Cytomorphological characteristics of Polypodium hydriforme and problems of myxozoan and cnidarian phylogeny

The present review analyses cytomorphological characters of the parasitic cnidarian Polypodium hydriforme, discriminating between those of bilateral (triploblastic) animals, common characters shared with the Myxozoa, and the unique characters of this species. Phylogenetic position of the group of parasitic cnidarians and of the class Polypodiozoa is discussed. A conclusion is made that the cytomorphological characters as well as 18S rDNA analysis of P. hydriforme and Myxozoa justify establishment of a new taxonomic group (a clade) of parasitic cnidarians (Endocnidozoa) uniting Polypodiozoa and Myxozoa (Zrzavy, Hypsa, 2003). The unique characters of P. hydriforme suggest that the phylum Cnidaria is more diverse than commonly supposed, and that P. hydriforme is not an aberrant cnidarian species but a relic organism, which might originally belong to the cnidarian class Polypodiozoa, which underwent reduction in the course of adaptation to parasitism.     

The joint evolution of the Myxozoa and their alternate hosts: A cnidarian recipe for success and vast biodiversity

The relationships between parasites and their hosts are intimate, dynamic and complex; the evolution of one is inevitably linked to the other. Despite multiple origins of parasitism in the Cnidaria, only parasites belonging to the Myxozoa are characterized by a complex life cycle, alternating between fish and invertebrate hosts, as well as by high species diversity. This inspired us to examine the history of adaptive radiations in myxozoans and their hosts by determining the degree of congruence between their phylogenies and by timing the emergence of myxozoan lineages in relation to their hosts. Recent genomic analyses suggested a common origin of Polypodium hydriforme, a cnidarian parasite of acipenseriform fishes, and the Myxozoa, and proposed fish as original hosts for both sister lineages. We demonstrate that the Myxozoa emerged long before fish populated Earth and that phylogenetic congruence with their invertebrate hosts is evident down to the most basal branches of the tree, indicating bryozoans and annelids as original hosts and challenging previous evolutionary hypotheses. We provide evidence that, following invertebrate invasion, fish hosts were acquired multiple times, leading to parallel cospeciation patterns in all major phylogenetic lineages. We identify the acquisition of vertebrate hosts that facilitate alternative transmission and dispersion strategies as reason for the distinct success of the Myxozoa, and identify massive host specification‐linked parasite diversification events. The results of this study transform our understanding of the origins and evolution of parasitism in the most basal metazoan parasites known.     

Molecular evidence of regression in evolution of metazoa

Molecular data permit to construct phylogenetic trees independently of morphological characters. It allows to consider their evolution without the frames of a priori hypothesis of regularities of morphological evolution and independently of palaeontological data. Cladistic analysis of elements of secondary structure of varible areas V7 and V2 in 18S rRNA with different Protozoa as "external" groups shows that Bilateria + Cnidaria are monophyletic, Ctenophora and Porifera are early derivatives of Metazoa, Trichoplax (Placozoa) is a form related to Cnidaria, while Rhombozoa, Orthonectida and Myxozoa were branched within Bilateria. Morphological reduction with losses of any organs and tissues took place many times in early evolution of Metazoa and Bilateria not only in parasitic species. It occurred both at early and late stages of embryonic development and differentiation. Two alternative scenario of morphological degeneration in Trichoplax and the way of their testing are suggested. The similarity of Ctenophora and Calcarea is discussed. Meridional or oblique position of the third cleavage furrow of ovule can be considered as an evidence of their origin from common ancestor.     

