Claire L. Vogan

Prostaglandins in non-insectan invertebrates: recent insights and unsolved problems

SUMMARY Prostaglandins (PG) are oxygenated derivatives of C20 polyunsaturated fatty acids including arachidonic and eicosapentaenoic acids. In mammals, these compounds have been shown to play key roles in haemostasis, sleep-wake regulation, smooth muscle tone, and vaso-, temperature and immune regulation. In invertebrates, PGs have been reported to perform similar roles and are involved in the control of oogenesis and spermatogenesis, ion transport and defence. Although there is often a detailed understanding of the actions of these compounds in invertebrates such as insects, knowledge of their mechanism of biosynthesis is often lacking. This account provides a critical review of our current knowledge on the structure and modes of biosynthesis of PGs in invertebrates, with particular reference to aquatic invertebrates. It emphasises some of the most recent findings, which suggest that some PGs have been misidentified. Prostaglandins in invertebrates can be categorised into two main types; the classical forms, such as PGE2 and PGD2 that are found in mammals, and novel forms including clavulones, bromo- and iodo-vulones and various PGA2 and PGE2 esters. A significant number of reports of PG identification in invertebrates have relied upon methods such as enzyme immunoassay that do not have the necessary specificity to ensure the validity of the identification. For example, in the barnacle Balanus amphitrite, although there are PG-like compounds that bind to antibodies raised against PGE2, mass spectrometric analysis failed to confirm the presence of this and other classical PGs. Therefore, care should be taken in drawing conclusions about what PGs are formed in invertebrates without employing appropriate analytical methods. Finally, the recent publication of the Ciona genome should facilitate studies on the nature and mode of biosynthesis of PGs in this advanced deuterostomate invertebrate.

Shell disease syndrome in the edible crab, Cancer pagurus--isolation, characterization and pathogenicity of chitinolytic bacteria.

Chitinolytic bacteria are believed to be the primary aetiological agents of shell disease syndrome in marine crustaceans. The disease principally results from the breakdown of their chitinous exoskeletons by the shell disease pathogens, but pathogenicity may also manifest internally should a breach of the carapace occur. The current study looks at the pathogenicity of a number of bacteria (predominantly from the genus Vibrio) isolated from the edible crab, Cancer pagurus. All chitinase-producing bacteria investigated were capable of growth in a minimal medium consisting of chitin powder from crab shells, but differed in their speed of growth and nature of chitinolytic activity, suggesting that they have different roles within the lesion community. Two isolates (designated I4 and I7) were chosen for studies on internal pathogenicity, which included the effect of the pathogen on crab tissues, the ability of the host to remove the bacteria from circulation and the antibacterial activity of crab blood. Initially, I4 was rapidly removed from circulation, but began to reappear in the blood after 24 h. By 100 h, 100% of crabs were moribund. The septicaemic effects of the isolate were reflected in the low levels of its killing by blood-cell lysate and serum. By contrast, I7 was only slowly removed from circulation and caused the rapid mortality of all crabs in <3 h. A large decline in the number of circulating blood cells following injection of I7 was mirrored by an accumulation of these cells in the gills. Initial experiments suggest that the death of the crabs following injection with I7 may be caused by toxic extracellular bacterial products that exert their effects on the blood cells and nervous system of the crabs.

Effects of shell disease syndrome on the haemocytes and humoral defences of the edible crab, Cancer pagurus

Crustaceans affected by shell disease syndrome display characteristic black-spot lesions in regions of exoskeletal degradation. Vogan et al. [Dis. Aquat. Org. (2001)] have shown that the severity of shell disease in the edible crab, Cancer pagurus, correlates with an increase in haemocoelic bacterial infections and may therefore serve as an external marker for the internal health of the animal. Therefore, this present study examined if the same crabs with shell disease lesions and varying degrees of bacterial septicaemia also displayed differences in the haemogramme, haemolymph phenoloxidase activity, total protein, copper and urea, as well as haemolymph-derived antibacterial activity, compared to uninfected individuals. A strong correlation was found between the severity of shell disease and a reduction in serum protein, which was further indicative of haemocoelic infection. Levels of copper, urea, phenoloxidase, and antibacterial activity in the haemolymph showed no correlations with the proportion of exoskeletal lesion cover. However, the degree of melanisation of haemolymph samples taken from shell-diseased individuals was less than that seen in disease-free crabs. Total haemocyte counts were unaffected by the disease, although some minor changes were found in the differential counts. Overall, despite having intrahaemocoelic infections, shell disease-affected individuals displayed few changes in the cellular or humoral defence parameters examined.

