Objective To study cellular immune responses induced by DNA vaccine against Chlamydia trachomatis (Ct) serotype E. Methods BALB/c mice were divided into three groups to be intramuscularly immunized by blank plasmid (negative control group), DNA vaccine against Ct serotype E (vaccine group), and inactivated Ct elementary body (positive control group), respectively. Two weeks after the last immunization,delayed-type hypersensitivity (DTH) response was evaluated; MTT assay was performed to detect the proliferation of spleen lymphocytes, ELISA to measure the serum level of interferon-γin mice. Some immunized mice underwent a genital challenge with Ct elementary body followed by isolation of Ct from exfoliated epithelial cells in genital tract and pathological examination of cervical tissue from the challenged mice. Results Compared to negative control group, vaccine group and positive control group experienced a stronger DTH response.The lymphocyte stimulating index and serum level of IFN-γwere highest in the positive control group (3.81 ±0.30, 2891.7 ± 1048.8 μg/L), followed by vaccine group (2.35 ± 0.25, 593.3 ± 342.6 μg/L) and negative control group (1.48 ± 0.15, 309.2 ± 157.9 μg/L), and significant difference was observed between the three groups (P ＜ 0.05 or 0.01 ). After Ct challenge, Ct was isolated from exfoliated epithelial cells and cervical tissue was damaged in the negative control group, while in the other two groups, Ct was undetected and genital tract tissue was intact. Conclusions The DNA vaccine against Ct serotype E could induce Ct-specific cellular immune responses to some extent, and offer a protection against vaginal challenge with Ct.

Objective To detect the antibodies against recombinant chlamydial plasmid-encoded protein 3(rPgp3),chlamydial protease-like activity factor(rCPAF),Ct143 encoded protein(rCT143), Ct101 encoded protein(rCT101),Ct694 encoded protein(rCT694),Ct813 encoded protein(rCT813), Chlamydia membrane protein A(rIncA),Ct875 encoded protein(rCT875),major outer membrane protein (rMOMP)and heat shock protein 60( rHsp60)in serum samples collected form patients with urogenital Chlamydia trachomatis(Ct)infection and to evaluate the antigenicity of those proteins. Methods The re-combinant plasmids expressing the 10 proteins and a blank plasmid were transformed into E. coli BL21 strains,respectively. The transformed E. coli BL21 strains were induced by isopropyl β-D-1-thiogalactopyr-anoside(IPTG)to express recombinant proteins. The glutathione pre-coated 96-well ELISA plates were coa-ted with lysates. Serum samples were collected from 50 patients with Ct infection and 10 patients without Ct infection. ELISA was performed to detect the antibodies against 10 recombinant proteins. Results The anti-bodies against rPgp3,rCPAF,rCT143,rCT101,rCT694,rCT875,rCT813,rMOMP,rIncA and rHsp60 proteins were respectively detected in 44 cases(88% ),38 cases(76% ),37 cases(74% ),36 cases (72% ),33 cases(66% ),31 cases(62% ),30 cases(60% ),26 cases(52% ),24 cases(48% )and 17 cases(34% )out of 50 serum samples. No antibodies against 10 recombinant proteins were detected in the serum samples collected from patients without Ct infection. Conclusion The rPgp3 protein showed the strongest antigenicity among all of the studied proteins,followed by rCPAF and rCT143 proteins. The rHsp60 protein showed the lowest antigenicity.

ObjectiveTo study the effect of a recombinant plasmid encoding mouse interleukin 2 (mlL-2) on the immunogenicity of DNA vaccine against Chlamydia trachomatis(Ct) serovar E.Methods BALB/c mice were divided into 4 groups to be intramuscularly inoculated with blank plasmid(negative control group),DNA vaccine against Ct serovar E(DNA vaccine group),DNA vaccine against Ct serovar E and a recombinant plasmid containing mIL-2(combination group),and inactivated Ct serovar E elementary bodies (positive control group),respectively.The immunological effects were evaluated by posterior foot pad thickness,proliferation level of spleen lymphocytes,serum level of IL-4 and interferon (IFN)-γ in mice,and the capability to clear Ct genital tract infection.ResultsThe proliferation index of spleen lymphocytes in the combination group and positive control group was similar(3.64 ± 0.41 vs.3.77 ± 0.34),but was significantly different from that in the blank control group and DNA vaccine group (1.37 ± 0.21 and 2.52 ± 0.30).The serum level of IL-4 was(38.49 ± 12.24) pg/ml in the positive control group,significantly higher than in the negative control group,DNA vaccine group and combination group ((25.37 ± 18.93),(24.75 ± 8.49),(21.74 ± 6.43) pg/ml,respectively).With respect to the serum level of IFN-γ,the combination group and positive control group were similar ((1923.3 ± 518.1) pg/ml vs.(2712.5 ± 887.2) pg/ml),but were significantly different from the negative control group and vaccine group((310.8 ± 160.7) pg/ml and(601.3 ± 357.9) pg/ml).Six days after Ct challenge,the exfoliated cells from genital tract were positive for Ct culture in the negative control group,but negative in the other 3 groups.ConclusionIL-2 genetic adjuvant can enhance the immune response,especially Th1 type response,induced by the DNA vaccine against Ct serovar E.

