Pregnancy-related pharmacokinetic changes.

The pharmacokinetics of many drugs are altered by pregnancy. Drug distribution and protein binding are changed by pregnancy. While some drug metabolizing enzymes have an apparent increase in activity, others have an apparent decrease in activity. Not only is drug metabolism affected by pregnancy, but renal filtration is also increased. In addition, pregnancy alters the apparent activities of multiple drug transporters resulting in changes in the net renal secretion of drugs.

In vitro cultivation of Hematodinium sp. isolated from Atlantic snow crab, Chionoecetes opilio: partial characterization of late developmental stages.

Hematodinium is a parasitic dinoflagellate of numerous crustacean species, including the economically important Atlantic snow crab, Chionoecetes opilio. The parasite was cultured in vitro in modified Nephrops medium at 0 °C and a partial characterization of the life stages was accomplished using light and transmission electron microscopy (TEM). In haemolymph from heavily infected snow crabs two life stages were detected; amoeboid trophonts and sporonts. During in vitro cultivation, several Hematodinium sp. life stages were observed: trophonts, clump colonies, sporonts, arachnoid sporonts, sporoblasts and dinospores. Cultures initiated with sporonts progressed to motile dinospores; however, those initiated with amoeboid trophonts proliferated, but did not progress or formed schizont-like stages which were senescent artefacts. Plasmodial stages were associated with both trophonts and sporonts and could be differentiated by the presence of trichocysts on TEM. Macrodinospores were observed but not microdinospores; likely due to the low number of Hematodinium sp. cultures that progressed to the dinospore stage. No early life stages including motile filamentous trophonts or gorgonlocks were observed as previously noted in Hematodinium spp. from other crustacean hosts. All Hematodinium sp. life stages contained autofluorescent, membrane-bound electron dense granules that appeared to degranulate or be expelled from the cell during in vitro cultivation.

Ultrastructure of trichocysts in Hematodinium spp. infecting Atlantic snow crab, Chionoecetes opilio.

Trichocyst morphology and development were explored using transmission electron microscopy in Hematodinium spp. isolated directly from Atlantic snow crab (Chionoecetes opilio) hemolymph and from in vitro cultures. Appearance of trichocysts defines the initiation of a morphological transition in the parasites life cycle from vegetative stage to the transmission stage. Trichocysts within sporonts were found in distinct clusters near the nucleus in close apposition to the Golgi. As cells transitioned to more mature dinospores however, trichocysts were found randomly distributed throughout the cytoplasm. Clusters contained both primordial and maturing trichocysts at various stages indicating an asynchronous development. The random distribution of mature trichocysts suggests deployment to the cell membrane for future extrusion. Mature trichocysts of Hematodinium spp. appeared structurally similar to trichocysts from photosynthetic dinoflagellates. Hematodinium spp. trichocysts differed by the presence of peripheral tubules associated with novel cuboidal appendages in the apical region rather than a network of central electron dense fibres as found in photosynthetic dinoflagellates. Additionally, the trichocyst membrane of Hematodinium spp. was in close apposition to the square crystalline core. Trichocyst expulsion was not observed during our study which along with features of development and maturation within Hematodinium life stages should provide insight into proposed roles in host attachment or defense that could further our understanding of the mechanisms of pathogenesis and transmission of the parasite.

Ichthyophonus-infected walleye pollock Theragra chalcogramma (Pallas) in the eastern Bering Sea: a potential reservoir of infections in the North Pacific.

In 2003, the Alaska walleye pollock industry reported product quality issues attributed to an unspecified parasite in fish muscle. Using molecular and histological methods, we identified the parasite in Bering Sea pollock as Ichthyophonus. Infected pollock were identified throughout the study area, and prevalence was greater in adults than in juveniles. This study not only provides the first documented report of Ichthyophonus in any fish species captured in the Bering Sea, but also reveals that the parasite has been present in this region for nearly 20 years and is not a recent introduction. Sequence analysis of 18S rDNA from Ichthyophonus in pollock revealed that consensus sequences were identical to published parasite sequences from Pacific herring and Yukon River Chinook salmon. Results from this study suggest potential for Ichthyophonus exposures from infected pollock via two trophic pathways; feeding on whole fish as prey and scavenging on industry-discharged offal. Considering the notable Ichthyophonus levels in pollock, the low host specificity of the parasite and the role of this host as a central prey item in the Bering Sea, pollock likely serve as a key Ichthyophonus reservoir for other susceptible hosts in the North Pacific.

Development and validation of a quantitative PCR assay for Ichthyophonus spp.

