Analysis of polymorphisms in the circumsporozoite protein gene of Plasmodium vivax isolates from Henan Province, China

Abstract

Background: Plasmodium vivax malaria has historically been a major source of disease in Henan, China. In the 1970s, the morbidity of malaria was highest in the country. With support from the government and the efforts of healthcare personnel, the reported malaria cases have declined dramatically and a national elimination programme was launched in 2010. To achieve the goal, it is essential to study the diversity of autochthonous malaria and transmission of Plasmodium parasites, which will provide baseline data for disease control and management.

Methods: Thirty-two P. vivax isolates from Henan province were collected from 2008 to 2011, and circumsporozoite protein (csp) genes were analysed to estimate the genetic diversity of this parasite.

Results: The assessment of csp sequences indicated that all the isolates were the VK210 type, however, none of them was identical to the VK210 strain. The sequences displayed variations in the central region, and eight sub-types were observed. Among the sub-types, HN7 was the most prevalent (37.5%), followed by HN3 (34.4%). A total of 653 repeat units were discovered in 32 Henan isolates. Nucleotide sequences were grouped in 13 unique repeat nucleotide sequence allotypes that coded for 7 different repeated amino acid allotypes. B (GNGAGGQAA) and D (GDRAAGQPA) were more frequent based on the results; they represented 53.9% (352/653) of the total. In comparison to the basic repeat units of VK210, more than 75% of the central repeat units had at least one non-synonymous nucleotide change.

Conclusions: Recent P. vivax populations in Henan province showed some degree of genetic diversity in csp, with 8 sub-types among 32 samples. Meantime, the results also suggested its relative conserved parasite populations. This could provide interesting baseline data that allow identifying whether potential new cases differ from the parasites already circulating in the area.

Keywords: CSP; China; Diversity; Henan; Plasmodium vivax.

Fig. 1

Malaria Incidence in Henan Province during 2005–2011

Fig. 2

Malaria situation and Sub-types distribution in Henan province, China. The constituent ratio of the reported cases in different cities during 2005–2011 was shown in the map. The cities with the most reported cases was Shangqiu city, the following was Nanyang city. a The eight sequences from Nanyang (HNNY) parasites represent four sub-types. b The twenty-two sequences from Shangqiu (HNSQ) parasites represent six sub-types

Fig. 3

Sequence alignments of amino acid of 8 CSP distinct allelic variants found from 32 Pv isolates. Sequences were lined up to the amino acid sequences of the reference strainVK210(M28746) using ClustalW and Bio Edit software. Dashes and dots represent identical deletions and residues, respectively

Fig. 4

Schematic representation of the alignment of the amino acid motifs for the eight allelic CSP variants with the VK210 repeat type. A different variant is shown in each row. Dashes and dots represent identical deletions and residues, respectively

Fig. 5

Dendrogram of csp gene based on the amino acid from 32 Pv Henan isolates and 70 published CSP sequences collected around the world. The tree was conducted by using the Neighbor-Joining method in MEGA version 7.0 software. The optimal tree with the sum of branch length = 1.73200559 is shown. The tree is drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the Poisson correction method and are in the units of the number of amino acid substitutions per site. The bootstrap values are indicated on the branches and these values show the number of times out of 500 replications. The geographical origin of the 70 CSP published sequences were as follows: Brazil (FJ845390), China (U08977–U08978, AAC46499-AAC46501, FJ601732, FJ601755, FJ601725), Columbia (KC154046, GU339085), El-Salvador (J02751), Gabon (U09737), Greece (KC896384), Honduras (DQ156131), India (KM099683, JQ308526, ACN69860, ACB38259), Indonesia (JQ308527), Iran (AAT73115, AY632261, AY632300), Mauritania (AY674050, AAV80840), Myanmar (EU048255), North Korea (AF316580, AF316581, M20670, DQ859734), Papua New Guinea (ABX71468, EU031828), Philippine (U08980–U08981, AAC46502), Solomon Island (U08982–U08983, AAC46505), South Korea (AF215741, AF316583, AF436890, AY137777, AY137778, AJ295636, AF164605), Sri Lanka (JQ362650), Thailand (M34697, HQ011320, AAA29507), Vietnam (ACB38262, EU401929), VK210 (M28746), VK247 (M28745) and Hainan and Anhui province sequences