219 Records out of 22207 Records

Understanding resistance in inter-specific rice cultivars to the parasitic witchweed Striga

Author: Cissoko, Mamadou

Awarding University: University of Sheffield, England

Level : PhD

Year: 2012

Holding Libraries: DA. H1c 62-11517 ; Institute of Commonwealth Studies Library ;

Subject Terms: Striga ; Rice ; Oryza sativa ; Oryza glaberrima ; Striga hermonthica ; Striga asiatica ; Genetics ; Weeds ;

Abstract:

Both cultivated rice species, Oryza sativa (L.) and Oryza glaberrima (Steud.), are grown in Africa. To take advantage of superior traits from each species, AfricaRice Center and partners developed inter-specific rice cultivars called NERICA (NEw RICe for Africa) for rain-fed upland ecosystems. NERICA rice cultivars showed different susceptibilities to both S. hermonthica and S. asiatica species under controlled environment conditions. Some cultivars showed good broad-spectrum resistance against several Striga ecotypes and species whilst others showed intermediate resistance or were very susceptible. In addition, some cultivars showed resistance to a particular ecotype of Striga but were susceptible to others. The phenotype of a resistant interaction was often characterized by necrosis at the host parasite interface and an inability of the parasite to penetrate the host root endodermis. In general, the most resistant NERICA cultivars grew better than the very susceptible cultivars although even a small number of parasites caused a reduction in above ground host biomass. There was however, genetic variation for tolerance to Striga (the ability to grow and yield well in the presence of Striga) amongst the NERICA cultivars. The NERICA cultivars were also grown in field trials at Kyela in Tanzania (under S. asiatica infestation) and at Mbita Point in Kenya (under S. hermonthica infestation) in 2010 and 2011 to determine the impact of environment on the expression of resistance. The resistance of the NERICA cultivars against S. hermonthica and S. asiatica, in the field, was broadly similar to that observed in the laboratory although there were some exceptions. These results allow us to recommend particular cultivars for Striga-infested regions but they also illustrate the necessity of understanding the genetic basis of resistance to different ecotypes of Striga for breeding of durable resistance (and pyramiding of appropriate resistance genes) in host cultivars adapted to different rice agro-ecosystems in sub-Saharan Africa. Sixty four lines of an inter-specific CSSL population and the parent cultivars O. glaberrima MG12 and O. sativa Caiapo were phenotyped for resistance to S. hermonthica. MG12 showed good resistance to S. hermonthica whilst Caiapo was very susceptible. The CSSLs showed a range of susceptibility to the parasite, however, only two CSSLs showed the same strong resistance phenotype as MG12. Graphical genotyping and a Quantitative Trait Loci (QTL) analysis revealed a large QTL on chromosome 12 (designated STR12.1) which explained at least 80 % of the variation for resistance in the population and suggests that resistance to S. hermonthica (in MG12) is due to one (or a few genes) of major effect. This finding opens the way for the identification of candidate Striga resistance genes (through fine mapping approaches) and their transfer to farmer-preferred cultivars via marker assisted breeding.

Strategy implementation at the Kenya Animal Genetics Resource Center

Author: Kiarie, Richard Mwaura

Awarding University: University of Nairobi, Kenya

Level : MBA

Year: 2012

Holding Libraries: University of Nairobi Jomo Kenyatta Memorial Library ;

Subject Terms: Strategic management/Kenya Animal Genetics Resource Center ;

Abstract:

This project studied the strategy implementation process at the Kenya Animal Genetics Resource Center. The process of strategy implementation has many challenges leading to most organizations failing to achieve the objectives of the strategies. The findings revealed the strategy had seven SMART (specific, measurable, attainable, realistic and time bound) clear objectives developed with participation by stakeholders both internally and externally. The organization employed multiple methods to ensure adequate and clear communication to everybody involved. The Board and the MD had taken ownership of the strategy document and were spearheading its implementation in the organization and therefore provided effective leadership for the implementation process. A new organization structure that clearly defined the roles and responsibilities of various departments without overlap or conflict had been created. Performance contracts that were linked to the strategy objectives had also been introduced and embraced by all the employees. The implementation process also faced some key challenges. These were inadequate financial and human resources, organization culture that had persisted from the traditional civil service that was not responsive to change, compensation systems that had not been linked to strategy and lack of adequate information systems for monitoring the implementation progress. The management had responded to the challenges by phasing activities and prioritizing activities such that those within the current financial reach and with high probability of success are implemented first. A new scheme of service has been developed to enable the organization recruit, motivate and retain highly qualified personnel. Organizational values have been communicated to employees and placed in conspicuous places throughout the institution. Finally the acquisition of modem information technology system has been given priority. The study recommended capacity building for employees, implementation of a robust Information Technology system and a Knowledge Management System to enable it support creation, transfer and application of knowledge in the organization.

