The only observed morphological difference between Spirometra erinacei and S. mansonoides is the uterine shape of the mature proglottid. Two species of worms are thought to be evolutionarily closely related. Biomolecular comparison of the two worms by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis was conducted to observe the genetic distance. The 28s rDNA, mitochondrial cytochrome c oxidase subunit I (mCOI), and ribosomal internal transcribed spacer 1 (ITS1) fragments were obtained from the worms by PCR. The PCR products were cleaved by 5 four-base pair restriction enzyme combinations (Msp I, Hae III, Alu I, Cfo I, Rsa I), electrophoresed and analyzed with PAUP 3.1.1. The fragment patterns of 28S rDNA and ITS1 demonstrated that two worms were in identical systematic tree with bootstrap number 94 and 100, respectively. As for mCOI, bootstrap number was 74 in a different tree. Above results are indicative of recent common ancestry between S. erinacei and S. mansonoides.
Fig. 1 Two percent agarose gel electrophoresis of PCR-RFLP for 28S rDNA D1 using the restriction endonuclease. Lane M, molecular weight markers; lane 1, S. erinacei; lane 2, S. mansonoides; lane 3, T. asiatica; lane 4, T. taeniaeformis; lane 5, H. diminuta; lane 6, D. latum.
Fig. 2 Two percent agarose gel electrophoresis of PCR-RFLP for mCOI using the restriction endonuclease. Lane M, molecular weight markers; lane 1, S. erinacei; lane 2, S. mansonoides; lane 3, T. asiatica; lane 4, T. taeniaeformis; lane 5, H. diminuta; lane 6, D. latum.
Fig. 3 Two percent agarose gel electrophoresis of PCR-RFLP for rDNA ITS1 using the restriction endonuclease. Lane M, molecular weight markers; lane 1, S. erinacei; lane 2, S. mansonoides; lane 3, T. asiatica; lane 4, T. taeniaeformis; lane 5, H. diminuta; lane 6, D. latum.
Fig. 4 Molecular phylogenetic analyses of PCR-RFLP obtained using 28S rDNA, mCOI and the rDNA ITS1 gene. Branch lengths and ranges of bootstrap values were obtained using statistical package (Power Macintosh 6100/66, MacClade V 3.0., Clustal V and PAUP Ver.3.1.1). Bootstrap values are given as a percent of the above corresponding branch. A, 28S rDNA; mCOI; C, rDNA ITS1 gene.
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