Spirorchiid LSR and ITS2 alignments, de Buron et al. 2018
Description
Life cycles of spirorchiids that infect the vascular system of turtles are poorly understood. Few life cycles of these blood flukes have been elucidated and all intermediate hosts reported are gastropods (Mollusca), regardless of whether the definitive host is a freshwater or a marine turtle. During a recent survey of blood fluke larvae in polychaetes on the coast of South Carolina USA, spirorchiid-like cercariae were found to infect the polychaetes Amphitrite ornata (Terebellidae) and Enoplobranchus sanguineus (Polycirridae). Partial sequences of D1-D2 domains of the large ribosomal subunit (LSR) and the internal transcribed spacer 2 (ITS2) genes allowed the identification of sporocysts and cercariae as belonging to two unidentified Neospirorchis species reported from green turtle, Chelonia mydas, in Florida: Neospirorchis sp. (Neogen 13) in A. ornata and Neospirorchis sp. (Neogen 14) in E. sanguineus. Phylogenetic analysis suggests that infection of annelids by blood flukes evolved separately in aporocotylids and spirorchiids. Our results support the contention that Spirorchiidae is not a valid family and suggest that Neospirorchis belongs to a distinct family. Since specificity of spirorchiids for their intermediate hosts is broader than it was thus far assumed, surveys of annelids in turtle habitats are necessary to further our understanding of the life history of these pathogenic parasites. (de Buron et al. 2018, International Journal of Parasitology)
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Select representative spirorchiid, aporocotylid and schistosomatid blood fluke sequences available from GenBank were aligned with the Neogen13-SC and Neogen14-SC sequences obtained in this study using MUSCLE (v. 3.8) (Robert, 2004) and edited using Jalview (v 2.8) (Waterhouse et al., 2009). Uninformative alignment positions were discarded using Gblocks (Talavera et al., 2007) with relaxed parameters allowing gaps in up to half of the sequences, a minimum conserved-block size of 5 positions, and less-strict flanking positions. The LSR and ITS2 alignments comprised 38 sequences aligned over 752 positions and 22 sequences aligned over 302 positions, respectively. Modelgenerator (v. 0.85.1) (Keane et al., 2006) was used to determine appropriate substitution models, and Bayesian inference was performed with MrBayes (v. 3.2.6) (Ronquist et al., 2012) using a GTR + I + G substitution model. The LSR analyses were run over 10,000,000 generations, with trees sampled at every 1000 generations. ITS2 analyses were run over 2,000,000 generations, with trees sampled at every 100. The first 25% of trees for each analysis were discarded as burn-in. Fifty-percent majority-rules consensus trees were obtained from the remaining trees and visualized using FigTree (v. 1.4.2; http://tree.bio.ed.ac.uk/software/figtree/). Following Orelis-Ribeiro et al. (2014), diplostomoid species Alaria alata, Cardiocephaloides longicollis, Ichthyocotylurus erraticus, and Hysteromorpha triloba served as outgroups for the LSR tree, which only included blood fluke species with a known intermediate host. A few exceptions were made when addition of species (without known intermediate hosts) rendered weak node support considerably stronger.