A Simple Model to Rank Shellfish Farming Areas Based on the Risk of Disease Introduction and Spread.

The European Union Council Directive 2006/88/EC requires that risk-based surveillance (RBS) for listed aquatic animal diseases is applied to all aquaculture production businesses. The principle behind this is the efficient use of resources directed towards high-risk farm categories, animal types and geographic areas. To achieve this requirement, fish and shellfish farms must be ranked according to their risk of disease introduction and spread. We present a method to risk rank shellfish farming areas based on the risk of disease introduction and spread and demonstrate how the approach was applied in 45 shellfish farming areas in England and Wales. Ten parameters were used to inform the risk model, which were grouped into four risk themes based on related pathways for transmission of pathogens: (i) live animal movement, (ii) transmission via water, (iii) short distance mechanical spread (birds) and (iv) long distance mechanical spread (vessels). Weights (informed by expert knowledge) were applied both to individual parameters and to risk themes for introduction and spread to reflect their relative importance. A spreadsheet model was developed to determine quantitative scores for the risk of pathogen introduction and risk of pathogen spread for each shellfish farming area. These scores were used to independently rank areas for risk of introduction and for risk of spread. Thresholds were set to establish risk categories (low, medium and high) for introduction and spread based on risk scores. Risk categories for introduction and spread for each area were combined to provide overall risk categories to inform a risk-based surveillance programme directed at the area level. Applying the combined risk category designation framework for risk of introduction and spread suggested by European Commission guidance for risk-based surveillance, 4, 10 and 31 areas were classified as high, medium and low risk, respectively.

PAH metabolites in fish bile: From the Seine estuary to Iceland.

Polycyclic aromatic hydrocarbons (PAH) are environmental contaminants that pose significant risk to health of fish. The International Workshop on Integrated Assessment of Contaminant Impacts on the North Sea (ICON) provided the framework to investigate biomarker responses as well as contaminant concentrations side by side in marine ecosystems. Concentrations of the main PAH metabolites 1-hydroxypyrene, 1-hydroxyphenanthren and 3-hydroxybenzo(a)pyrene were determined in bile by HPLC with fluorescence detection. Fish species under investigation were dab (Limanda limanda), flounder (Platichthys flesus) and haddock (Melanogrammus aeglefinus). A contamination gradient was demonstrated from the low contaminated waters of Iceland and off-shore regions of the North Sea towards higher concentrations in coastal areas. Concentrations of PAH metabolites differed primarily according to sampling region and secondarily to species.

F factor conjugation is a true type IV secretion system.

The F sex factor of Escherichia coli is a paradigm for bacterial conjugation and its transfer (tra) region represents a subset of the type IV secretion system (T4SS) family. The F tra region encodes eight of the 10 highly conserved (core) gene products of T4SS including TraAF (pilin), the TraBF, -KF (secretin-like), -VF (lipoprotein) and TraCF (NTPase), -EF, -LF and TraGF (N-terminal region) which correspond to TrbCP, -IP, -GP, -HP, -EP, -JP, DP and TrbLP, respectively, of the P-type T4SS exemplified by the IncP plasmid RP4. F lacks homologs of TrbBP (NTPase) and TrbFP but contains a cluster of genes encoding proteins essential for F conjugation (TraFF, -HF, -UF, -WF, the C-terminal region of TraGF, and TrbCF) that are hallmarks of F-like T4SS. These extra genes have been implicated in phenotypes that are characteristic of F-like systems including pilus retraction and mating pair stabilization. F-like T4SS systems have been found on many conjugative plasmids and in genetic islands on bacterial chromosomes. Although few systems have been studied in detail, F-like T4SS appear to be involved in the transfer of DNA only whereas P- and I-type systems appear to transport protein or nucleoprotein complexes. This review examines the similarities and differences among the T4SS, especially F- and P-like systems, and summarizes the properties of the F transfer region gene products.

Isolation of a rhabdovirus during outbreaks of disease in cyprinid fish species at fishery sites in England.

