High prevalence and extended deletions in Plasmodium falciparum hrp2/3 genomic loci in Ethiopia.

Deletions in Plasmodium falciparum histidine rich protein 2(pfhrp2) gene threaten the usefulness of the most widely used HRP2-based malaria rapid diagnostic tests (mRDTs) that cross react with its structural homologue, PfHRP3. Parasites with deleted pfhrp2/3 genes remain undetected and untreated due to 'false-negative' RDT results. As Ethiopia recently launched malaria elimination by 2030 in certain selected areas, the availability of RDTs and the scale of their use have rapidly increased in recent years. Thus, it is important to explore the presence and prevalence of deletion in the target genes, pfhrp2 and pfhrp3. From a total of 189 febrile patients visited Adama Malaria Diagnostic centre, sixty-four microscopically-and polymerase chain reaction (PCR)-confirmed P. falciparum clinical isolates were used to determine the frequency of pfhrp2/3 gene deletions. Established PCR assays were applied to DNA extracted from blood spotted onto filter papers to amplify across pfhrp2/3 exons and flanking regions. However, analysis of deletions in pfhrp2, pfhrp3 and flanking genomic regions was successful for 50 of the samples. The pfhrp2 gene deletion was fixed in the population with all 50(100%) isolates presenting a deletion variant. This deletion extended downstream towards the Pf3D7 0831900 (MAL7PI.230) gene in 11/50 (22%) cases. In contrast, only 2/50 (4%) of samples had deletions for the Pf3D7 0831700 (MALPI.228) gene, upstream of pfhrp2. Similarly, the pfhrp3 gene was deleted in all isolates (100%), while 40% of the isolates had an extension of the deletion to the downstream flanking region that codes for Pf3D7 13272400 (MAL13PI.485).The pfhrp3 deletion also extended upstream to Pf3D7 081372100 (MAL13PI.475) region in 49/50 (95%) of the isolates, exhibiting complete absence of the locus. Although all samples showed deletions of pfhrp2 exon regions, amplification of an intron region was successful in five cases. Two different repeat motifs in the intron regions were observed in the samples tested. Pfhrp2/3 gene deletions are fixed in Ethiopia and this will likely reduce the effectiveness of PfHRP2-based mRDTs. It will be important to determine the sensitivity PfHRP 2/3-based RDTs in these populations and conduct a countrywide survey to determine the extent of these deletions and its effect on routine RDT-based malaria diagnosis

Genome-wide association studies of severe P. falciparum malaria susceptibility: progress, pitfalls and prospects

Abstract Background P. falciparum malaria has been recognized as one of the prominent evolutionary selective forces of human genome that led to the emergence of multiple host protective alleles. A comprehensive understanding of the genetic bases of severe malaria susceptibility and resistance can potentially pave ways to the development of new therapeutics and vaccines. Genome-wide association studies (GWASs) have recently been implemented in malaria endemic areas and identified a number of novel association genetic variants. However, there are several open questions around heritability, epistatic interactions, genetic correlations and associated molecular pathways among others. Here, we assess the progress and pitfalls of severe malaria susceptibility GWASs and discuss the biology of the novel variants. Results We obtained all severe malaria susceptibility GWASs published thus far and accessed GWAS dataset of Gambian populations from European Phenome Genome Archive (EGA) through the MalariaGen consortium standard data access protocols. We noticed that, while some of the well-known variants including HbS and ABO blood group were replicated across endemic populations, only few novel variants were convincingly identified and their biological functions remain to be understood. We estimated SNP-heritability of severe malaria at 20.1% in Gambian populations and showed how advanced statistical genetic analytic methods can potentially be implemented in malaria susceptibility studies to provide useful functional insights. Conclusions The ultimate goal of malaria susceptibility study is to discover a novel causal biological pathway that provide protections against severe malaria; a fundamental step towards translational medicine such as development of vaccine and new therapeutics. Beyond singe locus analysis, the future direction of malaria susceptibility requires a paradigm shift from single -omics to multi-stage and multi-dimensional integrative functional studies that combines multiple data types from the human host, the parasite, the mosquitoes and the environment. The current biotechnological and statistical advances may eventually lead to the feasibility of systems biology studies and revolutionize malaria research

Sequence analysis of Plasmodium vivax Duffy binding proteins reveals the presence of unique haplotypes and diversifying selection in Ethiopian isolates.

