Mahbubani M H, Bej A K, Perlin M, Schaefer F W, Jakubowski W, Atlas R M
Department of Biology, University of Louisville, Kentucky 40292.
Appl Environ Microbiol. 1991 Dec;57(12):3456-61. doi: 10.1128/aem.57.12.3456-3461.1991.
A method was developed for the detection of Giardia cysts by using the polymerase chain reaction (PCR) and the giardin gene as the target. DNA amplification by PCR, using giardin DNA as the target, resulted in detection of both live and dead cysts. When giardin mRNA was used as the target, the ability to amplify cDNA by PCR depended on the mode of killing. Cysts killed by freezing were not detected by PCR when giardin mRNA was the target. Cysts killed by heating or exposure to monochloramine, however, gave positive detection signals for both DNA and giardin mRNA targets. The amount of giardin mRNA and total RNA was significantly increased in live cysts following the induction of excystation. Cysts killed by freezing, heating, or exposure to monochloramine did not show a change in RNA content. The detection of the giardin gene by PCR permits a sensitive and specific diagnosis for Giardia spp. Discrimination between live and dead cysts can be made by measuring the amounts of RNA or PCR-amplified product from the giardin mRNA target before and after the induction of excystation.
开发了一种利用聚合酶链反应（PCR）并以贾第虫肌动蛋白基因作为靶标来检测贾第虫包囊的方法。以贾第虫肌动蛋白DNA为靶标通过PCR进行DNA扩增，可检测到活的和死的包囊。当以贾第虫肌动蛋白mRNA为靶标时，通过PCR扩增cDNA的能力取决于杀灭方式。以贾第虫肌动蛋白mRNA为靶标时，冷冻杀死的包囊不能通过PCR检测到。然而，加热或暴露于一氯胺杀死的包囊，对于DNA和贾第虫肌动蛋白mRNA靶标均给出阳性检测信号。诱导脱囊后，活包囊中贾第虫肌动蛋白mRNA和总RNA的量显著增加。冷冻、加热或暴露于一氯胺杀死的包囊RNA含量未显示变化。通过PCR检测贾第虫肌动蛋白基因可对贾第虫属进行灵敏且特异的诊断。通过测量诱导脱囊前后来自贾第虫肌动蛋白mRNA靶标的RNA量或PCR扩增产物量，可区分活包囊和死包囊。
