[Comparative analysis of nucleotide sequences of lactate dehydrogenase (LDH) gene and LDH epitopes of Plasmodium vivax and Plasmodium falciparum].
OBJECTIVE
To analyze the difference of nucleotide sequences of lactate dehydrogenase (LDH) gene and LDH epitopes of Plasmodium vivax and P. falciparum.


METHODS
Specific primers were designed to amplify the full-length LDH gene sequence of P. vivax and P. falciparum (GenBank accession number: DQ198262 and DQ060151 respectively). The PCR products were sequenced and compared. The epitopes of objective LDH antigens were predicted by SYFPEITHI software. Pv-LDH and Pf-LDH genes were cloned into prokaryotic plasmid pET28a, then expressed in E. coli BL21(DE3) with isopropyl beta-D-1-thiogalactopyranoside (IPTG) induction. The immunogenicity of the recombinants Pv-LDH and Pf-LDH was analyzed by Western blotting and neutralization ELISA assays.


RESULTS
Pf-LDH gene was same to reference sequences(DQ198262), while there is a single nucleotide difference at the position 666 between Pv-LDH gene and reference sequences (DQ060151). The coding region of the two genes contained 951 bp encoding a 316-amino-acid residue. Compared with Pf-LDH, Pv-LDH showed a nucleotide sequence identity of 75.1%, and an amino acid sequence identity of 90.2%. T cell epitope prediction indicated that there were 28 human leukocyte antigen (HLA) types which could recognize pLDH antigen epitopes. The common or similar epitopes accounted for about 75% of the predicted 180 epitopes. The number of specific epitopes of Pv-LDH and Pf-LDH proteins was 38 and 45, respectively. Western blotting analysis showed that the Pv-LDH recombinant antigen reacted with the sera of malaria patients, and the reactivity was much lower than that of sera of immunized rabbit. Neutralization ELISA showed that about 70.3% reactivity of Pv-LDH polyclonal antibodies could be suppressed by Pv-LDH, while only 30.5% by Pf-LDH.


CONCLUSIONS
There are differences in DNA sequences of LDH gene and LDH epitopes between P. vivax and P. falciparum. The antibodies induced by the specific epitopes account for a small proportion in the antibody repertoire.
Jiang
Li
L
Changhai Municipal Center for Disease Control and Prevention, Shanghai 200336, China.
Wang
Zhen-yu
ZY
Ma
Xiao-jiang
XJ
Zhang
Xiao-ping
XP
Cai
Li
L
D000913:Antibodies, Protozoan / Q000276:immunology; D000953:Antigens, Protozoan / Q000276:immunology; D016054:DNA, Protozoan / Q000235:genetics; D000939:Epitopes / Q000235:genetics*; D017125:Genes, Protozoan; D007770:L-Lactate Dehydrogenase / Q000235:genetics* / Q000276:immunology; D010963:Plasmodium falciparum / Q000201:enzymology / Q000235:genetics* / Q000276:immunology; D010966:Plasmodium vivax / Q000201:enzymology / Q000235:genetics* / Q000276:immunology; D017422:Sequence Analysis, DNA
D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D016054:DNA, Protozoan; D000939:Epitopes; D007770:L-Lactate Dehydrogenase