Cytological paradoxes in the developmental cycle of the coelenterate Polypodium hydriforme--an intracellular parasite from the oocytes of acipenserid fishes

Successive stages of the embryonic development of Polypodium hydriforme, occurring at the parasitic phase of its life cycle, are considered. The development of a new parasitic generation starts without fertilization, i. e. parthenogenetically. The embryo develops from aberrant binucleate gametes formed in the result of meiosis within entodermal gonads of free-living animals. This type of gametogenesis, earlier considered as spermatogenesis (Raikova, 1961), is now interpreted as oogenesis. A conclusion is drawn about a change of the sexual orientation of the male gonad which becomes a female one in the course of evolution of Polypodium. As to the gonads of free-living animals, which were formerly interpreted as female ones, they seem to be abortive rudimentary organs since they produce no mature sex cells. A long-lasting block of cytokinesis of the 2nd meiotic division, as well as utilization of the polar body of this division as a phorocyte and, later, as a trophamnion, are important adaptations of Polypodium to parasitism. It is the larger nucleus with a voluminous cytoplasm, rather than the smaller nucleus, that becomes here the 2nd polar body. Polypodium differs from other coelenterates by the presence of highly polyploid feeding cells at both the parasitic (the trophamnion, 500 c) and free-living phases of the life cycle (trophocytes in the rudimentary female gonad, 8c-32c).     

The nervous system of parasitic cnidarian Polypodium hydriforme

The nervous system of intracellular parasitic cnidarian Polypodium hydriforme at various stages of its life cycle has been studied by the immunocytochemical method using antibodies to FMRF-amide and by electron microscopy. Neurosecretory, sensory, and ganglion cells have been identified both at the parasitic stage (planula and stolon stages, when body layers are inverted) and in free-living animals. These cells are characterized by the presence of round neurosecretory granules about 80–120 nm in diameter. Gap junctions have been detected between nerve cells. Most of the neurosecretory and sensory cells have been observed in the epidermis of sensory tentacles of free-living animals. Sensory cells possess immobile flagella. The chains of ganglion cells are located under the epidermis and penetrate mesoglea. A centriole encircled by a fragment of nuclear envelope, which is a marker of ectodermal lineage cells in Polypodium, has been described in the cytoplasm of the sensory cells, thus proving the ectodermal nature of the nervous system. Like in most cnidarians, the nervous system of Polypodium hydriforme is a network containing FMRF-amide-like neuropeptides. Neither sense organs, nor ring-shaped nerve concentrations have been observed.     

The parasitic coelenterate, Polypodium hydriforme USSOV, from the eggs of the American acipenseriform Polyodon spathula.

No significant differences in macro- and micromorphology were found between the parasitic stolon and free-living polyps of Polypodium sp. obtained from infected eggs of the North American acipenseriform fish Polyodon spathula and corresponding developmental stages of Polypodium hydriforme Ussov, parasitic in the Volga sterlet (Acipenser ruthenus). Therefore, both the American and the European forms of Polypodium belong to the species P. hydriforme Ussov.     

Long-Branch Abstractions

Recent attention has been focused on the sensitivities of various tree reconstructing algorithms to sequence rate heterogeneity (long-branch attraction). Phylogenetic conclusions from two recent empirical studies have been indicted as artifacts attributable to long-branch attraction. Siddall et al. (1995) concluded that Myxozoa are cnidarians and sister group to Polypodium based on 18S rDNA and morphology. Hanelt et al. (1996) argued that this result is due to long-branch attraction. Whiting et al. (1997) concluded that the Strepsiptera are sister group to Diptera based on parsimony analysis of 18S rDNA, 28S rDNA, and morphology. Huelsenbeck (1997) argued that this result also is attributable to long-branch attraction. We demonstrate that the analyses and arguments dismissing these results as the effects of long-branch attraction are fundamentally flawed. The criteria employed by these authors were applied arbitrarily by them to the groups that they did not want, and yet using those same criteria, there is more reason to exclude other taxa besides Polypodium and there is more reason to disbelieve monophyly of Diptera than monophyly of Strepsiptera with Diptera. Moreover, it is asserted, long-branch attraction cannot explain the presence of nematocysts in Myxozoa and halteres in Strepsiptera. For these reasons, and in light of the demonstration that long branches cannot attract each other in their mutual absence, we conclude that the monophyly of Myxozoa + Polypodium and Strepsiptera + Diptera is not due to long-branch attraction. We suggest that maximum likelihood methods are extremely sensitive to taxon and character sampling and that these data sets are demonstrative of the long-branch repulsion problem.     