Pharmacogenetics of drug metabolizing enzymes and clinical implications in selected Kenyan populations

Author: Oluka, Margaret Ng'wono

Awarding University: University of Nairobi, Kenya

Level : PhD

Year: 2012

Holding Libraries: University of Nairobi Jomo Kenyatta Memorial Library ;

Subject Terms: Pharmacology ; Genetics ; Drug therapy ; Enzymes ; Clinical outcomes ; Minority and ethnic groups ;

Abstract:

Much of the inter-individual and interethnic variability in drug response is now attributable to genetic differences in drug metabolism. Drug metabolizing enzymes exhibit genetic polymorphism with certain allelic variants displaying striking variable ethnic distribution. The pharmacogenetics of drug metabolism has been extensively studied in Caucasian and Asian populations, yet much remains to be done in African populations. The main objective of this thesis was to investigate the genetic polymorphisms and clinical implications of genes encoding drug metabolizing enzymes namely, CYP2B6. CYP2C19, CYP2D6. N-acetyl transferase (NAT2) and Glutathione- S- transferases (GSTs) in selected Kenyan populations. The first specific objective was to determine the distribution of pharmacogenetically relevant single nucleotide polymorphisms (SNPs) of CYP2B6. CYP2C19. CYP2D6. NAT2, GSTMl and GSTTl in selected Kenyan populations. A population of 350 Kenyans belonging to three ethno-linguistically distinct populations was recruited. The populations studied were the Western Nilotes (Luo) (100), the Eastern Nilotes (Maasai) (152) and the Bantu (Kikuyu) (102). Genotyping of allelic variants CYP2B6*6(516G>T); CYP2C19*2(681G>A); CYP2C19*3(638G>A); CYP2D6*4(1846G>A); CYP2D6*5(deletion); CYP2D6* 17(1 023C>T); CYP~D6*29(3183G>A); NAT2*5(341T>C);.NAT2*6(590(G>A); NAT2*7(857G>A); .NAT2*14 (191G>A); GSTMl and GSTTl was undertaken by polymerase chain reaction coupled with restriction fragment length polymorphism (PCR-RFLP). Results showed that the distribution of CYP2D6, GSTMl and GSTTl allelic variants was highly variable between the three Kenyan ethnic populations. CYP 2D6*4 was surprisingly higher in the Eastern Nilotes (9%) than in the Western Nilotes (3%) and the Bantu (2%) (P=0.002). CYP2D6*17 was more prevalent in the Bantu (34%) than in the two Nilotic populations (18-23%) (P= 0.003). The frequency of GSTMl*O/*O (deletion) in the Eastern Nilotes (16%) was nearly half that in the Western Nilotes and the Bantu (30%) (P=0.009) whereas that of GSTTl *01*0 (deletion) in the Eastern Nilotes (41 %) was nearly double that found in the Bantu and Western Nilotes (22-26%) (~=0.005). The null allele CYP2C19*3 was undetected in the Bantu but was found at <1 % in the Nilotes. There was homogeneity in the distribution of CYP2C19*2 (10-18%), CJP2B6*6 (34-37%) and the NAT2 slow acetylator alleles (NAT2*5 (33-38%), NAT*6 (23- 27%), NAT2*7(3-6%), NAT2*14 (9-14%) between the three Kenyan ethnic populations. The intermediate metabolizer (lM) was: for CYP2D6 (20-27%), CYP2C19 (20-27%), CYP2B6 516TT (12-16%) whereas NAT2 slow acetylation was 54-64%. The second specific objective was to analyze clinically relevant genetic variants of CYP2D6 and CYP2C19 and evaluate their impact on psychotropic medication patterns in Kenyan psychiatric in-patients. A total of 193 psychiatric in-patients at Mathari Hospital were recruited and genotyped for CYP2C19*2 and CYF2C19*3; CYP2D6*2, CYP2D6*4, CYP2D6*5, CYP2D6*17 and CYP2D6*29 using PCRRFLP methods. CYP2D6 and CYP2C 19 metabolic phenotypes were predicted from observed genotypes according to published methods and related to the pattern and CYP metabolic profiles of psychotropic medications in the patient population. The distribution of SNPs of CYP2C19 and CYP2D6 in psychiatric inpatients was similar to that in the general population except for CYP2C 19*3 which was higher in patients (8%) than in the general population ?1%). Predicted intermediate metabolizer (IM) of CYP2D6 and CYP2C19 was 24% and 26% respectively. Predicted CYP2D6 lM phenotype was strongly associated with the use of high doses (15-20mg) of anticholinergic medication (68%) (P<O.OOI) indicative of serious extrapyramidal side effects; frequent hospital admission of 5-6 times (67%) per year (P<O.OOI) and exposure to antipsychotic polypharmacy of >5 drugs (P = 0.002). Psychotropic dosage regimens in the in-patients were higher than the defined daily doses (D

