Interferon Regulatory Factor 8 Regulates Pathways for Antigen Presentation in Myeloid Cells and during Tuberculosis

IRF8 (Interferon Regulatory Factor 8) plays an important role in defenses against intracellular pathogens, including several aspects of myeloid cells function. It is required for ontogeny and maturation of macrophages and dendritic cells, for activation of anti-microbial defenses, and for production of the Th1-polarizing cytokine interleukin-12 (IL-12) in response to interferon gamma (IFNγ) and protection against infection with Mycobacterium tuberculosis. The transcriptional programs and cellular pathways that are regulated by IRF8 in response to IFNγ and that are important for defenses against M. tuberculosis are poorly understood. These were investigated by transcript profiling and chromatin immunoprecipitation on microarrays (ChIP-chip). Studies in primary macrophages identified 368 genes that are regulated by IRF8 in response to IFNγ/CpG and that behave as stably segregating expression signatures (eQTLs) in F2 mice fixed for a wild-type or mutant allele at IRF8. A total of 319 IRF8 binding sites were identified on promoters genome-wide (ChIP-chip) in macrophages treated with IFNγ/CpG, defining a functional G/AGAAnTGAAA motif. An analysis of the genes bearing a functional IRF8 binding site, and showing regulation by IFNγ/CpG in macrophages and/or in M. tuberculosis-infected lungs, revealed a striking enrichment for the pathways of antigen processing and presentation, including multiple structural and enzymatic components of the Class I and Class II MHC (major histocompatibility complex) antigen presentation machinery. Also significantly enriched as IRF8 targets are the group of endomembrane- and phagosome-associated small GTPases of the IRG (immunity-related GTPases) and GBP (guanylate binding proteins) families. These results identify IRF8 as a key regulator of early response pathways in myeloid cells, including phagosome maturation, antigen processing, and antigen presentation by myeloid cells

Top-Down Control of Herbivory by Birds and Bats in the Canopy of Temperate Broad-Leaved Oaks (Quercus robur)

The intensive foraging of insectivorous birds and bats is well known to reduce the density of arboreal herbivorous arthropods but quantification of collateral leaf damage remains limited for temperate forest canopies

Methodological study to evaluate the psychometric properties of FACIT-CD in a sample of Brazilian women with cervical intraepithelial neoplasia

Background: The occurrence of cervical intraepithelial neoplasia (CIN) is associated with changes in health-related quality of life, including psychological factors, such as fear and shame, and changes in sexuality and sexual satisfaction, such as decreased sexual desire and frequency of sexual intercourse. Personal relationships are the most affected because CIN is sexually transmitted and many women tend to blame their partner for disease transmission. The aim of this study was to evaluate the psychometric properties of the FACIT-CD questionnaire in Brazilian women diagnosed with CIN. Methods: The properties of the FACIT-CD questionnaire were tested on a sample of 439 women seen at the Department of Prevention of Barretos Cancer Hospital, including 329 patients who were diagnosed with CIN and 110 women who were not diagnosed with the disease. The analysed parameters included internal consistency (Cronbach's alpha), reproducibility (intraclass correlation coefficient), structural validity, convergent validity (correlation with the SF-12 and EORTC QLQ-CX24 questionnaires), discriminant validity (according to disease status, and self-rating of health), sensitivity, and responsiveness. Results: The Cronbach alpha values of the FACIT-CD scales were higher than 0.70 with the exception of the relationship scale (0.66). The FACIT-CD reproducibility was satisfactory, with variation in the intraclass correlation coefficients ranging between 0.50 and 0.83, although the 95% confidence interval (CI) was lower than 0.40 (0.33-0.64) on the treatment satisfaction scale. Regarding structural validity, only one item on the physical well-being scale was not kept in the original domain. The expected correlations between the FACIT-CD and SF-12 were not confirmed, whereas the correlations between the FACIT-CD and EORTC QLQ-CX24 were confirmed. The questionnaire was able to discriminate the groups according to disease status and self-rating of health. The sensitivity was low for the relationship scale and moderate for the other scales. The responsiveness of the FACIT-CD questionnaire varied between the groups that denominate the self-perception of health as no change, improvement or worsening. Conclusion: Our results are encouraging and indicate that the FACIT-CD questionnaire is a promising tool for the analysis of the quality of life of women with CIN.The postdoctoral fellowship was supported by Sao Paulo Research Foundation (Fundacao de Amparo a Pesquisa do Estado de Sao Paulo - FAPESP, Sao Paulo, Brazil). Process number: FAPESP 2014/10158-3. The funding body had no role in the design of the study and collection, analysis and interpretation of data and in writing the manuscript.info:eu-repo/semantics/publishedVersio

