An alternative DNA extraction method for detection of Blastocystis spp. in human fecal samples

Abstract

Polymerase chain reaction (PCR) is an effective technique for diagnosis of Blastocystis infection. Notably, DNA isolation procedure is extremely critical for the PCR step. In the present study, a recently described extraction procedure, named as the "sand method" was modified and adapted for isolation of Blastocystis DNA. To evaluate its efficacy, the current method and QIAamp DNA Stool Mini Kit (Qiagen) were applied to fresh human stool samples. Our results indicated that, the mean DNA concentrations obtained by the sand method and the commercial kit were 48 and 55 ng/mu l, respectively. Also, no DNA inhibitors were detected in two methods. The sand method was capable of detecting 16 parasites per 50 mg feces. DNA samples extracted by both methods were subjected to PCR. Blastocystis spp. were detected in 11 (31.4%) of 35 samples, and perfect agreement (kappa: 1.000) was found between the PCR-sand method and PCR-commercial kit method. The samples that were detected positive by PCR-sand method were successfully sequenced, and Blastocystis subtypes (STs) were identified as ST3, ST2 and ST1. In conclusion, the present study indicates that the sand method provides a simple, rapid and inexpensive procedure for reliable extraction of Blastocystis DNA from stool samples. (C) 2018 Elsevier Inc. All rights reserved.