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Molecular analysis of reticulocyte binding protein-2 gene in Plasmodium vivax isolates from India.

Prajapati SK, Kumari P, Singh OP - BMC Microbiol. (2012)

Bottom Line: ApoI-RFLP was found to be more efficient in identifying the extent of genetic polymorphism in pvrbp-2 compared to AluI-RFLP.The study suggests that pvrbp-2 is highly polymorphic genetic marker which can be used for population genetic analyses.RFLP analysis suggests presence of nearly similar proportion of Sal-1 and Belem alleles in Indian P. vivax populations.

View Article: PubMed Central - HTML - PubMed

Affiliation: Molecular Biology Division, National Institute of Malaria Research, Sector 8, Dwarka, Delhi, India. surendramrc@gmail.com

ABSTRACT

Background: Plasmodium vivax reticulocyte binding protein-2 (PvRBP-2) is a promising candidate for development of vaccine against parasite. DNA sequence polymorphism in pvrbp-2 which may hamper the vaccine development program has been identified in laboratory strains. Therefore, unraveling genetic polymorphism in pvrbp-2 from field isolates is a prerequisite for success in vaccine development. This study was designed with a primary aim to uncover genetic polymorphism in pvrbp-2 among P. vivax field isolates.

Results: Using virtual restriction mapping of pvrbp-2 sequences, two restriction enzymes (AluI and ApoI) were selected for the development of pvrbp-2 as a PCR-RFLP marker. Restriction fragment length polymorphism (RFLP) analysis revealed a high degree of genetic polymorphism in the pvrbp-2 gene among field isolates of P. vivax. ApoI-RFLP was found to be more efficient in identifying the extent of genetic polymorphism in pvrbp-2 compared to AluI-RFLP. Combined genotyping/haplotyping of RFLP pattern revealed a total of 36 distinct RFLP patterns among 83 P. vivax isolates analyzed. DNA sequence analysis also supports high degree of genetic polymorphism among field isolates of P. vivax. Pvrbp-2 PCR-RFLP method is able to distinguish multiple infection up to 16.86% and it revealed a low level of shared genetic pool between more than two populations.

Conclusion: The study suggests that pvrbp-2 is highly polymorphic genetic marker which can be used for population genetic analyses. RFLP analysis suggests presence of nearly similar proportion of Sal-1 and Belem alleles in Indian P. vivax populations. The larger extent of genetic polymorphism identified from limited samples advocates to screen genetic polymorphism in pvrbp-2 from malaria endemic geographical regions and countries for designing pvrbp-2 based anti-malarial control measures.

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Distribution of AluI and ApoI RFLP genotypes among Indian field isolates of Plasmodium vivax.
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Figure 4: Distribution of AluI and ApoI RFLP genotypes among Indian field isolates of Plasmodium vivax.

Mentions: A substantial number of RFLP pattern for both enzymes were observed with respect to the Sal-1 strain based pvrbp-2 gene sequence. In total, 13 distinct AluI and 30 distinct ApoI RFLP patterns were observed among 83 samples. RFLP analysis revealed mainly two distinct digestion patterns in field isolates by both enzymes. This suggests that each enzyme has two major types of digestion pattern. RFLP pattern of six samples was confirmed by DNA sequencing. Among six samples, DNA sequences of five samples were in agreement with RFLP data, however in a single sample (Pv-7) RFLP pattern of only ApoI enzyme was not matched. This may be due to the sequencing of only one clone from each cloning experiment. The numbers of RFLP pattern of individual enzymes from all samples are listed in Table 1. The frequencies of AluI and ApoI genotypes varied in field isolates (Figure 4). Further, combination of AluI and ApoI RFLP patterns revealed a total of 36 distinct haplotypes/genotypes suggesting a high degree of genetic diversity in pvrbp-2 sequences in the field isolates of P. vivax.


Molecular analysis of reticulocyte binding protein-2 gene in Plasmodium vivax isolates from India.

Prajapati SK, Kumari P, Singh OP - BMC Microbiol. (2012)

Distribution of AluI and ApoI RFLP genotypes among Indian field isolates of Plasmodium vivax.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3539960&req=5

Figure 4: Distribution of AluI and ApoI RFLP genotypes among Indian field isolates of Plasmodium vivax.
Mentions: A substantial number of RFLP pattern for both enzymes were observed with respect to the Sal-1 strain based pvrbp-2 gene sequence. In total, 13 distinct AluI and 30 distinct ApoI RFLP patterns were observed among 83 samples. RFLP analysis revealed mainly two distinct digestion patterns in field isolates by both enzymes. This suggests that each enzyme has two major types of digestion pattern. RFLP pattern of six samples was confirmed by DNA sequencing. Among six samples, DNA sequences of five samples were in agreement with RFLP data, however in a single sample (Pv-7) RFLP pattern of only ApoI enzyme was not matched. This may be due to the sequencing of only one clone from each cloning experiment. The numbers of RFLP pattern of individual enzymes from all samples are listed in Table 1. The frequencies of AluI and ApoI genotypes varied in field isolates (Figure 4). Further, combination of AluI and ApoI RFLP patterns revealed a total of 36 distinct haplotypes/genotypes suggesting a high degree of genetic diversity in pvrbp-2 sequences in the field isolates of P. vivax.

Bottom Line: ApoI-RFLP was found to be more efficient in identifying the extent of genetic polymorphism in pvrbp-2 compared to AluI-RFLP.The study suggests that pvrbp-2 is highly polymorphic genetic marker which can be used for population genetic analyses.RFLP analysis suggests presence of nearly similar proportion of Sal-1 and Belem alleles in Indian P. vivax populations.

View Article: PubMed Central - HTML - PubMed

Affiliation: Molecular Biology Division, National Institute of Malaria Research, Sector 8, Dwarka, Delhi, India. surendramrc@gmail.com

ABSTRACT

Background: Plasmodium vivax reticulocyte binding protein-2 (PvRBP-2) is a promising candidate for development of vaccine against parasite. DNA sequence polymorphism in pvrbp-2 which may hamper the vaccine development program has been identified in laboratory strains. Therefore, unraveling genetic polymorphism in pvrbp-2 from field isolates is a prerequisite for success in vaccine development. This study was designed with a primary aim to uncover genetic polymorphism in pvrbp-2 among P. vivax field isolates.

Results: Using virtual restriction mapping of pvrbp-2 sequences, two restriction enzymes (AluI and ApoI) were selected for the development of pvrbp-2 as a PCR-RFLP marker. Restriction fragment length polymorphism (RFLP) analysis revealed a high degree of genetic polymorphism in the pvrbp-2 gene among field isolates of P. vivax. ApoI-RFLP was found to be more efficient in identifying the extent of genetic polymorphism in pvrbp-2 compared to AluI-RFLP. Combined genotyping/haplotyping of RFLP pattern revealed a total of 36 distinct RFLP patterns among 83 P. vivax isolates analyzed. DNA sequence analysis also supports high degree of genetic polymorphism among field isolates of P. vivax. Pvrbp-2 PCR-RFLP method is able to distinguish multiple infection up to 16.86% and it revealed a low level of shared genetic pool between more than two populations.

Conclusion: The study suggests that pvrbp-2 is highly polymorphic genetic marker which can be used for population genetic analyses. RFLP analysis suggests presence of nearly similar proportion of Sal-1 and Belem alleles in Indian P. vivax populations. The larger extent of genetic polymorphism identified from limited samples advocates to screen genetic polymorphism in pvrbp-2 from malaria endemic geographical regions and countries for designing pvrbp-2 based anti-malarial control measures.

Show MeSH
Related in: MedlinePlus