阿米巴原虫LAMP检测方法的建立及其应用
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摘要
为了探索环介导等温扩增技术(LAMP)在牦牛粪便阿米巴原虫检测中的实用性,利用阿米巴原虫的18S rRNA基因序列进行引物设计,在优化LAMP检测方法的同时,对该方法在进行阿米巴原虫检测时的特异性、敏感性、重复性进行分析。结果显示:已建立的阿米巴原虫LAMP检测方法具有一定的特异性,敏感性优于PCR,并具有良好的重复性;在临床应用性方面,其检出率与利用传统PCR得到的检出率完全相同。结果表明:该LAMP检测方法可用于牦牛粪便中阿米巴原虫的检测。
In order to explore the applicability of loop-mediated isothermal amplification(LAMP) in detection of Entamoeba and to improve the detection efficiency of Entamoeba in Yak feces, primers were designed using the 18S rRNA gene of Entamoeba to establish the detection method of LAMP and to analyze the specificity, sensitivity, repeatability and clinical application of the method. The results showed the LAMP method established here only had a certain specificity, better sensitivity and repeatability, but also had clinical applicability, because the positive rate of the yak feces samples detected by LAMP was the same as that detected by traditional PCR. Therefore, the LAMP method can be applied to the detection of Entamoeba in feces of yak.
In order to explore the applicability of loop-mediated isothermal amplification(LAMP) in detection of Entamoeba and to improve the detection efficiency of Entamoeba in Yak feces, primers were designed using the 18S rRNA gene of Entamoeba to establish the detection method of LAMP and to analyze the specificity, sensitivity, repeatability and clinical application of the method. The results showed the LAMP method established here only had a certain specificity, better sensitivity and repeatability, but also had clinical applicability, because the positive rate of the yak feces samples detected by LAMP was the same as that detected by traditional PCR. Therefore, the LAMP method can be applied to the detection of Entamoeba in feces of yak.
引文
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[12] 李迎晓,焦凤超,赵瑜,等.沙门菌LAMP检测方法的建立[J].畜牧与兽医,2017,49(12):111-114.
[2] Ngui R,Angal L,Fakhrurrazi S A.Differentiating Entamoeba histolytica,Entamoeba dispar and Entamoeba moshkovskii using nested polymerase chain reaction(PCR) in rural communities in Malaysia[J].Parasit Vector,2012,5(1):187.
[3] 詹希美.人体寄生虫学[M].北京:北京人民卫生出版社,2001:35-47.
[4] 刘慧,孙晓东,聂仁华,等.3种方法检测粪便中溶组织内阿米巴的结果比较[J].中国病原生物学杂志,2008,3(9):679-680.
[5] 朱学军,刘晓燕,苏晓霁,等.18S rDNA的PCR扩增技术在棘阿米巴角膜炎临床诊断中的应用[J].国际眼科杂志,2009,9(4):715-718.
[6] Notomi T,Okayama H,Masubuchi H,et al.Loop-mediated isothermal amplification of DNA[J].Nucleic Acids Res,2000,28(12) :e63.
[7] Qiao J,Wang J,Meng Q,et al.Rapid detection of Akabane virus by a novel reverse transcription loop-mediated isothermal amplification assay(RT-LAMP)[J].Virol J,2013,10:288.
[8] 马国兰.浅谈青海的牦牛资源现状及保护措施[J].中国牛业科学,2014,40(3):89-91.
[9] Verweij J J,Laeijendecker D,Brienen E A,et al.Detection and identification of entamoeba species in stool samples by a reverse line hybridization assay[J].J Clin Microbiol,2003,41(11):5041-5045.
[10] Chung B S,Zhang H,Choi M H,et al.Development of resistance to reinfection by Clonorchis sinensis in rates[J].Korean J Parasitol,2004,42:19-26
[11] 肖璐,邬旭龙,王印,等.环介导等温扩增技术及其应用[J].动物医学进展,2015,36(7):113-117.
[12] 李迎晓,焦凤超,赵瑜,等.沙门菌LAMP检测方法的建立[J].畜牧与兽医,2017,49(12):111-114.