Localization and functional analysis of an Aquaporin gene (AQP 4886_gp) from Tsetse fly, Glossina pallidipes

Author: Bargul, Joel Ltilitan

Awarding University: Jomo Kenyatta University of Agriculture and Technology, Kenya

Level : MSc

Year: 2011

Holding Libraries: Jomo Kenyatta University of Agriculture and Technology Library ;

Subject Terms: Tsetse flies ; Glossina pallidipes ; Trypanosomiasis ; Rhipicephalus sanguineus ; Glossina morsitans morsitans ; Genetics ;

Abstract:

Tsetse flies are vectors of African trypanosomes, the protozoan agent of devastating disease, trypanosomiasis that afflicts both humans and animals. Currently, there is no promising vaccine in the horizon and treatment efforts are further constrained by the rapid increase in parasite drug resistance observed in patients. In addition, little effort is being made to develop new and effective drugs. Alternative methods to control trypanosomiasis and its transmission are therefore required. The trypanosome parasite develops into its infective metacyclic stage in the salivary glands of the tsetse fly, where the saliva provides a specific medium for its maturation and also becomes the fluid vehicle for the transfer of the parasites to the host through a blood meal. Water exchange across the salivary gland membrane occurs through aquaporin (AQP) water channels in brown dog tick, Rhipicephalus sanguineus. This study focused on the role(s) played by a putative water channel protein identified in the salivary glands of tsetse fly, Glossina pal/idipes, in relation to feeding and survival. The salivary gland AQP gene (herein named AQP 4886_gp), a homolog of GMsg 4886 gene from the transcriptome of Glossina morsitans morsitans, was PCRamplified and cloned from G. pallidipes. The AQP 4886_gp protein has a predicted molecular mass of 25.222 KDa. Topographic analysis suggested that AQP 4886_gp has the general aquaporin topology and possesses two conserved 'NPA' signature motifs (Asn-Pro-Ala) found in aquaporins. Multiple sequence alignment and protein distant tree plotted using Neighbour-Joining method shows that AQP 4886_gp is more closely related to many insect AQPs than vertebrates'. The AQP 4886_gp transcript was localized to the salivary glands, malpighian tubules, and midgut. These tissues are involved in high rates of water exchange in insects. The gene was detected in different life-cycle stages of the fly; larva, pupa, unfed teneral fly and adult tsetse fly using semi-quantitative reverse transcription (RT)-PCR. Functional studies of AQP 4886 ~ were carried out using RNA interference (RNAi) technique, where gene-specific double stranded RNA (dsRNA) was injected into experimental flies. The control group was injected with nuclease-free water (NFW). The effects of transient gene silencing were monitored by semi-quantitative RT-PCR, and relevant bioassays (survival, feeding success). AQP 4886~ gene knockdown was not lethal to the flies as they continued to survive and feed. The survival rates of 83% were achieved in both injected test and control groups. Binomial test of proportions showed no significant differences in the feeding success between the test (dsRNAinjected) and control (NFW-injected) flies at p<O.OS.

