التشخيص الجزيئي لطفيلي الاميبا الحالة للنسيج في منطقة الفرات الاوسط

Abstract

This study aimed to find the prevalence of E. histolytica parasite in Middle Euphrates region – Iraq, using one of the molecular methods. The study was conducted on the (1350) stool samples from diarrhea - suffering patients in three provinces of Middle Euphrates region: Karbala, Babel and Al- Najaf (450 in each province). Patients ranged in age (1 to more than 60) years and from both sexes and different regions who were attended to some hospitals, dispensaries and private laboratories in these governorates during the period from the first of May 2013 till the end of April 2014. All collected fresh samples were examined microscopically by wet-mounts method for E. histolytica detection. Around 0.5 g of each positive and some of negative stool samples in microscopic examination was preserved at -20Cº until use in molecular study. One third of positive samples in microscopic examination (which was selected to be representative for all positive samples) from each province and five of negative one as control group were submitted to the nucleic acid (DNA) extraction. One pairs of specific primers was designed to be used for molecular diagnosis of E. histolytica using PCR technique. Primers materials were supplied by Bioneer Co. and prepared according to its instructions. Molecular diagnosis of E. histolytica were performed using PCR reaction with amplification of the 355-bp fragments from the internal transcribed spacer region1 ITS1. The PCR was conducted using Accupower® PCR Premix kit/ Bioneer as DNA polymerase and the reaction was performed in a thermocycler. The products were submitted to 1.5 % agarose gel electrophoresis, stained with ethidium bromide and the gel images were recorded under transilluminator UV light. Based on the results of this study, microscopic examination showed that the total percentages of infection with E. histolytica were (15.1, 13.3 and 16.7) % in Karbala, Babel and Al- Najaf respectively. As infection rates are close in the three provinces and the absence of vast differences between these percentages, so the three provinces were treated as one (Middle Euphrates region). For the molecular diagnosis by using PCR technique the true prevalence of E. histolytica in Middle Euphrates region was found to be %12.8.