Members of the genus Ichthyophonus are trophically transmitted, cosmopolitan parasites that affect numerous fish species worldwide. A quantitative PCR (qPCR) assay specific for genus Ichthyophonus 18S ribosomal DNA was developed for parasite detection and surveillance. The new assay was tested for precision, repeatability, reproducibility, and both analytical sensitivity and specificity. Diagnostic sensitivity and specificity were estimated using tissue samples from a wild population of walleye pollock Theragra chalcogramma. Ichthyophonus sp. presence in tissue samples was determined by qPCR, conventional PCR (cPCR), and histology. Parasite prevalence estimates varied depending upon the detection method employed and tissue type tested. qPCR identified the greatest number of Ichthyophonus sp.-positive cases when applied to walleye pollock skeletal muscle. The qPCR assay proved sensitive and specific for Ichthyophonus spp. DNA, but like cPCR, is only a proxy for infection. When compared to cPCR, qPCR possesses added benefits of parasite DNA quantification and a 100-fold increase in analytical sensitivity. Because this novel assay is specific for known members of the genus, it is likely appropriate for detecting Ichthyophonus spp. DNA in various hosts from multiple regions. However, species-level identification and isotype variability would require DNA sequencing. In addition to distribution and prevalence applications, this assay could be modified and adapted for use with zooplankton or environmental samples. Such applications could aid in investigating alternate routes of transmission and life history strategies typical to members of the genus Ichthyophonus.

Protistan parasites as mortality drivers in cold water crab fisheries.

From a historical perspective, several protistan taxa, including the recently re-aligned Microsporidia, have been associated with or identified as causes of mortalities in crustacean populations. Depending upon the host species, associated protistan prevalences could be as low as 5% or approach 100%. It has generally been assumed that reported prevalences translated directly into significant mortalities that could impact the distribution and abundance of affected populations. However, this assumption may be incorrect especially when the dynamics of host-pathogen-environment interactions are not entirely understood. We will discuss the presumed impact of several protistan pathogens on temperate and cold water commercial crab species. By using selected examples such as a ciliate in the Dungeness crab (Cancer magister) and Hematodinium sp. infections in North Pacific crabs, we will attempt to contrast differences between prevalence and mortality, acute and chronic infections/mortalities, age or size selectivity of affected population, and geographically restricted and widespread epizootics. We will also briefly discuss the potential impact of environmental changes such as climate change and ocean acidification on both host and protistan pathogen.

Protistan diseases of commercially important crabs: a review.

Protists are a diverse group of eukaryotes that possess a unicellular level of organization. As unicellular organisms, the differentiation of cells into tissues does not occur, although when cell differentiation does occur, it is limited to sexual reproduction, alternate vegetative morphologies or quiescent life history stages. Protistan parasites may possess simple or complex life histories that are important factors to consider when investigating protistan diseases of decapods. Unfortunately, the life histories of many protistan parasites of decapods are insufficiently described, resulting in the fact that modes of infection and transmission are often unidentified. This is surprising considering the economic importance of many marine decapods and the ability of protistan parasites to produce significant, but generally transient and area limited mortalities. However, the marine disease landscape is changing and will continue to change as climate change and ocean acidification will play important roles in disease occurrence and distribution. As a result, the following discussion attempts to summarize current knowledge on several crab diseases, their protistan etiological agents, the impact of disease on economically important crab populations and draw attention to areas of needed research. The discussion is not complete as only selected diseases are addressed, or perfect as the Microsporidia are included in the discussion (a traditional error continued in this summary) despite the recent, but controversial placement of the taxon with the fungi.

Molecular detection of Hematodinium sp. in Northeast Pacific Chionoecetes spp. and evidence of two species in the Northern Hemisphere.

Hematodinium is a genus of parasitic dinoflagellates that infects crustaceans worldwide including Tanner crabs Chionoecetes bairdi and snow crabs C. opilio in the Northeast Pacific Ocean. The present study describes the optimization of a PCR-based assay for the detection and monitoring of Hematodinium sp. in snow and Tanner crabs. Two fragments, 1682 and 187 bp, were amplified from the 18S ribosomal DNA region of the parasite. The assay performed well in 6 additional decapod species (1 lobster and 5 crabs) infected with Hematodinium spp., suggesting that it could be used to detect Hematodinium spp. in other decapods. We also report Hematodinium spp. infections in the majid crab, Hyas coarctatus, and the lithodid crab, Lithodes couesi. Sequencing of 18S rDNA and the adjacent internal transcribed spacer 1 (ITS1) region of Hematodinium spp. isolated from 7 host species in the present study revealed the presence of 2 Hematodinium clades, one in the blue crab Callinectes sapidus and a second in all other host species. The ITS1 sequences of the 2 clades could not be aligned, but showed a conserved secondary structure that may be related to a functional diversification during a host switch. Comparison of our data with 18S and ITS1 sequence data available in GenBank placed the north Pacific Hematodinium sp. in a clade separate from the Hematodinium sp. infecting the portunoids, C. sapidus, Liocarcinus depurator and Scylla serrata, and within a second clade that infected all other decapod hosts located in the North Pacific and North Atlantic Oceans.