Genetic characterisation of Schistosoma mansoni populations from Mwea in Central Kenya using microsatellite markers with reference to host related factors, infection intensity and praziquantel treatment

Author: Agola, Eric Lelo

Awarding University: Jomo Kenyatta University of Agriculture and Technology, Kenya

Level : PhD

Year: 2011

Holding Libraries: Jomo Kenyatta University of Agriculture and Technology Library ;

Subject Terms: Schistosoma mansoni ; Mwea-Tebere Irrigation Scheme ; Bilharzia USE Schistosomiasis ; Deoxyribonucleic acid--DNA ; Genetics ;

Abstract:

Human schistosomiasis (or bilharzia) is a debilitating disease caused by blood flukes in the genus Schistosoma, and about 207 million people worldwide, mostly children, are infected. Schistosoma mansoni causes human intestinal schistosomiasis in Africa, Asia, parts of South America, and the Caribbean region; and is endemic in Kenya. Although S. mansoni is considered a uniform species morphologically, different isolates or strains may differ in a number of biological characteristics such as fecundity, infectivity, virulence, and response to drugs. It has been suggested that genetic variability may be responsible for these variations. A better understanding of population genetic characteristics of pathogens may help us understand mechanisms involved in several biological phenomenon including parasite response to chemotherapy. The present study was undertaken to determine the genetic diversity of S. mansoni miracidia, the larval forms hatched from parasite eggs isolated from fecal samples of infected school children living in Mwea, central Kenya; to assess the effect of host age, host sex, parasite infection intensity, and praziquantel (PZQ) treatment on genetic characteristics of S. mansoni miracidia populations; and finally, to determine the effect of passing S. mansoni through hosts on parasite genetic characteristics. Using a high-throughput technique that allows multi-locus micro satellite analysis of individual miracidia, 12 micro satellite markers were amplified from miracidial DNA by polymerase chain reaction (PCR) and the products analysed by micro satellite fragment analysis on a DNA sequencer. The mean number of alleles (MNA) observed per locus was in the range 8.22-10.22, expected heterozygosity in Hardy-Weinberg equilibrium was 0.68-0.70, and pairwise FST values ranged from 0.16% - 3.98% for the 12 populations analysed. Private alleles were found in 8 of the 12 populations, and the highest number recorded per population was 3. With respect to host age, there were no significant host agerelated differences in the MNA (r: = 0.333, d.f. = 1, P = 0.5640). However, there was a gradual decrease in the mean number of private alleles, with increase in host age. With regard to host sex, male children had a slightly higher mean number of private alleles than the female children, but the difference was not statistically significant (r: = 0.000, d.f. = 1, P = 1.000). With respect to parasite infection intensities, the mean number of private alleles increased with increase in infection intensities, but again, these increases were not statistically significant (X2 = 0.0000, d.f. = 2, P = 1.000). When the effect of PZQ treatment was examined, an increased MNA were observed in S. mansoni reinfections, 6-12 months after PZQ treatment, but within a 24-hr period, PZQ treatment slightly decreased MNA present in an individual child. Passing the parasite through the hosts, reduced overall MNA and mean number of private alleles observed in the adult parasite population, compared with those in the miracidial populations, suggesting a reduction in the genetic diversity of the adult parasite population with passaging. These results suggest that the S. mansoni population in Mwea is genetically diverse, with a low level of genetic structuring. Age, gender, infection intensity had an effect on private alleles but not other population parameters. More genotypes seem to have been acquired in re-infections, but on a short term, PZQ treatment tended to decrease the mean number of private alleles. Private alleles could potentially be useful for monitoring S. mansoni re-infections, or as markers for PZQ resistance.

Examining the relationship between genetic variation at G6PD and severe malaria.

