Mycobacterium salmoniphilum infection in farmed Atlantic salmon, Salmo salar L

Multiple greyish‐white visceral nodules containing abundant rapidly growing and acid‐fast bacteria, subsequently identified as Mycobacterium salmoniphilum, were detected in moribund and newly dead market‐sized fish during a period of increased mortality in an Atlantic salmon, Salmo salar, farm in western Norway. Isolates cultured from diseased fish were phenotypically consistent with Mycobacterium sp. previously isolated from Atlantic salmon [MT 1890 (= NCIMB13533), MT1892, MT1900 and MT1901] in the Shetland Isles, Scotland. Partial sequences of 16S rDNA, ribosomal RNA internal transcribed spacer (ITS1), 65‐kDa heat‐shock protein (Hsp65) and β subunit of RNA polymerase (rpoB) revealed 97‐99% similarity with M. salmoniphilum type strain ATCC 13758^T. The source of infection was not confirmed. Koch’s postulates were fulfilled following experimental challenge of Atlantic salmon with field isolate NVI6598 ( FJ616988 ). Mortality was recorded in experimentally infected fish; however, the infection remained subclinical in the majority of affected fish over the 131‐day challenge period.     

Protein displacement by DExH/D "RNA helicases" without duplex unwinding

Members of the DExH/D superfamily of nucleic acid-activated nucleotide triphosphatases are essential for virtually all aspects of RNA metabolism, including pre-messenger RNA splicing, RNA interference, translation, and nucleocytoplasmic trafficking. Physiological substrates for these enzymes are thought to be regions of double-stranded RNA, because several DExH/D proteins catalyze strand separation in vitro. These "RNA helicases" can also disrupt RNA-protein interactions, but it is unclear whether this activity is coupled to duplex unwinding. Here we demonstrate that two unrelated DExH/D proteins catalyze protein displacement independently of duplex unwinding. Therefore, the essential functions of DExH/D proteins are not confined to RNA duplexes but can be exerted on a wide range of ribonucleoprotein substrates.     

Fluorescence of muscle and connective tissue from cod and salmon

Autofluorescence of salmon and cod muscle was measured and compared with autofluorescence of collagen type I and type V. Similarities between fluorescence of fish muscle and collagen were found in that the same peaks were obtained around 390, 430, and 480 nm. These similarities are supported by principal component analyses. Texture and gaping score were predicted from the fluorescence spectra by partial least-squares regression. However, the predictions did not perform well. Relating fluorescence to the gaping score gave a prediction error of 0.91 and a correlation of 0.43 when measuring gaping on a scale from 0 to 5. There was no relation between texture and fluorescence spectra. Fluorescence of fish muscle could be related to the storage time. However, this relation seemed not to be induced by changes in collagen.     

Effect of CO2 enrichment on photosynthesis,growth and yield of tomato

Net photosynthesis of tomato plants was measured as CO₂ uptake in various light intensities, CO₂ concentrations, O₂ concentrations and temperatures during short-term experiments. Net photosynthesis increased significantly with increasing CO₂ concentration at all light intensities, even at the lowest one. The optimal temperature for photosynthesis increased with CO₂ enrichment. The changes in photosynthesis when the $\text{CO}{}_2\text{O}{}_2$ ratio was varied suggest that the effect of CO₂ enrichment is a result of a reduction in photorespiration.To determine whether the increase in photosynthesis caused by CO₂ enrichment would produce a greater yield, tomato plants were cultivated from seed to harvest in a tightly-closed greenhouse that was enriched with CO₂ continuously during the entire growing-period. The control greenhouse was ventilated by openings in the roof and door. The temperature of the 2 greenhouses was kept the same. Plants enriched with CO₂ showed a significant increase in fresh and dry weight and yield of tomatoes. The results are discussed in relation to earlier work in which CO₂ enrichment was discontinued when there was a need for ventilation.     

