title: Multiplex-Touchdown PCR to Simultaneously DetectCryptosporidium parvum,Giardia lamblia, andCyclospora cayetanensis, the Major Causes of Traveler's Diarrhea.
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, andCyclospora cayetanensis, the major causes of traveler's diarrhea and are resistant to standard antimicrobial treatments. The target genes included theCryptosporidiumoocyst wall protein forC. parvum, Glutamate dehydrogenase forG. lamblia, and 18S ribosomal RNA (18S rRNA) forC. 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 >1×103oocysts forC. parvum, >1×104cysts forG. lamblia, and >1 copy of the 18S rRNA gene forC. cayetanensis. Taken together, our protocol convincingly demonstrated the ability to simultaneously detectC. parvum,G. lamblia, andC. cayetanenesisin stool samples.
authors.affiliation: 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.
authors.forename: Ji-Hun
authors.initials: JH
authors.lastname: Shin
authors.affiliation: Division of Malaria and Parasitic Diseases, Korea National Research Institute of Health, Korea Center for Disease Control and Prevention, Cheongju 28159, Korea.
authors.forename: Sang-Eun
authors.initials: SE
authors.lastname: Lee
authors.affiliation: Department of Tropical Medicine and Inha Research Institute for Medical Sciences, Inha University School of Medicine, Incheon 22212, Korea.
authors.forename: Tong Soo
authors.initials: TS
authors.lastname: Kim
authors.affiliation: Division of Malaria and Parasitic Diseases, Korea National Research Institute of Health, Korea Center for Disease Control and Prevention, Cheongju 28159, Korea.
authors.forename: Da-Won
authors.initials: DW
authors.lastname: Ma
authors.affiliation: 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.|Institute of Parasitic Diseases, Korea Association of Health Promotion, Seoul 07653, Korea.
authors.forename: Jong-Yil
authors.initials: JY
authors.lastname: Chai
authors.affiliation: 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.|Seoul National University Bundang Hospital, Seongnam 13620, Korea.
authors.forename: Eun-Hee
authors.initials: EH
authors.lastname: Shin
mesh_terms: D016785:Cryptosporidium parvum / Q000235:genetics / Q000302:isolation & purification*; D021744:Cyclospora / Q000235:genetics / Q000302:isolation & purification*; D017931:DNA Primers / Q000235:genetics; D003967:Diarrhea / Q000175:diagnosis* / Q000469:parasitology; D005243:Feces / Q000469:parasitology*; D016829:Giardia lamblia / Q000235:genetics / Q000302:isolation & purification*; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D015800:Protozoan Proteins / Q000235:genetics; D012337:RNA, Ribosomal, 18S / Q000235:genetics; D014195:Travel
chemical_list: D017931:DNA Primers; D015800:Protozoan Proteins; D012337:RNA, Ribosomal, 18S
keywords: Cryptosporidium parvum; Cyclospora cayetanensis; Giardia lamblia; multiplex PCR; stool sample; touchdown PCR