EVALUATION OF FREQUENCY OF GIARDIA LAMBLIA SUB SPECIES BY PCR-RFLP IN HOSPITALIZED CHILDREN,S STOOL SPECIMENS IN URMIA MUTAHHARI HOSPITAL

Manafi, G.R and Hazrati Tappeh, KH and Diba, K and Mohammadzadeh, H and Asgharzadeh, M and Gheibi, SH (2013) EVALUATION OF FREQUENCY OF GIARDIA LAMBLIA SUB SPECIES BY PCR-RFLP IN HOSPITALIZED CHILDREN,S STOOL SPECIMENS IN URMIA MUTAHHARI HOSPITAL. The Journal of Urmia University of Medical Sciences, 24 (6). pp. 414-422.

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Abstract

Giardia lamblia is one of the most prevalent intestinal flagellate protozoan that infects a wide range of vertebrate hosts. Giardia is one of the most common intestinal pathogens in children causing sever intestinal disorder and growth retardation. This study was performed to determine subspecies of Giardia lamblia by the PCR-RFLP method, targeting the glutamate dehydrogenase(gdh) locus, in hospitalized children at the Urmia Mutahhari hospital, West Azarbaijan Province, Iran Methods: In this study, thirty four stool specimen were collected from the hospitalized children in Urmia Motahhari hospital and dgiardia cysts were detected using microscopy. Cysts were partially purified by the sucrose density gradient method and then washed with sterile distilled water to effectively remove the PCR inhibitors. Genomic DNA of G.lamblia isolates was extracted by freeze-thaw cycles followed by phenol/ chloroform/isoamyl alcohol method. A single step PCR-RFLP assay was used to differentiate the assemblages A and B which been found in humans. In this method, 432 bp expected size were amplified and then for detection sub species were used of specific restriction RsaI and BspLI enzymes. Findings: From 720 examined samples, totally 34 samples were positive in term of giardia cyst so the parasite spread rate was reported 4.72%. Analysis of PCR-RFLP on these samples revealed that 28 samples (93.3%) had the genotype BIII and two samples (6.7%) belonged to the subgroup BIV. Conclusions: PCR-RFLP is a rapid and reliable method that enables us to effective genotype discrimination within Giardia assemblages using glutamate dehydrogenase gene, and makes it possible to identify the presence of mixed genotypes. In the base results, an animal source of infection is suggested.

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