Multiplex-Touchdown PCR to Simultaneously Detect Cryptosporidium parvum, Giardia lamblia, and Cyclospora cayetanensis, the Major Causes of Traveler's Diarrhea.
10.3347/kjp.2016.54.5.631
- Author:
Ji Hun SHIN
1
;
Sang Eun LEE
;
Tong Soo KIM
;
Da Won MA
;
Jong Yil CHAI
;
Eun Hee SHIN
Author Information
1. Department of Parasitology and Tropical Medicine, Seoul National University College of Medicine, and Institute of Endemic Diseases, Seoul National University Medical Research Center, Seoul 03080, Korea. ehshin@snu.ac.kr
- Publication Type:Original Article
- Keywords:
Cryptosporidium parvum;
Giardia lamblia;
Cyclospora cayetanensis;
multiplex PCR;
touchdown PCR;
stool sample
- MeSH:
Cryptosporidium parvum*;
Cryptosporidium*;
Cyclospora*;
Diarrhea*;
Genes, rRNA;
Giardia lamblia*;
Giardia*;
Glutamate Dehydrogenase;
Humans;
Methods;
Multiplex Polymerase Chain Reaction;
Oocysts;
Parasites;
Polymerase Chain Reaction*;
RNA, Ribosomal, 18S
- From:The Korean Journal of Parasitology
2016;54(5):631-636
- CountryRepublic of Korea
- Language:English
-
Abstract:
This study aimed to develop a multiplex-touchdown PCR method to simultaneously detect 3 species of protozoan parasites, i.e., Cryptosporidium parvum, Giardia lamblia, and Cyclospora cayetanensis, the major causes of traveler’s diarrhea and are resistant to standard antimicrobial treatments. The target genes included the Cryptosporidium oocyst wall protein for C. parvum, Glutamate dehydrogenase for G. lamblia, and 18S ribosomal RNA (18S rRNA) for C. cayetanensis. The sizes of the amplified fragments were 555, 188, and 400 bps, respectively. The multiplex-touchdown PCR protocol using a primer mixture simultaneously detected protozoa in human stools, and the amplified gene was detected in >1×10³ oocysts for C. parvum, >1×10⁴ cysts for G. lamblia, and >1 copy of the 18S rRNA gene for C. cayetanensis. Taken together, our protocol convincingly demonstrated the ability to simultaneously detect C. parvum, G. lamblia, and C. cayetanenesis in stool samples.
