Wilson S M
London School of Hygiene and Tropical Medicine, U.K.
Ann Trop Med Parasitol. 1995 Dec;89 Suppl 1:95-100.
As conventional methods for the detection and/or diagnosis of infections with Leishmania parasites have limitations, the DNA-based alternatives have received much attention. By targeting multi-copy sequences such as kinetoplast DNA, ribosomal RNA genes, mini-exon-derived RNA genes or genomic repeats, the sensitivity of these systems can be increased. Similarly, by targeting conserved or variable regions of these targets, the specificity can be tailored to the genus, complex, species or even the individual isolate level. There are two main approaches to DNA-based detection: DNA probes involving hybridization; and amplification approaches such as PCR. DNA probes are less sensitive than amplification but are useful for large-scale screening of sandfly vectors or reservoir hosts, for example. PCR is much more sensitive and has been used for patient diagnosis with a sensitivity greater than microscopy or culture. The application of DNA probes and PCR can be simplified using chemiluminescent or colorimetric end-points, respectively, but both techniques require some specialized equipment and a certain degree of technical expertise. For this reason, their use is limited to research laboratories or central diagnostic facilities.
由于检测和/或诊断利什曼原虫寄生虫感染的传统方法存在局限性，基于DNA的替代方法受到了广泛关注。通过靶向多拷贝序列，如动基体DNA、核糖体RNA基因、小外显子衍生RNA基因或基因组重复序列，可以提高这些系统的灵敏度。同样，通过靶向这些靶点的保守或可变区域，可以将特异性定制到属、复合体、物种甚至单个分离株水平。基于DNA的检测有两种主要方法：涉及杂交的DNA探针；以及PCR等扩增方法。DNA探针的灵敏度低于扩增，但例如可用于大规模筛查白蛉媒介或储存宿主。PCR的灵敏度要高得多，已用于患者诊断，其灵敏度高于显微镜检查或培养。分别使用化学发光或比色终点可以简化DNA探针和PCR的应用，但这两种技术都需要一些专门设备和一定程度的技术专长。因此，它们的使用仅限于研究实验室或中央诊断设施。
