Polymorphism analysis of propeller domain of k13 gene in Plasmodium ovale curtisi and Plasmodium ovale wallikeri isolates original infection from Myanmar and Africa in Yunnan Province, China

Abstract

Background: Eighteen imported ovale malaria cases imported from Myanmar and various African countries have been reported in Yunnan Province, China from 2013 to 2018. All of them have been confirmed by morphological examination and 18S small subunit ribosomal RNA gene (18S rRNA) based PCR in YNRL. Nevertheless, the subtypes of Plasmodium ovale could not be identified based on 18S rRNA gene test, thus posing challenges on its accurate diagnosis. To help establish a more sensitive and specific method for the detection of P. ovale genes, this study performs sequence analysis on k13-propeller polymorphisms in P. ovale. Methods: Dried blood spots (DBS) from ovale malaria cases were collected from January 2013 to December 2018, and the infection sources were confirmed according to epidemiological investigation. DNA was extracted, and the coding region (from 206th aa to 725th aa) in k13 gene propeller domain was amplified using nested PCR. Subsequently, the amplified products were sequenced and compared with reference sequence to obtain CDS. The haplotypes and mutation loci of the CDS were analysed, and the spatial structure of the amino acid peptide chain of k13 gene propeller domain was predicted by SWISS-MODEL. Results: The coding region from 224th aa to 725th aa of k13 gene from P. ovale in 83.3% of collected samples (15/18) were amplified. Three haplotypes were observed in 15 samples, and the values of Ka/Ks, nucleic acid diversity index (π) and expected heterozygosity (He) were 3.784, 0.0095, and 0.4250. Curtisi haplotype, Wallikeri haplotype, and mutant type accounted for 73.3% (11/15), 20.0% (3/15), and 6.7% (1/15). The predominant haplotypes of P. ovale curtisi were determined in all five Myanmar isolates. Of the ten African isolates, six were identified as P. o. curtisi, three were P. o. wallikeri and one was mutant type. Base substitutions between the sequences of P. o. curtisi and P. o. wallikeri were determined at 38 loci, such as c.711. Moreover, the A > T base substitution at c.1428 was a nonsynonymous mutation, resulting in amino acid variation of T476S in the 476th position. Compared with sequence of P. o. wallikeri, the double nonsynonymous mutations of G > A and A > T at the sites of c.1186 and c.1428 leads to the variations of D396N and

Comparison of Plasmodium ovale curtisi and Plasmodium ovale wallikeri infections by a meta-analysis approach

“…After screening the full texts, 200 articles were excluded due to the following reasons: 135 not related to species, 17 genetic studies unrelated to P. ovale spp., 15 review articles, 14 case reports or case series, 10 methodology studies, 4 studies using the same participants, 2 letters, 1 with full-text that could not be retrieved, 1 with data that could not be extracted and 1 systematic review. Finally, a total of 36 studies 5 – 15 , 23 , 25 , 26 , 28 – 32 , 34 , 35 , 39 – 53 were found to meet the eligibility criteria and thus included in the present analysis (Fig. 1 ).…”

“…by Sutherland et al 25 , and 2020. The majority of included studies (14/36, 38.9%) were conducted in African countries (1 in Angola 9 , 1 in Equatorial Guinea 26 , 2 in Ethiopia 5 , 8 , 2 in Gabon 43 , 51 , 1 in Ghana 15 , 1 in Kenya 46 , 1 in Namibia 44 , 2 in the Republic of Congo/Congo 26 , 42 , 2 in Senegal 6 , 7 and 1 in Uganda 26 ), Asian countries (9 in China 12 , 14 , 32 , 35 , 39 , 49 , 50 , 52 , 53 , 2 in India 10 , 11 , 1 in Bangladesh 41 , 1 in Myanmar 12 and 1 in Thailand 13 ), Europe (3 in Italy 29 – 31 , 2 in France 23 , 45 , 2 in Spain 47 , 48 , 2 in the United Kingdom 25 , 34 and 1 in Germany 40 ) and Northern America (1 in Canada 28 ). Most of the included studies 23 , 25 , 28 , 30 – 32 , 34 , 35 , 39 , 40 , 45 , 47 – 50 , 52 , 53 (19/36, 52.8%) reported indigenous cases of malaria caused by P. ovale curtisi or P. ovale wallikeri , and the remaining studies (17/36, 47.2%) reported imported cases.…”

“…The first identification of the species P. ovale wallikeri and P. ovale curtisi was reported by Sutherland et al 25 . Since then, differences in morphology, clinical characteristics, laboratory parameters and genetic differences between the two species have been observed in Africa 5 – 9 , 15 , 26 , 42 – 44 , 46 , 51 , Asia 10 – 14 , 32 , 35 , 39 , 41 , 49 , 50 , 52 , 53 , Europe 23 , 25 , 29 – 31 , 34 , 40 , 45 , 47 , 48 , and North America 28 . This widespread distribution of P. ovale spp.…”

“…This widespread distribution of P. ovale spp. has necessitated the expansion of its research to other parts of the world, due to the increasing numbers of imported cases in non-endemic countries 23 , 25 , 28 , 30 – 32 , 34 , 35 , 39 , 40 , 45 , 47 – 50 , 52 , 53 .…”

“…The genetic polymorphisms that can distinguish between P. ovale curtisi and P. ovale wallikeri , including P. ovale spp. tryptophan-rich antigen ( potra ) 6 , 7 , P. ovale reticulate binding protein 2 ( porbp2 ) 11 , 25 , 34 , lactate dehydrogenase ( ldh ) 23 , cytochrome ( cytb b) 12 , cytochrome oxidase subunit 1 ( cox1 ) 12 , glyceraldehyde-3-phosphatase ( pog3p ) 26 , dihydrofolate reductase-thymidylate synthase ( podhfr-ts ) 12 and the k13 gene 35 , were initially identified to differentiate between the two P. ovale species.…”