Hepoxilins and trioxilins in barnacles: an analysis of their potential roles in egg hatching and larval settlement

SUMMARY The barnacle life cycle has two key stages at which eicosanoids are believed to be involved in cellular communication pathways, namely the hatching of nauplii and the settlement of cypris larvae. Barnacle egg-hatching activity has previously been reported to reside in a variety of eicosanoids, including 8-hydroxyeicosapentaenoic acid and a number of tri-hydroxylated polyunsaturated fatty acid derivatives, the trioxilins. The production of the eicosapentaenoic acid metabolite trioxilin A4 (8,11,12-trihydroxy-5,9,14,17-eicosatetraenoic acid) by the barnacles Balanus amphitrite and Elminius modestus was confirmed using a combination of high-performance liquid chromatography and gas chromatography, both linked to mass spectrometry. In addition, both species also generated trioxilin A3 (8,11,12-trihydroxy-5,9,14-eicosatrienoic acid; an arachidonic acid-derived product), 8,11,12-trihydroxy-9,14,17-eicosatrienoic acid (a ω3 analogue of trioxilin A3; derived from ω3 arachidonic acid) and 10,13,14-trihydroxy-4,7,11,16,19-docosapentaenoic acid (a docosahexaenoic acid-derived product). In contrast to earlier reports, trioxilin A3 had no E. modestus egg-hatching activity at any of the concentrations tested (10–9–10–6 mol l–1). The unstable epoxide precursor hepoxilin A3, however, caused significant levels of hatching at 10–6 mol l–1. Furthermore, the stable hepoxilin B3 analogue PBT-3 stimulated hatching at 10–7 mol l–1. Neither trioxilin A3, hepoxilin A3 or PBT-3 at 0.25–30 μmol l–1 served as settlement cues for B. amphitrite cypris larvae.

Increased Disease Calls for a Cost-Benefits Review of Marine Reserves

Marine reserves (or No-Take Zones) are implemented to protect species and habitats, with the aim of restoring a balanced ecosystem. Although the benefits of marine reserves are commonly monitored, there is a lack of insight into the potential detriments of such highly protected waters. High population densities attained within reserves may induce negative impacts such as unfavourable trophic cascades and disease outbreaks. Hence, we investigated the health of lobster populations in the UK’s Marine Conservation Zone (MCZ) at Lundy Island. Comparisons were made between the fished, Refuge Zone (RZ) and the un-fished, No-Take Zone (NTZ; marine reserve). We show ostensibly positive effects such as increased lobster abundance and size within the NTZ; however, we also demonstrate apparent negative effects such as increased injury and shell disease. Our findings suggest that robust cost-benefit analyses of marine reserves could improve marine reserve efficacy and subsequent management strategies.

Twelve tips for providing effective student support in undergraduate medical education

Abstract Medical students often require high levels of specialised institutional and personal support to facilitate success. Contributory factors may include personality type, course pressures and financial hardship. Drawing from research literature and the authors’ experience, 12 tips are listed under five subheadings: policy and systems; people and resources; students; delivering support; limits of support. The 12 tips provide guidance to organisations and individual providers that encourages implementation of good practice and helps them better visualise their role within the system. By following the tips, medical schools can make more effective provisions for the expected, diverse and sometimes specialist needs of their students. Schools must take a proactive, anticipatory approach to provide appropriately for their entire student body. This ensures that students receive the best quality support, are more likely to succeed and are adequately prepared for their medical careers.

Characterization and Molecular Epidemiology of a Fungal Infection of Edible Crabs (Cancer pagurus) and Interaction of the Fungus with the Dinoflagellate Parasite Hematodinium

ABSTRACT This study reports on an emerging fungal disease of the edible crab, Cancer pagurus. Juvenile (prerecruit) crabs were found to be subject to this disease condition during the months of May to September at two intertidal sites in South Wales, United Kingdom. Histopathology revealed that the fungi overwhelm the host response in the tissues, leading to progressive septicemia. The causative agent of this infection was isolated and grown in pure culture and was identified as a member of the Ophiocordyceps clade by sequencing of the small subunit of the fungal ribosomal DNA (rDNA). Of the crabs naturally infected with the fungus, 94% had a coinfection with the parasitic dinoflagellate Hematodinium species. To determine if there was any interaction between the two disease-causing agents, apparently fungus-free crabs, both with and without natural Hematodinium infections, were challenged with the fungal isolate. The presence of Hematodinium caused a significant reduction in fungal multiplication in the hemocoel of the crabs in comparison to that in Hematodinium-free individuals. Histopathology of coinfected crabs showed a systemic multiplication of Hematodinium within host tissues, leading to a rapid death, while Hematodinium-free crabs experimentally infected with the fungal isolate died due to fungal sepsis (septicemia) with the same characteristic pathology as seen in natural infections.

