) and multi-drug resistant (PfK1) strains simultaneously targeting PI4K and PKG were identified. Docking studies using a PfPI4K homology model and a PvPKG crystal structure discerned the molecular features responsible for the high affinity of the inhibitors for these Plasmodium targets. Benzyl analogues containing a fluoro or chloro group at the meta or para positions displayed high anti-plasmodium activity with potent PvPI4K inhibition but weak PfPKG inhibition. Notable analogues included 7 (PfNF54 IC50 = 0.029 µM; PvPI4K IC50 = 0.007 µM; PfPKG IC50 > 2 µM) and 35 (PfNF54 IC50 = 0.086 µM; PvPI4K IC50 = 0.008 µM; PfPKG IC50 > 10 µM). Introduction of basic or pyridyl substituents proved important for dual Plasmodium kinase activity as exemplified by the active anti-plasmodium pyridyl analogues 44 (PfNF54 IC50 = 0.104 µM; PvPI4K IC50 = 0.004 µM; PfPKG IC50 = 0.834 µM) and 49 (PfNF54 IC50 = 0.189 µM; PvPI4K IC50 = 0.006 µM; PfPKG IC50 = 0.384 µM). In addition, the two compounds displayed low cytotoxicity against the Chinese Hamster Ovarian cell line, with a favorable selectivity index (CHO; SI > 100), low human ether-a-go-go-related gene (hERG) activity (IC50 > 10 µM) and high metabolic stability against human, rat, and mouse (H/R/M) liver microsomes (> 75% remaining after 30-min incubation). Selected compounds from the series also showed the potential for transmission blockade with specificity for stage IV/V gametocytes (IC50 100 µM). Compounds displayed potent PvPI4K inhibition but weak PfPKG inhibition (IC50 > 1 µM) in enzyme assays. Four compounds, including one sulfoxide analogue, displayed high stability when incubated with H/R/M liver microsomes in microsomal metabolic stability assays. These features also mitigated hERG activity as five analogues tested displayed an IC50 > 10 µM. Ultimately, a front-runner lead compound (86; GS1 16) with high biological activity and a good safety profile (PfNF54/PfK1 = 0.063/0.100 µM; PvPI4K IC50 = 0.003 µM; CHO SI > 793), optimal solubility (195 µM), favorable microsomal metabolic stability (H/R/M = 96/85/88%) and low affinity on the hERG-encoded potassium channel (IC50 = 44.80 µM), was identified for further progression

Contribution of Entrepreneurial Education and Training to Performance of Small Enterprises Run by Women in Kenya

Women run Small Enterprises have become increasingly important in creating employment and wealth and in growth of many economies in the developing countries. However it is only a few of these enterprises that record consistent growth. Access to entrepreneur education and training is one of the challenges that women run small enterprises encounter leading to bleak performance and sluggish growth. This paper presents evidence on why small enterprises run by women experience low grow. The research set out to investigate the extent to which entrepreneurial education and training contribute to performance of women run small enterprises in Kenya. The paper is as a result of primary data collected from enterprises run by women in the vibrant Langata Business District in Nairobi Kenya. Through use of ordinary least squares regression model, the study has derived sufficient evidence in explaining the variation in performance of women run small enterprises. The paper posits that there is a positive relationship between entrepreneurial education and training and performance of women run small enterprises. The study recommends that women running small enterprises should be encouraged to seek more entrepreneurial education and training which would enable them to run their enterprises in an effective manner, usher in growth and lead to optimal performance. Key words: Entrepreneurial education, Entrepreneurial training, Women run small enterprise

Influence of Government Support on Performance of Women Run Small Enterprises in Kenya: Langata Sub-County

