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A new species of Dermocystidium is reported to be the causative agent of three outbreaks of gill disease in carp observed in recent years in Czechoslovakia. No other parasites or influences could be held responsible for the disease. It is suggested that at least some of gill disease of unknown aetiology in carp is due to dermocystidia, and the disease might be distributed in many regions of Europe. A description of a greater part of the developmental cycle of the species D. cyprini n.sp. Cervinka et Lom, is presented, supplemented by a discussion of other species of the genus. A summary of known facts on the host-parasite relation in these parasites is also given.

Electron-microscope studies demonstrate that presporangial cells (hypnospores) of the fungus Dermocystidium marinum are characterized by an apparent lack of mitochondria, by a cell wall homogeneous with respect to electron density, and by an electron-light nucleus. In precleavage sporangia, morphogenesis of mitochondria appears to occur from myelin figures, the cell walls acquire a three-layered appearance, and the nuclei become electron-dense with evidences of condensed chromatin. A discharge pore and tube are formed by rupture of the cell wall and subsequent unfolding of the tube from the innermost part of the cell wall. The pore is blocked by a plug which is a thickened section of the inner cell wall. Sporulation proceeds by successive bipartition of the protoplast involving alternating karyokinesis and cytokinesis. The nuclear membrane persists during karyokinesis. The process terminates with the formation of billagellated planonts, enclosed by two unit membranes. Evaginations of one or both of the membranes form a row of tinsels along one side of the anterior flagellum. Golgi bodies and a previously undescribed, electrondense body are found next to the planont nucleus. D. marinum is concluded to be a member of the Saprolegniales on the basis of planont morphology and previously described stages.

Dermocystidium koi sp. nov. This species is found as a parasite; in one case it is parasiting to the integument and in another to the muscle of each separate Cyprinus carpis L. The specimen of the former was captured at the Yamada Fish Culture Farm (Locality-Yamada Village, Hyogo Pref., Date-July 23, 1940), and that of the latter obtained from the Seto Fish Culture Farm. (Locality-Vicinity of Odawara City, Kanagawa Pref., Date-May 11, 1959). In the case of integumentary parasitim, the parasites grow between the epidermis and the cutis in uarious places of the body surface without scales; and the characteristic lesions a e formed arising from the integument, due to the increase of the parasites and host's tissues such as fibrous connective tissueand blood vessels. (see text-fig. 1 and 2, A, B.) The muscular parasitism occurs when the parasites are parasitic to the scaley host; and in this case the parasites grow in the muscle near the body surface; on the infected parts, the large and hard tumor like inflammatory swelling arise from the increase of the parasites and the same host's tissues as the former. The parasite is filiform in external character; its transuerse section is round; the diameter 0.04-0.30mm., and it contains innumerable spares. Spare subspherical, 6-14μ in diameter; the diameter of the large characteristic spherical enclosure is 4.5-10.0μ. The details are compiled in the table 1. The spares are figured in text-fig. 3, NOS. 1-9. The present species is distinguishable from all other known species, in external form of the parasite or the cyst, the modus parasitic, and the dimension of the spare etc. The lesions may be ruptured naturally in a certa'n period; and the contents fall out from them; and then they seem to be healed gradually. Therefore, the host should not suffer great influence from the parasite. But the histological reactions of the host are marked, which are chiefly proliferocis inflammation, rarely accompanied with phagocytosis. (Text.-fig. 3, No. 10).

PAGE INTRODUCTIONa. Studies in the Gulf of Mexicox. Discovery of Dermocystidiumn marinurm in Virginia 2 METHODS OF STUDY 2 The Thioglycollate Culture Technique 2 Rating the Intensity of Infections 3 Definitions of Terms 3 THE OCCURRENCE OF Dermocystidium marinuim IN LIVE OYSTERS 3 Seasonal Incidence of Infections 3 Intensity of Infections 5 Recapitulation ........ 6 THE OCCURRENCE OF Der-mocystidium marinumn IN GAPERS 6 Collection of Gapers 6 Seasonal Incidence ..... 6 Incidence in Gapers from Outlying Trays and Natural Habitats ..... 8 Recapitulation ........ 8 DISTRIBUTION OF Dermocystidium marinum IN CHESAPEAKE BAY 11 FUNGUS INFECTIONS IN RELATION TO AGE AND SouRCE OF OYSTEST.11 Data from Live Oysters . ..... 11 Data from Gapers ......... 12 Recapitulation. 13 EXPERIMENTAL INFECTION OF OYSTERS 14 Infection of Old Oysters ...... 14 Infection of Young Oysters I ........ 14 PAGE

