Browsing by Author "Malisa, Allen L"

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Drug resistance to sulphadoxine-pyrimethamine in Plasmodium falciparum malaria in Mlimba, Tanzania
(Malaria Journal, 2006-10) Mbugi, Erasto V; Mutayoba, Benezeth M; Malisa, Allen L; Balthazary, Sakurani T; Nyambo, Thomas B; Mshinda, Hassan
Background: Sulphadoxine-pyrimethamine (SP) has been and is currently used for treatment of uncomplicated Plasmodium falciparum malaria in many African countries. Nevertheless, the response of parasites to SP treatment has shown significant variation between individuals. Methods: The genes for dihydrofolate reductase (dhfr) and dihydropteroate synthase (dhps) were used as markers, to investigate parasite resistance to SP in 141 children aged less than 5 years. Parasite DNA was extracted by Chelex method from blood samples collected and preserved on filter papers. Subsequently, polymerase chain reaction (PCR) and restriction fragment length polymorphism (PCR-RFLP) were applied to detect the SP resistance-associated point mutations on dhfr and dhps. Commonly reported point mutations at codons 51, 59, 108 and 164 in the dhfr and codons 437, 540 and 581 in the dhps domains were examined. Results: Children infected with parasites harbouring a range of single to quintuple dhfr/dhps mutations were erratically cured with SP. However, the quintuple dhfr/dhps mutant genotypes were mostly associated with treatment failures. High proportion of SP resistance-associated point mutations was detected in this study but the adequate clinical response (89.4%) observed clinically at day 14 of follow up reflects the role of semi-immunity protection and parasite clearance in the population. Conclusion: In monitoring drug resistance to SP, concurrent studies on possible confounding factors pertaining to development of resistance in falciparum malaria should be considered. The SP resistance potential detected in this study, cautions on its useful therapeutic life as an interim firstline drug against malaria in Tanzania and other malaria-endemic countries.
The evolution of pyrimethamine resistant dhfr in Plasmodium falciparum of south-eastern Tanzania: comparing selection under SP alone vs SP +artesunate combination
(Malaria Journal, 2011) Malisa, Allen L; Pearce, Richard J; Mutayoba, Ben M; Abdullah, Salim; Mshinda, Hassan; Kachur, Patrick S; Bloland, Peter; Roper, Cally
Background: Sulphadoxine-pyrimethamine (SP) resistance is now widespread throughout east and southern Africa and artemisinin compounds in combination with synthetic drugs (ACT) are recommended as replacement treatments by the World Health Organization (WHO). As well as high cure rates, ACT has been shown to slow the development of resistance to the partner drug in areas of low to moderate transmission. This study looked for evidence of protection of the partner drug in a high transmission African context. The evaluation was part of large combination therapy pilot implementation programme in Tanzania, the Interdisciplinary Monitoring Programme for Antimalarial Combination Therapy (IMPACT-TZ) Methods: The growth of resistant dhfr in a parasite population where SP Monotherapy was the first-line treatment was measured for four years (2002-2006), and compared with the development of resistant dhfr in a neighbouring population where SP + artesunate (SP+AS) was used as the first-line treatment during the same interval. The effect of the differing treatment regimes on the emergence of resistance was addressed in three ways. First, by looking at the rate of increase in frequency of pre-existing mutant dhfr alleles under monotherapy and combination therapy. Second, by examining whether de-novo mutant alleles emerged under either treatment. Finally, by measuring diversity at three dhfr flanking microsatellite loci upstream of the dhfr gene Results: The reduction in SP selection pressure resulting from the adoption of ACT slowed the rate of increase in the frequency of the triple mutant resistant dhfr allele. Comparing between the two populations, the higher levels of genetic diversity in sequence flanking the dhfr triple mutant allele in the population where the ACT regimen had been used indicates the reduction in SP selection pressure arising from combination therapy. Conclusion: The study demonstrated that, alleles containing two mutations at the dhfr have arisen at least four times independently while those containing triple mutant dhfr arose only once, and were found carrying a single unique Asian-type flanking sequence, which apparently drives the spread of pyrimethamine resistance associated dhfr alleles in east Africa. SP+AS is not recommended for use in areas where SP cure rates are less than 80% but this study reports an observed principle of combination protection from an area where pyrimethamine resistance was already high.
