publications

2025

1. Vaccine

   

   Unveiling mismatch of RTS S AS01 and R21 Matrix M malaria vaccines haplotype among Ethiopian Plasmodium falciparum clinical isolates

   Aynalem Mandefro, Alebachew Messele Kebede, Mitchel Katsvanga, and 4 more authors

   Scientific reports, 2025

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   Malaria vaccines, RTS, S/AS01 and R21/Matrix which are based on the Plasmodium falciparum circumsporozoite protein (Pfcsp) have been approved by WHO for broad use in children in Africa. However, the extensive genetic diversity of Pfcsp limited its effectiveness, as vaccine efficacy reduced against non-vaccine strains. Using Oxford Nanopore Technology, we conducted amplicon sequencing of the full-length Pfcsp gene from 96 clinical isolates collected from three health centers in Ethiopia and compared the results against a reference genome. The result showed absence of population differentiation among the Ethiopian isolates. The N-terminal region was relatively conserved, with a KLKQP motif was present across all isolates. However, mutation at position A98G and an insertion of amino acids (DGNNNNGDNGREGKDEDKR) were identified in this region. The number of NANP and NVDP repeats of the central region per haplotype ranged from 39 to 42. Additionally, the Th2R and Th3R epitopes in the C-terminal region exhibited extensive polymorphism with at least one amino acid substitution compared to the reference strains. Notably, none of the Ethiopian Pfcsp haplotypes matched the vaccine haplotype. Furthermore, haplotype network and phylogenetic tree analyses shown considerable similarity among local and global isolates. The findings of this study revealed a high Pfcsp genetic diversity highlighting the need for further studies to inform allele selection for universal or region-specific vaccine development as this may influence vaccine efficacy.

2024

1. 

   Novel Plasmodium falciparum histidine-rich protein 2/3 repeat type in Ethiopian malaria infection: does this affect performance of HRP2-based malaria RDT?

   Aynalem Mandefro, Alebachew Messele Kebede, Bacha Mekonen, and 6 more authors

   Malaria Journal, 2024

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   Rapid diagnostic tests (RDTs) provide quick, easy, and convenient early diagnosis of malaria ensuring better case management particularly in resource-constrained settings. Nevertheless, the efficiency of HRP2-based RDT can be compromised by Plasmodium falciparum histidine-rich protein 2/3 gene deletion and genetic diversity. This study explored the genetic diversity of PfHRP2/3 in uncomplicated malaria cases from Ethiopia.

2023

1. Adaptation

   

   Genomic analysis of Plasmodium vivax describes patterns of connectivity and putative drivers of adaptation in Ethiopia

   Alebachew Messele Kebede, Edwin Sutanto, Hidayat Trimarsanto, and 8 more authors

   Scientific Reports, 2023

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   Ethiopia has the greatest burden of Plasmodium vivax in Africa, but little is known about the epidemiological landscape of parasites across the country. We analysed the genomic diversity of 137 P. vivax isolates collected nine Ethiopian districts from 2012 to 2016. Signatures of selection were detected by cross-country comparisons with isolates from Thailand (n = 104) and Indonesia (n = 111), representing regions with low and high chloroquine resistance respectively. 26% (35/137) of Ethiopian infections were polyclonal, and 48.5% (17/35) of these comprised highly related clones (within-host identity-by-descent > 25%), indicating frequent co-transmission and superinfection. Parasite gene flow between districts could not be explained entirely by geographic distance, with economic and cultural factors hypothesised to have an impact on connectivity. Amplification of the duffy binding protein gene (pvdbp1) was prevalent across all districts (16–75%). Cross-population haplotype homozygosity revealed positive selection in a region proximal to the putative chloroquine resistance transporter gene (pvcrt-o). An S25P variant in amino acid transporter 1 (pvaat1), whose homologue has recently been implicated in P. falciparum chloroquine resistance evolution, was prevalent in Ethiopia (96%) but not Thailand or Indonesia (35–53%). The genomic architecture in Ethiopia highlights circulating variants of potential public health concern in an endemic setting with evidence of stable transmission.

