Atlantic salmon paramyxovirus (ASPV) infection contributes to proliferative gill inflammation (PGI) in seawater-reared Salmo salar

Proliferative gill inflammation (PGI) causes significant losses in farmed Atlantic salmon Salmo salar L. in Norway, especially during the first months following seawater transfer. The aetiology is apparently multifactorial, including infection with chlamydia-like bacteria and Atlantic salmon paramyxovirus (ASPV). In the present study, gills from diseased fish from 3 farms on the western coast of Norway were sampled. The pathological changes were briefly described and the aetiological significance of ASPV studied by immunofluorescent staining of cryosections and by immunohistochemistry on sections of formalin-fixed and paraffin-embedded tissue. The pathological changes were macroscopically characterized by palour of the gills, and histologically by inflammation, circulatory disturbances, cell death and epithelial cell proliferation. ASPV was demonstrated in fish from all farms studied, as immunostaining consistent with ASPV was obtained in lamellar epithelial and endothelial cells of pathologically altered tissues. It is concluded that ASPV is at least a contributing cause of PGI. As far as we know, this is the first demonstration of fish disease related to infection with a paramyxovirus.     

Distinct mechanisms of CD4+ and CD8+ T-cell activation and bystander apoptosis induced by human immunodeficiency virus type 1 virions

Apoptosis of uninfected bystander T cells contributes to T-cell depletion during human immunodeficiency virus type 1 (HIV-1) infection. HIV-1 envelope/receptor interactions and immune activation have been implicated as contributors to bystander apoptosis. To better understand the relationship between T-cell activation and bystander apoptosis during HIV-1 pathogenesis, we investigated the effects of the highly cytopathic CXCR4-tropic HIV-1 variant ELI6 on primary CD4(+) and CD8(+) T cells. Infection of primary T-cell cultures with ELI6 induced CD4(+) T-cell depletion by direct cell lysis and bystander apoptosis. Exposure of primary CD4(+) and CD8(+) T cells to nonreplicating ELI6 virions induced bystander apoptosis through a Fas-independent mechanism. Bystander apoptosis of CD4(+) T cells required direct contact with virions and Env/CXCR4 binding. In contrast, the apoptosis of CD8(+) T cells was triggered by a soluble factor(s) secreted by CD4(+) T cells. HIV-1 virions activated CD4(+) and CD8(+) T cells to express CD25 and HLA-DR and preferentially induced apoptosis in CD25(+)HLA-DR(+) T cells in a CXCR4-dependent manner. Maximal levels of binding, activation, and apoptosis were induced by virions that incorporated MHC class II and B7-2 into the viral membrane. These results suggest that nonreplicating HIV-1 virions contribute to chronic immune activation and T-cell depletion during HIV-1 pathogenesis by activating CD4(+) and CD8(+) T cells, which then proceed to die via apoptosis. This mechanism may represent a viral immune evasion strategy to increase viral replication by activating target cells while killing immune effector cells that are not productively infected.     

Thermo-optically Induced Reorganizations in the Main Light Harvesting Antenna of Plants. II. Indications for the Role of LHCII-only Macrodomains in Thylakoids

We have investigated the circular dichroism spectral transients associated with the light-induced reversible reorganizations in chirally organized macrodomains of pea thylakoid membranes and loosely stacked lamellar aggregates of the main chlorophyll a/b light harvesting complexes (LHCII) isolated from the same membranes. These reorganizations have earlier been assigned to originate from a thermo-optic effect. According to the thermo-optic mechanism, fast local thermal transients due to dissipation of the excess excitation energy induce elementary structural changes in the close vicinity of the dissipation [Cseh et al. (2000) Biochemistry 39: 15250–15257]. Here we show that despite the markedly different CD spectra in the dark, the transient (light-minus-dark) CD spectra associated with the structural changes induced by high light in thylakoids and LHCII are virtually indistinguishable. This, together with other close similarities between the two systems, strongly suggests that the gross short-term, thermo-optically induced structural reorganizations in the membranes occur mainly, albeit probably not exclusively, in the LHCII-only domains [Boekema et al. (2000) J Mol Biol 301: 1123–1133]. Hence, LHCII-only domains might play an important role in light adaptation and photoprotection of plants.     

E. coli virulence factor hemolysin induces neutrophil apoptosis and necrosis/lysis in vitro and necrosis/lysis and lung injury in a rat pneumonia model

Enteric gram-negative bacilli, such as Escherichia coli are the most common cause of nosocomial pneumonia. In this study a wild-type extraintestinal pathogenic strain of E. coli (ExPEC)(CP9) and isogenic derivatives deficient in hemolysin (Hly) and cytotoxic necrotizing factor (CNF) were assessed in vitro and in a rat model of gram-negative pneumonia to test the hypothesis that these virulence factors induce neutrophil apoptosis and/or necrosis/lysis. As ascertained by in vitro caspase-3/7 and LDH activities and neutrophil morphology, Hly mediated neutrophil apoptosis at lower E. coli titers (1 x 10(5-6) cfu) and necrosis/lysis at higher titers (> or =1 x 10(7) cfu). Data suggest that CNF promotes apoptosis but not necrosis or lysis. We also demonstrate that annexin V/7-amino-actinomycin D staining was an unreliable assessment of apoptosis using live E. coli. The use of caspase-3/7 and LDH activities and neutrophil morphology supported the notion that necrosis, not apoptosis, was the primary mechanism by which neutrophils were affected in our in vivo gram-negative pneumonia model using live E. coli. In addition, in vivo studies demonstrated that Hly mediates lung injury. Neutrophil necrosis was not observed when animals were challenged with purified lipopolysaccharide, demonstrating the importance of using live bacteria. These findings establish that Hly contributes to ExPEC virulence by mediating neutrophil toxicity, with necrosis/lysis being the dominant effect of Hly on neutrophils in vivo and by lung injury. Whether Hly-mediated lung injury is due to neutrophil necrosis, a direct effect of Hly, or both is unclear.     

