Discrimination of the ITS1 types of Fasciola spp. based on a PCR-RFLP method.
Molecular characterization is important for discriminating Fasciola specimens having the deoxyribonucleic acid (DNA) sequences of Fasciola hepatica, Fasciola gigantica, and both Fasciola species, since three Fasciola forms coexist in Asian countries. We have developed a restriction fragment length polymorphism of amplified DNA (PCR-RFLP) of the nuclear ribosomal internal transcribed spacer 1 (ITS1) region in Fasciola species. The band patterns of the fragments digested with a restriction enzyme, Rsa I, were accurately distinguished among the three forms of Fasciola. Amplicons with the sequences of F. hepatica and F. gigantica were divided into fragments of about 360, 100, and 60 bp, and 360, 170, and 60 bp, respectively, and amplicons with the sequences of both Fasciola species yielded fragments of 360, 170, 100, and 60 bp. The results of PCR-RFLP completely coincided with those of sequence analysis, and thus PCR-RFLP is a useful technique for determining the ITS1 type in Fasciola species.
Ichikawa
Madoka
M
Laboratory of Veterinary Parasitology, Faculty of Agriculture, Iwate University, Ueda 3-18-8, Morioka 020-8550, Japan.
Itagaki
Tadashi
T
D000818:Animals; D001208:Asia; D001483:Base Sequence; D016172:DNA Fingerprinting / Q000379:methods*; D018509:DNA, Helminth / Q000737:chemistry / Q000235:genetics*; D021903:DNA, Ribosomal Spacer / Q000737:chemistry / Q000235:genetics*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D005209:Fasciola / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D005211:Fascioliasis / Q000469:parasitology; D005838:Genotype; D006801:Humans; D008969:Molecular Sequence Data; D010274:Parasitology / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D018509:DNA, Helminth; D021903:DNA, Ribosomal Spacer; D015252:Deoxyribonucleases, Type II Site-Specific; C040158:GTAC-specific type II deoxyribonucleases