[Construction of the DNA vaccine of major outer membrane protein of Neisseria gonorrhoeae and investigation of immune effects after vaccination].
OBJECTIVE
To clone PIB gene of Neisseria gonorrhoeae, and to construct a recombinant eukaryotic expression vector pCI-PIB and to understand the effects of pCI-PIB vaccination in mice to induce specific humoral and cellular immune responses.


METHODS
The entire PIB gene of Neisseria gonorrhoeae (960 bp) was amplified by using PCR. An eukaryotic eukaryotic vector pCI-PIB was then constructed. BALB/c mice (n = 65, 100 microg/time/mouse) were immunized with pCI-PIB by intramuscular injection. ABC assay was employed to examine the PIB expression in muscular cells of the pCI-PIB-immunized mice (n = 10). ELISA and MTT assays were used to measure the effects of humoral and cellular immune responses of the remaining pCI-PIB-immunized mice. By using slide agglutination test and complement bacteriolytic test, the serum anti-bacterial activity of the pCI-PIB immunized mice was determined.


RESULTS
The entire PIB gene amplification fragment of the expected size (960 bp) was successfully obtained by PCR. In comparison with the reported PIB gene sequence (GenBank No: AF090801), the homology of nucleotide sequence of the target inserted fragment in the recombinant plasmid pCI-PIB was as high as 99.28%. The muscular cells of the immunized mice could take in pCI-PIB and then express PIB. In the pCI-PIB immunized mice, the higher titer (1:4000) of specific serum IgG and the specific T lymphocyte response were found. The proliferation index (4.031) was significantly higher than that of the controls (1.127) (t = 71.71, P < 0.05). The sera and washings from the pCI-PIB immunized mice could agglutinate Neisseria gonorrhoeae and kill this microbe in presence of complements.


CONCLUSIONS
In this study we successfully constructed a recombinant eukaryotic expression vector pCI-PIB. The mice inoculated with pCI-PIB might efficiently produce the specific humoral and cellular immune responses, suggesting that pCI-PIB should be potential service as a candidate of Neisseria gonorrhoeae DNA vaccines.
Liao
Fang
F
Department of Microbiology, School of Basic Medical Sciences, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
He
Chao
C
Liu
Hai-Peng
HP
Song
Qi-Fa
QF
Yan
Jie
J
D000818:Animals; D000917:Antibody Formation; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000276:immunology*; D004274:DNA, Recombinant / Q000276:immunology; D005260:Female; D007111:Immunity, Cellular; D051379:Mice; D008807:Mice, Inbred BALB C; D009344:Neisseria gonorrhoeae / Q000235:genetics / Q000276:immunology*; D010957:Plasmids; D019444:Vaccines, DNA / Q000276:immunology*
D001425:Bacterial Outer Membrane Proteins; D004274:DNA, Recombinant; D019444:Vaccines, DNA