MSX, a pathogen of oysters (Crassostrea virginica), produced a drastic epizootic in high—salinity areas of Chesapeake Bay from 1959 to 1963. The patterns of infection and mortality were determined by imports from disease—free seed—oyster areas. Winter and spring imports became infected in early summer and began dying in late summer. Late—summer imports apparently became infected promptly but infections remained subclinical until the following May. Death rates were highest during warm months but losses occurred throughout the year. MSX was assigned as the cause of a high percentage of deaths by sampling live and dead oysters. Mortality for the first and second years after import was usually 50 to 60% annually. A fungus disease caused by Dermocystidium marinum was also prevalent in some areas. Prevalence of MSX did not decline as oyster populations were decimated; hence nearly half of Virginia's private beds have been forced out of production.

A serious outbreak of mortalities occurred among adult salmon held in the Priest Rapids Dam spawning channel in November 1965. Gross examination revealed small, white cysts on and in the gill filaments of otherwise normal-appearing fish. Approximately 25% of the 5000 mature adult salmon in the channel died prior to spawning from this disease caused by a fungus identified as the genus Dermocystidium. This is the first record of an epidemic among fish of North America associated with Dermocystidium. The primary cause of death appeared to be anoxia due to subacute inflammation and granulation of the gill tissue. A description of the histopathology associated with these parasites is given. Chondrococcus columnaris, a pathogen of warm-water fishes, was present as a secondary invader on the gill filaments of several salmon and an unknown fungal or yeast-like organism was present in the testes of the male fish examined.

1. 1. Fatty acids of four marine fungi Thraustochytrium roseum, T. aureum, Schizochytrium aggregatum and Dermocystidium sp. and one fresh-water fungus Phlyctochytrium punctatum were determined. 2. 2. These fungi synthesized ω 3 and ω 6 polyunsaturated fatty acids. They contained large quantities of C20 and C22 polyenes; three of the marine fungi contained unusually large amounts of 22:6 fatty acid. 3. 3. Major fatty acids for T. roseum and S. aggregatum were palmitic, oleic and docosahexaenoic acids; for T. aureum, palmitic, docosapentaenoic and docosahexaenoic acids; for Dermocystidium, palmitic, palmitoleic and eicosadienoic acids; for P. punctatum, palmitic, stearic, oleic and linoleic acids. 4. 4. The pattern of biosynthesis of polyunsaturated fatty acids by these aquatic fungi as well as their morphology supports the hypothesis that they evolved from a primitive phytoflagellate (monad) ancestor.

Serious outbreaks of a pernicious disease caused by the fungus Dermocystidium are reported in both adult and juvenile salmon in the Columbia River and its tributaries. The disease evokes a characteristic histopathology and appears to cause death in a large percentage of the infected fish. The disease appears to be most virulent at water temperatures below 60 F. Some prophylactic relief is given by treatment with the organic herbicide, Diquat, at a dilution of 1:500,000 for 1 hr.

Twelve antifungal antibiotics were screened to determine their potential for controlling Dermocystidium marinum, a lethal fungus parasite of oysters. Two antibiotics, cycloheximide and streptimidone, completely inhibited enlargement of D. marinum in thioglycollate cultures at concentrations of 0.25–1.0 μg/ml. Cycloheximide shows promise for use in arresting D. marinum infections in oysters held in closed laboratory systems. Twenty-two antibacterial antibiotics were screened for their effect on the enlargement of D. marinum in thioglycollate cultures, and for their efficacy in retarding growth in bacteria associated with oyster tissues. Three tetracyclines (tetracycline, chlortetracycline, and oxytetracycline), bacitracin, and gramicidin showed effective antibacterial properties at levels of 250–500 units or mcg/ml. However, all of these agents except gramicidin markedly inhibited parasite enlargement at concentrations of 250–750 units or mcg/ml. Further studies proved gramicidin to be inferior to either chloramphenicol or the penicillin-dihydrostreptomycin combination for inhibiting bacterial growth and tissue putrefaction in the thioglycollate culture technique for detecting D. marinum.

Dermocystidium Gasterostei n. sp., A Parasite of Gasterosteus Aculeatus L. and Gasterosteus Pungitius L.