Parasitization of juvenile edible crabs (Cancer pagurus) by the dinoflagellate, Hematodinium sp.: pathobiology, seasonality and its potential effects on commercial fisheries

This study reports on the prevalence and severity of infections caused by the parasitic dinoflagellate, Hematodinium in juvenile edible crabs (Cancer pagurus) found in 2 intertidal survey sites (Mumbles Head and Oxwich Bay) in the Bristol Channel, UK. Crabs were assessed for the presence and severity of Hematodinium infections by the histological examination of infected tissues. Such infections were found to exhibit a seasonal trend in the 2 study areas with high numbers of animals (ca. 30%) infected in the spring to summer but with low severity. Conversely, in November only ca. 10% of crabs were infected but these animals had large numbers of parasites in their haemolymph and other tissues. At this time, the carapace and underlying tissues of infected crabs had the chalky, pinkish-orange appearance that is characteristic of this disease. Hematodinium-infected crabs ranged in size from 12 to 74 mm carapace width. Overall, it is concluded that the high prevalence of infection of juvenile crabs in this area may have implications for the sustainability of the edible crab fishery in the Bristol Channel.

A comparison of the structure of American (Homarus americanus) and European (Homarus gammarus) lobster cuticle with particular reference to shell disease susceptibility.

The integument of arthropods is an important first-line defence against the invasion of parasites and pathogens. Once damaged, this can be subject to colonisation by microbial agents from the surrounding environment, which in crustaceans can lead to a condition termed shell disease syndrome. This condition has been reported in several crustacean species, including crabs and lobsters. The syndrome is a progressive condition where the outer cuticle becomes pitted and eroded, and in extreme cases is compromised, leaving animals susceptible to septicaemia. This study examined the susceptibility of juvenile American (Homarus americanus) and European (Homarus gammarus) lobsters to shell disease, as a result of mechanical damage. Scanning electron microscopy was used as a method to identify differences in the cuticle structure and consequences of mechanical damage. Claw regions were aseptically punctured, whilst carapaces were abraded using sterile sandpaper, to mimic natural damage. After a period of between 10 and 12 weeks, lobsters were sacrificed, fixed and stored for later examination. The carapace and claws of juvenile American lobsters were shown to be thinner and more vulnerable to abrasion damage than their European counterparts. In addition, the number and distribution of setal pits and pore canal openings also differed between the two species of lobster. Mechanical damage resulted in the formation of shell disease lesions on the claw and carapace of both lobster species. However, American lobsters, unlike their European counterparts, had extensive bacterial colonisation on the margins of these lesions. Overall, it is concluded that the cuticle of the American lobster is more susceptible to damage and resulting microbial colonisation. This may have implications for susceptibility of both species of lobster to shell disease syndrome.

Bacterial septicaemia in prerecruit edible crabs, Cancer pagurus L.

Juvenile edible crabs, Cancer pagurus L., were surveyed from Mumbles Head and Oxwich Bay in South Wales, UK, and the number of heterotrophic bacteria and vibrios in the hemolymph was determined. The percentage of crabs with hemolymph containing bacteria was variable over the survey with higher numbers of animals affected in summer than in winter. Post-moult crabs contained significantly higher numbers of heterotrophic bacteria in the hemolymph than pre- and intermoult animals. Crabs with cuticular damage to the gills also had significantly higher numbers of bacteria in the hemolymph. Crabs were found to have a high prevalence of infection by the dinoflagellate, Hematodinium. Such animals had significantly fewer bacteria in the blood in comparison with Hematodinium-free animals. Of the 463 crabs surveyed, only 3 individuals had hemolymph containing 2000 + CFU mL(-1). Based on 16S rRNA gene sequences, two of these crabs contained a Vibrio pectenicida-like isolate, while the other had a mixed assemblage of vibrios. Although 59% of the crabs surveyed had culturable bacteria in the hemolymph, the majority only had small numbers (<2000 CFU mL(-1) ), suggesting that such infections may be of limited importance to the sustainability of the crab fishery in this region.