Phylogenetic analysis of the outer-membrane-protein genes of Chlamydiae, and its implication for vaccine development

Examination of 18 complete and 6 partial sequences of the major outer- membrane protein from 24 chlamydiae isolates was used to reconstruct their evolutionary relationships. From this analysis, assuming that the clades with 100% bootstrap support are correct, come the following conclusions: (1) The tree of these sequences is not congruent with the phylogeny of the hosts, and thus host switching would seem to have occurred, thereby limiting the extent to which there has been coevolution of parasite and host. (2) The tree is also noncongruent with clustering by type of cell infected, thereby limiting the extent to which there has been coevolution of parasite and the cell type that it infects. (3) The tree is also noncongruent with clustering by the organ infected (eyes or genitalia), thereby limiting the extent to which there has been coevolution of parasite and the organ that it infects. (4) The tree is also noncongruent with genital strains arising from lymphogranuloma venereum strains. (5) The tree is also noncongruent with the geographic site at which the isolates were obtained, thereby limiting the extent of divergence explained by geographic separation. (6) There are estimated to be 185 amino acid positions that are invariable (as opposed to unvaried) in the major outer-membrane protein. There are 10 unvaried positions in the variable domains, of which 9 appear to be invariable, giving some reason to hope that development of a vaccine might be possible. (7) The rate of change of this protein is too small to see increased divergence over the time span of isolation of these genes, giving hope to any vaccine having longevity. Bootstrapping supports those portions of the tree on which the first five conclusions above depend. The picture that these results provide is more one of pathogen versatility than one of coevolutionary constraints. In addition, we examined 10 60-KDa, outer-membrane protein- 2 genes, all but one of which were from these same strains. The tree was not, among the trachomatis strains, congruent with the major-outer- membrane protein tree, suggesting that gene exchange could be occurring among strains. Moreover, there is an apparent slowdown in divergence in this gene, among the trachomatis strains.      

A Prominent Role for DC-SIGN+ Dendritic Cells in Initiation and Dissemination of Measles Virus Infection in Non-Human Primates

Measles virus (MV) is a highly contagious virus that is transmitted by aerosols. During systemic infection, CD150⁺ T and B lymphocytes in blood and lymphoid tissues are the main cells infected by pathogenic MV. However, it is unclear which cell types are the primary targets for MV in the lungs and how the virus reaches the lymphoid tissues. In vitro studies have shown that dendritic cell (DC) C-type lectin DC-SIGN captures MV, leading to infection of DCs as well as transmission to lymphocytes. However, evidence of DC-SIGN-mediated transmission in vivo has not been established. Here we identified DC-SIGN^hi DCs as first target cells in vivo and demonstrate that macaque DC-SIGN functions as an attachment receptor for MV. Notably, DC-SIGN^hi cells from macaque broncho-alveolar lavage and lymph nodes transmit MV to B lymphocytes, providing in vivo support for an important role for DCs in both initiation and dissemination of MV infection.     

EFFECTS OF MORINGA OLEIFERA SEED EXTRACT ON RUMEN FERMENTATION IN VITRO

Moringa oleifera is a pantropical tree of the family Moringaceae. A previously undescribed property of an aqueous extract from the seeds of this plant is the modulation of ruminal fermentation patterns, especially protein degradation, as demonstrated in a short-term batch incubation system. Gas, short chain fatty acids (SCFA) and cellulolytic enzyme activities were determined as general fermentation parameters. A dot blot assay able to directly detect true protein in rumen fluid samples was used to quantify protein degradation. For complex substrates the interpretation of protein degradation profiles was amended by polyacrylamide gel electrophoresis (PAGE) of the samples. When incubated with pure carbohydrates at a concentration of 1 mg ml^–1, the extract reduced microbial degradation of the model protein, bovine serum albumin (BSA), such that its concentration was at least 40% above the control after 12 h of incubation. Total protein degradation was thus delayed by approximately 9 h. When fermented along with wheat straw, leaf protein (Rubisco) was almost entirely protected during 12 h of fermentation. The degradation of soy proteins was retarded by at least 4- 6 h, depending on the protein band. There were strong side effects on the fermentation of pure cellulose (SCFA yield -60% after 12 h), whereas cellobiose and starch fermentation were less affected (-18 and -8%, respectively). When the complex substrates were fermented, SCFA yield was reduced by approximately 30% after 12 h. In our work we clearly demonstrate the efficacy of the new substance, which is neither a tannin nor a saponin, in an in vitro system, using pure as well as complex substrates. The properties shown in vitro for the crude extract suggest that it could have a positive effect on the protein metabolism of ruminants under intensive management and that negative side effects can be overcome by an optimized dosage. If the chemical nature of the active substance and its mechanism of action can be clarified, it may provide an alternative to replace critical synthetic feed additives (such as antibiotics) for high yielding dairy cows.     

