A study on difference of transcription level of thermostable direct hemolysin in Vibrio parahaemolyticus / 预防医学

Objective To measure the transcription level of thermostable direct hemolysin gene (TDH)in 24 strains of Vibrio parahaemolyticus.Methods Total RNA was extracted from strains of Vibrio parahaemolyticus which were isolated from patients,seafood and environment.The RNA was proved TDH positive with routine PCR method;then the real -time fluorescent quantitative PCR was carried out to obtain the cycle of threshold (Ct)of THD and internal standard of 16s rRNA.Transcription level of THD compared with 16s rRNA was designated as ΔCt which was calculated as Ct value of THD minus Ct value of 16s rRNA.Results Ct values of THD,16s rRNA and the difference between them of the 24 strains was 18.04 ~25.95,8.30 ~10.93 and 8.28 ~15.34 respectively.The difference between the maximum and the minimum of ΔCt was 7.06;the highest transcription level was 133 (ΔΔCt =27.06 )times of the lowest one.Conclusion A great difference of transcription level of THD in Vibrio parahaemolyticus has been proved and further study is needed to clarify the possible molecular mechanisms and relationship between the transcription level of THD and pathogenic mechanism.

Analysis of beta-lactams-resistance genes in Pseudomonas aeruginosa in burn ward / 中华烧伤杂志

<p><b>OBJECTIVE</b>To investigate the resistance genes and antibiotic resistance patterns against beta-lactams in Pseudomonas aeruginosa prevalent in burn ward.</p><p><b>METHODS</b>K-B method was performed to test bacterial resistance patterns against 9 species of beta-lactams in Pseudomonas aeruginosa isolated from wounds and dressings of the patient in burn wards. Seven species of resistance genes against beta-lactams were detected with PCR. Tazobactam-inhibited piperacillin resistance test was performed to study whether the above strains produce extended spectrum beta-lactams.</p><p><b>RESULTS</b>All 12 strains of bacteria with resistance genes detected were resistant to penicillin and cephalosporins (100%), among them 11 were resistant to all antibiotics. Tazobactam-inhibited piperacillin resistance test demonstrated that all strains with resistance genes were ESBLs.</p><p><b>CONCLUSION</b>High incidence of beta-lactams resistance genes is found in Pseudomonas aeruginosa isolated from burn ward, and they have close relationship with the occurrence of multiple drug-resistance.</p>

Construction of the DNA vaccine of major outer membrane protein of Neisseria gonorrhoeae and investigation of immune effects after vaccination / 中华预防医学杂志

<p><b>OBJECTIVE</b>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.</p><p><b>METHODS</b>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.</p><p><b>RESULTS</b>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.</p><p><b>CONCLUSION</b>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.</p>

Comparison of class 1 integrons detected in Pseudomonas aeruginosa in different periods / 中华检验医学杂志

Objective To study the structure of class 1 integrons in 90 strains of Pseudomonas aeruginosa isolated during two periods of 1992-1996 and 2003-2005,and to get information about the structure changing of class 1 integrons by comparing their structures in two different periods.Methods Routine PCR and long PCR were performed to amplify the class 1 integrons and the gene cassettes they carried, followed with sequencing and blast via GenBank.Results Thirteen out of 41 strians ioslated during the period of 1992-1996 were positive on class 1 intergrons.Long PCR showed that the class 1 integron was 1868 bp in length and contained 2 resistance genes averagely.Six types of resistance genes of qacEA1 (n=6), sull (n=14),aadA1 (n=2),aadB (n=1),PSE-1 (n=2) and tetA (n=1) were found in these integrons, which consisted of 5 patterns of resistance cassette arrangements.Nineteen strains were proved to carry class 1 integrons in 49 isolates from 2003-2005.The mean DNA sequence length of them was 3383 bp with 3.6 resistant genes in averagely,10 types of resistance genes,qacEA1 (n=18),sull (n=25),aadA1 (n=6), aadB (n=7),aacA4 (n=2),PSE-1 (n=3),VEB-1 (n=4),OXA10 (n=1),cm1 A (n=1) and tetA (n =2),were identified in these integrons,which were composed of 9 patterns of resistance cassette arrangements.Conclusion In terms of produce length and resistance cassettes carried in the integrons, greater complexity is found in the structure of class 1 integrons in strains isolated during 2003-2005 than those during 1992-1996.