Objective To detect antibodies against chlamydial plasmid-encoded protein 3 (Pgp3),outer membrane complex protein B C-terminal peptide (OmcBc),CT841 protein and heat shock protein 60 (HSP60) in the sera of patients with urogenital Chlamydia trachomatis infection.Methods Recombinant plasmids encoding the aforementioned four proteins and an empty plasmid were transformed into Escherichia coli separately followed by 2-hour isopropyl-1-thio-β-galactopyranoside (IPTG) induction and cell lysis.The expressed proteins were purified with glutathione magnetic beads and then used to coat 96-well enzyme-linked immunosorbent assay (ELISA) plates precoated by glutathione.Serum samples were collected from 20 patients with and 20 clients without urogenital C.trachomatis infection attending the sexually transmitted disease (STD) clinic of Tianjin Medical University General Hospital.ELISA with the expressed protein-coated plates was adopted to detect antibodies against these proteins in the serum samples.Results Of the 20 serum samples from C.trachomatis-infected patients,14 (70％)had anti-Pgp3 antibody,9 (45％) anti-OmcBc antibody,8 (40％) anti-CT841 antibody,and 5 (25％) anti-HSP60 antibody.Meanwhile,no antibody was detected in any of the serum samples from uninfected clients except for one with anti-HSP60 antibody.Conclusions Of the four studied C.trachomatis proteins,Pgp3 appears to have the strongest antigenicity with the highest antibody-detection rate,while HSP60 exhibits the weakest antigenicity with the lowest antibody-detection rate.

Objective To investigate the morbility and epidemic characteristics of hair dye dermatitis in individuals who dyed their hair in Tianjin.Methods Questionnaires were distributed to the outpatients in the Gerneral Hospital of Tianjin Medical University,students and teachers in Tianjin Medical University,residents in the community and customers in barber shop from Aug.2007 to Mar.2008.The personal data including the methods and site of coloring hair and something correlated to hair dyes were investigated.Results A total of 597 cases with the history of coloring hair were enrolled in the study,including 485 women and 112 men,with mean age of 41 years (ranged from 16- 74years).Among 597 cases,69 cases had allergic reactions to hair dye,including 51 women and 18 men,with mean age of 44 years (ranged from 19-65 years).The median age of the first coloring hair was 40 years (ranged from 3-50 years).The comparison between the sites of coloring hair had no statistic significance (P ＞0.05),but there was a significant difference between black dyed hairs and col or dyed hairs (P ＜0.05).Conclusions People with black hair dye are prone to be allergic.

Objecfive To detect serum antibodies to Pmp in patients with urogenital Chlamydia trachomatis infection and to assess the relationship between Pmp and urogenital C.traehomatis infection.Methods Twenty healthy adults and 77 patients with urogenital C. trachomatis infection were recruited into this study.A 3-month foilow-up was carried out in 43 patients,who were classified into persistent infection group(n=19)and negative-conversion group(n=24).Western-blot was performed to detect serum antibodies to Pmp in all subjects.Results The positivity rate of anti-Pmp antibodies was 90.20% (71/77) in patients,significantly higher than that in the normal controls[20% (4/20),P<0.05].All the 9 types of anti-Pmp antibodies were detected in patients with a varying positivity rates,which were 61.04% (47/77),88.31% (68/77),63.63% (49/77),28.57% (22,77),63.63% (49/77),75.32% (58/77),62.34% (48/77),77.92% (60/77)and 70.13% (54/77) for antibodies against PmpA,PmpB,PmpC,PmpD,PmpE,PmpF,PmpG,PmpH and PmpI respectivelyThe prevalence was highest for anti-Pmp B antibodies and lowest for anti-Pmp D antibodies.There was no significant difference in the positivity rate of anti-Pmp antibodies between persistent infection group and negativeconversion group.Conclusions Anti-Pmp antibodies could be generated in patients infected with C. trachomatis.The immunogenicity of different Pmps is different,and the immunoprotective activity of Pmps is rather weak.Individual differences exist in serum anti-Pmp antibodies among patients.Nine types of Pmps are expressed in patients with urogenital C. trachomatis infection.

