Stephen W. Feist

Recent advances in our knowledge of the Myxozoa

Abstract 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.

Biodiversity: Disease threat to European fish

The deliberate introduction of new species can have unexpected negative consequences and we show here how a recently introduced fish, the invasive Asian cyprinid Pseudorasbora parva, is causing increased mortality and totally inhibiting spawning in an already endangered native fish, the European cyprinid Leucaspius delineatus. This threat is caused by an infectious pathogen, a rosette-like intracellular eukaryotic parasite that is a deadly, non-specific agent. It is probably carried by healthy Asian fish, and could decrease fish biodiversity in Europe, as well as having implications for commercial aquaculture.

Disease threat to European fish.

The deliberate introduction of new species can have unexpected negative consequences and we show here how a recently introduced fish, the invasive Asian cyprinid Pseudorasbora parva, is causing increased mortality and totally inhibiting spawning in an already endangered native fish, the European cyprinid Leucaspius delineatus. This threat is caused by an infectious pathogen, a rosette-like intracellular eukaryotic parasite that is a deadly, non-specific agent. It is probably carried by healthy Asian fish, and could decrease fish biodiversity in Europe, as well as having implications for commercial aquaculture.

The British river of the future: How climate change and human activity might affect two contrasting river ecosystems in England

The possible effects of changing climate on a southern and a north-eastern English river (the Thames and the Yorkshire Ouse, respectively) were examined in relation to water and ecological quality throughout the food web. The CLASSIC hydrological model, driven by output from the Hadley Centre climate model (HadCM3), based on IPCC low and high CO(2) emission scenarios for 2080 were used as the basis for the analysis. Compared to current conditions, the CLASSIC model predicted lower flows for both rivers, in all seasons except winter. Such an outcome would lead to longer residence times (by up to a month in the Thames), with nutrient, organic and biological contaminant concentrations elevated by 70-100% pro-rata, assuming sewage treatment effectiveness remains unchanged. Greater opportunities for phytoplankton growth will arise, and this may be significant in the Thames. Warmer winters and milder springs will favour riverine birds and increase the recruitment of many coarse fish species. However, warm, slow-flowing, shallower water would increase the incidence of fish diseases. These changing conditions would make southern UK rivers in general a less favourable habitat for some species of fish, such as the Atlantic salmon (Salmo salar). Accidental or deliberate, introductions of alien macrophytes and fish may change the range of species in the rivers. In some areas, it is possible that a concurrence of different pressures may give rise to the temporary loss of ecosystem services, such as providing acceptable quality water for humans and industry. An increasing demand for water in southern England due to an expanding population, a possibly reduced flow due to climate change, together with the Water Framework Directive obligation to maintain water quality, will put extreme pressure on river ecosystems, such as the Thames.

A New Class and Order of Myxozoans to Accommodate Parasites of Bryozoans with Ultrastructural Observations on Tetracapsula bryosalmonae (PKX Organism)

Abstract Tetracapsula bryosalmonae, formerly PKX organism, is a myxozoan parasite that causes proliferative kidney disease in salmonid fish. Its primary hosts, in which it undergoes a sexual phase, are phylactolaemate bryozoans. It develops in the bryozoan coelomic cavity as freely floating sacs which contain two types of cells, stellate cells and sporoplasmogenic cells, which become organised as spores. Eight stellate cells differentiate as four capsulogenic cells and four valve cells which surround a single sporoplasmogenic cell. The sporoplasmogenic cell undergoes meiosis and cytoplasmic fission to produce two sporoplasms with haploid nuclei. Sporoplasms contain secondary cells. The unusual development supports previously obtained data from 18S rDNA sequences, indicating that species of Tetracapsula form a clade. It diverged early in the evolution of the Myxozoa, before the radiation that gave rise to the better known genera belonging to the two orders in the single class Myxosporea. The genus Tetracapsula as seen in bryozoans shares some of the characters unique to the myxosporean phase and others typical of the actinosporean phase of genera belonging to the class Myxosporea. However, it exhibits other features which are not found in either phase. A new class Malacosporea and order Malacovalvulida are proposed to accommodate the family Saccosporidae and genus Tetracapsula. Special features of the new class are the sac-like proliferative body, valve cells not covering the exit point of the polar filament, lack of a stopper-like structure sealing the exit, maintenance of valve cell integrity even at spore maturity, absence of hardened spore walls and unique structure of sporoplasmosomes in the sporoplasms.

Microsporidia: diverse, dynamic, and emergent pathogens in aquatic systems

Microsporidia cause important diseases in aquatic organisms. Susceptible hosts span a remarkable taxonomic spectrum, from protists to mammals. Known genera represent a small fraction of extant taxa in global aquatic systems. Transmitting horizontally or vertically, effects range from cryptic to catastrophic; individual level effects being extrapolated to populations and ecosystems. Debate over positioning within the eukaryotes and internal structuring of the phylum has benefited from molecular phylogenetic approaches to taxonomy. Similar tools offer new perspectives on transmission between hosts of differing trophic status and habitats. Accessible models for host-pathogen interaction are emerging from full genome sequencing projects. Hyperparasitism and the close phylogenetic relationship between taxa infecting invertebrates and vertebrates not only underline a ubiquity in aquatic systems but also potential for zoonotic transfer.