Immunohistochemical studies of GLWamides in Cnidaria

GLWamides are a recently described, novel family of neuropeptides in Cnidaria. Antibodies specific for the GLWamide terminus have been raised and used to evaluate the occurrence and localisation of immunopositive material in various Cnidaria in order to determine whether GLWamides are present and to obtain a first impression of the possible regulatory role of these neuropeptides. GLWamide immunoreactivity has been found in all species tested and is not confined to distinct life stages but is present during most of the life cycle of the Cnidaria. Additionally, GLWamides are expressed by different nerve cells at different life stages. GLWamide-immunoreactive cells constitute a subset of the neural equipment. Overall our data suggest that GLWamides generally occur in the nervous system of Cnidaria and that these peptides are multifunctional. Putative functions other than the control of development include the regulation of nematocyst discharge, muscle contraction and the regulation of gastric function.     

Phylogenetic comparison of the myxosporea based on an actin cDNA isolated from Myxobolus cerebralis

The full-length actin gene from Myxobolus cerebralis (McerAct-1), the first characterized from representatives in the phylum Myxozoa, encodes a 378-amino acid polypeptide with an estimated molecular weight of 41,580-Da. A phylogenetic comparison found M. cerebralis to branch outside the metazoans. This finding contrasts with previous reports that suggest an evolutionary affinity of the Myxozoa with either the Bilateria or Cnidaria.     

Peculiarities of the embryonic development of Polypodium hydriforme Ussov (Coelenterata), a parasite of acipenserid oocytes

"Unicellular" stages (107 specimens) and multicellular stages (64 specimens) of embryogenesis of Polypodium, found in 14 sterlet (Acipenser ruthenus L.) females, have been studied with light microscopy, cytophotometry, and autoradiography following incubation with 3H-uridine. All stages of the embryonic development occur inside host oocytes. The "unicellular" stage includes a binucleate cell with unequally sized nuclei; separation inside it of a small cell around the smaller nucleus, i.e. transformation of the single cell into a complex of 2 cells, the larger one enveloping the smaller; formation of a cavity inside the nucleus of the large (outer) cell, and migration of the small cell into it, and "cell-in-a-cell" stage, the small (generative) cell being inside the cavity formed by the nucleus of the large (trophic) cell. The latter gives rise to a hypertrophied but still unicellular envelope around the embryo, the trophamnion. The multicellular stages start with segmentation of the generative cell into blastomeres. These form a morula lying inside the cavity of the trophamnion. Gastrulation occurs by morular delamination. The inversion of the germ layers, typical of parasitic Polypodium stages, apparently arises during gastrulation. Both the generative cell ("egg") and the blastomeres are haploid, at least until the morula stage. The eggs of Polypodium are the smallest ones among coelenterates; they lack yolk and develop without fertilization. Diploidy seems to be restored during segmentation. The trophamnion cell grows, its nucleus becomes highly polypoid, and its cytoplasm accumulates mucoprotein inclusions. Both the blastomere nuclei and the trophamnion nucleus have large nucleoli and actively synthesize RNA. The stages of embryogenesis of Polypodium closely correspond to stages of the host oogenesis. The embryonic development of Polypodium lasts several years and is the slowest among coelenterates. However, it has some features typical of the class Hydrozoa.     

Sacculogenesis of Buddenbrockia plumatellae (Myxozoa) within the invertebrate host Plumatella repens (Bryozoa) with comments on the evolutionary relationships of the Myxozoa