Objective To investigate the efficacy of widely used antibiotics for urogenital Chlamydia trachomatis infection in recent 5 years.Methods A total of 2 809 cases of Chlamydia trachomatis urogenital infected patients who visited STD clinics of Tianjin Medical University General Hospital from 2006 to 2010 were collected.All the patients had accomplished a course of treatment of azithromycin, minocycline, moxifloxacin or clarithromycin and followed up for 3 months (once every month).Cochran-Armitage trend test was used to analyzed the antibiotics effect changing trends overtime.Results From 2006 to 2010, the etiology clearance rates of azithromycin were 76.70% (79/103), 74.19% (92/124), 74.13% (106/143), 71.43% (100/140) and 70.77% (92/130), respectively;those of minocycline were 75.31% (61/81), 64.67% (97/150), 66.53% (159/239), 65.05% (188/289) and 63.03% (104/165), respectively;those of moxifloxacin were 88.82% (167/188), 86.23% (119/138), 82.96% (185/223), 81.19% (233/287) and 81.03% (158/37), respectively;those of clarithromycin were 82.93% (34/41), 80.49% (33/41), 79.25% (42/53), 78.18% (43/55) and 75.00% (18/24), respectively.Ochran-Armitage trend test showed that antimicrobial efficacy of moxifloxacin for urogenital Chlamydia trachomatis infection rates declined year by year (P<0.05), while those of azithromycin, clarithromycin and minocycline did not decline significantly (all P>0.05).Conclusions The etiology clearance rate of moxifloxacin is the highest but gradually declines by years, and that of azithromycin takes the second place, while the treatment efficacy of minocycline is lower but quite stable.The number of cases treated with clarithromycin is too small to draw a conclusion.

Objective To clone, express, and identify chlamydial GPIC capsid Vpl gene and pro- tein. Methods The complete sequence of Vp1 gene from?CPG1 phage was amplified. The amplicor was cut by restriction endonuclease, linked to plasmid vector, and transformed into E. coli.The expressed pro- tein of recombinant Vp1 was purified and identified. Results The recombinant 1 661 bp gene was se- quenced and proved to be?CPG1 Vp1 by searching Genebank. A 62 kDa capsid protein was expressed and confirmed by SDS-PAGE and Western blot. Conclusion The recombinant Vp1 seems to be a highly con- served and specific marker for chlamydial phage.

Objective To optimize immunodominant protein combinations for serological screening for Cblamydia trachomatis (Ct) infection.Methods Both serum and genital swab samples were collected from 50 patients with Ct infection confirmed by colloidal gold immunochromatographic assay (GICA),and 30 GICA-negative clients without Ct infection at a sexually transmitted disease (STD) clinic in Tianjin Medical University General Hospital.The 30 serum samples from GICA-negative clients were also negative for microimmunofluorescence (MIF) assay.Eight Ct immunodominant proteins,including Pgp3,CPAF,CT143,CT101,CT694,CT875,CT813 and IncA,were selected as antigens to detect corresponding antibodies in the serum samples by enzyme-linked immunosorbent assay (ELISA) with the Ct proteins Hsp60 and major outer membrane protein (MOMP) as references.The results of ELISA were compared with those of the traditional gold standard method MIF assay to determine the immunodominant protein combination with the highest sensitivity and specificity.Results Of the 50 serum samples from patients with Ct infection,44 were positive and 6 negative by MIF.The results of ELISA with the combination of immunodominant proteins Pgp3,CT694 and CT875 as antigens were 97.73％ (43/44) consistent to those of MIF assay.Of the 30 serum samples from GICA-negative clients,all were negative by MIF.Meanwhile,no antibody was detected against any of the immunodominant proteins Pgp3,CT694 and CT875 in any of the serum samples from GICA-negative clients.Conclusions The ELISA with the combination of immunodominant proteins Pgp3,CT694 and CT875 as antigens has good sensitivity and specificity for serological screening for Ct infection,and is simple to operate and easy to popularize.

Objective To evaluate the susceptibility of Chlamydia trachomatis clinical isolates to rifampin, and assess the relationship between rpoB mutations and antibiotic resistance in them.Methods A microculture method was used to determine the minimal inhibitory concentration (MIC) of rifampin in 52 Chlamydia trachomatis clinical isolates.The rpoB gene was amplified from all the clinical isolates and a standard strain of Chlamydia trachomatis followed by single-strand conformation polymorphism (SSCP)analysis.Sequencing of PCR products was carried out for two clinical isolates.Results No rifampin-resistant strain was found among these clinical isolates.The MIC of rifampin varied from 0.004 to 0.030 mg/L Neither SSCP analysis nor sequencing showed rpoB mutations.Conclusions No rpoB mutations were found in Chlamydia trachomatis isolates from patients unresponsive to rifampin.The unresponsiveness to rifampin may be attributed to multiple factors.