Surface properties of sulfur- and ether-linked phosphonolipids with and without purified hydrophobic lung surfactant proteins

Two novel C16:0 sulfur-linked phosphonolipids (S-lipid and SO(2)-lipid) and two ether-linked phosphonolipids (C16:0 DEPN-8 and C16:1 UnDEPN-8) were studied for surface behavior alone and in mixtures with purified bovine lung surfactant proteins (SP)-B and/or SP-C. Synthetic C16:0 phosphonolipids all had improved adsorption and film respreading compared to dipalmitoyl phosphatidylcholine, and SO(2)-lipid and DEPN-8 reached maximum surface pressures of 72mN/m (minimum surface tensions of <1mN/m) in compressed films on the Wilhelmy balance (23 degrees C). Dispersions of DEPN-8 (0.5mg/ml) and SO(2)-lipid (2.5mg/ml) also reached minimum surface tensions of <1mN/m on a pulsating bubble surfactometer (37 degrees C, 20cycles/min, 50% area compression). Synthetic lung surfactants containing DEPN-8 or SO(2)-lipid+0.75% SP-B+0.75% SP-C had dynamic surface activity on the bubble equal to that of calf lung surfactant extract (CLSE). Surfactants containing DEPN-8 or SO(2)-lipid plus 1.5% SP-B also had very high surface activity, but less than when both apoproteins were present together. Adding 10wt.% of UnDEPN-8 to synthetic lung surfactants did not improve dynamic surface activity. Surfactants containing DEPN-8 or SO(2)-lipid plus 0.75% SP-B/0.75% SP-C were chemically and biophysically resistant to phospholipase A(2) (PLA(2)), while CLSE was severely inhibited by PLA(2). The high activity and inhibition resistance of synthetic surfactants containing DEPN-8 or SO(2)-lipid plus SP-B/SP-C are promising for future applications in treating surfactant dysfunction in inflammatory lung injury.     

Human transformations of the Wadden Sea ecosystem through time: a synthesis

Todays Wadden Sea is a heavily human-altered ecosystem. Shaped by natural forces since its origin 7,500 years ago, humans gradually gained dominance in influencing ecosystem structure and functioning. Here, we reconstruct the timeline of human impacts and the history of ecological changes in the Wadden Sea. We then discuss the ecosystem and societal consequences of observed changes, and conclude with management implications. Human influences have intensified and multiplied over time. Large-scale habitat transformation over the last 1,000 years has eliminated diverse terrestrial, freshwater, brackish and marine habitats. Intensive exploitation of everything from oysters to whales has depleted most large predators and habitat-building species since medieval times. In the twentieth century, pollution, eutrophication, species invasions and, presumably, climate change have had marked impacts on the Wadden Sea flora and fauna. Yet habitat loss and overexploitation were the two main causes for the extinction or severe depletion of 144 species (~20% of total macrobiota). The loss of biodiversity, large predators, special habitats, filter and storage capacity, and degradation in water quality have led to a simplification and homogenisation of the food web structure and ecosystem functioning that has affected the Wadden Sea ecosystem and coastal societies alike. Recent conservation efforts have reversed some negative trends by enabling some birds and mammals to recover and by creating new economic options for society. The Wadden Sea history provides a unique long-term perspective on ecological change, new objectives for conservation, restoration and management, and an ecological baseline that allows us to envision a rich, productive and diverse Wadden Sea ecosystem and coastal society.     

Molecular cloning, functional expression in Escherichia coli and enzymatic characterisation of a cysteine protease from white clover (Trifolium repens)

This paper presents the cloning and biochemical characterisation of the cysteine protease Tr-cp 14 from white clover (Trifolium repens). The predicted amino acid sequence of Tr-cp 14 is 71%, 74% and 74% identical to the cysteine proteases XCP1 and XCP2 from Arabidopsis thaliana, and p48h-17 from Zinnia elegans, respectively. These cysteine proteases have previously been shown to be involved in programmed cell death during tracheary element differentiation. The precursor polypeptide of Tr-cp 14 was expressed in Escherichia coli, purified from inclusion bodies and refolded. The precursor polypeptide could be processed to its active mature form autocatalytically at pH 5.0 and had a requirement for 20 mM l-cysteine for optimal activity. Mature Tr-cp 14 showed a preference for synthetic aminomethylcoumarin substrates with either Leu or Phe in the P2 position when tested with Arg in P1. A substrate with Arg in both the P1 and P2 position was not accepted as substrate.     

Stress sensor triggers conformational response of the integral membrane protein microsomal glutathione transferase 1

Microsomal glutathione (GSH) transferase 1 (MGST1) is a trimeric, integral membrane protein involved in cellular response to chemical or oxidative stress. The cytosolic domain of MGST1 harbors the GSH binding site and a cysteine residue (C49) that acts as a sensor of oxidative and chemical stress. Spatially resolved changes in the kinetics of backbone amide H/D exchange reveal that the binding of a single molecule of GSH/trimer induces a cooperative conformational transition involving movements of the transmembrane helices and a reordering of the cytosolic domain. Alkylation of the stress sensor preorganizes the helices and facilitates the cooperative transition resulting in catalytic activation.