Developmental Studies on the Laboratory diagnosis at an early stage of Mycoplasam pneumoniae-infection (mycoplasmal pneumonia)

我们建立了间接免疫荧光试验（IF）和乳胶凝集试验（LAT）检测呼吸道渗出液中肺炎支原体（MP）的方法，用于M。肺炎感染。并将IF和LAT与目前唯一的商业化快速检测方法DNA探针试验（DP）进行了比较。好吧该抗体与多种支原体交叉反应，但与正常人血清或健康人呼吸道分泌物不发生交叉反应。抗体与除支原体以外的支原体种属的交叉反应性。当与马血清和酵母提取物一起吸收时，Renitalium完全减少。培养基的成分。-M.用吸附的抗体检测生殖器的LAT和DP有交叉反应。但与M. Renitalium不太可能引起重大的诊断混淆，因为M。生殖器是 关于我们 明显低于。，的M。肺炎和M.生殖器是生殖道的唯一殖民者。因此，IF和LAT结合吸附的抗体和DP具有足够的特异性，可用于检测M。在体外，IF的检测限约为2 × 10^5 cfu/ml，LAT的检测限为2 × 10^5 cfu/ml，cif DP的检测限为5 × 10^4 cfu/ml。DP法敏感性最高。肺炎感染者IF阳性率为85.7%。临床标本中M.培养法检测肺炎支原体，IF阳性率为73.3%，LAT阳性率为63.3%，DP阳性率为26.6%。检查肺炎悬液。悬浮液和混合物（M.呼吸道分泌物）在37 ℃下孵育。随时间推移取出一等份，并检查LAT和DP。在LAT中，样品在三天内显示相同滴度。各样品的滴度（样品-cpm与阴性对照-cpm的比值）在5 h内显著下降。认为LAT和IF中的靶SS'分子积聚在咽部。但DP中的靶分子很快被破坏，不能预期积累。原因F。临床标本中DP的低阳性率可能是由于靶分子降解快所致，因此IF和LAT应适用于M.在临床实验室中从患者的呼吸道渗出物中检测肺炎克雷伯菌。特别推荐LAT，因为LAT的程序可以在半小时内完成，无需复杂的方式。临床标本中LAT的阳性率略低于IF。现在我们正在研究一种处理临床样品以提高LAT检测限的方法。少

We developed an indirect immunofluorescence test (IF) and a latex agglutination test (LAT) for detection of Mycoplasma pneumonias in respiratory exudated as rapid diagnosis at an early stage of M. pneumoniae-infection. Further, IF and LAT were compared with DNA-probe test (DP) which was the only commercially available test for the rapid detection of the organism.Firstly, we prepared polyclonal antibody specific to M. Pneumoniae. The antibody cross-react with many species of mycoplasmas, but not with normal human serum or with respiratory exudates from healthy person. Cross-reactivity of the antibody with species of mycoplasmas other than M. Renitalium was fully diminished when absorbed with horse serum and yeast extract. components of culture medium. -M. genitalium showed a cross-reaction in LAT with using the absorbed antibody and also in DP. However, the cross-reactivity with M. Renitalium, is unlikely to cause significant diagnostic confusion because the titer of M. genitalium was s … More ignificantly lower than., that of M. pneumonia, and M. genitalium is exclusively a genital tract colonizer. Therefore, IF and LAT with using the absorbed antibody, and DP are specific enough to be used for the detection of M. pneumonia in respiratory exudates.The detection limit of IF was about 2 x 10^5 cfu/ml, that-, of LAT was 2 x 10^5 cfu/ml and that cif DP was 5 x 10^4 cfu/ml in vitro. DP had the highest sensitivity among three methods.Among clinical specimens(throat smears)from patients with serologically confirmed M. pneumoniae-infection, 85.7% gave a positive test in IF. Among clinical specimens in which M. pneumoniae was detected by culture method, positive rate in IF was 73.3%, that in LAT was 63.3% and that in DP was 26.6%.A effect of the incubation of M. pneumonia suspension was examined. The suspension and the mixture (M. Pneumoniae suspension and respiratory exudates) were incubated at 37 ﾟC. An aliquot was removed with time course, and examinedin LAT and DP. The samples showed the same titer within three days in LAT. The titer (rate of sample-cpm to negativp- control-cpm) of each sample decreased remarkably within 5 h. It was considered that target SS'olecules in LAT and IF were accumulated in the pharyngeal portion. However, target molecule in DP (ribosomal RNA) was destructed much sooner, and the accumulation could not be expected. The reason f. or the low positive rate of DP in clinical'specimens may be due to the fast breakdown of target molecule.Consequently, IF and LAT must be applicable to the detection of M. pneumoniae in respiratory exudates from the patients in clinical laboratories. Especially, LAT was recommended because the procedure of LAT can be completed within a half hour without complicated manner. though the positive rate of LAT in clinical specimens was slightly lower than that of IF. Now we are examinating a process of treating clinical samples for rising the detection limit of LAT. Less