2. 

   Amplicon deep sequencing of five highly polymorphic markers of Plasmodium falciparum reveals high parasite genetic diversity and moderate population structure in Ethiopia

   Abeba Gebretsadik Reda, Tiffany Huwe, Cristian Koepfli, and 5 more authors

   Malaria Journal, 2023

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   Background* Plasmodium falciparum genetic diversity can add information on transmission intensity and can be used to track control and elimination interventions.Methods* Dried blood spots (DBS) were collected from patients who were recruited for a P. falciparum malaria therapeutic efficacy trial in three malaria endemic sites in Ethiopia from October to December 2015, and November to December 2019. qPCR-confirmed infections were subject to amplicon sequencing of polymorphic markers ama1-D3, csp, cpp, cpmp, msp7. Genetic diversity, the proportion of multiclonal infections, multiplicity of infection, and population structure were analysed. Results* Among 198 samples selected for sequencing, data was obtained for 181 samples. Mean MOI was 1.38 (95% CI 1.24–1.53) and 17% (31/181) of infections were polyclonal. Mean He across all markers was 0.730. Population structure was moderate; populations from Metema and Metehara 2015 were very similar to each other, but distinct from Wondogent 2015 and Metehara 2019. Conclusion* The high level of parasite genetic diversity and moderate population structure in this study suggests frequent gene flow of parasites among sites. The results obtained can be used as a baseline for additional parasite genetic diversity and structure studies, aiding in the formulation of appropriate control strategies in Ethiopia.

2022

1. 

   Temporal dynamics of Plasmodium falciparum population in Metehara, east-central Ethiopia

   Abeba Gebretsadik Reda, Alebachew Messele, Hussein Mohammed, and 3 more authors

   Malaria Journal, 2022

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   Plasmodium falciparum is the most serious, genetically most complex and fastest-evolving malaria parasite. Information on genetic diversity of this parasite would guide policy decision and malaria elimination endeavors. This study explored the temporal dynamics of P. falciparum population in two time points in Metehara, east-central Ethiopia.

2. 

   Equine histoplasmosis in Ethiopia: phylogenetic analysis by sequencing of the internal transcribed spacer region of rRNA genes

   Gobena Ameni, Alebachew Messele Kebede, Aboma Zewude, and 7 more authors

   Frontiers in cellular and infection microbiology, 2022

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   Equine histoplasmosis commonly known as epizootic lymphangitis (EL) is a neglected granulomatous disease of equine that is endemic to Ethiopia. It is caused by Histoplasma capsulatum variety farciminosum, a dimorphic fungus that is closely related to H. capsulatum variety capsulatum. The objective of this study was to undertake a phylogenetic analysis of H. capsulatum isolated from EL cases of horses in central Ethiopia and evaluate their relationship with H. capsulatum isolates in other countries and/or clades using the internal transcribed spacer (ITS) region of rRNA genes. Clinical and mycological examinations, DNA extraction, polymerase chain reaction (PCR), Sanger sequencing, and phylogenetic analysis were used for undertaking this study. Additionally, sequence data of Histoplasma isolates were retrieved from GenBank and included for a comprehensive phylogenetic analysis. A total of 390 horses were screened for EL and 97 were positive clinically while H. capsulatum was isolated from 60 horses and further confirmed with PCR, of which 54 were sequenced. BLAST analysis of these 54 isolates identified 29 H. capsulatum isolates and 14 isolates from other fungal genera while the remaining 11 samples were deemed insufficient for further downstream analysis. The phylogenetic analysis identified five clades, namely, African, Eurasian, North American 1 and 2, and Latin American A and B. The Ethiopian isolates were closely aggregated with isolates of the Latin American A and Eurasian clades, whereas being distantly related to isolates from North American 1 and 2 clades as well as Latin American B clade. This study highlights the possible origins and transmission routes of Histoplasmosis in Ethiopia.

2021

1. 