The life cycle of a nonfilamentous marine phycomycete, Dermocystidium sp., is described from its growth in axenic culture under controlled conditions. Reproduction occurs by vegetative means involving a process of cell enlargement and ecdysis modifiable by the methods of cultivation. Zoospores and sexuality were not observed. The results of inoculation studies with algae, phanerogam debris, and molluscs suggest that the fungus may be a free-living saprophyte in coastal waters.

The microstructure of the protist Labyrinthomyxa marina as found in a host organism, Crassostrea virginica, is described. Most of the organelles were similar to those of many eucaryotic protists; however, a few unusual structural features were observed. Viruslike particles (460–530 A diameter) were found in the nucleus and cytoplasm of the Virginia, but not the Florida, pathogens. Cell-wall formation appeared to occur, in part, by formation of material in vesicles derived from the endoplasmic reticulum. The vesicles appeared to fuse with the plasmalemma and release material into the cell wall. Funguslike lomasomes were found between the cell wall and plasmalemma. Vacuoplast formation occurred by synthesis of droplets of electrondense material through the membranes of large vacuoles followed by coalescence to form one or two large vacuoplasts. Mitochondria were involved in the formation of vesicles which contained electron-dense material and which were delimited by two membranes. Since L. marina has previously been allied with the Labyrinthulia (Protozoa) and the Saprolegniales (Phycomycetes), comparisons are made with the ultra-structure of representatives from both groups. Affinities with the fungi seem to be the closest on the strength of lomasome formation and the repeated formation of cell walls in all vegetative stages.

In the skin of Rana temporaria from Czechoslovakia two unicellular parasites have been found: Dermocystidium ranae Guyénot & Naville and Dermosporidium granulosum n.sp.Dermocystidium ranae. Two stages were found, one of which is new. An account is given of the morphology, development and occurrence of both stages, including details which have hitherto not been recorded.Dermosporidium granulosum n.sp. This species is represented by cysts situated in the stratum spongiosum corii. They are divided by septa into small chambers containing groups of spherical spores with a central nucleus and with numerous small cytoplasmic inclusions. Within the cyst, the sporoblasts occupy the periphery, while the ripe spores are in the centre.

THE examination and control of oyster (Crassostrea virginica Gmelin) pathogens have received much impetus in recent years with the recognition of Haplosporidium costale Wood and Andrews, ‘MSX’ an undescribed haplosporidian, and Dermocystidium marinum Mackin, Owen and Collier along the Atlantic and Gulf coasts of the United States. Research in the problems of isolation, identification and control of oyster pathogens could bo facilitated if techniques for the culture of oyster cells were available.

Zoospores were obtained from a protist, parasitic in the clamMacoma balthica. It has been previously suggested that the parasite is either a closely related species or is the same species asLabyrinthomyxa marina (Dermocystidium marinum), which is found in the oysterCrassostrea virginica. The demonstration of zoosporulation and the description of zoospore micromorphology are here offered as evidence of similarity between the two parasites. The zoospores were biflagellated, pyriform cells with mastigonemes along one side of the anterior flagellum. Negative staining revealed substructure in the mastigonemes.

Dermocystidium marinum is a pathogen which kills oysters in Chesapeake Bay from July through October. The fungus proliferates readily only at temperatures above 25°C but overwinters in oysters as sub-clinical cases. Experiments were conducted in Nature to demonstrate the effects of proximity of infected oysters on epizootiology in isolated disease-free populations. Prevalence ofDermocystidium in the York River was moderate in 1957, heavy in 1959, and extremely low in 1963. In 1957, with 40 feet of isolation, groups mixed with infected oysters showed large increases in deaths and prevalences over controls. In 1959, the fungus was so widely prevalent in York River during a long hot summer that relatively minor reductions in death rates were obtained by 50 feet of isolation. In 1963, with the fungus nearly absent, a failure in laboratory infection techniques caused late timing of the experiment, and results were inconclusive. A new disease caused by “MSX”, a protistan parasite, developed high prevalences in the 1963 lots. This organism, by destroying oyster, populations in high-salinity waters (above 15‰ in late summer) has effectively reducedDermocystidium to a minor role as an oyster pathogen in lower Chesapeake Bay. Losses fromDermocystidium in Chesapeake Bay are dependent upon duration of warm seasons and survival of cases through winter. MSX, is slightly more sensitive to low salinities but much more tolerant of low temperatures thanDermocystidium. The decline ofDermocystidium with the disappearance of oyster populations suggests that the pathogen may be dependent solely upon transmission from oyster, to oyster.