Molecular characterization of myxozoan parasites from Lake Sasajewun, Algonquin Park, Ontario, by riboprinting

The small subunit-rRNA genes of 18 myxozoans from Lake Sasajewun, Algonquin Park were amplified and digested with restriction endonucleases for riboprinting analysis. Identical riboprints were not found between the myxosporeans and the actinosporeans. The distinct riboprinting patterns observed among these myxozoans indicate considerable genetic diversity within this group. Identical riboprints were found between Myxobolus pendula and Myxobolus pellicides, and between triactinomyxon 'C' and Triactinomyxon ignotum. Parsimony analysis of the riboprints demonstrated that neither the myxosporeans nor the actinosporeans formed a monophyletic group. Some species of Myxobolus are more closely related to forms of triactinomyxon, echinactinomyxon or raabeia than to other Myxobolus species. These results are consistent with the two-host life cycle hypothesis of myxozoans that myxosporeans and actinosporeans are alternating stages of the same organisms.     

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.     

Molecular relationships and phylogeny in a community of myxosporeans and actinosporeans based on their 18S rDNA sequences

The community of myxosporeans and actinosporeans inhabiting a typical Scottish highland stream and the outflow area of an adjacent salmon hatchery was analysed on the basis of their 18S rDNA sequences. Nine myxosporeans belonging to the genera Sphaerospora, Chloromyxum, Zschokkella, Myxidium, Hoferellus and Myxobilatus were identified from mature spores in different organs of the fish species present. Twelve actinosporean types belonging to the collective groups of neoactinomyxum, aurantiactinomyxon, raabeia, echinactinomyxon and synactinomyxon were found to be released from oligochaete worms collected from sediments. Twenty of the 21 sequences obtained from these myxozoans are new entries to the myxozoan database, and the genera Chloromyxum, Hoferellus and Myxobilatus were entered for the first time. Study of the molecular relationships between the different taxa and with other myxozoan sequences available showed that the myxosporeans inhabiting the urinary system clearly cluster together, independently of host species or spore morphology. However, the sequences of the two Sphaerospora species encountered show considerable differences from other members of this group and all other freshwater myxosporeans, and they were found to occupy an ancestral marine position. Three actinosporeans, i.e. Neoactinomyxum eiseniellae, Aurantiactinomyxon pavinsis and Raabeia 'type 3' were found to represent alternate life cycle stages of Chloromyxum sp., Chloromyxum truttae and Myxidium truttae, respectively (approximately 1400 identical base pairs each). Three other actinosporeans encountered (two echinactinomyxon and one raabeia type) showed over 92% sequence identity with myxosporeans from GenBank, whereas all other actinosporeans formed a closely related group devoid of any known myxosporeans.     

Fine Structure of Triactinomyxon Early Stages and Sporogony: Myxosporean and Actinosporean Features Compared

ABSTRACT. The first ultrastructural study of the actinosporean genus Triactinomyxon was carried out on Triactinomyxon legeri from the intestinal epithelium of Tubifex tubifex. The developmental cycle starts with bi- and uninucleate cells. We propose that these cells may be an early proliferative phase of the cycle and may unite to give rise to the four-cell stage, initiating pansporoblast formation. Valvogenic cells transform in the long stylus and anchor-like projections of the spore. In the capsulogenic cells, the primordium of the polar capsules originates as a simple, dense, club-shaped structure not observed in other actinosporeans. In all other respects, actinosporean ultrastructure follows more or less similar patterns. Comparison of actinosporean and myxosporean species gives evidence of considerable structural similarity, exemplified in both classes by the occurrence of cell junctions in their multicellular spores, identical polar capsules and their morphogenesis, cell-in-cell condition, pansporoblast formation, and presence of dense bodies (sporoplasmosomes) primarily in the sporoplasm. This unity of patterns speaks in favor of the postulated actinosporean-myxosporean transformation, which warrants further study.     

Alternation of Actinosporean and Myxosporean Phases in the Life Cycle of Zschokkella nova (Myxozoa)

ABSTRACT. Experimental evidence has been gathered to show that the life cycle of the myxozoan gallbladder parasite Zschokkella nova Klokacewa, 1914, which infects the fish Carassius carassius , has a complex life cycle with alternation of two hosts (fish and Oligochaeta) and two developmental phases (myxosporean and actinosporean). The gut epithelium of the oligochaete, Tubifex tubifex , exposed experimentally to Z. nova , obtained from C. carassius , became infected with organisms resembling Actinosporea. The spore structure and cube-like network of the interconnected spores is reminiscent of Siedleckiella silesica Janiszewska, 1952, although the spores are very different in size and number of sporoplasm nuclei. The life cycle of Z. nova resembles that of the whirling disease agent Myxosoma cerebralis described by Wolf and Markiw, which also alternates between fish and oligochaete hosts.