All over the world, it is generally accepted that women run small enterprises (SEs) are becoming increasingly important in employment, wealth creation, and development of innovation. Small Enterprises encounter many challenges one of which is insufficient government support which hinder their performance and growth. This study sought to investigate why women run small enterprises grow less rapidly and shut down more often than men run enterprises. The objective of the study was to determine the extent to which government support influence the performance of women run small enterprises operating in Langata Sub-County, Kenya. The study adopted a descriptive research design and targeted a population of 118 women run small enterprises in Langata Sub-County in Nairobi County, Kenya. To collect primary data, a sample size of 91 enterprises was randomly selected for administering of questionnaire. The ordinary least squares (OLS) regression model derived from the analysis sufficiently explained the variation in performance of women run small enterprises. The findings indicated significant positive relationship between government support and performance of women run small enterprises in Langata sub-county. The study recommends that the government should increase its support to women run small enterprises because such support would greatly improve their performance. Keywords: Government support, Performance, Women-Run, Enterpris

Determinants of Foreign Direct Investment Growth: Kenya’s Manufacturing Sector

The FDI indicators in Kenya show a mixed signal and although institutional indexes for Kenya have been worsening over the years, foreign direct investment inflows (FDI) though sluggish has been on the rise. While many theories of FDI have been put forward, mostly ownership, location and internationalization (OLI), the extent to which institutional determinants influence growth of FDI in the Kenyan manufacturing sector has not gotten adequate attention. The combination of institutional determinants and OLI framework in determining flows of FDI has been avoided in this sector. This study using FDI inflows in the Kenyan manufacturing sector and governance indicators data performed a cross-sectional analysis for the period 2009-2013. The findings indicated significant positive relationship between governance and FDI growth which implies that governance determines growth of FDI in the Kenyan manufacturing sector. The evidence presented confirms that a good political climate and good corporate governance are important for foreign investment to flow into a Country. Keywords: foreign direct investment; corporate governance; political risk; manufacturing secto

Ceftriaxone-Resistant Salmonella enterica Serotype Typhimurium Sequence Type 313 from Kenyan Patients Is Associated with the blaCTX-M-15 Gene on a Novel IncHI2 Plasmid

Multidrug-resistant bacteria pose a major challenge to the clinical management of infections in resource-poor settings. Although nontyphoidal Salmonella (NTS) bacteria cause predominantly enteric self-limiting illness in developed countries, NTS is responsible for a huge burden of life-threatening bloodstream infections in sub-Saharan Africa. Here, we characterized nine S. Typhimurium isolates from an outbreak involving patients who initially failed to respond to ceftriaxone treatment at a referral hospital in Kenya. These Salmonella enterica serotype Typhimurium isolates were resistant to ampicillin, chloramphenicol, cefuroxime, ceftriaxone, aztreonam, cefepime, sulfamethoxazole-trimethoprim, and cefpodoxime. Resistance to β-lactams, including to ceftriaxone, was associated with carriage of a combination of blaCTX-M-15, blaOXA-1, and blaTEM-1 genes. The genes encoding resistance to heavy-metal ions were borne on the novel IncHI2 plasmid pKST313, which also carried a pair of class 1 integrons. All nine isolates formed a single clade within S. Typhimurium ST313, the major clone of an ongoing invasive NTS epidemic in the region. This emerging ceftriaxone-resistant clone may pose a major challenge in the management of invasive NTS in sub-Saharan Africa

A putative, novel coli surface antigen 8B (CS8B) of enterotoxigenic Escherichia coli

Enterotoxigenic Escherichia coli (ETEC) strains harbor multiple fimbriae and pili to mediate host colonization, including the type IVb pilus, colonization factor antigen III (CFA/III). Not all colonization factors are well characterized or known in toxin positive ETEC isolates, which may have an impact identifying ETEC isolates based on molecular screening of these biomarkers. We describe a novel coli surface antigen (CS) 8 subtype B (CS8B), a family of CFA/III pilus, in a toxin producing ETEC isolate from a Kenyan collection. In highlighting the existence of this putative CS, we provide the sequence and specific primers, which can be used alongside other ETEC primers previously described

Finite Difference Analysis on Magnetohydrodynamic Fluid Flow Past an Infinite Continuous Moving Surface Embedded in Porous Medium.