EPPO Study on the Risk of Imports of Plants for Planting: description and details of the first outcomes

EPPO member countries requested that a study be conducted to identify and better address the risks presented by the trade of plants for planting, which has led to numerous introductions of pests into the EPPO region in recent years. Concerns were raised about the efficacy of the current plant health systems in place in the EPPO region to deal with the risks presented by plants for planting. The EPPO Study on the Risk of Imports of Plants for Planting was launched by the EPPO Council in 2010. The first part of the Study was completed in spring 2012. It was published as EPPO Technical Document 1061 ( http://www.eppo.int/QUARANTINE/EPPO_Study_on_Plants_for_planting.pdf ). Examples of pest outbreaks in the EPPO region suspected to be caused by international trade of plants for planting were analyzed. This analysis identified characteristics of the pest/crop/trade patterns associated with the risks of importing pests. These characteristics are described as criteria that are intended to be used in a screening process to enable identification of commodities that require an assessment prior to import in the EPPO region. The further elaboration of the screening process is briefly outlined.     

A novel IS element, ISMpa1, in Mycobacterium avium subsp. paratuberculosis

A novel insertion element belonging to the IS110 family was identified in Mycobacterium avium subsp. paratuberculosis. The IS element, ISMpa1, is 1500 bp and has one ORF encoding a putative transposase. Three copies of ISMpa1 were identified in the M. avium subsp. paratuberculosis genome. The element had inserted into the 3' end of the highly conserved mycobacterial genes prrB and a homologue of M. tuberculosis Rv1593c, and between a putative cytochrome p450 oxygenase and a putative hydrolase. The IS element was present in all (n = 11) M. avium subsp. paratuberculosis strains but not detected in most other mycobacterial species examined, including 10 M. avium subsp. avium isolates of human, avian and porcine origin. However two porcine isolates of M. avium subsp. avium and the reference strain IWGMT49 did harbour ISMpa1. These three strains belong to a previously described subgroup of M. avium subsp. avium based on IS1245 restriction fragment length polymorphism (RFLP) pattern and serovars. All of the M. avium subsp. paratuberculosis strains examined had an identical RFLP pattern when probed with sequences corresponding to the 5' end of ISMpa1, whereas a different pattern was seen in the positive M. avium subsp. avium strains. This novel IS element might be a useful tool in strain classification of M. avium subsp. avium and also for the identification of M. avium subsp. paratuberculosis when used in combination with IS900.     

Detection of Mycobacterium avium subsp. paratuberculosis by buoyant density centrifugation, sequence capture PCR and dot blot hybridisation

Detection of Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) by polymerase chain reaction (PCR) is often hampered by the lack of efficient methods for sample treatment. We report a protocol for analysis of faecal samples based on buoyant density centrifugation in Percoll and IS900 sequence capture PCR combined with a dot blot assay for detection of low-grade infection of M. paratuberculosis. Serial dilutions of M. paratuberculosis genomic DNA and M. paratuberculosis bacteria were used to assess the sensitivity of the method. The final evaluation was performed with spiked faecal samples, which also were analysed by culture. The presence of PCR inhibitory substances in processed faecal samples was evaluated by including a PCR internal control. By using buoyant density centrifugation, sequence capture PCR, and dot blot hybridisation, we achieved a sensitivity of 10(3)CFU (colony forming units)/g of faeces. The detection limit by culture was assessed to 10(2)CFU/g of faeces. We conclude that the described protocol is a fast and sensitive alternative to bacterial culture of faecal samples.     

Efficient Screening of Marine Extracts for Protease Inhibitors by Combining FRET Based Activity Assays and Surface Plasmon Resonance Spectroscopy Based Binding Assays

The screening of extracts from marine organisms is a widely used strategy to discover new drug leads. A common problem in the screening process is the generation of false positive hits through unspecific effects from the complex chemical composition of the crude extracts. In this study, we explored a combination of a fluorescence resonance energy transfer (FRET) based activity assay and a surface plasmon resonance (SPR) based binding assay to avoid this problem. An aqueous extract was prepared from rest raw material of the Norwegian spring spawning herring, and further fractionated by methanol solubility and solid phase extraction. FRET based activity assays were used to determine the influence of each extract on the activity of different proteases. Several extracts showed more than 50% inhibition. The inhibition mechanisms were elucidated by SPR based competition experiments with known inhibitors. For the secreted aspartic proteases 1, 2, 3 and HIV-1 protease, the results indicated that some extracts contain inhibitors interacting specifically with the active site of the enzymes. The study shows that a combination of an activity assay and an SPR based binding assay is a powerful tool to identify potent inhibitors in marine extracts. Furthermore, the study shows that marine vertebrates offer an interesting source for new bioactive compounds, although they have rarely been explored for this purpose.