Author: Shah, S S

Awarding University: University of Oxford, England

Level : DPhil

Year: 2011

Holding Libraries: ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; ; Institute of Commonwealth Studies Library ;

Subject Terms: Malaria/Genetics/Biochemistry/ ;

Abstract:

Glucose-6-phosphate dehydrogenase (G6PD) deficiency, a common heritable trait whose prevalence mirrors geographic patterns of historic malaria endemicity, is thought to confer a selective advantage owing to partial protection conferred against malaria. This thesis examines genetic diversity at the G6PD locus, and the considers how such variation impacts both immediate molecular phenotypes and multifactorial clinical phenotypes. First, Chapter 3 presents a survey of variation at G6PD in several malaria-endemic areas, while Chapter 4 describes a novel technique for polymorphism discovery using pooled massively parallel sequencing. Next, in Chapters 5 and 6, I evaluate the link between genetic variation at the locus and G6PD enzyme activity, identifying major and minor determinants of G6PD deficiency state in an association study conducted in Kenya, and demonstrating a new technique for assaying G6PD deficiency at the level of an individual erythrocyte in a pilot project in Mali. Finally, Chapter 7 addresses the malaria protection hypothesis directly by conducting a fine-mapping case-control association study of severe malaria in the Gambia, where I found that G6PD deficiency alleles exhibited differential direction of association with respect to two important clinical syndromes trending towards risk conferred to severe malarial anemia, and protection with respect to cerebral malaria. Overall, these findings suggest that future clinical association studies should consider heterogeneity at the genetic level, as well as the level of molecular and clinical phenotypes in order to achieve a better mechanistic understanding of the relationship between G6PD deficiency and severe malaria.

Identification and tissue localization of olfactory proteins in the antenna and head of Glossina species

Author: Nyanjom, Steven Reuben Ger

Awarding University: Jomo Kenyatta University of Agriculture and Technology, Kenya

Level : PhD

Year: 2011

Holding Libraries: Jomo Kenyatta University of Agriculture and Technology Library ;

Subject Terms: Glossina pallidipes ; Tsetse flies ; Glossina tachinoides ; Glossina pallidipes gambiense ; Drosophila melanogaster ; Anopheles gambiae ; Aedes aegypti ; Culex quinquefasciatus ; Genetics ;

Abstract:

Tsetse flies use olfaction in search for food, mates and larviposition sites. Olfactory proteins [(odorant binding proteins .(OBPs), pheromone binding proteins (PBPs), chemosensory protein (CSPs), odorant degrading enzymes (ODEs) and odorant receptors (Ors)], located within the antennae, play key role in this process. In this work, presence of olfactory proteins was investigated by constructing and sequencing eDNA libraries from Glossina pallidipes Austen, antennae; Glossina palpalis gambiensis Vanderplank, head and Glossina tachinoides Westwood, head. The Expressed sequence tags (ESTs) were clustered using cDNA Annotationr? Software (CAS) and annotated by blast searches. ESTs were generated from the antennal (1127) and head (906 for G. p. gambiensis and 830 for G. tachinoides) libraries, composed of 296 clusters (18 contigs and 278 singletons for G. pallidipes antennae), 305 clusters (36 contigs and 269 singletons for G. p. gambiensis head) and 232 clusters (54 contigs and 178 singletons for G. tachinoides). The analyses implicated ten (l0) sequences in olfaction (2 OBPs from G. pallidipes, 5 OBPs from G. p. gambiensis, 2 OBPs and 1 CSP from G. tachinoides). Clustal alignment revealed a diverse multigene family while phylogenetic analysis supports the existence of different olfactory protein subfamilies. To identify Dipteran orthologs, the three Glossina ESTs (G. pallidipes antennae, G. p. gambiensis head and G. tachinoides head) were clustered using StackP ACK, then compared to G. morsitans morsitans, Drosophila melanogaster, Anopheles gambiae, Aedes aegypti and Culex quinquefasciatus proteomes. A total of 663 clusters for G. pallidipes (45 contigs and 618 singletons), 930 clusters for G. p. gambiensis (43 contigs and 387 singletons) and 444 clusters for G. tachinoides (40 contigs and 404 singletons) were generated. Nine putative aBPs (G. pallidipes: 2, G. p, gambiensis: 5, G. tachinoides: 2) and one putative CSP (G. tachinoides) were identified by BLAST search against the dipteran protein databases. Multiple sequence alignments revealed a diverse aBP gene family and a conserved CSP. Phylogenetic analysis revealed a closely related multigene family that could have evolved separately along different evolutionary time. Reverse Transcription Polymerase Chain Reaction (RT-PCR) screening of male and female G. pallidipes tissues (antennae, head, thorax and abdomen) for presence of aBPs and CSPs homologs identified in G. pallidipes, G. tachinoides and G. p. gambiensis, and similar ones (putative aBPs from G. m. morsitans), revealed 7 none sex specific (tissue dependent), and 2 sex (male) specific G. m morsitans aBP homologues, one specific to the thorax tissue (Gmm_cnI4014) and the other to both thorax and abdomen tissues (Gmm_GLAAS20TVB). Two putative aBPs identified in G. palidipes and G. p. gambiensis (Gpacontig266 and Gphcontigl84) were localised to the antennae tissue. Alignment of the sequenced amplicons revealed a diverse OBP and conserved CSP gene families. These results indicates that olfactory process in tsetse is a complex and interactive process involving established olfactory and non olfactory tissues, suggesting that tissues, other than antennae should also be targeted/considered in development of novel odor based technologies for tsetse control. shade and larviposit (Rogers and Robinson, 2006). The distribution patterns of fusca group takes three main types. G. longipennis is confined to dry parts of South East Sudan, southern boarder of Ethiopia and north-eastern parts of Somalia, Kenya and Tanzania. G. brevipalpis is widely scattered throughout eastern parts of Africa, from Ethiopia and Somalia in the north, to Mozambique and South Africa in the south. The remainder of the fusca group are limited to thickly forested areas of Africa with G. tabaniformis, G. nashi and G.? haningtoni being restricted to rain forests; G. fusca, G. medicorum, G. fuscipleuris and G. schwetzi being forest edge species while G. medicorum