Host Genetic Background Strongly Influences the Response to Influenza A Virus Infections

The genetic make-up of the host has a major influence on its response to combat pathogens. For influenza A virus, several single gene mutations have been described which contribute to survival, the immune response and clearance of the pathogen by the host organism. Here, we have studied the influence of the genetic background to influenza A H1N1 (PR8) and H7N7 (SC35M) viruses. The seven inbred laboratory strains of mice analyzed exhibited different weight loss kinetics and survival rates after infection with PR8. Two strains in particular, DBA/2J and A/J, showed very high susceptibility to viral infections compared to all other strains. The LD50 to the influenza virus PR8 in DBA/2J mice was more than 1000-fold lower than in C57BL/6J mice. High susceptibility in DBA/2J mice was also observed after infection with influenza strain SC35M. In addition, infected DBA/2J mice showed a higher viral load in their lungs, elevated expression of cytokines and chemokines, and a more severe and extended lung pathology compared to infected C57BL/6J mice. These findings indicate a major contribution of the genetic background of the host to influenza A virus infections. The overall response in highly susceptible DBA/2J mice resembled the pathology described for infections with the highly virulent influenza H1N1-1918 and newly emerged H5N1 viruses

Deletion Study of DNA Topoisomerase IB from Leishmania donovani: Searching for a Minimal Functional Heterodimer

The substantial differences between trypanosomal and leishmanial DNA topoisomerase IB concerning to their homologues in mammals have provided a new lead in the study of the structural determinants that can be effectively targeted. Leishmania donovani, the causative agent of visceral leishmaniasis, contains an unusual heterodimeric DNA topoisomerase IB. The catalytically active enzyme consists of a large subunit (LdTopIL), which contains the non-conserved N-terminal end and the phylogenetically conserved “core” domain, and of a small subunit (LdTopIS) which harbors the C-terminal region with the characteristic tyrosine residue in the active site. Heterologous co-expression of LdTopIL and LdTopIS genes in a topoisomerase I deficient yeast strain, reconstitutes a fully functional enzyme LdTopIL/S which can be used for structural studies. An approach by combinatorial cloning of deleted genes encoding for truncated versions of both subunits was used in order to find out structural insights involved in enzyme activity or protein-protein interaction. The role played by the non-conserved N-terminal extension of LdTopIL in both relaxation activity and CPT sensitivity has been examined co-expressing the full-length LdTopIS and a fully active LdTopIΔS deletion with several deletions of LdTopIL lacking growing sequences of the N-terminal end. The sequential deletion study shows that the first 26 amino acids placed at the N-terminal end and a variable region comprised between Ala548 to end of the C-terminal extension of LdTopIL were enzymatically dispensable. Altogether this combinatorial approach provides important structural insights of the regions involved in relaxation activity and for understanding the atypical structure of this heterodimeric enzyme

Differences in Walking Pattern during 6-Min Walk Test between Patients with COPD and Healthy Subjects

