Hemocytic sarcoma of the body wall in a California king crab Paralithodes californiensis.

Neoplasia is rarely reported in decapod crustaceans, and sarcoma has not been previously reported in any crab species. A California king crab Paralithodes californiensis with a recent history of autotomy (4 legs lost) and anorexia was found dead. Grossly, the crab had a pigmented ulcer on the right cheliped merus. Necropsy tissue samples were placed in 10% neutral buffered formalin and processed routinely for histology. Both histochemical (i.e. Brown and Brenn Gram, Fite-Faraco acid fast, Fontana-Masson, Giemsa, hematoxylin and eosin, Masson's trichrome, periodic acid-Schiff [PAS], phosphotungstic acid-hematoxylin, and von Kossa) and immunohistochemical (i.e. cytokeratin, vimentin, and lysozyme) stains were performed. The body wall (presumably of the right cheliped merus) was ulcerated and subtended by a densely cellular, unencapsulated, invasive neoplasm composed of spindle cells arranged in intersecting streams and bundles embedded in a small to moderate amount of fibromatous stroma. Neoplastic cells were oval to elongate with fibrillar, pale eosinophilic cytoplasm that occasionally contained moderate numbers of small, spherical, brightly eosinophilic granules that were highlighted with PAS and Giemsa stains. Neoplastic cells had mild atypia and no evident mitoses. Immunohistochemical stains were noncontributory. This neoplasm is consistent with hemocytic sarcoma of semi-granulocytic origin. Decapod crustaceans have 3 types of hemocytes: hyalinocytes, granulocytes, and semi-granulocytes. Neoplastic cells had PAS- and Giemsa-positive granules, which are present in both semi-granulocytes and granulocytes. Semi-granulocytes can elongate and are associated with deposition of extracellular matrix during some immune responses. Neoplastic cells were elongate and associated with deposition of matrix. These findings suggest neoplastic cells were of semi-granulocytic origin.

A review of current state of knowledge concerning Perkinsus marinus effects on Crassostrea virginica (Gmelin) (the eastern oyster).

The eastern oyster, Crassostrea virginica (Gmelin), is both an important component of our estuaries and an important farmed food animal along the east and south coasts of the United States. Its populations have been significantly diminished in the wild due to decades of overfishing beginning in the 1890 s. Unfortunately, in 1950, a new disease in eastern oysters caused by the protistan agent, Perkinsus marinus, was identified. The disease, resulting from infection with this protozoan, leads to high mortality of both wild and cultured eastern oysters. Current restoration efforts are hampered by the disease, as is the aquaculture of this economically important food. The parasite infects hemocytes and causes hemolytic anemia and general degeneration of the tissues, leading to death. Ongoing research efforts are attempting to develop oysters resistant to the disease. Transport regulations exist in may states. Infection with P. marinus is listed as a reportable disease by the World Health Organization.

DGGE-based detection method for Quahog Parasite Unknown (QPX).

Quahog Parasite Unknown (QPX) is a significant cause of hard clam Mercenaria mercenaria mortality along the northeast coast of the United States. It infects both wild and cultured clams, often annually in plots that are heavily farmed. Subclinically infected clams can be identified by histological examination of the mantle tissue, but there is currently no method available to monitor the presence of QPX in the environment. Here, we report on a polymerase chain reaction (PCR)-based method that will facilitate the detection of QPX in natural samples and seed clams. With our method, between 10 and 100 QPX cells can be detected in 1 l of water, 1 g of sediment and 100 mg of clam tissue. Denaturing gradient gel electrophoresis (DGGE) is used to establish whether the PCR products are the same as those in the control QPX culture. We used the method to screen 100 seed clams of 15 mm, and found that 10 to 12% of the clams were positive for the presence of the QPX organism. This method represents a reliable and sensitive procedure for screening both environmental samples and potentially contaminated small clams.

Diagnosis and management of atypical Mycobacterium spp. infections in established laboratory zebrafish (Brachydanio rerio) facilities.

