A molecular phylogeny of the genus Ichthyocotylurus (Digenea, Strigeidae)

Three nucleotide data sets, two nuclear (ribosomal internal transcribed spacer regions 1 and 2, ITS1 and ITS2) and one mitochondrial (cytochrome c oxidase subunit 1, CO1), were analysed using distance matrix and maximum likelihood methods to determine the inter-relationships amongst the four species attributed to the genus Ichthyocotylurus Odening, 1969. Sequence data obtained from all gene loci investigated supported the position of Ichthyocotylurus variegatus as a species discrete from Ichthyocotylurus platycephalus. Phylogenetic analyses yielded congruent trees, with I. variegatus isolates comprising a common clade to which I. platycephalus constitutes a sister taxon. Ichthyocotylurus erraticus and Ichthyocotylurus pileatus were found to demonstrate a similarly close inter-specific relationship. The greatest intra-generic divergence occurred in the CO1 region (16% variability), with resultant disparities in three to eight encoded amino acids. PCR amplification yielded multiple ITS1 products for all Ichthyocotylurus spp. Analyses of equivalent-sized amplicons showed 5.4% intra-generic variation and several point mutations between I. variegatus isolates from different geographical localities and from different piscine hosts. The ITS2 locus was extremely conserved, with less than 1% variation between species. No intra-specific variation was recorded for any CO1 or ITS2 sequences.     

Systematic analysis of the Diplostomidae and Strigeidae (Trematoda)

A systematic analysis of the genera in the Diplostomidae and Strigeidae was made using the Proterodiplostomidae as the outgroup. The Proterodiplostomidae was the family with the greatest preponderance of primitive characters and its monophyly was supported by the unique paraprostate gland. However, no character state supported the monophyly of the Diplostomidae sensu Dubois, 1970. That paraphyletic taxon was composed of 3 monophyletic groups: the pseudosuckerless Neodiplostomidae n. fam. had the most primitive character states of the 3 and its monophyly was based on characters in the neascus/neodiplostomulum metacercariae; the Bolbophoridae n. fam., with pseudosuckers, had its monophyly supported by characters present in the newly named prodiplostomulum metacercaria; and the emended Diplostomidae, also with pseudosuckers, had the most derived states and its monophyly was supported by characters present in the diplostomulum. The presence of pseudosuckers united the Bolbophoridae n. fam., the emended Diplostomidae, and the Strigeidae as a monophyletic assemblage. The Strigeidae had the most derived characters of these 3 taxa and its monophyly was supported by characters in the tetracotyle and the cup-shaped forebody and bilobed tribocytic organ of the adult. In general, the adult stages of these strigeoid families showed very conserved morphology and it was the metacercariae that possessed the innovations. The conserved adult morphology was typical of what one might expect if the intramolluscan stages were analyzed. Thus, the data were concordant with the view that the mollusc and vertebrate definitive host were the original hosts to the Digenea and that the second intermediate host and metacercaria were more recently intercalated. More specifically, the phylogeny of these groups suggested that ancestral bisegmented strigeoids originally infected reptiles, they subsequently radiated into birds with which they coevolved extensively, and on 5 separate occasions they radiated into mammals. The radiations into mammals were, by all available evidence, preceded by second intermediate host shifts from fish to amphibians.     

Chaetotaxy and surface structures of the miracidia of Ichthyocotylurus erraticus (Rudolphi, 1809), I. variegatus (Creplin, 1825) and Apatemon gracilis (Szidat, 1928) (Digenea,Strigeidae)

Silver nitrate staining was used to investigate the number and arrangement of argentophillic structures (epidermal plates, sensilla, excretory pores) of three species of strigeid miracidia, Ichthyocotylurus erraticus (Rudolphi, 1809), I. variegatus (Creplin, 1825) and Apatemon gracilis (Szidat, 1928). The epidermal plates of all three species of miracidia were arranged in four tiers according to the formula (6 + 9 + 4 + 3) = 22, while the number and distribution of sensilla were also found to be identical. Modifications to the nomenclature of Dimitrov et al. (1989) for miracidial chaetotaxy are proposed.Scanning electron microscopy (SEM) was also employed to examine the surface structures of deciliated miracidia of Ichthyocotylurus spp., confirming the position and revealing the form of body sensilla.     

Sensory apparatus of the cercaria of 2 species of the genus Apatemon (Trematoda: Strigeidae)

The sensory apparatus of cercariae of two species of trematodes (A. cobitidis and A. minor) is described. The species differ both morphologically and biologically. The only common feature in the sensory apparatus of the above species is the topography of sensillae in the posterior half of the lateral complex of the caudal trunk and of the furcal complex.     

Diplostomiasis (Trematoda: Strigeidae) in Arctic charr (Salvelinus alpinus) from Charr Lake, northern Labrador

The eyes of 273 Arctic charr (Salvelinus alpinus) taken from Charr Lake, the Ikarut River, and Hebron Fiord in northern Labrador were examined for diplostomula. All anadromous charr taken from the Ikarut river and Hebron Fiord were uninfected. The resident fish from Charr Lake were parasitized with large numbers of diplostomula and intensity of infection increased with age and length. Infections of up to 2,852 diplostomula in one fish are the highest recorded. Such infections did not appear to affect the vision of the charr.     