Members of the phylum Myxozoa are obligate parasites, primarily of aquatic organisms. Their phylogeny has remained problematic, with studies placing them within either the Bilateria or Cnidaria. The discovery that the enigmatic Buddenbrockia plumatellae is a myxozoan that possesses distinct bilaterian features appeared to have finally resolved the debate. B. plumatellae is described as a triploblastic 'worm-like' organism, within which typical myxozoan malacospores form. Using EM we examined the early development of the B. plumatellae 'worms' within the bryozoan host Plumatella repens. The initial development involved numerous unicellular, amoeboid pre-saccular stages that were present within the basal lamina of the host's body wall. These stages migrate immediately beneath the peritoneum where a significant host tissue reaction occurs. The stages aggregate, initiating the formation of a 'worm'. The base of a developing 'worm' forms a pseudosyncytium which resolves into an ectoderm surrounding a mesendoderm. The pseudosyncytium is directly anchored into neighbouring host cells via masses of striated fibres. The replication of the ectodermal and mesendodermal cells extends the developing 'worm' into the coelom of the host. The mesendoderm resolves to form a mesoderm and an endoderm. Myogenesis appears to be initiated from the anchored end of the 'worm' and develops along the mesoderm. The aggregation and differentiation of amoeboid pre-saccular stages to initiate the 'worm' draws analogies to the sacculogenesis observed for Tetracapsuloides bryosalmonae, B. plumatellae's sister taxon within the class Malacosporea. The development of a multicellular, spore forming organism, from single cells does not correlate to any bilaterian or cnidarian species. Current phylogenies indicate the Myxozoa are basal bilaterians along with the Acoela and Mesozoa. Comparison with these other basal groups may help to resolve the placement of Myxozoa within the tree of life.     

On Some Features of Early Embryonic Development Stages of Cnidaria

Division of the life cycle of Cnidaria (except for Anthozoa) into two independent generations, polypoid and medusoid, i.e., metagenesis, is considered to be unjustified. Like other Metazoa, their life cycle can be divided into three periods: embryonic, postembryonic, and definitive, i.e., according to the age [9, 10]. An important feature of Cnidaria is the transition of some postembryonic stages to the sedentary mode of life. As in other animals, this change results in a substantial reduction in organismic integrity and an anarchical type of cell division. Some researchers [3, 5, 7] regard this type of cell division as original. However, the anarchical type of cell division itself is secondary and, for this reason, cannot be ancestral to other types. The statement that the spiral type of cell division originated from the pseudo-spiral type also arouses serious criticism. The spiral type is caused by a change in the structure of the blastomere ooplasm rather than the appropriate arrangement of blastomeres.     

Small subunit ribosomal RNA sequence of Henneguya exilis (class Myxosporea) identifies the actinosporean stage from an oligochaete host

Several transmission studies, as well as recent molecular data, have indicated that the two classes Myxosporea and Actinosporea represent different life cycle stages of Myxozoa. To evaluate the life cycles of myxozoa in catfish aquaculture systems, the small subunit (18S) ribosomal RNA gene sequences of Henneguya exilis, a myxosporean from channel catfish Ictalurus punctatus, and an actinosporean (previously designated as Aurantiactinomyxon janiszewskai) from the aquatic oligochaete Dero digitata were determined. The sequences were identical, indicating that H. exilis and the actinosporean are alternate life stages of a single species. This is the first report identifying the actinosporean stage of the genus Henneguya.     

Fine structure of the nematocytes of Polypodium hydriforme Ussov (Cnidaria)

The fine structure of the stinging cells (nematocytes) and stinging capsules (nematocysts) is described for Polypodium hydriforme. a freshwater coclenterate with a prominent endoparasitie stage in its life cycle. All the nematocysts belong to the type of lesser glutinants (atrichous isorhiza) and fall into three size classes. The internal structure of the capsules is similar in the three classes. A novel type of organization of the cnidocil apparatus of the nematocysts is described. The cnidocil lacks a root fibre and its kinctosome sits directly on the operculum of the nematocyst, so that the entire cnidocil apparatus has a radial rather than bilateral symmetry. It is compared with that of other types of nematocytes and its similarity with the mechanoreccptors of the coelentcratcs is noted. The possible place of the Polypodium nematocytes in the evolution of the collar receptors of the Metazoa is discussed.     

Life cycles of the trematodes Parasymphilodora japonica (Yamaguti, 1938) and P. markewitschi (Kulakowskaja, 1947) (Monorchidae) in the conditions of Primorye land

In contrasts to formerly known data, it is found that two species of the Parasymphilodora trematodes, Parasymphilodora japonica (Yamaguti, 1938) and P. markewitschi (Kulakowskaja, 1947) are actually distributed in the territory of Primorye land. The first intermedial hosts of the former species are snails of the genus Parafossarulus; in the second species, these hosts are snails of Boreoelona. A life cycle and stages of development of both trematodes are described.     

