Structure of the Fe-heme in the homodimeric hemoglobin from Scapharca inaequivalvis and in the T72I mutant: an X-ray absorption spectroscopic study at low temperature

The Fe site structure in the recombinant wild-type and T721 mutant of the cooperative homodimeric hemoglobin (HbI) of the mollusc Scapharca itnaequivalvis has been investigated by measuring the Fe K-edge X-ray absorption near edge structure (XANES) spectra of their oxy, deoxy and carbonmonoxy derivatives, and the cryogenic photoproducts of the carbonmonoxy derivatives at T = 12 K. According to our results, the Fe site geometry in T72I HbI-CO is quite similar to that of human carbonmonoxy hemoglobin (HbA-CO), while in native HbI-CO it seems intermediate between that of HbA-CO and sperm whale MbCO. The XANES spectra of oxy and deoxy derivatives are similar to the homologous spectra of human HbA, except for T72I HbI, for which the absorption edge is blue-shifted (about + 1 eV) towards the spectrum of the oxy form. XANES spectra of the cryogenic photoproducts of HbA-CO (HbA*), HbI-CO (HbI*) and mutant HbI-CO (T72I HbI*) were acquired under continuous illumination at 12 K. The Fe-heme structures of the three photoproducts are similar; however, while in the case of HbA* and HbI* the data indicate incomplete structural relaxation of the Fe-heme towards its deoxy-like (T) form, the relaxation in T72I HbI* is almost completely towards the proposed "high affinity" Fe-heme structure of T72I HbI. This evidence suggests that minor tertiary restraints affect the Fe-heme dynamics of T72I HbI, corresponding to a reduction of the energy necessary for the T --> R structural transition, which can contribute to the observed dramatic enhancement in oxygen affinity of this hemoprotein, and the decreased cooperativity.     

Role of GM3-enriched microdomains in signal transduction regulation in T lymphocytes

Gangliosides, sialic acid containing glycosphigolipids, are ubiquitous constituents of cell plasma membranes. Each cell type shows a peculiar ganglioside expression pattern. In human T lymphocytes monosialoganglioside GM3 represents the main ganglioside constituent of cell plasma membrane where it is concentrated in glycosphingolipid-enriched microdomains (GEM). The presence of tyrosine kinase receptors, mono- (Ras, Rap) and heterotrimeric G proteins, Src-like tyrosine kinases (lck, lyn, fyn), PKC isozymes, glycosylphosphatidylinositol (GPI)-anchored proteins and, after T cell activation, the Syk-family kinase Zap-70, prompts these portions of the plasma membrane to be considered as “glycosignaling domains.” In particular, during T cell activation and/or other dynamic functions of the cell, such as apoptosis, key signaling molecules are recruited to these microdomains, where they strictly interact with GM3. The association of transducer proteins with GM3 in microdomains suggests that this ganglioside is the main marker of GEM in human lymphocytes and is a component of a cell plasma membrane multimolecular signaling complex involved in cell-cell interaction, signal transduction, and cell activation. Published in 2004. This revised version was published online in August 2006 with corrections to the Cover Date.     

Sarcocystis caninum and Sarcocystis svanai n. spp. (Apicomplexa: Sarcocystidae) Associated with Severe Myositis and Hepatitis in the Domestic Dog (Canis familiaris)

There are several reports of Sarcocystis sarcocysts in muscles of dogs, but these species have not been named. Additionally, there are two reports of Sarcocystis neurona in dogs. Here, we propose two new names, Sarcocystis caninum, and Sarcocystis svanai for sarcocysts associated with clinical muscular sarcocystosis in four domestic dogs (Canis familiaris), one each from Montana and Colorado in the USA, and two from British Columbia, Canada. Only the sarcocyst stage was identified. Most of the sarcocysts identified were S. caninum. Sarcocysts were studied using light microscopy, transmission electron microscopy (TEM), and polymerase chain reaction. Based on collective results two new species, S. caninum and S. svanai were designated. Sarcocystis caninum and S. svanai were structurally distinct. Sarcocystis caninum sarcocysts were up to 1.2 mm long and up to 75 μm wide. By light microscopy, the sarcocyst wall was relatively thin and smooth. By TEM, the sarcocyst wall was “type 9”, 1–2 μm thick, and contained villar protrusions that lacked microtubules. Bradyzoites in sections were 7–9 μm long. Sarcocysts of S. svanai were few and were identified by TEM. Sarcocystis svanai sarcocysts were “type 1”, thin walled (< 0.5 μm), and the wall lacked villar protrusions but had tiny blebs that did not invaginate. DNA was extracted either from infected frozen muscle biopsies or formalin‐fixed paraffin‐embedded sections. Dogs were either singly infected with S. caninum or multiply co‐infected with S. caninum and S. svanai (the result of a mixed infection) based on multilocus DNA sequencing and morphology. BLASTn analysis established that the sarcocysts identified in these dogs were similar to, but not identical to Sarcocystis canis or Sarcocystis arctosi, parasites found to infect polar bears (Ursus maritimus) or brown bears (Ursus arctosi), respectively. However, the S. caninum sequence showed 100% identify over the 18S rRNA region sequenced to that of S. arctica, a parasite known to infect Arctic foxes (Vulpes lagopus).     

