The online version of this article (https://doi.org/10.1186/s12936-017-2157-5) contains supplementary material, which is available to authorized users.
Antigen-detecting rapid diagnostic tests (RDTs) have been recommended by the World Health Organization for use in remote areas to improve malaria case management. Lactate dehydrogenase (LDH) of Plasmodium falciparum is one of the main parasite antigens employed by various commercial RDTs. It has been hypothesized that the poor detection of LDH-based RDTs is attributed in part to the sequence diversity of the gene. To test this, the present study aimed to investigate the genetic diversity of the P. falciparum ldh gene in Thailand and to construct the map of LDH sequence diversity in P. falciparum populations worldwide.
The ldh gene was sequenced for 50 P. falciparum isolates in Thailand and compared with hundreds of sequences from P. falciparum populations worldwide. Several indices of molecular variation were calculated, including the proportion of polymorphic sites, the average nucleotide diversity index (π), and the haplotype diversity index (H). Tests of positive selection and neutrality tests were performed to determine signatures of natural selection on the gene. Mean genetic distance within and between species of Plasmodium ldh was analysed to infer evolutionary relationships.
Nucleotide sequences of P. falciparum ldh could be classified into 9 alleles, encoding 5 isoforms of LDH. L1a was the most common allelic type and was distributed in P. falciparum populations worldwide. Plasmodium falciparum ldh sequences were highly conserved, with haplotype and nucleotide diversity values of 0.203 and 0.0004, respectively. The extremely low genetic diversity was maintained by purifying selection, likely due to functional constraints. Phylogenetic analysis inferred the close genetic relationship of P. falciparum to malaria parasites of great apes, rather than to other human malaria parasites.
This study revealed the global genetic variation of the ldh gene in P. falciparum, providing knowledge for improving detection of LDH-based RDTs and supporting the candidacy of LDH as a therapeutic drug target.
Additional file 1. Nucleotide sequence IDs of the ldh gene of Plasmodium falciparum in Thailand.
Additional file 2. Nucleotide sequence IDs of the ldh gene of Plasmodium falciparum from different geographical locations.
Additional file 3. Nucleotide sequence IDs of the ldh gene of the malaria parasites of humans and non-human primates.
Additional file 4. Neighbour Joining tree of 61 allelic sequences of the gene encoding lactate dehydrogenase ( ldh) from 12 Plasmodium parasite species. The sequences are named according to parasite species and allelic type. The first two letters indicate parasite species: Pf ( Plasmodium falciparum), Pm ( Plasmodium malariae), Po ( Plasmodium ovale), Pv ( Plasmodium vivax), Pp ( Plasmodium praefalciparum), Pr ( Plasmodium reichenowi), Pbi ( Plasmodium billcollinsi), Pbl ( Plasmodium blacklocki), Pa ( Plasmodium alderi), Pg ( Plasmodium gaboni), Pk ( Plasmodium knowlesi) and Pc ( Plasmodium cynomolgi). Species showed on the right hand site are labelled with color representing parasite host: Homo sapiens (blue), Gorilla gorilla (black), Pan troglodytes (green) and Macaca fascicularis (red). The tree was constructed using the aligned sequences of 768 nucleotides, corresponding to nucleotide position 52–819 after P. falciparum strain 3D7. Bootstrap values are shown next to the nodes. Scale bar shows nucleotide substitution per site.
Additional file 5. Sequence alignment of Plasmodium falciparum LDH epitopes in immunodiagnostic assays and LDH sequences of human Plasmodium species. Only LDH sequences that were different from LDH epitopes were shown. Letters in grey indicate polymorphic amino acid residues. Dot (.) indicates an amino acid residue identical to that of LDH epitopes.
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- Global sequence diversity of the lactate dehydrogenase gene in Plasmodium falciparum
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