Recent Advances in Our Knowledge of the Myxozoa

In the last few years two factors have helped to significantly advance our understanding of the Myxozoa. First, the phenomenal increase in fin fish aquaculture in the 1990s has lead to the increased importance of these parasites; in turn this has lead to intensified research efforts, which have increased knowledge of the development, diagnosis. and pathogenesis of myxozoans. The hallmark discovery in the 1980s that the life cycle of Myxobolus cerebralis requires development of an actinosporean stage in the oligochaete. Tubifex tubifex, led to the elucidation of the life cycles of several other myxozoans. Also, the life cycle and taxonomy of the enigmatic PKX myxozoan has been resolved: it is the alternate stage of the unusual myxozoan, Tetracapsula bryosalmonae, from bryozoans. The 18S rDNA gene of many species has been sequenced, and here we add 22 new sequences to the data set. Phylogenetic analyses using all these sequences indicate that: 1) the Myxozoa are closely related to Cnidaria (also supported by morphological data); 2) marine taxa at the genus level branch separately from genera that usually infect freshwater fishes; 3) taxa cluster more by development and tissue location than by spore morphology; 4) the tetracapsulids branched off early in myxozoan evolution, perhaps reflected by their having bryozoan, rather than annelid hosts; 5) the morphology of actinosporeans offers little information for determining their myxosporean counterparts (assuming that they exist); and 6) the marine actinosporeans from Australia appear to form a clade within the platysporinid myxosporeans. Ribosomal DNA sequences have also enabled development of diagnostic tests for myxozoans. PCR and in situ hybridisation tests based on rDNA sequences have been developed for Myxobolus cerebralis, Ceratomyxa shasta, Kudoa spp., and Tetracapsula bryosalmonae (PKX). Lectin-based and antibody tests have also been developed for certain myxozoans, such as PKX and C. shasta. We also review important diseases caused by myxozoans, which are emerging or re-emerging. Epizootics of whirling disease in wild rainbow trout (Oncorhynchus mykiss) have recently been reported throughout the Rocky Mountain states of the USA. With a dramatic increase in aquaculture of fishes using marine netpens, several marine myxozoans have been recognized or elevated in status as pathological agents. Kudoa thyrsites infections have caused severe post-harvest myoliquefaction in pen-reared Atlantic salmon (Salmo salar), and Ceratomyxa spp., Sphaerospora spp., and Myxidium leei cause disease in pen-reared sea bass (Dicentrarchus labrax) and sea bream species (family Sparidae) in Mediterranean countries.     

The contrariwise life of a parasitic, pedomorphic copepod with a non-feeding adult: ontogenesis, ecology, and evolution

Abstract. The extraordinary parasitic metanauplius larva of Caribeopsyllus amphiodiae is sexually dimorphic, with conspicuous gonads, and elaborate lens-bearing eyes. The parasites usually occur singly within their host, and grow for ≤5 months within the stomach of burrowing ophiuroids ( Amphiodia urtica ). They transform into free-living, semelparous, non-feeding adults that live only 2 weeks. The species' life-history pattern, with a larval period ∼10 × longer than the adult life span, is contrariwise to that of other copepods but not for animals with non-feeding adults of both sexes. It appears that the life cycle of C. amphiodiae is pedomorphic, and probably evolved through a delay of metamorphosis regulated by developmental hormones. We attribute the dominance of the larval phase to the greater potential for survival and growth of the enterozoic parasitic stages than of the free-living, post-metamorphic stages. We note that among marine invertebrates, non-feeding adults of both sexes occur exclusively in taxa with a complex life cycle, and that non-feeding adults of both sexes are never found in taxa that have small larvae and delayed maturation. They occur only when there is a large larva that can provide the adult stage with sufficient nutrient reserves for reproduction.     

