International trade in live molluscs: perspective from the Americas.

There is an active world trade in seed and edible molluscs, including such species as oysters, abalones, clams, scallops, and mussels. The supply of new and effectively-produced products, such as triploid oyster seed, is likely to increase due to greater world-wide demand for these high-value products, together with improved technology. Transfer to live molluscs is driven by need an availability, with concern over infectious disease risks often playing subordinate role or serving to support economic trade barriers. Experience shows that serious risks from infectious disease transfer exist. Technology and balanced management, however, can limit such risks. Separation of biological risks from trade control objectives is a necessary first step. A systematic method of risk analysis is urgently needed, along with predefined actions which can be adapted to the various risk levels. Critical to the effective development and implementation of this risk analysis is the participation of the shellfish industry in defining risks and actions. Organization and infrastructure to improve recognition of infectious diseases and decrease response time are necessary improvements, together with appropriately-targeted research on impacts, diagnosis, epidemiology, carrier status, and geographical distribution of diseases.

Disseminated neoplastic cells in Mytilus trossulus: verification of host species origin by (16S-like) rRNA sequence comparison.

Disseminated neoplasia is a leukemia-like disease that occurs in many species of bivalve molluscs worldwide, including the bay mussel (Mytilus trossulus). The etiology of the disease is undetermined, but an early report proposed that the anomalous bivalve cells were actually an invasive parasite rather than cancerous cells of host origin. Comparison of partial sequences of small subunit rRNA from normal and putative cancer cells was performed to resolve this issue. These studies showed a close phylogenetic relationship of the different forms of cancer cells to each other (similarity coefficient, 0.982), to the normal hemocytes (similarity coefficient, 0.990, 0.992), and to the oyster, Crassostrea virginica (similarity coefficient, 0.895-0.927). A large phylogenetic distance separates all 3 mussel hemocyte types from several representative protists (similarity coefficient, 0.702-0.761). These results indicate that the disseminated neoplastic cells in mussels are indeed proliferative host cells and not unicellular parasites.

Alternate pathogenesis of systemic neoplasia in the bivalve mollusc Mytilus.

The proliferative disease systemic neoplasia, also termed hemic neoplasia or disseminated sarcoma, was studied in four Puget Sound, Washington populations of the bay mussel (Mytilus sp.). Using flow cytometric measurement of DAPI-stained cells withdrawn from the hemolymph, DNA content frequency histograms were generated for 73 individuals affected by the disease. The cells manifesting systemic neoplasia were found to exist as either of two separate types, characterized by G0G1 phase nuclear DNA contents of either approximately 4.9 x haploid (pentaploid form) or approximately 3.8 x haploid (tetraploid form). The two disease forms were found to coexist in all four mussel populations sampled, with overall relative prevalences of 66% pentaploid form, 29% tetraploid form, and 5% exhibiting both disease forms simultaneously. These findings represent the first unequivocal demonstration of multiple cell types in a bivalve neoplasia. The two forms appear to represent separate pathogenetic processes rather than sequential stages of a single pathogenesis. Two cell cycling parameters associated with proliferative activity were employed to compare the alternate forms: (i) the percentage of cells assigned to the DNA Synthesis (S) phase of the neoplastic cell cycle, and (ii) the proportion of neoplastic cell mitotic figures in hemocytological preparations. Mean values for both parameters were significantly higher for mussels with the tetraploid form of the disease, suggesting a higher rate of proliferation relative to the pentaploid form. Qualitatively, cells of the tetraploid form contained slightly lower nuclear and cytoplasmic volumes compared to those of the pentaploid form. An observed wide variation in neoplastic cell nuclear size within either disease form may reflect the distribution of cells in the G0G1, S, and G2M phases of the cell cycle. Potential etiologic relationships between the two forms are discussed.

Postoperative endophthalmitis.

Postoperative endophthalmitis is a rare complication of ophthalmic surgery. The use of prophylactic antibiotics and scrupulous surgical techniques will help to reduce its incidence, but sporadic cases may still occur. If postoperative endophthalmitis is a diagnostic possibility aggressive investigation and initial management improves the prognosis. More work needs to be done to evaluate the penetration of newer antibiotics into the eye and their therapeutic role. The use of corticosteroids also needs to be more carefully defined in the treatment of this potentially devastating disease.

Impaired defense mechanisms in bay mussels, Mytilus edulis, with hemic neoplasia.

Immunocompetence of bay mussels, Mytilus edulis, with hemic neoplasia was investigated with an in vitro yeast phagocytosis assay and by in vivo clearance from the blood of injected Cytophaga sp. bacteria. The yeast phagocytosis assay was conducted with hemocytes maintained in 90% plasma. Neoplastic hemocytes, characterized by enlarged nuclei and scant cytoplasm, failed to phagocytose yeast cells. In contrast, greater than 90% of hemocytes from unaffected animals and morphologically normal hemocytes from mussels with the disease phagocytosed yeast. Substitution of normal plasma with that from a mussel with advanced disease (essentially 100% neoplastic hemocytes) did not affect the phagocytic capability of normal hemocytes. Conversely, normal plasma did not enhance the phagocytic capabilities of neoplastic cells. Mussels with advanced disease showed reduced bacterial clearance; control or lightly affected mussels (less than 11% neoplastic hemocytes) cleared greater than 90% of injected bacteria in 4 hr, while mussels with advanced disease cleared 44-83%. These experiments indicate that mussels with advanced hemic neoplasia have compromised defense systems. This may account for the reported mortality in mussels and other bivalve molluscs with hemic neoplasia.