Two established zebrafish colonies experienced increased mortality and decreased reproductive performance. Initial examination of several fish from one facility revealed hyperemic gills, petechia around the opercula, abdominal distention, and emaciation. Affected fish had congested liver with inflammation and multifocal hepatic necrosis. Large numbers of acid-fast-positive, rod-shaped bacteria were evident in multiple tissues and the blood. Mycobacterium fortuitum was subsequently isolated from several fish. Zebrafish from the second facility had skin erosions and ulceration along the flank just caudal to the pectoral fins. Large numbers of acid-fast-positive, rod-shaped bacteria were observed within the necrotic centers of well-demarcated, multifocal granulomas in gonads, liver, and peritoneum from affected fish. Mycobacterium abscessus and M. chelonae were isolated and identified biochemically. Definitive diagnosis in these outbreaks was obtained by culture on selective media. Because Mycobacterium spp. grow extremely slowly and positive confirmation may require 45 to 60 days, Mycobacterium species-specific polymerase chain reaction analysis was used to provide a rapid screening assay for Mycobacterium spp. as well as for verification of culture results. To our knowledge, this is the first documentation of mycobacterial infection in laboratory-maintained zebrafish and provides guidelines for diagnosis, management, and prevention of atypical mycobacteriosis in laboratory zebrafish colonies.

Observations of a Protistan Disease Similar to QPX in Mercenaria mercenaria (Hard Clams) from the Coast of Massachusetts

During the summer and fall of 1995, in clam aquaculture leases at two locations on the coast of Massachusetts, significant mortalities were observed to occur primarily in 1½- to 2-year old hard clams (Mercenaria mercenaria, quahog) planted in the leases. Examination of hard clams from those leases suggested a parasite similar to QPX (Quahog Parasite Unknown), as described by Whyte, S. K., Cawthorn, R. J., and McGladdery, S. E. (1994, Can. Dis. Ag. Org. 19, 129-136), was responsible for the poor condition of affected clams and the resulting high mortality. In clam tissues, the QPX-like parasite formed thalli surrounded by a mucofilamentous net. Larger thalli underwent endosporulation producing sporangia that contained approximately 40 endospores. Rupture of the sporangium released the endospores into the surrounding tissue. Inflammatory response by the clam to infection was chronic, active, and granulomatous. The occurrence both of phagocytic multinucleated giant inflammatory cells and inflammatory encapsulation of parasites were commonly identified as part of the response. Moderately to severely infected clams showed significantly reduced growth and a poorer condition index as compared to uninfected or mildly infected clams. Copyright 1998 Academic Press. Copyright 1998 Academic Press

Correlation of 2,3,7,8-tetrachlorodibenzo-p-dioxin induction of cytochrome P4501A in vascular endothelium with toxicity in early life stages of lake trout.

Edema and cardiovascular dysfunction occur in vertebrates exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) during early development. This study examined cytochrome P4501A (CYP1A) induction in endothelium and its possible association with mortality due to the edema and vascular effects of TCDD in lake trout early life stages. Lake trout (Salvelinus namaycush) eggs were injected at 24-50 hr postfertilization with 0.2 microl of 50 mM phosphatidylcholine liposomes or liposomes containing TCDD to give seven doses ranging from 11 to 176 pg TCDD/g egg. Doses of TCDD greater than 44 pg/g egg elicited hemorrhages; yolk sac, pericardial, and meningial edema; craniofacial malformations; regional ischemia; growth retardation; and mortality at the sac fry stage of development. Expression of CYP1A was assessed at four developmental stages, by immunohistochemical analysis of serial sections of individual fish with monoclonal antibody 1-12-3 to teleost CYP1A. CYP1A staining occurred in endothelial cells of many organs of TCDD-exposed but not vehicle-exposed embryos at 1 week prehatch and sac fry at 2 weeks posthatch. Earlier developmental stages examined were negative for CYP1A expression at any dose of TCDD. The strongest response occurred in sac fry at TCDD doses greater than 88 pg TCDD/g egg but was detected at doses as low as 22 pg TCDD/g egg. CYP1A staining in endothelium appeared at lower doses and was stronger than that in other cell types, in both prehatch embryos and posthatch sac fry. Thus, the vascular system is a major initial site affected by TCDD in lake trout early life stages, and the vascular endothelium is a cell type uniquely sensitive to induction of CYP1A in these developing animals. Based on an index of immunohistochemical staining of CYP1A, endothelial CYP1A induction in sac fry by TCDD occurred with an ED50 of 64-69 pg TCDD/g egg, similar to the dose-response for mortality occurring during the sac fry stage of development (LD50 = 47 pg TCDD/g egg). The correlations seen here suggest that CYP1A or aryl hydrocarbon receptor (AhR) in the endothelium may be linked to early lesions that result in TCDD-induced vascular derangements leading to yolk sac, pericardial, and meningial edema that is associated with lake trout sac fry mortality, but the precise mechanism remains to be determined.