Discovery of a ciliate parasitoid of euphausiids off Oregon, USA: Collinia oregonensis n. sp. (Apostomatida: Colliniidae).

An apostome ciliate, Collinia oregonensis n. sp., is reported inhabiting the cephalothorax and abdomen of 3 euphausiid species from the Oregon-Washington coast: Euphausia pacifica Hansen, 1911, Thysanoessa spinifera Holmes, 1900, and Thysanoessa gregaria G.O. Sars, 1883. This ciliate is the 7th species described for the genus Collinia and the 2nd species known to infect euphausiids. Disease progression and ciliate morphology are described using (1) modified protargol stain, (2) hematoxylin counterstained with Fast Green, and (3) Scanning Electron Microscopy (SEM). All endoparasitic developmental stages (trophont, tomont, tomitogenesis, protomite, and tomite) of C. oregonensis are astomatous and possess between 14 and 22 kineties. C. oregonensis is smaller than C. beringensis Capriulo & Small, 1986, which infects the euphausiid Thysanoessa inermis Krøyer, 1846 in the Bering Sea and which possesses between 24 and 80 kineties. The ciliate is a parasitoid because it must kill the host to complete its life cycle. Infections and mortalities in multiple host species likely reflect the virulent nature of the ciliate. Adult euphausiids infected with this parasitoid possess a swollen and bright orange cephalothorax. C. oregonensis feeds and proliferates inside euphausiids, producing fulminating infections that rupture the cephalothorax and release large numbers of tomites into the surrounding water. After several hours in the free swimming stage under shipboard conditions in the present study, the tomites adhered to each other, forming filaments. Infection rates ranged between 3 and 20% within individual euphausiid aggregations, but infected aggregations were randomly and sparingly distributed. Infected euphausiids were found at 6.7% of 316 stations sampled during 3 summer cruises. No infected euphausiids were collected in winter. Because E. pacifica and T. spinifera account for about 90% of the euphausiid standing stock in the northern California Current System, this parasitoid ciliate may have a significant impact on euphausiid population abundance, distribution and secondary productivity.

Characterization of intracytoplasmic prokaryote infections in Dreissena sp. (Bivalvia: Dreissenidae).

This study characterizes intracytoplasmic infections with prokaryote microorganisms in Dreissena sp. (near Dreissena polymorpha) from northeastern Greece and represents the first report of such infections in freshwater bivalves. Light microscope observations of stained tissues revealed basophilic, cytoplasmic inclusion bodies in 87.5% (28/32) of the mussels sectioned. Inclusions in epithelial cells and connective tissues were noted, respectively, in 34.4 and 71.9% of the sample, with 5 mussels (15.6%) having both tissue types infected. Epithelial cell infections were observed in histological sections only in digestive gland tubules and ducts; within tubules, inclusions were present more often in secretory than digestive cells. Connective tissue infections, however, were systemic; among the 32 mussels sectioned, inclusions were found in the gills (65.6%), foot (12.5%), mantle (9.4%), labial palps (6.3%), digestive gland (6.3%), stomach (6.3%), and gonads (3.1%). Cytoplasmic inclusions (maximum dimension, 138 microm) were prominent enough in the gills to be visible in 17.0% of the 247 mussels dissected. Ultrastructurally, prokaryote cells in gill connective tissues were clearly characteristic of Chlamydiales-like organisms, with each intracytoplasmic inclusion containing a loosely packed mixture of elementary, reticulate, intermediate bodies, and blebs. Prokaryote colonies in digestive gland epithelial cells exclusively contained 1 of 4 morphological cell types and were considered Rickettsiales-like. Hexagonal, virus-like particles were present in the cytoplasm of the largest of these Rickettsiales-like prokaryotes. Although host stress was evident from localized cell necrosis and dense hemocyte infiltration, overall infection was fairly benign, with no major, adverse impact on body condition evident among sectioned or dissected mussels. A possible negative effect was partial constriction of gill water tubes, but at the infection intensity observed (typical range 1 to 7 inclusion bodies per section), significant interference with respiration and other metabolic functions of the gills was highly unlikely.