Currently in China, the trend of Plasmodium vivax cases imported from Southeast Asia was increased especially in the China-Myanmar border area. Driven by the increase in P. vivax cases and stronger need for vaccine and drug development, several P. vivax isolates genome sequencing projects are underway. However, little is known about the genetic variability in this area until now. The sequencing of the first P. vivax isolate from China-Myanmar border area (CMB-1) generated 120 million paired-end reads. A percentage of 10.6 of the quality-evaluated reads were aligned onto 99.9% of the reference strain Sal I genome in 62-fold coverage with an average of 4.8 SNPs per kb. We present a 539-SNP marker data set for P. vivax that can identify different parasites from different geographic origins with high sensitivity. We also identified exceptionally high levels of genetic variability in members of multigene families such as RBP, SERA, vir, MSP3 and AP2. The de-novo assembly yielded a database composed of 8,409 contigs with N50 lengths of 6.6 kb and revealed 661 novel predicted genes including 78 vir genes, suggesting a greater functional variation in P. vivax from this area. Our result contributes to a better understanding of P. vivax genetic variation, and provides a fundamental basis for the geographic differentiation of vivax malaria from China-Myanmar border area using a direct sequencing approach without leukocyte depletion. This novel sequencing method can be used as an essential tool for the genomic research of P. vivax in the near future.

The current trend of Plasmodium vivax cases imported from Southeast Asia into China has sharply increased recently, especially from the China–Myanmar border (CMB) area. High recombination rates of P. vivax populations associated with varied transmission intensity might cause distinct local selective pressures. The information on the genetic variability of P. vivax in this area is scant. Hence, this study assessed the genetic diversity of P. vivax genome sequence in CMB area and aimed to provide information on the positive selection of new gene loci. This study reports a genome-wide survey of P. vivax in CMB area, using blood samples from local patients to identify population-specific selective processes. The result showed that considerable genetic diversity and mean pair-wise divergence among the sequenced P. vivax isolates were higher in some important gene families. Using the standardized integrated haplotype score (|iHS|) for all SNPs in chromosomal regions with SNPs above the top 1% distribution, it was observed that the top score locus involved 356 genes and most of them are associated with red blood cell invasion and immune evasion. The XP-EHH test was also applied and some important genes associated with anti-malarial drug resistance were observed in high positive scores list. This result suggests that P. vivax in CMB area is facing more pressure to survive than any other region and this has led to the strong positive selection of genes that are associated with host-parasite interactions. This study suggests that greater genetic diversity in P. vivax from CMB area and positive selection signals in invasion and drug resistance genes are consistent with the history of drug use during malaria elimination programme in CMB area. Furthermore, this result also demonstrates that haplotype-based detecting selection can assist the genome-wide methods to identify the determinants of P. vivax diversity.

China has made progress in malaria control and aims to eliminate malaria nationwide, but implementing effective interventions along the border regions remain a huge task. The Plasmodium falciparum cases imported from Southeast Asia has frequently reported especially in the China-Myanmar border (CMB) area. Though, information is scant on P. falciparum genetic variability in this area. This study reported P. falciparum isolates genome sequence of six clinical isolates in the CMB area. Furthermore, we estimated the nucleotide diversity, Watterson’s estimator and Tajima’s D value for the whole genome mutation rate in slide window. Our data were aligned onto 96.05–98.61% of the reference 3D7 genome in high fold coverages. Principal component analysis result showed that P. falciparum clustered generally according to their geographic origin. A total of 91 genes were identified as positive selection with Ka/Ks ratio significantly higher than 1, and most of them were multigene families encoding variant surface antigens (VSAs) such as var, rif and stevor. The enrichment of the positive selection on VSA genes implied that the environment complexity subjected CMB’s P. falciparum to more pressure for survival. Our research suggests that greater genetic diversity in CMB area and the positive selection signals in VSA genes, which allow P. falciparum to fit the host immune system well and aggravate the difficulty of treatment. Meanwhile, results obtained from this study will provide the fundamental basis for P. falciparum population genomic research in CMB area.

