基于端粒序列的双重PCR检测法检测克氏锥虫和兰氏锥虫感染
Detection of Trypanosoma cruzi and Trypanosoma rangeli infection by duplex PCR assay based on telomeric sequences.
作者信息
Chiurillo Miguel Angel, Crisante Gladys, Rojas Agustina, Peralta Andreina, Dias Manuel, Guevara Palmira, Añez Néstor, Ramírez José Luis
机构信息
Decanato de Medicina, Universidad Centroccidental Lisandro Alvarado, Barquisimeto 3001, Venezuela.
出版信息
Clin Diagn Lab Immunol. 2003 Sep;10(5):775-9. doi: 10.1128/cdli.10.5.775-779.2003.
Abstract
We used the species specificity and repetitious nature of subtelomeric kinetoplastida sequences to generate a duplex PCR assay for the simultaneous detection of Trypanosoma cruzi and Trypanosoma rangeli in experimentally and naturally infected triatomine (Reduviid) bugs and in infected human subjects. The assay was species specific and was capable of detecting 1/20th of T. cruzi and 1/4th of T. rangeli cell equivalents without complementary hybridization. In addition, the PCR-based assay was robust enough for direct application to difficult biological samples such as Reduviid feces or guts and was capable of recognizing all T. cruzi and T. rangeli strains and lineages. Because the assay primers amplify entirely different target sequences, no reaction interference was observed, facilitating future adaptation of this assay to an automated format.
摘要
我们利用端粒下动基体序列的物种特异性和重复性，开发了一种双重PCR检测方法，用于同时检测实验感染和自然感染的锥蝽（猎蝽科）臭虫以及感染人类受试者体内的克氏锥虫和兰氏锥虫。该检测方法具有物种特异性，无需互补杂交即可检测到1/20的克氏锥虫细胞当量和1/4的兰氏锥虫细胞当量。此外，基于PCR的检测方法足够稳健，可直接应用于如猎蝽粪便或肠道等难以处理的生物样本，并且能够识别所有克氏锥虫和兰氏锥虫菌株及谱系。由于检测引物扩增的是完全不同的靶序列，未观察到反应干扰，这便于该检测方法未来采用自动化形式。
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