A recombinant CHSE-214 cell line expressing an Mx1 promoter-reporter system responds to both interferon type I and type II from salmonids and represents a versatile tool to study the IFN-system in teleost fish

A transgenic cell line for the detection of salmon interferons (IFNs) has been established. It is based on a CHSE-214 cell line containing a reporter construct expressing firefly luciferase under the control of the rainbow trout promoter for the IFN-induced Mx1 gene. This cell line, named CHSE-Mx10, showed IFN-induced luciferase expression after more than 80 passages, confirming the stability of this cell line. Interestingly, the Mx promoter was shown to respond to both salmon IFN-alpha/beta and trout IFN-gamma in a dose-dependent manner, while there was no response to TNF-alpha and IL-1beta. IFN-alpha/beta activity could be measured at a range of 9-150 U/ml, and IFN-gamma showed activity between 10 and 100 ng/ml. The reproducibility of both responses was good. The CHSE-Mx10 reporter system constitutes a versatile tool to study the induction and regulation of IFN signaling in teleost fish. A preliminary study presented herein suggests that both infectious pancreas necrosis virus (IPNV) and salmon pancreas disease virus (SPDV) may block activation of the Mx promoter in CHSE-Mx10 stimulated with IFN-alpha/beta.     

What is the Value of a Good Map? An Example Using High Spatial Resolution Imagery to Aid Riparian Restoration

Riparian areas contain structurally diverse habitats that are challenging to monitor routinely and accurately over broad areas. As the structural variability within riparian areas is often indiscernible using moderate-scale satellite imagery, new mapping techniques are needed. We used high spatial resolution satellite imagery from the QuickBird satellite to map harvested and intact forests in coastal British Columbia, Canada. We distinguished forest structural classes used in riparian restoration planning, each with different restoration costs. To assess the accuracy of high spatial resolution imagery relative to coarser imagery, we coarsened the pixel resolution of the image, repeated the classifications, and compared results. Accuracy assessments produced individual class accuracies ranging from 70 to 90% for most classes; whilst accuracies obtained using coarser scale imagery were lower. We also examined the implications of map error on riparian restoration budgets derived from our classified maps. To do so, we modified the confusion matrix to create a cost error matrix quantifying costs associated with misclassification. High spatial resolution satellite imagery can be useful for riparian mapping; however, errors in restoration budgets attributable to misclassification error can be significant, even when using highly accurate maps. As the spatial resolution of imagery increases, it will be used more routinely in ecosystem ecology. Thus, our ability to evaluate map accuracy in practical, meaningful ways must develop further. The cost error matrix is one method that can be adapted for conservation and planning decisions in many ecosystems.     

Immobilized pH gradient-driven paper-based IEF: a new method for fractionating complex peptide mixtures before MS analysis

Introduction  

The vast difference in the abundance of different proteins in biological samples limits the determination of the complete proteome of a cell type, requiring fractionation of proteins and peptides before MS analysis.     

Mitogenome rearrangement in the cold-water scleractinian coral Lophelia pertusa (Cnidaria, Anthozoa) involves a long-term evolving group I intron

Group I introns are genetic insertion elements that invade host genomes in a wide range of organisms. In metazoans, however, group I introns are extremely rare, so far only identified within mitogenomes of hexacorals and some sponges. We sequenced the complete mitogenome of the cold-water scleractinian coral Lophelia pertusa, the dominating deep sea reef-building coral species in the North Atlantic Ocean. The mitogenome (16,150 bp) has the same gene content but organized in a unique gene order compared to that of other known scleractinian corals. A complex group I intron (6460 bp) inserted in the ND5 gene (position 717) was found to host seven essential mitochondrial protein genes and one ribosomal RNA gene. Phylogenetic analysis supports a vertical inheritance pattern of the ND5-717 intron among hexacoral mitogenomes with no examples of intron loss. Structural assessments of the Lophelia intron revealed an unusual organization that lacks the universally conserved ωG at the 3′ end, as well as a highly compact RNA core structure with overlapping ribozyme and protein coding capacities. Based on phylogenetic and structural analyses we reconstructed the evolutionary history of ND5-717, from its ancestral protist origin, through intron loss in some early metazoan lineages, and into a compulsory feature with functional implications in hexacorals.     

Intracellular localisation and innate immune responses following Francisella noatunensis infection of Atlantic cod (Gadus morhua) macrophages

The facultative intracellular bacterium Francisella noatunensis causes francisellosis in Atlantic cod (Gadus morhua), but little is known about its survival strategies or how these bacteria evade the host immune response. In this study we show intracellular localisation of F. noatunensis in cod macrophages using indirect immunofluorescence techniques and green fluorescent labelled bacteria. Transmission electron microscopy revealed that F. noatunensis was enclosed by a phagosomal membrane during the initial phase of infection. Bacteria were at a later stage of the infection found in large electron-lucent zones, apparently surrounded by a partially intact or disintegrated membrane. Immune electron microscopy demonstrated the release of bacterial derived vesicles from intracellular F. noatunensis, an event suspected of promoting phagosomal membrane degradation and allowing escape of the bacteria to cytoplasm. Studies of macrophages infected with F. noatunensis demonstrated a weak activation of the inflammatory response genes as measured by increased expression of the Interleukin (IL)-1β and IL-8. In comparison, a stronger induction of gene expression was found for the anti-inflammatory IL-10 indicating that the bacterium exhibits a role in down-regulating the inflammatory response. Expression of the p40 subunit of IL-12/IL-17 genes was highly induced during infection suggesting that F. noatunensis promotes T cell polarisation. The host macrophage responses studied here showed low ability to distinguish between live and inactivated bacteria, although other types of responses could be of importance for such discriminations. The immunoreactivity of F. noatunensis lipopolysaccharide (LPS) was very modest, in contrast to the strong capacity of Escherichia coli LPS to induce inflammatory responsive genes. These results suggest that F. noatunensis virulence mechanisms cover many strategies for intracellular survival in cod macrophages.