Evidence for colonization and destruction of hinge ligaments in cultured juvenile Pacific oysters (Crassostrea gigas) by cytophaga-like bacteria.

Several strains of cytophaga-like gliding bacteria (CLB) were isolated as numerically dominant or codominant components of bacterial populations associated with proteinaceous hinge ligaments of cultured juvenile Pacific oysters, Crassostrea gigas. These bacteria were morphologically similar to long, flexible bacilli occurring within degenerative lesions in oyster hinge ligaments. Among bacteria isolated from hinge ligaments, only CLB strains were capable of sustained growth with hinge ligament matrix as the sole source of organic carbon and nitrogen. In vitro incubation of cuboidal portions of ligament resilium with ligament CLB resulted in bacterial proliferation on the surfaces and penetration deep into ligament matrices. Bacterial proliferation was accompanied by loss of resilium structural and mechanical integrity, including complete liquefaction, at incubation temperatures between 10 and 20 degrees C. The morphological, distributional, and degradative characteristics of CLB isolated from oyster hinge ligaments provide compelling, albeit indirect, evidence that CLB are the agents of a degenerative disease affecting juvenile cultured oysters. The motility, metabolic, and hydrolytic characteristics of hinge ligament CLB and the low moles percent G + C values (32.4 to 32.9) determined for three representative strains indicate that they are marine Cytophaga spp.

Goussia girellae n. sp. (Apicomplexa: Eimeriorina) in the opaleye, Girella nigricans.

Goussia girellae n. sp. is described from the opaleye fish, Girella nigricans. Merogonic stages were observed in the apices of intestinal epithelial cells, in the lamina propria, and in extra-intestinal sites including liver, gills, and spleen. Gamonts were observed in the intestinal epithelial cells. Only unsporulated oocysts were detected in the intestine, and sporulation occurred when feces containing oocysts were incubated for 48 h in seawater at 21 degrees C. Oocysts are elongated (24.8 x 14.7 micron) with a wall about 200 nm thick and have no residuum, micropyle, or polar granule. Sporocysts are ellipsoid (8.5 x 4.5 micron), have a thin two-layered wall approximately 30 nm thick, and consist of two valves joined by a suture. Although moribund opaleye were also infected with Gyrodactylus sp., Cryptobia sp., Cardicola sp., and epitheliocystis organisms (chlamydia), all fish were heavily infected with G. girellae and morbidity was thus attributed to the coccidium.

Transmission of hemic neoplasia in the bay mussel, Mytilus edulis, using whole cells and cell homogenate.

Experimental studies with hemic neoplasia in the bay mussel indicated that the condition can be transmitted allogeneically with intact whole cells and cell-free homogenate. A differential pathogenesis of the disease in mussels receiving the two different inocula supports the argument that actual cell transplantation occurred. In addition to the first demonstration of the infectious nature of the disease with cell-free homogenates, it was also shown that the disease is transmitted by cohabitation. Remission of the disease occurred in some mussels indicating individual variation in recognition mechanisms.

An internal X-cell pseudotumor in a black croaker (Cheilotrema saturnum).

Cells closely resembling X-cells were the primary cellular component of a large pseudotumor in the viscera of a black croaker (Cheilotrema saturnum). The fish was captured in coastal waters off San Diego, California and was maintained at the Scripps Aquarium. After 2 years in captivity the fish exhibited extreme abdominal swelling due to a visceral mass, which weighed approximately one-fifth the total body weight. The cells associated with the pseudotumor were identified as X-cells due to their great variability in size, vesicular nuclei which stained negative for DNA and their formation of a pseudotumor. This is the first report of an X-cell pseudotumor in the visceral cavity. We agree with the hypothesis that the X-cell is a protozoan, and the description here of X-cells in a visceral pseudotumor in a fish of the family Sciaenidae associates an additional type of lesion with this enigmatic parasite.

An intranuclear microsporidium associated with acute anemia in the chinook salmon, Oncorhynchus tshawytscha.

An intranuclear microsporidium is described from hemoblastic cells of the chinook salmon, Oncorhynchus tshawytscha. The infection is associated with an acute anemia in the fish. Up to 47% of the hemoblast nuclei were infected in anemic fish. The organisms, found only in spleen and kidney tissues, were 1-2 microns in diameter and consisted of vegetative and early sporulation forms. This microsporidium differs from known species which parasitize fish in its tissue location; however, the absence of mature spores and other life cycle stages precludes determination of its precise taxonomic identity.