Diagnosis of Blastocystis Sp., Cryptosporidium Sp., and Giardia Intestinalis by Multiplex PCR: an Optimization Study

Objective: It was aimed to develop a new Multiplex Polymerase Chain Reaction (PCR) protocol with isolates obtained from local patients for the diagnosis of Blastocystis sp., Cryptosporidium sp. and Giardia intestinalis, which can cause severe gastrointestinal system complaints especially in immunocompromised patients and children. Method: DNA isolation was performed with a commercial kit from three stool samples of different patients whose microscopic examination showed dense amounts of Blastocystis sp., Cryptosporidium sp. and Giardia intestinalis. First, a special PCR protocol has been developed for each protozoon. Then, the multiplex PCR protocol, in which these three protozoa can be diagnosed together, was optimized. Results: In the multiplex PCR protocol performed after DNA isolation, bands of 95 bp., 227 bp. and 258 bp. were obtained for Cryptosporidium sp., Blastocystis sp. and G. intestinalis, respectively. Conclusion: Blastocystis sp., Cryptosporidium sp. and Giardia intestinalis were diagnosed by multiplex PCR with the original protocol developed. Due to the difficulties in using different methods in parasitological examination, by adding other protozoa important for public health to this optimized protocol, it will be possible to detect a large number of parasites with a single molecular method.