BACKGROUND: To date, detailed analyses of walking patterns using accelerometers during the 6-min walk test (6MWT) have not been performed in patients with chronic obstructive pulmonary disease (COPD). Therefore, it remains unclear whether and to what extent COPD patients have an altered walking pattern during the 6MWT compared to healthy elderly subjects. METHODOLOGY/PRINCIPAL FINDINGS: 79 COPD patients and 24 healthy elderly subjects performed the 6MWT wearing an accelerometer attached to the trunk. The accelerometer features (walking intensity, cadence, and walking variability) and subject characteristics were assessed and compared between groups. Moreover, associations were sought with 6-min walk distance (6MWD) using multiple ordinary least squares (OLS) regression models. COPD patients walked with a significantly lower walking intensity, lower cadence and increased walking variability compared to healthy subjects. Walking intensity and height were the only two significant determinants of 6MWD in healthy subjects, explaining 85% of the variance in 6MWD. In COPD patients also age, cadence, walking variability measures and their interactions were included were significant determinants of 6MWD (total variance in 6MWD explained: 88%). CONCLUSIONS/SIGNIFICANCE: COPD patients have an altered walking pattern during 6MWT compared to healthy subjects. These differences in walking pattern partially explain the lower 6MWD in patients with COPD

Extracellular DNA Chelates Cations and Induces Antibiotic Resistance in Pseudomonas aeruginosa Biofilms

Biofilms are surface-adhered bacterial communities encased in an extracellular matrix composed of DNA, bacterial polysaccharides and proteins, which are up to 1000-fold more antibiotic resistant than planktonic cultures. To date, extracellular DNA has been shown to function as a structural support to maintain Pseudomonas aeruginosa biofilm architecture. Here we show that DNA is a multifaceted component of P. aeruginosa biofilms. At physiologically relevant concentrations, extracellular DNA has antimicrobial activity, causing cell lysis by chelating cations that stabilize lipopolysaccharide (LPS) and the outer membrane (OM). DNA-mediated killing occurred within minutes, as a result of perturbation of both the outer and inner membrane (IM) and the release of cytoplasmic contents, including genomic DNA. Sub-inhibitory concentrations of DNA created a cation-limited environment that resulted in induction of the PhoPQ- and PmrAB-regulated cationic antimicrobial peptide resistance operon PA3552–PA3559 in P. aeruginosa. Furthermore, DNA-induced expression of this operon resulted in up to 2560-fold increased resistance to cationic antimicrobial peptides and 640-fold increased resistance to aminoglycosides, but had no effect on β-lactam and fluoroquinolone resistance. Thus, the presence of extracellular DNA in the biofilm matrix contributes to cation gradients, genomic DNA release and inducible antibiotic resistance. DNA-rich environments, including biofilms and other infection sites like the CF lung, are likely the in vivo environments where extracellular pathogens such as P. aeruginosa encounter cation limitation

Genome Degradation in Brucella ovis Corresponds with Narrowing of Its Host Range and Tissue Tropism

Brucella ovis is a veterinary pathogen associated with epididymitis in sheep. Despite its genetic similarity to the zoonotic pathogens B. abortus, B. melitensis and B. suis, B. ovis does not cause zoonotic disease. Genomic analysis of the type strain ATCC25840 revealed a high percentage of pseudogenes and increased numbers of transposable elements compared to the zoonotic Brucella species, suggesting that genome degradation has occurred concomitant with narrowing of the host range of B. ovis. The absence of genomic island 2, encoding functions required for lipopolysaccharide biosynthesis, as well as inactivation of genes encoding urease, nutrient uptake and utilization, and outer membrane proteins may be factors contributing to the avirulence of B. ovis for humans. A 26.5 kb region of B. ovis ATCC25840 Chromosome II was absent from all the sequenced human pathogenic Brucella genomes, but was present in all of 17 B. ovis isolates tested and in three B. ceti isolates, suggesting that this DNA region may be of use for differentiating B. ovis from other Brucella spp. This is the first genomic analysis of a non-zoonotic Brucella species. The results suggest that inactivation of genes involved in nutrient acquisition and utilization, cell envelope structure and urease may have played a role in narrowing of the tissue tropism and host range of B. ovis