Abstract:

AIM: To provide a theoretical basis for designing safe and effective vaccines of human cerebral malaria. METHODS: Genomic DNA samples of two P. falciparum isolates were prepared directly from 5 cases of cerebral malaria patients’blood in Mengla County, Yunnan Province(CMH/YN ) and in Yingjiang County, Yunnan Province (CYJ/YN ). The samples were used for polymerase chain reaction (PCR) amplification and the two pairs of oligonucleotides for the highly conserved genes encoding of FC27 merozoite surface protein 2 (MSP2) and the regions 12- 17 in MAD20 merozoite surface protein 1 (MSP1) of Papua New Guinea strain of P. falciparum were used as primers. The PCR products were digested with EcoRI and KpnI, BamHI and HindIII, respectively, and the generated fragments were cloned into M13mp18 and M13mp19 vectors and transfected into Escherichia coli (E. coli) TG1. A single colorless plaque on the LB agar plate containing X-gal (5-bromo-4-chloro-3-indolyl-β-D-galactoside) and IPTG (isopropylthio-β-D-galactoside) was randomly picked and transformed into E. coli JM103. The replicative form (RF) DNA (RFDNA ) of M13 recombinant DNA extracted from E. coli by the method of alkali lysis were digested with EcoRI and KpnI, BamHI and Hind III, respectively, and the generated fragments were identical with inserted foreign DNA 0.918 kb and 0.8 kb designed by ourselves. RESULTS: It is proved that M13 recombinant DNA consists of M13 vectors with an insert of genes encoding MSP2 and the regions 16 - 17 in MSP1 from two isolates CMH/YN and CYJ/YN of P. falciparum from Chinese patients with cerebral malaria at its corresponding site. CONCLUSION: The results demonstrate for the first time that both isolates CMH/YN and CYJ/YN of P. falciparum from Chinese patients with cerebral malaria examined contain genes identical to those defined in known MAD20 MSP1 and FC27 MSP2 allelic dimorphic family. These findings provide valuable strategies both for the development of vaccines to prevent human cerebral malaria and for the establishment of a specific detection method of P.falciparum from patients with cerebral malaria.

Key words: Cerebral malaria patient, Plasmodium falciparum, MSP1, MSP2, clone, vaccine