Comparison of different primes for PCR-based diagnosis of cutaneous leishmaniasis
Comparison of different primes for PCR-based diagnosis of cutaneous leishmaniasis
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DOI：
10.1590/s1413-86702011000300004
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发表时间：
2011-06-01
影响因子：
3.4
通讯作者：
Silveira, Thais Gomes Verzignassi
中科院分区：
文献类型：
--
作者：
Oliveira, Diego Molina de;Lonardoni, Maria Valdrinez Campana;Silveira, Thais Gomes Verzignassi
目的：分析在流行病学研究中常用的不同的聚合酶链式反应检测利什曼原虫DNA的方法，并将其与传统的美国皮肤利什曼病(ACL)直接寄生虫搜索法进行比较。材料和方法：使用5对引物，其中4对来自利什曼原虫kDNA序列(MP3H-MP1L；B1-B2；LBF1-LBR1；13A-13B)，1对来自先前报道的SL RNA(微型外显子)基因重复序列(LU5A-LB3C)。结果：以MP3 H-MP1L为模板扩增利什曼原虫DNA的效果最好，可检出2株FG。DNA13A-13B的检测效果最差，检测出512×10(3)Fg的DNA。结论：本研究发现，不同方法的分析灵敏度差异较大，与常规的ACL诊断方法有显著差异，强调了规范PCR技术、分析灵敏度、选择合适的寡核苷酸引物的重要性。
Objective: The objective of this study was to analyze different primers that are commonly used in epidemiological studies for the detection of Leishmania DNA by PCR, and to compare them to the conventional direct parasite search for American cutaneous leishmaniasis (ACL) diagnosis. Material and methods: Five pairs of primers, four of them derived from Leishmania kDNA sequences (MP3H-MP1L; B1-B2; LBF1-LBR1; 13A-13B), and one derived from the SL RNA (mini-exon) gene repeat (LU5A-LB3C), reported previously, were used. Results: The MP3H-MP1L primers were the best at amplifying the DNA, detecting 2 fg of Leishmania spp. DNA. The 13A-13B primers presented the worst performance, detecting 512 x 10(3) fg of DNA. Conclusion: The wide variation in the analytical sensitivity of the primers used in the PCR, and the significant differences from the conventional method of ACL diagnosis found in this study, emphasize the importance of standardizing the PCR technique, analyzing sensitivity, and selecting suitable oligonucleotide primers.