Malaria incidence has declined dramatically over the past decade and China was certified malaria-free in 2021. However, the presence of malaria in border areas and the importation of cases of malaria parasites are major challenges for the consolidation of the achievements made by China. Plasmodium vivax Duffy binding protein (PvDBP) performs a significant role in erythrocyte invasion, and is considered a promising P. vivax vaccine. However, the highly polymorphic region of PvDBP (PvDBP-II) impedes the development of blood-stage vaccine against P. vivax. In this study, we investigated the genetic diversity and natural selection of PvDBP-II among 124 P. vivax isolates collected from the China-Myanmar border (CMB) in Yunnan Province, China, during 2009–2011. To compare genetic diversity, natural selection, and population structure with CMB isolates, 85 pvdbp-II sequences of eastern Myanmar isolates were obtained from GenBank. In addition, global sequences of pvdbp-II were retrieved from GenBank to establish genetic differentiation relationships and networks with the CMB isolates. In total, 22 single nucleotide polymorphisms reflected in 20 non-synonymous and two synonymous mutations were identified. The overall nucleotide diversity of PvDBP-II from the 124 CMB isolates was 0.0059 with 21 haplotypes identified (Hd = 0.91). The high ratio of non-synonymous to synonymous mutations suggests that PvDBP-II had evolved under positive selection. Population structure analysis of the CMB and eastern Myanmar isolates were optimally grouped into five sub-populations (K = 5). Polymorphisms of PvDBP-II display that CMB isolates were genetically diverse. Mutation, recombination, and positive selection promote polymorphism of PvDBP-II of P. vivax population. Although low-level genetic differentiation in eastern Myanmar was identified along with the more effective malaria control measures, the complexity of population structure in malaria parasites has maintained. In conclusion, findings from this study advance knowledge of the understanding of the dynamic of P. vivax population, which will contribute to guiding the rational design of a PvDBP-II based vaccine.

Introduction: Plasmodium vivax (Pv) and P. knowlesi account together for a considerable share of the global burden of malaria, along with P. falciparum (Pf). However, inaccurate diagnosis and undetectable asymptomatic/submicroscopic malaria infections remain very challenging. Blood-stage antigens involved in either invasion of red blood cells or sequestration/cytoadherence of parasitized erythrocytes have been immunomics-characterized, and are vital for the detection of malaria incidence. Areas covered: We review the recent advances in Plasmodium immunomics to discuss serological markers with potential for specific and sensitive diagnosis of malaria. Insights on alternative use of immunomics to assess malaria prevalence are also highlighted. Finally, we provide practical applications of serological markers as diagnostics, with an emphasis on dot immunogold filtration assay which holds promise for malaria diagnosis and epidemiological surveys. Expert commentary: The approach largely contributes to Pf and Pv research in identifying promising non-orthologous antigens able to detect malaria incidence and to differentiate between past and recent infections. However, further studies to profiling naturally acquired immune responses are expected in order to help discover/validate serological markers of no cross-seroreactivity and guide control interventions. More so, the application of immunomics to knowlesi infections would help validate the recently identified antigens and contribute to the discovery of additional biomarkers of exposure, immunity, or both.

Plasmodium vivax apical membrane antigen-1 (PvAMA-1) is an important vaccine candidate for vivax malaria. However, antigenic variation within PvAMA-1 is a major obstacle to the design of a global protective malaria vaccine. In this study, we analyzed the genetic polymorphism and selection of the PvAMA-1 gene from 152 P. vivax isolates from imported cases to China, collected in the China–Myanmar border (CMB) area in Yunnan Province (YP) during 2009–2011 (n = 71) and 2014–2016 (n = 81), in comparison with PvAMA-1 gene information from Myanmar (n = 73), collected from public data. The overall nucleotide diversity of the PvAMA-1 gene from the 152 YP isolates was 0.007 with 76 haplotypes identified (Hd = 0.958). Results from the population structure suggested three groups among the YP and Myanmar isolates with optimized clusters value of K = 7. In addition, YP (2014–2016) isolates generally lacked some K components that were commonly found in YP (2009–2011) and Myanmar. Meanwhile, PvAMA-1 domain I is found to be the dominant target of positive diversifying selection and most mutation loci were found in this domain. The mutation frequencies of D107N/A, R112K/T, K120R, E145A, E277K, and R438H in PvAMA-1 were more than 70% in the YP isolates. In conclusion, high genetic diversity and positive selection were found in the PvAMA-1 gene from YP isolates, which are significant findings for the design and development of PvAMA-1-based malaria vaccine.