The genetic change in P. falciparum populations of rural Tanzania resulting from national policy on firstline malaria treatment and pilot Sulfadoxine/pyrimethamine and Artesunate combination
(Malaria Journal, 2010-10) Malisa, Allen L; Pearce, Richard J; Abdullah, Salim; Mshinda, Hassan; Kachur, Patrick; Bloland, Peter; Roper, Cally
Theory predicts that we can protect the efficacy of future antimalarials by changing treatment practice or drug formulation, but the potential success of such interventions rests upon their impact on drug pressure in the field. So far, gathering field data on the relationship between policy, drug pressure, recombination and the evolution of resistance has been entirely challenging. To test these predictions, dhfr and dhps frequency changes were measured in two rural districts of Rufiji and Kilombero/Ulanga during 2000-2006, and the frequencies of the two genes compared prior, during and after antimalarial policy change from first line CQ to first line SP in 2001. Furthermore, while SP first line was maintained in Kilombero/Ulanga, pilot combination therapy of SP+Artesunate (ART) was introduced in Rufiji in 2002 to replace SP and dhfr and dhps frequency changes compared between the two districts. Size polymorphisms at three sets of microsatellite loci linked to dhfr and three other sets of unlinked microsatellite loci were analysed. Genetic analysis of SP resistance genes was carried out on 9,662 Plasmodium falciparum infections identified in a series of annual cross sectional surveys conducted in the two districts between 2000-2006. The frequency of dhfr and dhps resistance alleles did not change significantly while SP was the recommended second-line treatment, but highly significant changes occurred during the subsequent year after the switch to first line SP. The frequency of the triple mutant dhfr allele increased by 37% -63% and that of double mutant dhps allele increased 200%-300%. A strong association between these unlinked alleles also emerged; confirming that they are co-selected by SP. Distribution of major lineages indicates that there is extensive genetic exchange among the geographic regions. Combination therapy had visible effect on the frequencies of dhfr and dhps resistance alleles. The findings of this study provide insight on the interplay between policy, drug pressure, recombination and the evolution of resistance.
Media, health workers, and policy makers’ relationship and their impact on antimalarial policy adoption: a population genetics perspective
(Sokoine University of Agriculture, Tanzania., 2010-04) Malisa, Allen L; Pearce, Richard J; Mutayoba, Benezeth; Abdullah, Salim; Mshinda, Hassan; Kachur, Patrick; Bloland, Peter; Roper, Cally
Drug resistance negatively impacts malaria treatments, making treatment policy revision unavoidable. So far, studies relating sociopolitical and technical issues on policy change with malaria parasite genetic change are lacking.We have quantified the effect of malaria treatment policy on drug pressure and the influence of the media, policy makers, and health worker relationship on parasite population genetic change in Kilombro/Ulanga district. Cross-sectional surveys of asymptomatic infections conducted before, during and after the switch from chloroquine to sulphadoxine/pyrimethamine were used for genetic analysis of SP resistance genes in 4,513 asymptomatic infections identified, and their frequency change was compared with retrospective study of the documented process of policy change. Highly significant changes of dhfr and dhps resistance alleles occurred within one year of switch to SP first line, followed by a decline of their rate of selection caused by reduction of SP usage, as a result of negative media reports on SP usage and lack of adequate preparations.
Multiplicity of infections and level of recrudescence in Plasmodium falciparum malaria in Mlimba, Tanzania
(Academic Journals, 2006-09) Mbugi, Erasto V.; Mutayoba, Benezeth M; Balthazary, Sakurani T; Malisa, Allen L; Nyambo, Thomas B; Mshinda, Hassan
Polymorphism and antigenic variation are important biological survival strategies of malaria parasites determining the episode, outcome and implications of treatment interventions. In P. falciparum, polymorphic antigens are associated with the asexual blood-stage; merozoite surface protein 2 (MSP2). The MSP2 genes have been invaluable in post-treatment discrimination of parasite resurgence from new infection, especially in high transmission areas. We performed polymerase chain reaction (PCR) on DNA extracted from blood samples of 141 malaria-infected infants, followed by restriction fragment length polymorphism (RFLP) of PCR products. The findings showed multiplicity of infections of single to six infections with an average of 2.58 infections per patient. Single infections of either 3D7 or FC27 allelic families of the MSP2 gene occurred in 51 patients (50.5%) out of all PCR-RFLP successful samples (n = 101). Out of 15 (10.6%) follow up samples with resurgent parasitaemia, 3 (20%) samples had recrudescent infections while 12 (80%) had variable results. Our findings provide an insight on the prevalence of the genetic determinants of suphadoxine-pyrimethamine (SP) resistance in Mlimba during the study period, and in the face of rapidly spreading resistance, calls for the periodic surveillance in order to timely detect early warning signal of the deteriorating SP cure rate.
