A New Duplex Reverse Transcription PCR for Simultaneous Detection of Viable Cryptosporidium parvum Oocysts and Giardia duodenalis Cysts

Key words: 
• Cryptosporidium parvum /
• Giardia duodenalis /
• Heat-shock /
• RT-PCR

Abstract: Objective To simultaneously detect viable Cryptosporidium parvum oocysts and Giardia duodenalis cysts for the purpose of reducing time and cost spent. Methods A duplex reverse transcription polymerase chain reaction (RT-PCR) method was newly developed. Results Using duplex RT-PCR method for the hsp70 gene, viable (oo)cyst concentrations of 101 and 103 (oo)cysts/100 μL could be detected for C. Parvum and G duodenalis, respectively. However, after heat-shock stimulation the expression of hsp70 mRNAs was detectable at 100 and 101 (oo)cysts/100 μL concentrations of C parvum and G duodenalis, respectively. Thus, the detection sensitivity was significantly increased when the viable (oo)cysts were exposed to heat shock. Conclusion This study describes a new duplex RT-PCR method for hsp70 gene to detect the viable (oo)cysts of the C. Parvum and G duodenalis with less time consumed and at a lower cost. This newly developed duplex RT-PCR method may be used to detect these parasites not only in aquatic environments but also in clinical samples.