PATTERNS OF OCCURRENCE AND 18S rDNA SEQUENCE VARIATION OF PKX (TETRACAPSULA BRYOSALMONAE), THE CAUSATIVE AGENT OF SALMONID PROLIFERATIVE KIDNEY DISEASE

Recent progress in understanding the etiology of proliferative kidney disease (PKD) includes the identification of freshwater bryozoans as the natural hosts of the myxozoan parasite that causes the disease in salmonid fish and formal description of the parasite as Tetracapsula bryosalmonae. This paper presents data on patterns of occurrence of T. bryosalmonae and sequence variation among isolates. T. bryosalmonae infects bryozoans that range from primitive to more derived genera within the Phylactolaemata and that differ in growth form and habits. Infected bryozoans have been collected in diverse habitats including cold, clear streams and warm, eutrophic lakes. Temporal surveys reveal intra- and interannual variation in infection levels, and spatial variation in incidence of infection is implicit by the apparent absence of T. bryosalmonae from many bryozoan populations. The significance of minor variation in partial sequences of 18S rDNA requires further investigation. The information presented here provides the first significant insights into the ecology of T. bryosalmonae.

Recommendations for control of pathogens and infectious diseases in fish research facilities.

Concerns about infectious diseases in fish used for research have risen along with the dramatic increase in the use of fish as models in biomedical research. In addition to acute diseases causing severe morbidity and mortality, underlying chronic conditions that cause low-grade or subclinical infections may confound research results. Here we present recommendations and strategies to avoid or minimize the impacts of infectious agents in fishes maintained in the research setting. There are distinct differences in strategies for control of pathogens in fish used for research compared to fishes reared as pets or in aquaculture. Also, much can be learned from strategies and protocols for control of diseases in rodents used in research, but there are differences. This is due, in part, the unique aquatic environment that is modified by the source and quality of the water provided and the design of facilities. The process of control of pathogens and infectious diseases in fish research facilities is relatively new, and will be an evolving process over time. Nevertheless, the goal of documenting, detecting, and excluding pathogens in fish is just as important as in mammalian research models.

Susceptibility of the European common frog Rana temporaria to a panel of ranavirus isolates from fish and amphibian hosts

Ranaviruses are an emerging group of viruses and have been implicated in an increase of epidemics in susceptible species. They have a wide host range, infecting fish, amphibians and reptiles, with some isolates able to infect multiple species from different animal classes. Whilst some information exists on the pathogenicity of ranaviruses to novel hosts, there is none on the pathogenicity of fish ranaviruses to amphibians; this information is needed to develop measures to prevent the further spread of ranaviral disease in the aquatic environment. We undertook bath infection trials to assess the susceptibility of the European common frog Rana temporaria to 9 ranavirus isolates comprising doctor fish virus (DFV), European sheatfish virus (ESV), epizootic haematopoietic necrosis virus (EHNV), guppy virus 6 (GV6), pike-perch iridovirus (PPIV) and short-finned eel ranavirus (SERV) from fish hosts, and Bohle iridovirus (BIV), frog virus 3 (FV3) and Rana esculenta virus 282/I02 (REV) from amphibians. Animals were challenged as tadpoles at 15 and 20°C and as recent metamorphs at room temperature (20 ± 1°C) to investigate the effect of temperature and amphibian developmental stage on virus pathogenicity. Tadpoles were susceptible to FV3, PPIV and REV, but refractory to the other ranaviruses. Post-metamorphs were susceptible to FV3 and REV but refractory to BIV (the other ranaviruses were not tested). Significant mortality occurred in post-metamorphs and in tadpoles challenged at 20°C but was low in tadpoles challenged at 15°C. This study presents the first evidence of mortality in an amphibian species after challenge with ranavirus originally isolated from fish.

A RANKING SYSTEM FOR THE EVALUATION OF INTERSEX CONDITION IN EUROPEAN FLOUNDER (PLATICHTHYS FLESUS )

Intersex (ovotestis) is the condition whereby an individual possesses oocytes, at varying degrees of severity, within the normal testicular tissue and is regarded as an endpoint of endocrine disruption in male fish. The occurrence of intersex has been linked with exposure to endocrine-disrupting compounds present in the environment, which are thought to mimic the actions of the female sex hormone 17-beta-oestradiol. Using stained histological sections of archived cases of intersex from European flounder (Platichthys flesus), this study developed a grading method to assess the extent of the condition. This was termed the ovotestis severity index (OSI). The OSI utilizes the distribution pattern (focal, diffuse, cluster, zonal) and type of oocyte (previtellogenic, cortical alveolar, vitellogenic) to assign a severity rating to each intersex fish. A high OSI refers to fish with testicular material almost completely replaced with mature (vitellogenic) oocytes, and lower scores indicate those fish with small numbers of immature (previtellogenic) oocytes. The development of this index provides pathologists with a robust tool for the grading of the intersex condition in European flounder and potentially in other fish species.