   Genetic diversity and acquired drug resistance mutations detected by deep sequencing in Virologic failures among antiretroviral treatment experienced human immunodeficiency Virus-1 patients in a pastoralist region of Ethiopia

   Erdaw Tachbele, Samuel Kyobe, Fred Ashaba Katabazi, and 8 more authors

   Infection and Drug Resistance, 2021

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   Purpose*: This study was conducted to investigate the drug resistance mutations and genetic diversity of HIV-1 in ART experienced patients in South Omo, Ethiopia. Patients and Methods*: A cross-sectional study conducted on 253 adult patients attending ART clinics for ≥ 6 months in South Omo. Samples with VL ≥ 1000 copies/mL were considered as virological failures (VF) and their reverse transcriptase gene codons 90– 234 were sequenced using Illumina MiSeq. MinVar was used for the identification of the subtypes and drug resistance mutations. Phylogenetic tree was constructed by neighbor-joining method using the maximum likelihood model. Results*: The median duration of ART was 51 months and 18.6% (47/253) of the patients exhibited VF. Of 47 viraemic patients, the genome of 41 were sequenced and subtype C was dominant (87.8%) followed by recombinant subtype BC (4.9%), M-09-CPX (4.9) and BF1 (2.4%). Of 41 genotyped subjects, 85.4% (35/41) had at least one ADR mutation. Eighty-one percent (33/41) of viraemic patients harbored NRTI resistance mutations, and 48.8% (20/41) were positive for NNRTI resistance mutations, with 43.9% dual resistance mutations. Among NRTI resistance mutations, M184V (73.2%), K219Q (63.4%) and T215 (56.1%) complex were the most mutated positions, while the most common NNRTI resistance mutations were K103N (24.4%), K101E, P225H and V108I 7.5% each. Active tuberculosis (aOR=13, 95% CI= 3.46– 29.69), immunological failure (aOR=3.61, 95% CI=1.26– 10.39), opportunistic infections (aOR=8.39, 95% CI= 1.75– 40.19), and poor adherence were significantly associated with virological failure, while rural residence (aOR 2.37; 95% CI: 1.62– 9.10, P= 0.05), immunological failures (aOR 2.37; 95% CI: 1.62– 9.10, P= 0.05) and high viral load (aOR 16; 95% CI: 5.35 51.59, P < 0.001) were predictors of ADR mutation among the ART experienced and viraemic study subjects. Conclusion*: The study revealed considerable prevalence of VF and ADR mutation with the associated risk indicators. Regular virological monitoring and drug resistance genotyping methods should be implemented for better ART treatment outcomes of the nation.

2. 

   Genetic variation of Plasmodium falciparum histidine-rich protein 2 and 3 in Assosa zone, Ethiopia: its impact on the performance of malaria rapid diagnostic tests

   Gezahegn Solomon Alemayehu, Alebachew Messele, Kayla Blackburn, and 4 more authors

   Malaria Journal, 2021

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   Rapid diagnostic tests (RDT) are commonly used for the diagnosis of malaria caused by Plasmodium falciparum. However, false negative results of RDT caused by genetic variation of P. falciparum histidine-rich protein 2 and 3 genes (pfhrp2/3) threaten existing malaria case management and control efforts. The main objective of this study was to investigate the genetic variations of the pfhrp2/3 genes.

2020

1. 

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

   Lemu Golassa, Alebachew Messele, Alfred Amambua-Ngwa, and 1 more author

   PloS one, 2020

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   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.

2019

1. Vaccine-design

   

   Effect of low complexity regions within the PvMSP3αblock II on the tertiary structure of the protein and implications to immune escape mechanisms

   Alebachew Messele Kebede, Fitsum Girma Tadesse, Adey Desta Feleke, and 2 more authors

   BMC structural biology, 2019

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   Plasmodium vivax merozoite surface protein 3α (PvMSP3α) is a promising vaccine candidate which has shown strong association with immunogenicity and protectiveness. Its use is however complicated by evolutionary plasticity features which enhance immune evasion. Low complexity regions (LCRs) provide plasticity in surface proteins of Plasmodium species, but its implication in vaccine design remain unexplored. Here population genetic, comparative phylogenetic and structural biology analysis was performed on the gene encoding PvMSP3α.