Investigation of Cryptosporidium spp. Positivity in Formol-Fixed Stool Samples by Polymerase Chain Reaction Method

Abstract

Objective: It was aimed to determine Cryptosporidium spp., which is thought to cause digestive system complaints in primary school children, by Modified Acid-Fast (MAF) staining method and Polymerase Chain Reaction (PCR). Method: One hundred sixty four stool samples taken from children were examined under light microscopy with MAF. Positive samples were fixed in 10% formalin and stored at + 4 °C until analysis. These samples were then purified using the Sucrose Gradient method and, DNA isolation was performed using the modified QIAamp DNA Mini Kit (Qiagen) protocol. In order to increase sensitivity of the study, SSU rRNA gene region was amplified by PCR method. Results: Microscopic examination showed that 89 of the 164 stool samples (54,26%) were found to be positive for Cryptosporidium spp.oocysts. Eighty nine samples which were found to be positive by microscopic examination and fixed in %10 formalin were studied by PCR and 74 (45,12%) of them were positive. Conclusions: Cryptosporidiosis, which is known to cause severe infections, especially in immunocompromised individuals and children, is transmitted by fecal-oral route from infected humans and animals, and contaminated water and nutrients. Cryptosporidiosis, which is the result of fecal pollution, causes a great risk for human health even with a low number of oocysts due to the fact that oocysts are resistant to chlorine and they can survive for a long time in the outer environment. It was observed that, primary school children living in area of the study were under risk for cryptosporidiosis.