Nucleic acids: vaccines of the future

The recent successful immunization of experimental animals using nucleic acids has provided a revolutionary new approach in vaccinology. In this article, Gary Waine and Don McManus examine the potential of nucleic acid vaccines for their effectiveness not only against infectious and parasitic organisms exhibiting an intracellular phase during their life cycle, but also against parasitic helminths, whose life cycle stages are either predominantly or completely extracellular.     

Morphological and ultrastructural analysis of Turritopsis nutricula during life cycle reversal

The hydrozoa life cycle is characterized, in normal conditions, by the alternation of a post-larval benthic polyp and an adult pelagic medusa; however, some species of Hydrozoa react to environmental stress by reverting their life cycle: i.e. an adult medusa goes back to the juvenile stage of polyp. This very uncommon life cycle could be considered as some sort of inverted metamorphosis. A morphological study of different stages during the reverted life cycle of Turritopsis nutricula led to the characterization of four different stages: healthy medusa, unhealthy medusa, four-leaf clover and cyst. The ultrastructural study of the cellular modifications (during the life cycle reversion of T. nutricula) showed the presence of both degenerative and apoptotic processes. Degeneration was prevalent during the unhealthy medusa and four-leaf clover stages, while the apoptotic rate was higher during the healthy medusa and cyst stages. The significant presence of degenerative and apoptotic processes could be related to the occurrence of a sort of metamorphosis when an adult medusa transforms itself into a polyp.     

Phylogeny of the Metazoa Based on Morphological and 18S Ribosomal DNA Evidence

Cladistic analysis of traditional (i.e. morphological, developmental, ultrastructural) and molecular (18S rDNA) data sets (276+501 informative characters) provides a hypothesis about relationships of all meta-zoan higher taxa. Monophyly of Metazoa, Epith-eliozoa (= -03non-Porifera), Triploblastica, Mesozoa, Eutriploblastica (=Rhabditophora+Catenulida+“higher triploblasts”=Neotriploblastica, including Xeno- turbellida and Gnathostomulida), Rhabditophora, Syndermata (=“Rotifera”+Acanthocephala), Neotrichozoa (=Gastrotricha+Gnathostomulida), Nematozoa (=Nematoda+Nematomorpha), Panarthropoda (=Onychophora+Tardigrada+ Arthropoda), Cephalorhyncha, Deuterostomia, Ambulacralia (=Hemichordata+Echinodermata), Chordata, Phoronozoa (=Phoronida+“Brachiopoda”), Bryozoa, Trochozoa (=Eutrochozoa+Entoprocta+ Cycliophora), Eutrochozoa, and Chaetifera (=Annelida+ Pogonophora+Echiura) is strongly supported. Cnidaria (including Myxozoa), Ecdysozoa (=Cepha- lorhyncha + Nematozoa + Chaetognatha + Panarthropoda), Eucoelomata (=Bryozoa+Phoronozoa+Deuterostomia+Trochozoa, possibly including also Xenoturbellida), and Deuterostomia+Phoronozoa probably are monophyletic. Most traditional “phyla” are monophyletic, except for Porifera, Cnidaria (excluding Myxozoa), Platyhelminthes, Brachiopoda, and Rotifera. Three “hot” regions of the tree remain quite unresolved: basal Epitheliozoa, basal Triploblastica, and basal Neotriploblastica. A new phylogenetic classification of the Metazoa including 35 formally recognized phyla (Silicispongea, Calcispongea, Placozoa, Cnidaria, Ctenophora, Acoela, Nemertodermatida, Orthonecta, Rhombozoa, Rhabditophora, Catenulida, Syndermata, Gnathostomulida, Gastrotricha, Cephalorhyncha, Chaetognatha, Nematoda, Nematomorpha, Onychophora, Tardigrada, Arthropoda, Echinodermata, Hemichordata, Chordata, Phoronozoa, Bryozoa s. str., Xenoturbellida, Entoprocta, Cycliophora, Nemertea, Mollusca, Sipuncula, Echiura, Pogonophora, and Annelida) and few i ncertae sedis g roups (e.g. Myzostomida and Lobatocerebromorpha) is proposed.