An analysis has been carried out on the effects of variable viscosity on the problem of Magnetohydrodynamic fluid flow past an infinite continuous moving sheet embedded in porous media. The fluid viscosity is assumed to vary as an inverse linear function of temperature

A Comparative Study on the Quality of Life of Leprosy Patients in Kilifi and Kwale Counties in Kenya

Background:Kenya at present is in the post-elimination phase of leprosy having achieved a prevalence of <1 case per 10,000 persons in 1989. In 2019 Kenya notified 163 leprosy patients, highest being in Kilifi and Kwale counties. About a quarter (26%) of the notified leprosy patients had grade 2 disability at the time of diagnosis, this being the most severe form of disability that may indicate a late diagnosis. This study aimed at assessing the quality of life of leprosy patients to guide policies and programs intended to enhance the health and well-being of leprosy patients.Materials and Methods:This was a case-control study conducted in Kilifi and Kwale Counties. For every leprosy index patient enrolled, two controls were identified within the same village to match the case. Descriptive statistics were used to summarise demographic and clinical variables. The World Health Organisation (WHOQOL-BREF) tool was used to measure the quality of life. The tool derived four (4) domains of physical health, psychological status, social relationship, and environmental profile. These were transformed into a scale between 0 to 100 for analysis. The F-test was used to compare mean scores in the four domains between cases and controls. The quality of life among the index cases against their controls was further analysed using conditional logistic regression models.Results: A total of 98 leprosy patients and 167 controls were evaluated for quality of life. On the perception of quality of life, leprosy patients had significantly lower mean transformed scores of 39 (SD 25) versus 49 (SD 25) p= <0.0001 compared to controls. Similarly, index cases had lower health satisfaction scores of 42 (SD 26) compared to controls scoring 61(SD 27) p=<0.001. Overall leprosy patients had statistically significant poorer scores on physical health, psychological health, social relationships and environmental QoL domains. Differences were most remarkable in the psychological domain, with a mean transformed score of 53 (SD 20) versus 68 (SD 16) p= < 0.0001 for controls. The overall quality of life model revealed that leprosy patients who were found to have either diabetes or hypertension enjoyed a better overall quality of life with OR of 10.98 and 1.22 respectively with a p-value <0.00001. Patients with tuberculosis and HIV presented the poorest quality of life with ORs of 0.49 and 0.14 respectively.Conclusion: The quality of life of the leprosy patients was significantly lower than that of the community controls in all the domains. Governments and communities need to prioritize rehabilitation measures such as the provision of artificial limbs, cataract surgery, and social protection disbursements to help leprosy victims improve their quality of life

Ethnicity and Breast Cancer Characteristics in Kenya

Purpose: There are no published data from specific regions of sub-Saharan Africa describing the clinical and pathological characteristics and molecular subtypes of invasive breast cancer by ethnic group. The purpose of this study was to investigate these characteristics among the three major ethno-cultural groupings in Kenya. Methods: The study included women with pathologically confirmed breast cancer diagnosed between March 2012 and May 2015 at 11 hospitals throughout Kenya. Sociodemographic, clinical, and reproductive data were collected by questionnaire, and pathology review and immunohistochemistry were performed centrally. Results: The 846 cases included 661 Bantus (78.1%), 143 Nilotes (16.9%), 19 Cushites (2.3%), and 23 patients of mixed ethnicity (2.7%). In analyses comparing the two major ethnic groups, Bantus were more educated, more overweight, had an older age at first birth, and had a younger age at menopause than Nilotes (p \u3c 0.05 for all comparisons). In analyses restricted to definitive surgery specimens, there were no statistically significant differences in tumor characteristics or molecular subtypes by ethnicity, although the Nilote tumors tended to be larger (OR for ≥ 5 cm vs. \u3c 2 cm: 3.86, 95% CI 0.77, 19.30) and were somewhat more likely to be HER2 enriched (OR for HER2 enriched vs. Luminal A/B: 1.41, 95% CI 0.79, 2.49). Conclusion: This case series showed no significant differences in breast cancer tumor characteristics or molecular subtypes, but significant differences in sociodemographic characteristics and reproductive factors, among the three major ethnic groups in Kenya. We suggest further evaluation of ethnic differences in breast cancer throughout the genetically and culturally diverse populations of sub-Saharan Africa