Nuclear ribosomal transcription units in Asian Paragonimus species (Paragonimidae: Platyhelminthes): structure, polymorphism, and implications for intersubordal phylogeny
Author:
Nguyen Khue Thi1, Doan Huong Thi Thanh1, Pham Khanh Linh Thi1, Roan Do Thi1, Agatsuma Takeshi2, Doanh Pham Ngoc3, Le Thanh Hoa1
Affiliation:
1. Immunology Department, Institute of Biotechnology (IBT), Vietnam Academy of Science and Technology (VAST), 18. Hoang Quoc Viet Rd, Cau Giay, Hanoi, Vietnam. 2. Department of Environmental Health Sciences, Kochi Medical School, Kohasu, Oko-Cho 185-1, Nankoku, Kochi 783-8505, Japan 3. Institute of Ecology and Biological Resources, Viet Nam Academy of Science and Technology, 18 Hoang Quoc Viet, Cau Giay, Hanoi, Vietnam
Abstract
Abstract
The complete coding sequences (from the 5’ terminus of 18S to the 3’ terminus of 28S rRNA genes) of ribosomal transcription units (designated as rTU*) of seven strains of five Asian Paragonimus species, excluding the unsequenced flanking ETS and IGS regions, were genetically characterized. The rTU* length was 7,661 bp for P. heterotremus (LC strain, Vietnam), 7,543 bp for P. iloktsuenensis (Amami strain, Japan), 6,932 bp for P. skrjabini miyazakii (OkuST1 strain, Japan), 7,422 bp for P. ohirai (Kino strain, Japan), and 8,616 bp (Megha strain, India), 7,292 bp (Bogil strain, South Korea), and 7,052 bp (QT2 strain, Vietnam) for P. westermani geographical samples, respectively. All Paragonimus strains' rRNA genes were equal in length: 1,977 bp for 18S, 160 bp for 5.8S, and 3,881 bp for 28S, except for P. s. miyazakii’s 18S rRNA gene, which was 1,974 bp. The ITS2 region in all Paragonimus species was 283 − 285 bp. The ITS1 lengths varied from 634 bp (P. s. miyazakii) to 2,313 bp (Indian P. westermani, Megha strain) due to the presence of tandem repeats, except P. s. miyazakii. The structural polymorphism in ITS1 is defined by the existence of 120 bp- or 121 bp-repeat units. The ITS1 and ITS2 sequences were capable of forming de novo “hairpin” (or stem-loop) and “loop” secondary structures, and the ITS2 shared a characteristic “four fingers” motif, which is likely conserved in trematodes. The base composition of A, T, G, and C, as well as skewness values of AT and GC content for 18S, 28S, 18S + 28S, and rTU*, indicated that T, A, and C were used equally frequently, with G being more common than C. These usages resulted in low negative AT-skew and low positive GC-skew. The PhyML software package was used to construct a comprehensive maximum-likelihood phylogeny from the alignment of 161 sequences of 28S D1 − D3 rDNA (1.1–1.3 kb) from 101 species of 62 genera and 23 families. Of these, 75 sequences came from 18 species of 6 genera from the suborder Troglotremata (previously, Troglotrematoidea). The phylogenetic tree clearly showed a well-supported phylogeny and clarified relationships within Paragonimidae as well as between families within the Troglotremata and Xiphidiata, Echinostomata, and Haplosplanchnata suborders, where Paragonimidae is monophyletic. There were two significant clusters: one was mixed-Paragonimus with a very tight group of P. heterotremus strains, and the other was P. westermani/siamensis, which was seperated into eight geographical/country strain groupings. Xiphidiata appears to be polyphyletic. The ribosomal datasets presented here will be valuable for taxonomic reappraisal as well as evolutionary and population genetics studies in the superfamily Troglotrematoidea (or suborder Troglotremata).
Publisher
Research Square Platform LLC
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