Observations on the in vitro culture of Cotylurus erraticus (Trematoda: Strigeidae)

Mtitchell J. S., Halton D. W. and Smyth J. D. 1978. Observations on the in vitro culture of Cotylurus erraticus (Trematoda: Strigeidae). International Journal for Parasitology8: 389–397. Cotylurus erraticus metacercariae obtained from around the heart of rainbow trout were excysted and grown in vitro and in vivo to egg-producing adults. For in vitro development, tissue culture media M199 or NCTC 135 was used, together with varying amounts of chicken serum. Worms grown in media containing the highest concentration of serum (80% per volume) showed the fastest rate of development, measured by the time taken for the first eggs to appear in the uterus. The testes, ovaries and vitellaria of these worms were comparable in structure and histochemistry with those of worms reared in gulls. Eggs were produced by worms in all media containing chicken serum, but the eggs had abnormal shells and failed to embryonate.     

Histopathology, ultrastructure and immunohistochemistry of Coregonus lavaretus hearts naturally infected with Ichthyocotylurus erraticus (Trematoda)

Histopathological, ultrastructural and immunohistochemical investigations were conducted on 26 specimens of powan Coregonus lavaretus (L.) from Loch Lomond (Scotland). The hearts of all 26 powan (15 females and 11 males) investigated harboured metacercariae of the digenean trematode Ichthyocotylurus erraticus (Rudolphi, 1809). The vast majority of metacercariae were located either singly or as an aggregation of white cysts on the surface of the bulbus arteriosus. The intensity of infection ranged from 2 to 200 larvae heart(-1), although the number of metacercariae found on male powan did not exceed 13. Histochemically, the parasite cyst wall gave a strong positive reaction with periodic acid schiff (PAS) and a faint positive signal with Azan-Mallory stain. All the metacercariae cysts were embedded in a granulomatous proliferation of heart epicardium tissue, forming a reactive fibroconnective capsule around the parasite. The capsule enclosing the parasite (produced by the host's reaction to the parasite) measured 13.57 to 90.20 microm (37.43 +/- 3.56) in thickness. Within the capsule wall, eosinophilic granular cells (EGCs), granulocytes, melanocytes and, in some instances, partially degenerated or vacuolated epithelioid cells were observed in close proximity to the cyst wall. Pigment-bearing macrophages were scattered throughout the granulomatous host-tissue reaction and as macrophage aggregates (MAs) within the capsules surrounding parasites. Immunohistochemical tests were applied to infected heart sections using 12 different antisera. Nerve fibres immunoreactive to bombesin, substance P (SP), and atrial natriuretic peptide (ANP) antisera were observed in close proximity to the parasite larvae. The presence of a serotonin-like substance was also observed within host immune-cells surrounding trematode cysts. Large cells of the epicardium were found to be immunoreactive to met-enkephalin and vasoactive intestinal peptide (VIP) antisera but not immunoreactive to anti-protein gene-product 9.5 (PGP9.5) sera.     

Morphological and molecular differentiation of Parastrigea (Trematoda: Strigeidae) from Mexico,with the description of a new species

Parastrigea plataleae n. sp. (Digenea: Strigeidae) is described from the intestine of the roseate spoonbill Platalea ajaja (Threskiornithidae) from four localities on the Pacific coast of Mexico. The new species is mainly distinguished from the other 18 described species of Parastrigea based on the ratio of its hindbody length to forebody length. A principal component analysis (PCA) of 16 morphometric traits for 15 specimens of P. plataleae n. sp., five of Parastrigea cincta and 11 of Parastrigea diovadena previously recorded in Mexico, clearly shows three clusters, which correspond to the three species. DNA sequences of the internal transcribed spacers (ITSs) of ribosomal DNA and the mitochondrial gene cytochrome c oxidase subunit I (cox 1) were used to corroborate this morphological distinction. The genetic divergence estimated among P. plataleae n. sp., P. cincta and P. diovadena ranged from 0.5 to 1.48% for ITSs and from 9.31 to 11.47% for cox 1. Maximum parsimony (MP) and maximum likelihood (ML) analyses were performed on the combined datasets (ITSs + cox 1) and on each dataset alone. All of the phylogenetic analyses indicated that the specimens from the roseate spoonbill represent a clade with strong bootstrap support. The morphological evidence and the genetic divergence in combination with the reciprocal monophyly in all of the phylogenetic trees support the hypothesis that the digeneans found in the intestines of roseate spoonbills represent a new species.     

Karyological studies on three strigeid digeneans: Ichthyocotylurus erraticus (Rudolphi, 1809), I. variegatus (Creplin, 1825) and Apatemon gracilis (Rudolphi, 1819)

Previous work has shown that karyology may be a valuable technique for discriminating morphologically similar species. In the present study, chromosome number and morphology were described for three strigeid species, Ichthyocotylurus erraticus (Rudolphi, 1809), I. variegatus (Creplin, 1825) and Apatemon gracilis (Rudolphi, 1819). The results show that these species possess the same diploid number of chromosomes (2n = 20) in somatic cells. The relative length and the position of the centromere were determined, and significant inter-specific differences in the karyotypes are demonstrated, enabling the ready discrimination of I. erraticus and I. variegatus cercariae. A review of known karyotypes of the family Strigeidae suggests that possible routes of evolutionary changes involve pericentric inversions within chromosomes rather than Robertsonian translocations between elements.