TRAIL+ monocytes and monocyte‐related cells cause lung damage and thereby increase susceptibility to influenza–Streptococcus pneumoniae coinfection

Streptococcus pneumoniae coinfection is a major cause of influenza-associated mortality; however, the mechanisms underlying pathogenesis or protection remain unclear. Using a clinically relevant mouse model, we identify immune-mediated damage early during coinfection as a new mechanism causing susceptibility. Coinfected CCR2^−/− mice lacking monocytes and monocyte-derived cells control bacterial invasion better, show reduced epithelial damage and are overall more resistant than wild-type controls. In influenza-infected wild-type lungs, monocytes and monocyte-derived cells are the major cell populations expressing the apoptosis-inducing ligand TRAIL. Accordingly, anti-TRAIL treatment reduces bacterial load and protects against coinfection if administered during viral infection, but not following bacterial exposure. Post-influenza bacterial outgrowth induces a strong proinflammatory cytokine response and massive inflammatory cell infiltrate. Depletion of neutrophils or blockade of TNF-α facilitate bacterial outgrowth, leading to increased mortality, demonstrating that these factors aid bacterial control. We conclude that inflammatory monocytes recruited early, during the viral phase of coinfection, induce TRAIL-mediated lung damage, which facilitates bacterial invasion, while TNF-α and neutrophil responses help control subsequent bacterial outgrowth. We thus identify novel determinants of protection versus pathology in influenza–Streptococcus pneumoniae coinfection.     

Skeletal response of <Emphasis Type="Italic">Lophelia pertusa</Emphasis> (Scleractinia) to bioeroding sponge infestation visualised with micro-computed tomography

The skeleton morphology of the azooxanthellate cold-water coral Lophelia pertusa can be strongly influenced by invasive boring sponges that infest corallites in the still living part of the colony. Atypically swollen corallites of live Lophelia pertusa from the Galway Mound (Belgica Carbonate Mound Province, Porcupine Seabight, NE Atlantic), heavily excavated by boring organisms, have been examined with a wide range of non-destructive and destructive methods: micro-computed tomography, macro- and microscopic observations of the outer coral skeleton, longitudinal and transversal thin sections and SEM analyses of coral skeleton casts. As a result, three excavating sponge species have been distinguished within the coral skeleton: Alectona millari, Spiroxya heteroclita and Aka infesta. Furthermore, four main coral/sponge growth stages have been recognised: (1) cylindrical juvenile corallite/no sponge cavities; (2) flared juvenile corallite/linear sponge cavities (if present); (3) slightly swollen adult corallites/chambered oval sponge cavities; (4) very swollen adult corallites/widespread cavities. The inferred correlation between corallite morphology and boring sponge infestation has been detected in micro-computed tomography (micro-CT) images and confirmed in sponge trace casts and peculiar features of coral skeleton microstructure. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorised users.     

Inhibitory effect of Ocotea quixos (Lam.) Kosterm. and Piper aduncum L. essential oils from Ecuador on West Nile virus infection

West Nile virus (WNV) is a mosquito-borne flavivirus responsible of neuroinvasive manifestations. Natural products are well-known for their biological activities and pharmaceutical application. In this study, the inhibitory effects of essential oils (EOs) of Ocotea quixos (Lam.) Kosterm. and Piper aduncum L. on WNV replication were investigated. WNV was incubated with EOs before adsorption on Vero cells, viral replication was carried out in the absence or presence of EO. Cells were exposed to EO before the adsorption of untreated-virus. GC-MS and GC-FID were used for chemical characterization of EOs. Cell protection from infection was observed for both EOs. P. aduncum EO was characterized by dillapiole as main compound (48.21%) and O. quixos EO by 1,8-cineole (39.15%). Further investigations, such as the study of molecular and cellular mechanisms of action and in vivo evaluation, should be performed on these essential oils to derive new potential drugs against WNV.