التحري عن الاميبا الحالة للنسج/ الاميبا المتغايرة Entamoeba histolytica/Entamoeba dispar في عينات البراز للمرضى با ستعمال تقنية PCR == Detection of Entamoeba Histolytic /Entamoeba Dispar In Stool Specimens By Using PCR Technique
Author name:
بثينة عبد الحمزة حسون الزبيدي
Supervisor name:
اخلاص مشرف عيدان
General topic:
Biology
Specific topic:
Microbiology - Parasites
Degree:
Master
University:
University of Baghdad
Language:
Arabic
University location:
Baghdad
First pages:
24T2904 - p.pdf
Abstract:
تسلط الدراسة الحالية الضوء على تشخيص وتحديد نسبة الاصابة بطفيلي الاميبا المرضية المتمثلة بالاميبا الحالة للنسج Entamoeba histolytica وتمييزها عن الاميبا غير المرضية المتمثلة بالاميبا المتغايرة Entamoeba dispar اللتين تتمايزان بشكلهما المتماثل في كلا الطور | This study sheds light on the determination and diagnose of the percentage of infection of like Entamoeba histolytica and differentiate it from the non pathogenic Entamoeba dispar that were morphologically identical in both cysts and trophozoite phase in two different groups : the first group includes 50 stool samples of patients suffering from diarrhea and abdominal pain (symptomatic group) while the second group includes 95 stool samples of patients not suffering from diarrhea and abdominal pain (Asymptomatic group).In addition to that, 20 stool samples were collected from healthy individuals as a control these samples were collected patients of AL - Zafaranyia General Hospital and AL - Alwyia Childhood Teaching Hospital in Baghdad from July, 2011 to May, 2012.These samples belong to different ages (1 year - < 50 years) of different sexes (males and females). Entamoeba samples were diagnosed by using two methods : the first one is the wet mount as a General Stool Examination(GSE) after macroscopic examination for sample and classifying them according to their consistency (solid, semisolid and liquid) and color (brown and yellow) and presence of blood and mucous in addition to diagnosing them microscopically to detect the presence of pus cells and both phases trophozoite and cyst for Entamoeba histolytica.While the second method that was depended in diagnosis is duplex polymerase chain reaction (dPCR) by using two pairs of genes for detection of their amplification in DNA sample isolated from stool. The first pair is cystein proteinase which represents one of virulent factors in Entamoeba histolytica parasite as a tissue invader. The second pair encodes the surface antigen present on the surfaces of both parasites E. histolytica and E. dispar. It is represented by Actin gene (Act). The results of the wet mount showed that in percentage of infection of both species (E.histolytica/E.dispar) the (54%) of patient symptom and (34.37%) patient a symptom.While results of the specificity and sensitivity showed that in positive sample wet mount negative PCR while negative sample wet mount showed that positive PCR and higher percentage of infection the Entamoeba dispar. While the results of the dPCR showed that in spite of the presence of both species Entamoeba pathogenic (Entamoeba histolytica) and non pathogenic Entamoeba (Entamoeba dispar) , the percentage of infection of Entamoeba histolytica was lower than that of Entamoeba dispar. This was clear by the detection of amplification the Reaction producets (dPCR) by using both genes : Actin &Ehcp5 where only 8 samples were detected and they were found to be positive amplification Ehcp5 and 31 samples were positive amplification in both groups of patients (symptomatic and a symptomatic). The percentage of infection in females was higher than that in males for all ages when using both tests (wet mount & PCR). More over the patients with less than (10) years old had higher percentage of infection in both sexes.In conclusion, it should not be depended on direct wet mount technique for the identification of infected parasite Entamoeba histolytica and differentiating it from Entamoeba dispar because of their high homology. That the molecular examination is the only method that is able to determine the kind of infection with parasite Entamoeba