Phenotypic and genetic characterization of Vibrio cholerae 01 strains isolated from various regions of Kenya between 2007 and 2010

Author: Njeru, Mercy Njura

Awarding University: Jomo Kenyatta University of Agriculture and Technology, Kenya

Level : MSc

Year: 2011

Holding Libraries: Jomo Kenyatta University of Agriculture and Technology Library ;

Subject Terms: Cholera ; Vibrio cholerae ; Genetics ; Deoxyribonucleic acid--DNA ; Epidemiology ;

Abstract:

Cholera remains an important public health concern in developing countries that lack basic infrastructure especially access to safe drinking water and proper sanitation. In this laboratory based study, 76 Vibrio eholerae 01 isolates that represents 2007-2010 cholera epidemics in Kenya were phenotypically and genetically characterized. Fifty-six Vibrio eholerae 01 isolates were used to represent 2009 cholera outbreaks, whereas 20 Vibrio eholerae 01 isolates represented outbreaks from 2007, 2008 and 2010 in Kenya. The inclusion of Vibrio eholerae 01 strains from 2007, 2008 and 2010 was to allow for comparison with those from 2009. These were characterized by serotyping, biotyping, antimicrobial susceptibility testing, toxin and pathogenicity gene detection, ribotyping and Pulse Field Gel Electrophoresis (pFGE). V. eholerae 01 Inaba was the dominant serotype (88.2%) whereas Ogawa was isolated in 11.8%. All the strains were resistant to polymyxin B. The multiplex and simplex PCR assay showed that all the V. eholerae isolates were positive for etxA, tepA (EI Tor) and rtxC genes, showing Kenyan isolates belonged to biotype EI Tor. The MAMA-PCR assays for the differentiation of cholera toxin B subunit of classical and EI Tor biotypes of V. cholerae 01 indicated that the 76 strains were V. eholerae 01 biotype EI Tor variant that harbors the classical etxB gene. Based on CLSI guidelines of defining susceptibility, 1000.10 of the strains were susceptible to tetracycline, doxycycline, ofloxacin, azithromycin, norfloxacin and ceftriaxone. Susceptibility to ciprofloxacin was 97.7%. Intermediate susceptibility to ciprofloxacin, nalidixic acid, ampicillin and chloramphenicol was observed in 2.3%, 59.1%, 11.4%, and 93.2% of the strains respectively. The strains were 100% resistant to furazolidone, streptomycin and sulfamethoxazole-trimethoprim, while 88.6% were resistant to nalidixic acid, 40.9'10 resistant to ampicillin and 6.8% resistant chloramphenicol. The V. cholerae 01 Isolates from the different regions of Kenya had genetically similar pattern on ribotyping of Bgi I digested chromosomal DNA and Pulse Field Gel Electrophoresis (pFGE) patterns of Not 1- digested chromosomal DNA. All the tested strains belonged to ribotype RIll. Tetracycline and doxycycline currently used in treatment of cholera cases In Kenya were found to be effective and thus should continue being used. A coordinated multidisciplinary approach and working with communities to encourage behavioral change to diminish the risks of infection may be the most efficient way to prevent and contain cholera outbreaks, as increase in number of cholera cases reported in Kenya in 2009 may not have been as a result of change in the V.cholerae 01 strains in circulation. Further sequence based typing of V. cholera 01 strains may be useful in confirming epidemiological linkage of the cholera outbreaks in Kenya.