BACKGROUND: Red blood cell invasion by the Plasmodium vivax merozoite requires interaction between the Duffy antigen receptor for chemokines (DARC) and the P. vivax Duffy-binding protein II (PvDBPII). Given that the disruption of this interaction prevents P. vivax blood-stage infection, a PvDBP-based vaccine development has been well recognized. However, the polymorphic nature of PvDBPII prevents a strain transcending immune response and complicates attempts to design a vaccine. METHODS: Twenty-three P. vivax clinical isolates collected from three areas of Ethiopia were sequenced at the pvdbpII locus. A total of 392 global pvdbpII sequences from seven P. vivax endemic countries were also retrieved from the NCBI archive for comparative analysis of genetic diversity, departure from neutrality, linkage disequilibrium, genetic differentiation, PvDBP polymorphisms, recombination and population structure of the parasite population. To establish a haplotype relationship a network was constructed using the median joining algorithm. RESULTS: A total of 110 variable sites were found, of which 44 were parsimony informative. For Ethiopian isolates there were 12 variable sites of which 10 were parsimony informative. These parsimony informative variants resulted in 10 nonsynonymous mutations. The overall haplotype diversity for global isolates was 0.9596; however, the haplotype diversity was 0.874 for Ethiopia. Fst values for genetic revealed Ethiopian isolates were closest to Indian isolates as well as to Sri Lankan and Sudanese isolates but further away from Mexican, Papua New Guinean and South Korean isolates. There was a total of 136 haplotypes from the 415 global isolates included for this study. Haplotype prevalence ranged from 36.76% to 0.7%, from this 74.2% were represented by single parasite isolates. None of the Ethiopian isolates grouped with the Sal I reference haplotype. From the total observed nonsynonymous mutations 13 mapped to experimentally verified epitope sequences. Including 10 non-synonymous mutations from Ethiopia. However, all the polymorphic regions in Ethiopian isolates were located away from DARC, responsible for junction formation. CONCLUSION: The results of this study are concurrent with the multivalent vaccine approach to design an effective treatment. However, the presence of novel haplotypes in Ethiopian isolates that were not shared by other global sequences warrant further investigation

The biology of unconventional invasion of Duffy-negative reticulocytes by Plasmodium vivax and its implication in malaria epidemiology and public health.

Plasmodium vivax has been largely neglected over the past century, despite a widespread recognition of its burden across region where it is endemic. The parasite invades reticulocytes, employing the interaction between Plasmodium vivax Duffy binding protein (PvDBP) and human Duffy antigen receptor for chemokines (DARC). However, P. vivax has now been observed in Duffy-negative individuals, presenting a potentially serious public health problem as the majority of African populations are Duffy-negative. Invasion of Duffy-negative reticulocytes is suggested to be through duplication of the PvDBP and a novel protein encoded by P. vivax erythrocyte binding protein (EBP) genes. The emergence and spread of specific P. vivax strains with ability to invade Duffy-negative reticulocytes has, therefore, drawn substantial attention and further complicated the epidemiology and public health implication of vivax malaria. Given the right environment and vectorial capacity for transmission coupled with the parasite's ability to invade Duffy-negative individuals, P. vivax could increase its epidemiological significance in Africa. In this review, authors present accruing knowledge on the paradigm shift in P. vivax invasion of Duffy-negative reticulocytes against the established mechanism of invading only Duffy-positive individuals and offer a perspective on the epidemiological diagnostic and public health implication in Africa

Zingiber Officinale Roscoe and Echinops Kebericho Mesfin Showed Antiplasmodial Activities against Plasmodium Berghei in a Dosedependent Manner in Ethiopia