A virus was isolated during disease outbreaks in bream Abramis brama, tench Tinca tinca, roach Rutilis rutilis and crucian carp Carassius carassius populations at 6 fishery sites in England in 1999. Mortalities at the sites were primarily among recently introduced fish and the predominant fish species affected was bream. The bream stocked at 5 of the 6 English fishery sites were found to have originated from the River Bann, Northern Ireland. Most fish presented few consistent external signs of disease but some exhibited clinical signs similar to those of spring viraemia of carp (SVC), with extensive skin haemorrhages, ulceration on the flanks and internal signs including ascites and petechial haemorrhages. The most prominent histopathological changes were hepatocellular necrosis, interstitial nephritis and splenitis. The virus induced a cytopathic effect in tissue cultures (Epithelioma papulosum cyprini [EPC] cells) at 20 degrees C and produced moderate signals in an enzyme immunoassay (EIA) for the detection of SVC virus. The virus showed a close serological relationship to pike fry rhabdovirus in both EIA and serum neutralisation assays and to a rhabdovirus isolated during a disease outbreak in a bream population in the River Bann in 1998. A high degree of sequence similarity (> or = 99.5% nucleotide identity) was observed between the English isolates and those from the River Bann. Experimental infection of juvenile bream, tench and carp with EPC cell-grown rhabdovirus by bath and intraperitoneal injection resulted in a 40% mortality of bream in the injection group only. The virus was re-isolated from pooled kidney, liver and spleen tissue samples from moribund bream. The field observations together with the experimental results indicate that this rhabdovirus is of low virulence but may have the potential to cause significant mortality in fishes under stress.

Paralytic shellfish poisoning toxins induce xenobiotic metabolising enzymes in Atlantic salmon (Salmo salar).

Paralytic shellfish poisoning (PSP) toxins have been implicated as the causative agent of a number of fish kills. Exposure experiments indicate that fish are susceptible to PSPs by intraperitoneal (i.p.) and oral administration, while sampling of fish affected by toxic blooms reveals that these toxins can be accumulated. In spite of the potential impact to marine fisheries, little research has been conducted on the potential metabolism and detoxification of PSPs in marine fishes. Previous work by this group has shown that the xenobiotic metabolising enzyme (XME) cytochrome P-450 (CYP1A) is induced in Atlantic salmon (Salmo salar) following i.p. exposure to saxitoxin (STX). Salmon injected i.p. with sub-lethal doses of STX show a four- to eight-fold induction of hepatic CYP1A (as shown by ethoxyresorufin-O-deethylase activity) over controls after 96 h. Results presented here show that the phase II XME glutathione S-transferase (GST) is also induced in salmon following PSP exposure. Post smolts were exposed to three injections of PSPs (2 micrograms STXeq/kg) over 21 days. Injection of both STX and PSPs extracted from a toxic strain of dinoflagellate (Alexandrium fundyense, CCMP 1719) resulted in induction of hepatic GST, as measured by activity for 1-chloro 2,4-dinitrobenzene. Such inductions indicate a potential role for XMEs in PSP metabolism. Possible roles for other enzymes are also discussed.

Crystal structure of the bacterial conjugation repressor finO.

The conjugative transfer of F-like plasmids is repressed by FinO, an RNA binding protein. FinO interacts with the F-plasmid encoded traJ mRNA and its antisense RNA, FinP, stabilizing FinP against endonucleolytic degradation and facilitating sense-antisense RNA recognition. Here we present the 2.0 A resolution X-ray crystal structure of FinO, lacking its flexible N-terminal extension. FinO adopts a novel, elongated, largely helical conformation. An N-terminal region, previously shown to contact RNA, forms a positively charged alpha-helix (helix 1) that protrudes 45 A from the central core of FinO. A C-terminal region of FinO that is implicated in RNA interactions also extends out from the central body of the protein, adopting a helical conformation and packing against the base of the N-terminal helix. A highly positively charged patch on the surface of the FinO core may present another RNA binding surface. The results of an in vitro RNA duplexing assay demonstrate that the flexible N-terminal region of FinO plays a key role in FinP-traJ RNA recognition, and supports our proposal that this region and the N-terminus of helix 1 interact with and stabilize paired, complementary RNA loops in a kissing complex.

The FinO repressor of bacterial conjugation contains two RNA binding regions.

Conjugative transfer of F-like plasmids in Escherichia coli is repressed by a plasmid-encoded protein, FinO. FinO blocks the translation of TraJ, a positive activator of transcription of genes required for conjugation. FinO binds a traJ antisense RNA, FinP, thereby protecting it from degradation, and catalyzes FinP-traJ mRNA hybridization. Interactions between these two RNAs are predicted to block the traJ ribosomal binding site. In this paper, we use limited proteolysis, circular dichroism spectroscopy, and an electrophoretic mobility shift assay to map the regions within FinO that are required for interactions with RNA. Our results show that FinO is largely helical, binds to its highest affinity binding site within FinP as a monomer, and contains two distinct RNA binding regions, one of which is localized between residues 26 and 61, and a second which is localized between residues 62 and 186.