| Summary: | 碩士 === 國立臺灣大學 === 獸醫學研究所 === 88 === The microfilariae of Dirofilaria immitis and Dipetalonema reconditum could circulate in the blood of hosts. They are very similar in morphology and easy to confuse. There are disadvantages of traditional differential diagnosis techniques and the current study has differentiated them by PCR assay. Owing to the absence of DNA sequence of Dipetalonema reconditum, we chose rDNA to approach. A set of univrsal primer was used to amplify rDNA segments, including 18S, ITS1, 5.8S, ITS2, and 28S, from Dirofilaria immitis and Dipetalonema reconditum. A PCR product of 1.2kbp was amplified from Dirofilaria immitis, and of 1.1kbp from Dipetalonema reconditum. The two rDNA segments were sequenced and published on GenBank (AF21800 and AF21801). Because there is no significant similarity of ITS2 between Dirofilaria immitis and Dipetalonema reconditum, two sets of specific primers were designed: IS and IT. A PCR product of 302bp could be amplified by using IS for Dirofilaria immitis and of 348bp by using IT for Dipetalonema reconditum. The detection limits of the three sets of primers are all 1X10-2 microfilaria and could amplify predicted PCR product from Dirofilaria immitis and Dipetalonema reconditum mixed samples. We also apply the established technique to differentiate the microfilariae successfully for the clinical cases.
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