Polymorphisms analysis of the Plasmodium ovale tryptophan-rich antigen gene (potra) from imported malaria cases in Henan Province

Abstract

BackgroundPlasmodium ovale has two different subspecies: P. ovale curtisi and P. ovale wallikeri, which may be distinguished by the gene potra encoding P. ovale tryptophan-rich antigen. The sequence and size of potra gene was variable between the two P. ovale spp., and more fragment sizes were found compared to previous studies. Further information about the diversity of potra genes in these two P. ovale spp. will be needed.MethodsA total of 110 dried blood samples were collected from the clinical patients infected with P. ovale, who all returned from Africa in Henan Province in 2011–2016. The fragments of potra were amplified by nested PCR. The sizes and species of potra gene were analysed after sequencing, and the difference between the isolates were analysed with the alignment of the amino acid sequences. The phylogenetic tree was constructed by neighbour-joining to determine the genetic relationship among all the isolates. The distribution of the isolates was analysed based on the origin country.ResultsTotally 67 samples infected with P. o. wallikeri, which included 8 genotypes of potra, while 43 samples infected with P. o. curtisi including 3 genotypes of potra. Combination with the previous studies, P. o. wallikeri had six sizes, 227, 245, 263, 281, 299 and 335 bp, and P. o. curtisi had four sizes, 299, 317, 335 and 353 bp, the fragment sizes of 299 and 335 bp were the overlaps between the two species. Six amino acid as one unit was firstly used to analyse the amino acid sequence of potra. Amino acid sequence alignment revealed that potra of P. o. wallikeri differed in two amino acid units, MANPIN and AITPIN, while potra of P. o. curtisi differed in amino acid units TINPIN and TITPIS. Combination with the previous studies, there were ten subtypes of potra exiting for P. o. wallikeri and four subtypes for P. o. curtisi. The phylogenetic tree showed that 11 isolates were divided into two clusters, P. o. wallikeri which was then divided into five sub-clusters, and P. o. curtisi which also formed two sub-clusters with their respective reference sequences. The genetic relationship of the P. ovale spp. mainly based on the number of the dominant amino acid repeats, the number of MANPIN, AITPIN, TINPIN or TITPIS. The genotype of the 245 bp size for P. o. wallikeri and that of the 299 and 317 bp size for P. o. curtisi were commonly exiting in Africa.ConclusionThis study further proved that more fragment sizes were found, P. o. wallikeri had six sizes, P. o. curtisi had four sizes. There were ten subtypes of potra exiting for P. o. wallikeri and four subtypes for P. o. curtisi. The genetic polymorphisms of potra provided complementary information for the gene tracing of P. ovale spp. in the malaria elimination era.

Characterization of Plasmodium ovale spp. imported from Africa to Henan Province, China

“…Since P. ovale was classified into two different subspecies, P. o. curtisi and P. o. wallikeri 23 , more attention has been paid to the detection and genotype of the two subspecies 30–35 , and studies focusing on the prevalence and distribution of P. ovale spp. have been relatively few.…”

“…Nested PCR was performed using the methods published 42,43 . Subspecies of P. ovale were confirmed by nested PCR following sequencing according to previously published methods 23,30 .…”

Plasmodium ovale wallikeri and P. ovale curtisi Infections and Diagnostic Approaches to Imported Malaria, France, 2013–2018

“…We evaluated the polymorphism of potra genes in 80 samples and, as previously described, identified a limited number of polymorphisms (Table 4) (28). Our results, combined with those of Zhou et al (29), demonstrate that the potra gene is not a satisfying genetic marker of relapse. New genetic markers, such as microsatellite typing, need to be developed for P. ovale genotyping, as was previously done for P. falciparum (41,42) and P. vivax (43,44).…”

“…Isolates from GenBank under accession nos. HM594183 ( 28 ), MG588152, and MG588154 ( 29 ) were used as P. ovale curtisi reference sequences; HM594180 ( 28 ) and MG588148–150 ( 29 ) were used as P. ovale wallikeri reference sequences.…”

Immunogenicity analysis of genetically conserved segments in Plasmodium ovale merozoite surface protein-8

“…Ovale malaria in humans is caused by two closely related but distinct subspecies of the malarial parasite, P. ovale curtisi (the classic type) and P. ovale wallikeri (the variant type) [4]. Little attention has been paid to ovale malaria because it often presents with low parasitemia and mixed infection with other Plasmodium species, with mild clinical symptoms that are easily treated with the conventional antimalarial drug chloroquine [5–9].…”