Correlative light and electron microscopy imaging of autophagy in a zebrafish infection model

High-resolution imaging of autophagy has been used intensively in cell culture studies, but so far it has been difficult to visualize this process in detail in whole animal models. In this study we present a versatile method for high-resolution imaging of microbial infection in zebrafish larvae by injecting pathogens into the tail fin. This allows visualization of autophagic compartments by light and electron microscopy, which makes it possible to correlate images acquired by the 2 techniques. Using this method we have studied the autophagy response against Mycobacterium marinum infection. We show that mycobacteria during the progress of infection are frequently associated with GFP-Lc3-positive vesicles, and that 2 types of GFP-Lc3-positive vesicles were observed. The majority of these vesicles were approximately 1 μm in size and in close vicinity of bacteria, and a smaller number of GFP-Lc3-positive vesicles was larger in size and were observed to contain bacteria. Quantitative data showed that these larger vesicles occurred significantly more in leukocytes than in other cell types, and that approximately 70% of these vesicles were positive for a lysosomal marker. Using electron microscopy, it was found that approximately 5% of intracellular bacteria were present in autophagic vacuoles and that the remaining intracellular bacteria were present in phagosomes, lysosomes, free inside the cytoplasm or occurred as large aggregates. Based on correlation of light and electron microscopy images, it was shown that GFP-Lc3-positive vesicles displayed autophagic morphology. This study provides a new approach for injection of pathogens into the tail fin, which allows combined light and electron microscopy imaging in vivo and opens new research directions for studying autophagy process related to infectious diseases.     

Anti-carbamylated protein antibodies in the pre-symptomatic phase of rheumatoid arthritis,their relationship with multiple anti-citrulline peptide antibodies and association with radiological damage

IntroductionThe presence of a new autoantibody system, anti-carbamylated protein (anti-CarP) antibodies, has been identified in rheumatoid arthritis (RA). The presence of anti-CarP antibodies was evaluated in samples taken from individuals who subsequently developed RA before and after onset of symptoms and related to previously analysed antibodies against citrullinated peptides (ACPA specificities) and anti-CCP2.MethodsA total of 252 individuals, with 423 samples from before onset of symptoms of RA, and 197 population controls were identified as donors to the Medical Biobank of Northern Sweden; 192 of them were also sampled at the time of diagnosis. All samples were analysed for anti-CarP IgG and anti-CCP2 antibodies using ELISAs. Ten different antibody reactivities against citrullinated antigens (ACPA specificities) were analysed using a custom-made microarray based on the ImmunoCAP ISAC system (Phadia).ResultsThe concentration of anti-CarP antibodies was significantly increased in the pre-symptomatic individuals compared with controls (P <0.001) and also increased significantly after disease onset (P <0.001). The sensitivity for anti-CarP antibodies in the pre-symptomatic individuals was 13.9% (95% CI: 11 to 17.6) and 42.2% (95% CI: 35.4 to 49.3) following development of RA. Anti-CarP antibody positivity was found in 5.1% to 13.3% of individuals negative for anti-CCP2 or ACPA specificities. Presence of anti-CarP antibodies was significantly related to radiological destruction at baseline, at 24 months and also to radiological change (P <0.05, all).ConclusionsThe results indicate that anti-CarP antibodies are associated with disease development, even after adjusting for the presence of different ACPA fine specificities, and in anti-CCP2 negative individuals and contribute to the identification of a subset of patients with worse radiological progression of the disease independent of ACPA.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0536-2) contains supplementary material, which is available to authorized users.     

Thymol and Carvacrol Prevent Cisplatin‐Induced Nephrotoxicity by Abrogation of Oxidative Stress,Inflammation, and Apoptosis in Rats

The aim of the present study is to assess the possible protective effects of thymol and carvacrol against cisplatin (CP)‐induced nephrotoxicity. A single dose of CP {6 mg/kg, intraperitoneally (i.p.)} injected to male rats revealed significant increases in serum urea, creatinine, and tumor necrosis factor alpha levels. It also increased kidney contents of malondialdehyde and caspase‐3 activity with significant reduction in serum albumin, kidney content of reduced glutathione as well as catalase, and superoxide dismutase activity as compared to that of the control group. In contrast, administration of thymol {20 mg/kg, orally (p.o.)} and/or carvacrol (15 mg/kg, p.o.) for 14 days before CP injection and for 7 days after CP administration restored the kidney function and examined oxidative stress parameters. In conclusion, thymol was more effective nephroprotective than carvacrol. Moreover, a combination of thymol and carvacrol had a synergistic nephroprotective effect that might be attributed to antioxidant, anti‐inflammatory, and antiapoptotic activities.