猪鼻支原体3种核酸检测方法的临床应用比较

Evaluation on clinical application of three nucleic acid amplification assays for the detection of Mycoplasma hyorhinis

为研究猪鼻支原体(Mhr)核酸检测方法的优越性及在临床应用中的价值,本实验比较了常规PCR、套式PCR和荧光定量PCR 3种方法的敏感性,并进行了临床样品的检测。常规PCR敏感性为104拷贝/μL,套式PCR和荧光定量PCR敏感性均为10拷贝/μL;对74份肺组织、36份气溶胶、99份鼻拭子和47份气管支气管灌洗液样品进行检测,常规PCR、套式PCR、荧光定量PCR检出率分别为18.0%(46/256)、53.9%(138/256)和55.1%(141/256)。常规PCR与套式PCR、荧光定量PCR方法的检出率差异极显著(p<0.01),而套式PCR与荧光定量PCR方法的检出率差异无显著性(p>0.05)。结果表明,套式PCR敏感性高,具有较高的临床实用价值;荧光定量PCR敏感性高,最具有实验室诊断价值,但对操作者要求较高;常规PCR方法敏感性稍低,可能造成漏检,但易于操作,对条件简单的基层实验室仍有一定的实用性。在检测临床样品时,推荐采用套式PCR结合荧光定量PCR,结果更为准确可靠,可以提高Mhr的检出率。

To compared superiority and evaluate the conventional PCR, nested PCR and real-time PCR in clinical practice for Mycoplasma hyorhinis detections, the recombinant plasmid and clinical samples were detected parallelly by the three assay. The results showed that nested PCR and real-time PCR were the most sensitive method with the detection limit of 10 copies/IxL, but conventional PCR had a lower sensitivity with the detection limit of 10,000 copies/i.LL. In addition, a total of 256 clinical samples were detected by the three method including 74 lung tissue, 36 air samples, 99 nasal swab, 47 bronchoalveolar lavage fluids. The positive rates of nested PCR and real-time PCR were 53.9% （138/256） and 55.1% （141/256）, however, conventional PCR was 18.0% （46/256）. These results suggested that the method of nested PCR combining with real-time PCR is recommended in clinical detection of M.hyorhinis, which is not only cost efficient but also accurate.