Host specificity, molecular phylogeny and morphological differences of Phyllodistomum pseudofolium Nybelin, 1926 and Phyllodistomum angulatum Linstow, 1907 (Trematoda: Gorgoderidae) with notes on Eurasian ruffe as final host for Phyllodistomum spp

Abstract

Background: Host-specificity patterns are not well-defined for trematodes of the genus Phyllodistomum Braun, 1899. The Eurasian ruffe, Gymnocephalus cernuus L., has been recorded as a definitive host for Phyllodistomum folium (Olfers, 1816), P. angulatum Linstow, 1907 and P. megalorchis Nybelin, 1926 and as the type-host for P. pseudofolium Nybelin (1926). A wide range of other host fishes have been recorded for these species as well. All present host records have been based on light microscopy and the life-cycles of P. pseudofolium, P. angulatum and P. megalorchis are unknown. The validity of P. pseudofolium and P. megalorchis require verification. In this study, rDNA sequences generated from adult Phyllodistomum spp., as well as from larval stages developing in Pisidium amnicum Müller, were analysed to establish the real number of Phyllodistomum species utilizing G. cernuus, and to associate larvae with the corresponding adult forms.

Results: Phylogenetic analyses of adult and larval stages of Phyllodistomum spp. based on ITS2 and partial 28S rDNA data allowed the confirmation of the validity of P. pseudofolium. A macrocercous cercaria, known as Phyllodistomum sp. from P. amnicum is genetically identical to adult P. pseudofolium. Phyllodistomum megalorchis obtained from its type-host, Lota lota L., showed no genetic differences from P. angulatum parasitizing Sander lucioperca L. In our analysis, P. pseudofolium, P. angulatum and P. macrocotyle formed a highly supported clade despite the fact that these species appear to be associated with distinct patterns of first intermediate host identity and cercarial morphology. Some morphological differences between gravid specimens of P. pseudofolium and P. angulatum were observed and their SEM tegumental surface topography is described.

Conclusions: The results lead us to the perception that macroevolutionary host switching in the genus Phyllodistomum is independent of host phylogeny. This study suggests strict host-specificity (oioxeny) for P. pseudofolium using one first intermediate host species (P. amnicum) and one definitive host species (G. cernuus). Phyllodistomum megalorchis is to be regarded as a synonym of P. angulatum. The close phylogenetic relatives, P. pseudofolium and P. angulatum, can be differentiated by morphological traits, the micromorphology and tegumental surface topography of these two species is intended to provide useful data for their identification and support the use of such features as a valuable taxonomic criterion. Molecular data showed that G. cernuus is a definitive host for two species: the oioxenous P. pseudofolium and the euryxenous P. folium.

Keywords: 28S; Eurasian ruffe; Host specificity; ITS2 rDNA; Life-cycles; Morphological variation; P. angulatum; Phyllodistomum pseudofolium; SEM.

Fig. 1

Phylogenetic tree based on Maximum Likelihood analysis of the ITS2 nuclear rDNA region. Bootstrap support values lower than 70% are not shown. GenBank accession numbers of the collapsed clades are provided in Table 1

Fig. 2

Phylogenetic tree based on Maximum Likelihood analysis of partial sequences of the 28S nuclear rDNA gene. Bootstrap support values lower than 70% are not shown. GenBank accession numbers of the collapsed clades are provided in Table 1

Fig. 3

Morphological variability in species of Phyllodistomum. a P. angulatum Linstow, 1907 ex Sander lucioperca, b P. angulatum ex Lota lota, c P. pseudofolium Nybelin, 1926 ex Gymnocephalus cernuus, arrows show contractions of hindbodies at the level of caecal terminations. Scale-bars: 500 μm

Fig. 4

SEM micrographs of the surface topography of Phyllodistomum angulatum. a, c Body of a mature worm, ventral view. b Oral sucker rim; note a constant pattern of 20 sensory papillae: four papillae localised on each side of the frontal pit; six papillae arranged in an outer ring; six papillae arranged in an inner ring and four papillae are situated within the sucker cavity (inset, papilla of oral sucker, scale-bar: 10 μm). d Four papillae within oral sucker cavity. e Evaginated cirrus; note smooth tegumental surface around and on evaginated cirrus. f Posterior notch, dorsal view. g Ventral sucker rim showing six large constant papillae and four small papillae; note negligible ‘acetabular fold’ (inset, papilla of the ventral sucker, scale-bar: 10 μm). Abbreviations: af, acetabular fold; ec, evaginated cirrus; fb, forebody; fp, frontal pit; gp, genital pore; hb, hindbody; ip, papilla of the inner ring; lp, lateral papilla surrounding frontal pit; op, papilla of the outer ring; os, oral sucker; p, papilla; pn, posterior notch; rc, radial corrugations of the sucker rim; sa, sucker aperture; sp, small papilla within sucker cavity; tr, transverse tegumental ridges; vlp, large papilla of the ventral sucker rim; vs, ventral sucker; vsp, small papilla of the ventral sucker rim

Fig. 5

SEM micrographs of the surface topography of Phyllodistomum pseudofolium. a Body of a mature worm, ventral view. b Oral sucker rim; note two papillae situated on each side of the frontal tubercle, six papillae arranged in an inner ring, two lateral pairs of papillae, and four papillae situated within the sucker cavity (inset, papilla and secretory pore close to the oral sucker). c Forebody, ventral view showing five pairs of papillae arranged symmetrically in two ventro-lateral rows. d Four symmetrically arranged papillae on the internal surface of the oral sucker. e Partly retracted ventral sucker showing four symmetrically arranged papillae within the sucker cavity; note rather negligible ‘acetabular fold’. f Part of the ventral surface of the forebody showing the presence on each side of both ventro-lateral and lateral rows of papillae. g Retracted ventral sucker showing the lack of papillae on the sucker rim and pronounced ‘acetabular fold’. h Posterior notch, dorsal view; note the irregular surface and lateral papillae. Abbreviations: af, acetabular fold; ft, frontal tubercle; gp, genital pore; hb, hindbody; ifr, inner papilla on the ventral surface of the forebody; ip, papilla of the inner ring of the oral sucker; lfr, papilla of the lateral row on the ventro-lateral margin of the forebody; lp, lateral papilla beside the frontal pit; op, papilla of the outer ring of oral sucker; os, oral sucker; p, papilla; pn, posterior notch; sa, sucker aperture; sp, secretory pore; tr, transverse tegumental ridges; vs, ventral sucker; vsr, ventral sucker rim