BACKGROUND: The emergence and spread of Plasmodium falciparum resistance to antimalarial drugs necessitated the search for new drugs from natural products. Zingiber officinal Roscoe and Echinops Kebericho Mesfin are traditional herbal medicines widely used for the treatment of malaria in Ethiopia. The aim of the study was to assess the toxicity profile and in vivo antiplasmodial activities of 70% methanol crude extracts of both plant materials against Plasmodium berghei.METHODS: Healthy male Swiss Albino mice of age 4-5 weeks and weight 25-36 g were infected by P. berghei. The extracts were administered orally at doses 5000, 2500 and 1250 mg/kg for acute toxicity of E. kebericho Mesfin. Graded doses at 1000, 500 and 250 mg/kg used for four days suppressive studies. Parasitemia, body weight, packed cell volume (PCV) and survival time were determined. SPSS Version 20 was used for the analysis of data of parasitemia, body weight, PCV, and survival times. Statistical significance was determined by one-way ANOVA. Independent ttest was used to compare results. Results were presented as a mean ± standard error of the mean (M ± SEM). All data were analyzed at a 95% confidence interval (α= 0.05).RESULTS: At the dose of 5000 mg/kg, E. kebericho Mesfin showed no toxic effects. The LD50 of extract could go beyond the dose used. In vivo antiplasmodial activity of extracts showed excellent chemo suppression at 500 and 1000 mg/kg in a dose dependent manner compared with the negative control. The chemo suppressions of the 1000 mg/kg of both plant extracts were 49.53 ± 1.90% and 32.83 ± 1.03%, respectively. The survival times of P. berghei infected mice were also a dose dependent manner while failed to prevent weight loss.CONCLUSION: The extracts of both medicinal plants showed antiplasmodial activities against P. berghei. It confirmed the literature findings and their traditional uses.

Insilico Functional Analysis of Genome-Wide Dataset From 17,000 Individuals Identifies Candidate Malaria Resistance Genes Enriched in Malaria Pathogenic Pathways

Recent genome-wide association studies (GWASs) of severe malaria have identified several association variants. However, much about the underlying biological functions are yet to be discovered. Here, we systematically predicted plausible candidate genes and pathways from functional analysis of severe malaria resistance GWAS summary statistics (N = 17,000) meta-analysed across 11 populations in malaria endemic regions. We applied positional mapping, expression quantitative trait locus (eQTL), chromatin interaction mapping, and gene-based association analyses to identify candidate severe malaria resistance genes. We further applied rare variant analysis to raw GWAS datasets (N = 11,000) of three malaria endemic populations including Kenya, Malawi, and Gambia and performed various population genetic structures of the identified genes in the three populations and global populations. We performed network and pathway analyses to investigate their shared biological functions. Our functional mapping analysis identified 57 genes located in the known malaria genomic loci, while our gene-based GWAS analysis identified additional 125 genes across the genome. The identified genes were significantly enriched in malaria pathogenic pathways including multiple overlapping pathways in erythrocyte-related functions, blood coagulations, ion channels, adhesion molecules, membrane signalling elements, and neuronal systems. Our population genetic analysis revealed that the minor allele frequencies (MAF) of the single nucleotide polymorphisms (SNPs) residing in the identified genes are generally higher in the three malaria endemic populations compared to global populations. Overall, our results suggest that severe malaria resistance trait is attributed to multiple genes, highlighting the possibility of harnessing new malaria therapeutics that can simultaneously target multiple malaria protective host molecular pathways

Major subpopulations of Plasmodium falciparum in sub-Saharan Africa.

Understanding genomic variation and population structure of Plasmodium falciparum across Africa is necessary to sustain progress toward malaria elimination. Genome clustering of 2263 P. falciparum isolates from 24 malaria-endemic settings in 15 African countries identified major western, central, and eastern ancestries, plus a highly divergent Ethiopian population. Ancestry aligned to these regional blocs, overlapping with both the parasite's origin and with historical human migration. The parasite populations are interbred and shared genomic haplotypes, especially across drug resistance loci, which showed the strongest recent identity-by-descent between populations. A recent signature of selection on chromosome 12 with candidate resistance loci against artemisinin derivatives was evident in Ghana and Malawi. Such selection and the emerging substructure may affect treatment-based intervention strategies against P. falciparum malaria

