Résumé
Objective To evaluate the clinical value of recombinant major surface glycoprotein C(Msg C)consensus antigen of Pneumocystis jirovecii in serological diagnosis of Pneumocystis pneumonia(PCP), and explore serological diagnosis for PCP. Methods ELISA method was established for testing IgM antibody of Pneumocystis jirovecii Msg C consensus antigen. Serum antiMsg C consensus antigen IgM antibody and(1, 3)-β-D glucan were determined in 48 patients at high risk of PCP and 51 healthy subjects. The results of ELISA and(1, 3)-β-D glucan assay were compared with the results of PCR in bronchoalveolar lavage fluid. Results In a total of 99 specimens, Msg C consensus antigen IgM antibody detection(28.3％, 28/99)showed similar positive rate as(1, 3)-β-D glucan assay(25.3％, 25/99)(P>0.05). For the 48 patients at high risk of PCP, the positive rate of Msg C consensus antigen IgM antibody and(1, 3)-β-D glucan assay was 35.4％(17/48)and 33.3％(16/48), respectively(P>0.05). The two methods showed 67.7％ agreement in testing 99 specimens and 52.1％ agreement in testing 48 high-risk specimens. The bronchial lavage fluid samples of 48 patients at high risk were also tested by PCR. The result was positive in 15 cases(31.3％), showing no significant difference from Msg C consensus antigen IgM antibody test(P=0.665). The agreement between Msg C consensus antigen IgM antibody test and PCR was 58.3％. The agreement with PCR result increased to 84.0％ in the 25 specimens with the same result by two serological methods.When taking the positive result of either serological method as reference, serological method can detect majority of the PCR positive cases(86.7％, 13/15). Conclusions IgM antibody against Msg C consensus antigen in combination with serological marker(1, 3)-β-D glucan is valuable for PCP diagnosis. Further examination such as lower respiratory tract specimen PCR and conventional cytology should be carried out to confirm the diagnosis when both IgM antibody against Msg C consensus antigen and(1, 3)-β-D glucan are positive.
Résumé
Objective@#To obtain recombinant major surface glycoprotein(rMsg)of Pneumocystis jirovecii(Pj) and survey anti-Pj rMsg IgG and IgM antibodies using enzyme-linked immunosorbent assay (ELISA) among randomly selected sera from Beijing local children under 14 years of age.@*Methods@#ELISA method were established to detect anti-Pj rMsg IgG and IgM antibodies. We randomly selected sera of 201 cases who were under 14 years of age were screened.@*Results@#There were 78 cases in the age group of 1 day to 30 days, 33 cases in the group of 1 month to 12 months, and 90 cases in the group of 1 year to 14 years. The positive rates of anti-Pj rMsg IgG antibody in these three age groups were 10.3%(8/78), 21.2%(7/33), and 41.1%(37/90)respectively(χ2=21.190, P=0.000). The positive rates of anti-Pj rMsg IgM antibody were 6.4%(5/78), 39.4%(13/33), and 76.7%(69/90)respectively(χ2=84.260, P=0.000). The overall positive rates of anti-Pj rMsg IgG and IgM antibodies were 25.9%(52/201)and 43.3%(87/201)respectively(χ2=53.463, P=0.000). Among these, simultaneously IgG positive IgM positive, IgG positive IgM negative, IgG negative IgM positive and IgG negative IgM negative rates were 22.4%(45/201), 3.5%(7/201), 20.9%(42/201), and 53.2%(107/201)respectively.@*Conclusions@#The overall serological response rate after Pj exposure, either anti-Pj IgG or IgM antibody positive, was as high as 46.8% in children under 14 years of age in Beijing area. The positive rates of anti-rMsg IgG and IgM antibodies increased with age, with highest rate in age group of more than one years old.
Résumé
Objective:To construct and identify a plasmid vector of short interfering RNA(siRNA) on pneumocystis carinii(PC) major surface glycoprotein(MSG) gene.Methods:Short hairpin RNA(shRNA) oligonucleotide targeting PC MSG gene which was chemically synthesized and inserted into RNAi-Ready plasmid vector pTZU6+1 after annealing.the recombinant plasmid, pPC-MSG,transgormed into E.coil.TOP10 and amplified,was digested by restriction endonucleases HindⅢand EcoRⅠand identified by gelelectrophoresis and DNA sequencing.Results:Gel electrophoresis and DNA sequencing showed that the recombinant plasmid containing the correct and full shRNA oligonucleotide.Conclusion:The siRNA plasmid,pPC-MSG was constructed successfully.