Evaluation of HIV-1 Evolution and its role developement of Antiretroviral drug resistence in Nairobi Cohort

Author: Nyamache, Anthony Kebira

Awarding University: Jomo Kenyatta University of Agriculture and Technology, Kenya

Level : PhD

Year: 2011

Holding Libraries: Jomo Kenyatta University of Agriculture and Technology Library ;

Subject Terms: Antiretrovirals ; HIV (infection) ; AIDS (disease) ; Nairobi, Kenya ; Genetics ;

Abstract:

The treatment of HIV -1 infection with antiretroviral drugs has greatly improved the survival of those who are infected. However, HIV -1 diversity and drug resistance are major challenges in patient management, disease control and surveillance especially in resourcepoor countries. In this study, 188 blood samples were collected from Nairobi cohort and peripheral mononuclear cells (PBMCs) separated. Total proviral DNA was used in nested polymerase chain reaction to amplify 450bp HIV env C2V3, 288bp Integrase and env gp41 regions and directly sequenced. Generated sequences were aligned and phylogenetically analysed using known reference subtypes sequences and drug resistance mutations and substitutions determined. Phylogenetic analysis based on env C2V3 region revealed Al (59.6%), C (18.1%), D (10.6%), B (2.1%), G (2.1%), CRF02_AG (3.2%) and the rest of 6.9% were CRFs. In HIV- 1 co-receptor switch showed R5 tropism (69.6%) while X4 (30.4%). In addition, 2.4% T97 A that is associated with reduced susceptibility to Raltegravir and 26.2% had secondary mutations associated with resistance to intergrase inhibitors. In fusion inhibitors, the following mutations were detected; A316TII323V (2.6%) combination, A316T (63%), I323V (1.1%) for Maraviroc, (10%) K305R, (3.2%) G321E, (35.1%) R315Q, (4.5%) K305RfR315Q, (62.8%) T320R for Vicriviroc and (1.6%) A316T+ K305R+ R315Q, (12.7%) A316T+R315Q, (3.2%) R3I5Q+A316T+I323V, (0.5%) R315+A316T+G321E for Maraviroc and Vicriviroc combinations. In addition, 4.2% intermediate resistance associated to Enfuvirtide was detected. The point mutations at; N42S was detected in 16.7% of all the samples, while N42D was detected in 4.2%, S 138L IT 3.1 %, L44M 2.1 % and 1 % each for in the following mutations; N43I and L45V drug resistance mutations. In evolutionary rate; 12.5% had dNdS ration 1>, 88.5% dNdS ration < 1 and in those with dNdS ration =11. The results indicate that HIV -1 subtypes in Nairobi cohort like the rest of the country, is predominated by mv -1 subtype AI, though there could be possibity of an increase proportion of mv -1 subtype C prevalences. Existence of diverse HIV -1 recombinants indicated viral mixing among the population, possibly as a result of dual infections. Evolutionary rate of the virus showed natural selection with high proportions of R5 strains suggestive of feasibility of use of maraviroc (CCR5 antagonists) in Kenya. However, multiple drug resistance mutations observed in the intergrase inhibitors and CCR5 antagonists prior to their introduction. There is a need for constant monitoring of HIV-l genetic diversity and drug resistance. Drug resistance seen to common drugs was minimal (2 %), indicating that we need to continue to prescribe the currently used drugs. In addition, the new classes of entry, fusion and integrase inhibitors are feasible as firstline and thirdline drugs respectively. In addition, it was realised that it is not necessary to carry out resistance testing at baseline unless there is strong evidence of virological failure.

