中国实验诊断学
中國實驗診斷學
중국실험진단학
CHINESE JOURNAL OF LABORATORY DIAGNOSIS
2009年
7期
872-874
,共3页
王连明%辛晓敏%路娟%王晓慧%王永晨
王連明%辛曉敏%路娟%王曉慧%王永晨
왕련명%신효민%로연%왕효혜%왕영신
申克孢子丝菌%引物%巢式PCR%病理诊断
申剋孢子絲菌%引物%巢式PCR%病理診斷
신극포자사균%인물%소식PCR%병리진단
Sporothrix schenckii%primer%nested PCR%pathologic diagnosis.
目的 研究实验小鼠申克孢子丝菌感染的分子生物学鉴定方法,为建立检测人申克孢子丝菌感染的快速、敏感、特异方法提供依据.方法 建立申克孢子丝菌感染小鼠模型,应用合成的申克孢子丝菌种特异性引物ITS3-SSP进行巢式PCR扩增小鼠皮损组织内核糖体DNA的ITS2靶序列,将检测结果与标准形态学鉴定结果对比,观察种特异性引物PCR扩增结果的准确程度、敏感性和特异性.结果 组织学检查可以从11只感染小鼠尾组织中见到染成紫红色的圆形、卵圆形孢子;第一循环PCR检测实验小鼠申克孢子丝菌,只有3只小鼠呈阳性;而巢式PCR检测申克孢子丝菌特异性DNA,11只实验小鼠中9只呈阳性;而3只对照组小鼠无一呈阳性.结论 本实验结果说明巢式PCR结合种特异性引物ITS3-SSP可以有效检出实验小鼠皮损中的申克孢子丝菌,具有一定的敏感性和特异性,尤其是对于组织学检查和真菌培养阴性的标本,本方法是一个很有潜力的替代方法.
目的 研究實驗小鼠申剋孢子絲菌感染的分子生物學鑒定方法,為建立檢測人申剋孢子絲菌感染的快速、敏感、特異方法提供依據.方法 建立申剋孢子絲菌感染小鼠模型,應用閤成的申剋孢子絲菌種特異性引物ITS3-SSP進行巢式PCR擴增小鼠皮損組織內覈糖體DNA的ITS2靶序列,將檢測結果與標準形態學鑒定結果對比,觀察種特異性引物PCR擴增結果的準確程度、敏感性和特異性.結果 組織學檢查可以從11隻感染小鼠尾組織中見到染成紫紅色的圓形、卵圓形孢子;第一循環PCR檢測實驗小鼠申剋孢子絲菌,隻有3隻小鼠呈暘性;而巢式PCR檢測申剋孢子絲菌特異性DNA,11隻實驗小鼠中9隻呈暘性;而3隻對照組小鼠無一呈暘性.結論 本實驗結果說明巢式PCR結閤種特異性引物ITS3-SSP可以有效檢齣實驗小鼠皮損中的申剋孢子絲菌,具有一定的敏感性和特異性,尤其是對于組織學檢查和真菌培養陰性的標本,本方法是一箇很有潛力的替代方法.
목적 연구실험소서신극포자사균감염적분자생물학감정방법,위건립검측인신극포자사균감염적쾌속、민감、특이방법제공의거.방법 건립신극포자사균감염소서모형,응용합성적신극포자사균충특이성인물ITS3-SSP진행소식PCR확증소서피손조직내핵당체DNA적ITS2파서렬,장검측결과여표준형태학감정결과대비,관찰충특이성인물PCR확증결과적준학정도、민감성화특이성.결과 조직학검사가이종11지감염소서미조직중견도염성자홍색적원형、란원형포자;제일순배PCR검측실험소서신극포자사균,지유3지소서정양성;이소식PCR검측신극포자사균특이성DNA,11지실험소서중9지정양성;이3지대조조소서무일정양성.결론 본실험결과설명소식PCR결합충특이성인물ITS3-SSP가이유효검출실험소서피손중적신극포자사균,구유일정적민감성화특이성,우기시대우조직학검사화진균배양음성적표본,본방법시일개흔유잠력적체대방법.
Objective To study the molecular biology identification methods of Sporothrix schenkii from experimentally infected mice,and to establish specific,sensitive and reliable method.Methods setting up a mouse model of Sporothrix schenckii infection,Sporothrix schenckii strain specific primers ITS3-SSP was used to amplify the internal transcribed spacer region 2 target sequences of mouse skin lesions tissues ribosome DNA by nested PCR.The results of species-specific primer PCR amplification were compared with that of standard morphological identification to observe the accuracy,sensitivity and specificity.Results Histological examination of 11 mice tail tissues showed round,oval fuchsia spores.Three mice presented positive in the first cycle of PCR detection.However,nested-PCR detection of Sporothrix schenckii-specific DNA give out 9 positive from 11 experimental mice and none were positive in the control group.Conclusion The results show that nested PCR combined with species-specific primers ITS3-SSP can effectively detect Sporothrix schenckii in experimental infected mice lesions and present a certain sensitivity and specificity,especially for histological examination and fungal culture negative specimens.This method is a promising alternative for clinical molecular diagnosis.
