PCR detection of Giardia lamblia in stool: targeting intergenic spacer region of multicopy rRNA gene

A PCR based detection that amplifies the 552-bp intergenic spacer (IGS) reg ion of multicopy rRNA gene of Giardia lamblia and 320-bp internal sequences to first PCR product has been used in diagnosis of giardiasis in stool sam ple. The primers were found highly specific to Giardia spp. only, because n o amplification was observed with DNAs from other enteric pathogens like Es cherichia coli, Shigella dysenteriae and Entamoeba histolytica. The test co uld detect even less than 2 pg of genomic DNA from Giardia trophozoites. In direct diagnosis of Giardia lamblia in stool samples, it was observed that PCR amplification of IGS followed by nested PCR could enhance the sensitiv ity and specificity of the tests manifold and the system was able to detect as low as 10 parasites in 100 mu l of stool. The comparative evaluation of the present system with conventional microscopy, CIEP and ELISA in the dia gnosis of giardiasis from diarrhoeic stool samples and control subjects dem onstrated a 100% correlation among nested PCR, microscopic examination and ELISA in patients suggestive of giardiasis (Group I) and control subjects ( Group II). In Group cases (patients suffering from other than giardiasis), CIEP, ELISA and nested PCR showed better results than microscopic examinati on. However, among them, PCR was found most sensitive and specific because 20% positivity was noticed by PCR whereas CIEP and ELISA showed only 7.14% and 12.85%, respectively. Break-up results showed that all the samples whic h were positive by CIEP or ELISA, also found positive by PCR. The present o bservation clearly suggests the use of PCR that amplifies the intergenic sp acer region of multicopy rRNA gene of Giardia lamblia followed by nested PC R for routine, quick and reliable detection of Giardia lamblia in stool sam ples. (C) 2000 Academic Press.