Prevalence and constraints of typhoid fever and its control in an endemic area of Singida region in Tanzania: Lessons for effective control of the disease
(Journal of Public Health and Epidemiology, 2010-08) Malisa, Allen L; Nyaki, Honest
Typhoid fever (TF) is an important communicable disease that is endemic to Tanzania, quite often complicated with malaria co-infection leading to diagnostic complications and significant mortality. Despite considerable control efforts, the disease continues to persist in Singida urban leading to significant morbidity and mortality. This paper retrospectively examines morbidity and mortality trend of typhoid infection for the past five years in Singida urban. By using questionnaire, Focus Group Discussion (FGD) and direct observational methods, the authors report interview results of 120 respondents from the study community, regarding their awareness of TF and its control methods. Results showed that, the TF prevalence records revealed a fluctuating trend with annual incidence rate of 580 – 1,400/100,000 persons, and an overall increase from 771 – 942 cases/100,000 persons (p _ 0.0001) between 2003 and 2007. While 88% of the respondents were aware of TF disease, 53% were unaware of its control methods. The study also revealed an acute shortage of diagnostic laboratory services which indicated that, 75% of health facilities had no such services. In adequate knowledge about personal hygiene, scarcity or lack of access to safe water, improper drainage systems and problems of unsanitary toilets in Singida urban were some of the obstacles to effective TF control. Effective TF control measures in the study district, as in other areas in the tropics, requires integration of intensive health education as a public health tool, provision and access to safe water supply and adequate strengthening of health systems.
Reduced variatio around drug-resistant dhfr alleles in African plasmodium falciparum
(Oxford University Press, 2005-05-25) Pearce, Richard J; Malisa, Allen L; Kachur, Patrick; Barnes, Karen; Brian, Sharp; Roper, Cally
We have measured microsatellite diversity at 26 markers around the dhfr gene in pyrimethamine-sensitive and -resistant parasites collected in southeast Africa. Through direct comparison with diversity on sensitive chromosomes we have found significant loss of diversity across a region of 70 kb around the most highly resistant allele which is evidence of a selective sweep attributable to selection through widespread use of pyrimethamine (in combination with sulfadoxine) as treatment for malaria. Retrospective analysis through four years of direct and continuous selection from use of sulfadoxinepyrimethamine as first-line malaria treatment on a Plasmodium falciparum population in KwaZulu Natal, South Africa, has revealed how recombination significantly narrowed the margins of the selective sweep over time. A deterministic model incorporating selection coefficients measured during the same interval indicates that the transition was toward a state of recombination-selection equilibrium. We compared loss of diversity around the same resistance allele in two populations at either extreme of the range of entomological inoculation rates (EIRs), namely, under one infective bite per year in Mpumalanga, South Africa, and more than one per day in southern Tanzania. EIRs determine effective recombination rates and are expected to profoundly influence the dimensions of the selective sweep. Surprisingly, the dimensions were broadly consistent across both populations. We conclude that despite different recombination rates and contrasting drug selection histories in neighboring countries, the region-wide movement of resistant parasites has played a key role in the establishment of resistance in these populations and the dimensions of the selective sweep are dominated by the influence of high initial starting frequencies.
Species and gender differentiation between and among domestic and wild animals using mitochondrial and sex-linked DNA markers
(2005-11) Malisa, Allen L; Balthazary, Sakurani; Wasser, Sam; Mutayoba, Benezeth; Gwakisa, Paul
In many African countries accurate and reliable identification of poached wildlife products like carcasses or meat presents a big problem when morphological characters such as skin hair or bones are missing. We describe a molecular based approach that has a potential of serving as a forensic tool in game meat identification in Africa. A mitochondial DNA marker (mt700) and one restriction enzyme, Rsa1 were used in the PCR-RFLP species identification of game meat obtained from two National Parks in Tanzania. Species-specific reference DNA fragment patterns were obtained using fresh meat from ten wildlife and four domesticated species. All species except the zebra, produced unique monomorphic RFLP patterns. Collectively, these patterns demonstrate the potential ability of genetic techniques for discriminating between and among wildlife and domestic species. The reference PCR-RFLP fragments enabled species identification of about 79% of unknown meat samples. In addition, sex was also assigned to all of the samples following successful amplification of gender-specific, SRY and ZFY/X, chromosomal domains. Although the present study has been conducted on a limited range both in numbers and genetic diversity of wildlife species present in Africa, the results demonstrate the potential usefulness of the DNA approach in wildlife forensics in the continent.