Association between interferon-gamma promoter -183 (G/T), -1616 (A/G) and intronic +2200 (A/G) variants and malaria disease outcomes in children exposed to Plasmodium falciparum

Author: Okoth, Evans Ouma Raballah

Awarding University: Jomo Kenyatta University of Agriculture and Technology, Kenya

Level : MSc

Year: 2011

Holding Libraries: Jomo Kenyatta University of Agriculture and Technology Library ;

Subject Terms: Plasmodium falciparum ; Siaya District Hospital, Kenya ; Malaria ; Babies ; Children and youth ; Anemia ; Genetics ;

Abstract:

Infection with Plasmodium falciparum, the most common human plasmodial parasite causes significant global morbidity and mortality. Interferon-gamma (IFN-y) is a typeI cytokine with T helper (Th) I-biased immune responses for intracellular pathogen control. Elevated IFN-y levels contribute to protective immunity against clinical and cerebral malaria. High IFN-y responses are associated with reduced asexual parasite multiplication rates fOllowing a primary infection with P. falciparum. Previous studies have associated IFN-y single nucleotide polymorphisms (SNPs) with susceptibility to asthma, tuberculosis and functional changes in circulating IFN-y levels. However, the role of IFN-y polymorphisms in conditioning malarial disease outcomes in children residing in P. falciparum holoendemic transmission areas remains unexplored. The current study, using a hospital-based cross-sectional design, investigated the functional associations between IFN-y genotypic and haplotypic promoter [-183G/T and -1616A1G] and intronic [+2200AlG] variation in conditioning malarial disease outcomes [(severe malarial anaemia (SMA; Haemoglobin (Hb)<6.0 gldL). malarial anaemia (MA; Hb<8.0 gldL and high-density parasitaemia (HDP; ~lO,OOO parasites/JlL)] in infants and young children (age 3-36 months, n=574) enrolled at Siaya District Hospital, western Kenya. Blood samples (3mL) for malaria diagnosis and complete haematological measurements were obtained upon consent from parent/guardian. In addition, plasma was separated and frozen at -80?C until determination of circulating IFN-y levels. DNA was extracted from blood spotted on filter paper using Chelex method. Genotyping of the IFN-y SNPs was performed using a Taqman' 5' allelic discrimination Assay-By-Design method according to manufacturer's instructions and by PCR followed by restriction fragment length polymorphism (RFLP), IFN-y levels were measured as part of human 25-cytokine plex assay. Multivariate logistic regression models controlling for the confounders revealed that GT heterozygosity at the IFN-y -183Gff variants showed a trend of reduced risk of developing SMA (OR, 0.603; 95% CI, 0.286-1.271; P=0.183) relative to the wild type (GG homozygotes). Additionally, the IFN-y -1616A1G variants demonstrated that individuals with GG had a two-fold increased risk of developing MA (OR, 2.306; 95% CI, 1.141-4.660; P=0.020) relative to the homozygous A (wild type). The IFN-y +2200AIG variants were not associated with SMA, though heterozygous individuals (AG) demonstrated a tendency towards reduced risk of developing MA (OR, 0.597; 95% CI, 0.331-1.076; P=0.086). In addition, the GAA .haplotype tended to reduce risk of developing MA (OR, 0.635; 95% CI, 0.391-1.030; P=0.066), while the GAG haplotype tended towards increased risk of developing MA (OR, 1.794; 95% CI, 0.957-3.364; P=0.068). However, carriage of these genotypes and haplotypes did not influence the circulating levels of IFN-y. The results demonstrate that genetic variation in the three IFN-y genotypes investigated in this study may be important in regulating MA outcomes in paediatric P. falciparum infection. These findings provide an enhanced understanding of how variation in IFN-y conditions malaria pathogenesis and the important role that IFN-y plays in children with falciparum malaria. From these results it is recommended that future work should focus on comprehensive longitudinal studies to delineate the role ofIFN-y variants in modulating malarial outcomes in this population.