Climatic variables and malaria transmission dynamics in Jimma town, South West Ethiopia

<p>Abstract</p> <p>Background:-</p> <p>In Ethiopia, malaria is seasonal and unstable, causing frequent epidemics. It usually occurs at altitudes < 2,000 m above sea level. Occasionally, transmission of malaria occurs in areas previously free of malaria, including areas > 2,000 m above sea level. For transmission of malaria parasite, climatic factors are important determinants as well as non-climatic factors that can negate climatic influences. Indeed, there is a scarcity of information on the correlation between climatic variability and malaria transmission risk in Ethiopia in general and in the study area in particular. Therefore, the aim of this study was to determine the level of correlation between meteorological variables and malaria cases.</p> <p>Methods: -</p> <p>Time-series analysis was conducted using data on monthly meteorological variables and monthly total malaria in Jimma town, south west Ethiopia, for the period 2000-2009. All the data were entered and analyzed using SPSS-15 database program. Spearman correlation and linear regression analysis were used to asses association between the variables.</p> <p>Results: -</p> <p>During last ten years (2000-2009), a fluctuating trend of malaria transmission was observed with <it>P.vivax </it>becoming predominant species. Spearman correlation analysis showed that monthly minimum temperature, total rainfall and two measures of relative humidity were positively related with malaria but monthly maximum temperature negatively related. Also regression analysis suggested that monthly minimum (p = 0.008), monthly maximum temperature (p = 0.013) and monthly total rainfall (p = 0.040), at one month lagged effect, were significant meteorological factors for transmission of malaria in the study area.</p> <p>Conclusion: -</p> <p>Malaria incidences in the last decade seem to have a significant association with meteorological variables. In future, prospective and multidisciplinary cooperative research involving researchers from the fields of parasitology, epidemiology, botany, agriculture and climatology is necessary to identify the real effect of meteorological factors on vector- borne diseases like malaria.</p

Urban malaria and associated risk factors in Jimma town, south-west Ethiopia

<p>Abstract</p> <p>Background</p> <p>Malaria kills millions around the world. Until recently it was believed to be a disease of rural areas, since the <it>Anopheles </it>mosquito, which transmits <it>Plasmodium </it>species breeds in rural areas. Urban malaria is emerging as a potential, but "avertable" crisis, in Africa. In view of the rapidly growing number of small and medium-sized towns in Ethiopia there is a pressing need to improve the understanding of the epidemiology of malaria. Therefore, the aim of this study was to determine malaria prevalence and associated risk factors in Jimma town.</p> <p>Methods</p> <p>A cross-sectional study was carried out in Jimma town from April 1 to May 28, 2010. 804 study participants were included from 291 households for microscopic examination of malaria parasites. Socio-demography data and risk factors were collected using structured questionnaires. Logistic regression analysis was done using SPSS 15.0 statistical software.</p> <p>Results</p> <p>From a total of 804 study participants in current survey only 42 (5.2%) were positive for malaria parasites. <it>Plasmodium vivax, Plasmodium falciparum </it>and mixed infection accounted 71.4%, 26.2% and 2.4%, respectively. Higher malaria prevalence rate was observed among under-five children (11%). Those who do not use insecticide-treated bed nets (ITN) were more likely to be infected with malaria (OR = 13.6; 95% CI 4.9-37.2, p < 0.001) compared with those who use the ITN. Living in areas where stagnant water existed (OR = 2.1; 95% CI 1.00-4.2, p = 0.047) and its distance of existence <1 km from the house(OR = 2.1; 95% CI 2.0-15.8, p = 0.001) were more likely to be infected with malaria parasite compared with those who live away from stagnant at a distance greater than 1 km.</p> <p>Conclusion</p> <p>Malaria is a major health problem with <it>P. vivax </it>becoming a predominant species in the town. The prevalence was strongly associated with proximity of residence to potential mosquito breeding sites. Malaria is affecting significant proportions of the urban settlers and human activities nevertheless play an important role in bringing the mosquito breeding sites closer to residences.</p