Integrated MicroRNA-mRNA-Analysis of Human Monocyte Derived Macrophages upon Mycobacterium avium subsp. hominissuis Infection

Many efforts have been made to understand basal mechanisms of mycobacterial infections. Macrophages are the first line of host immune defence to encounter and eradicate mycobacteria. Pathogenic species have evolved different mechanisms to evade host response, e.g. by influencing macrophage apoptotic pathways. However, the underlying molecular regulation is not fully understood. A new layer of eukaryotic regulation of gene expression is constituted by microRNAs. Therefore, we present a comprehensive study for identification of these key regulators and their targets in the context of host macrophage response to mycobacterial infections.We performed microRNA as well as mRNA expression analysis of human monocyte derived macrophages infected with several Mycobacterium avium hominissuis strains by means of microarrays as well as quantitative reverse transcription PCR (qRT-PCR). The data revealed the ability of all strains to inhibit apoptosis by transcriptional regulation of BCL2 family members. Accordingly, at 48 h after infection macrophages infected with all M. avium strains showed significantly decreased caspase 3 and 7 activities compared to the controls. Expression of let-7e, miR-29a and miR-886-5p were increased in response to mycobacterial infection at 48 h. The integrated analysis of microRNA and mRNA expression as well as target prediction pointed out regulative networks identifying caspase 3 and 7 as potential targets of let-7e and miR-29a, respectively. Consecutive reporter assays verified the regulation of caspase 3 and 7 by these microRNAs.We show for the first time that mycobacterial infection of human macrophages causes a specific microRNA response. We furthermore outlined a regulatory network of potential interactions between microRNAs and mRNAs. This study provides a theoretical concept for unveiling how distinct mycobacteria could manipulate host cell response. In addition, functional relevance was confirmed by uncovering the control of major caspases 3 and 7 by let-7e and miR-29a, respectively

The non-clonality of drug resistance in Beijing-genotype isolates of Mycobacterium tuberculosis from the Western Cape of South Africa

Background. The Beijing genotype of M. tuberculosis is a virulent strain that is disseminating worldwide and has a strong association with drug resistance. In the Western Cape of South Africa, epidemiological studies have identified the R220 cluster of the Beijing genotype as a major contributor to a recent outbreak of drug-resistant tuberculosis. Although the outbreak is considered to be due to clonal transmission, the relationship among drug resistant isolates has not yet been established. Results. To better understand the evolution of drug resistance among these strains, 14 drug-resistant clinical isolates of the Beijing genotype were sequenced by whole-genome sequencing, including eight from R220 and six from a more ancestral Beijing cluster, R86, for comparison. While each cluster shares a distinct resistance mutation for isoniazid, mapping of other drug-resistance mutations onto a phylogenetic tree constructed from single nucleotide polymorphisms shows that resistance mutations to many drugs have arisen multiple times independently within each cluster of isolates. Thus, drug resistance among these isolates appears to be acquired, not clonally derived. This observation suggests that, although the Beijing genotype as a whole might have selective advantages enabling its rapid dissemination, the XDR isolates are relatively less fit and do not propagate well. Although it has been hypothesized that the increased frequency of drug resistance in some Beijing lineages might be caused by a mutator phenotype, no significant shift in synonymous substitution patterns is observed in the genomes. Conclusion. While MDR-TB is spreading by transmission in the Western Cape, our data suggests that further drug resistance (i.e. XDR-TB) at this stage is acquired.Peer Reviewe

Photo-neutron reaction cross-sections for Zr-nat in the bremsstrahlung end-point energies of 12-16 and 45-70 MeV

The Zr-nat(gamma, xn)Zr89-86 reaction cross-sections were experimentally determined at the bremsstrahlung end-point energies of 12, 14, 16, 45, 50, 55, 60 and 70 MeV by activation and off-line gamma-ray spectrometric technique using the 20 MeV electron linac (ELBE) at the Helmholtz-Zentrum Dresden-Rossendorf (HZDR), Dresden, Germany, and the 100 MeV electron linac at the Pohang Accelerator Laboratory (PAL), Pohang, Korea. The Zr-nat(gamma, xn)Zr89-86 reaction cross-sections as a function of photon energy were also calculated using the TALYS 1.4 computer code. The flux-weighted average cross-sections at the end-point energies of 12-16 and 45-70 MeV were obtained using the literature and the TALYS calculation data based on mono-energetic photons and are found to be in good agreement with the present data. It was also found that the present data and the flux-weighted literature and theoretical values for the Zr-nat (gamma, xn)Zr89-86 reaction cross-sections increase from the threshold values to a certain energy, where other reaction channels open. This indicates the role of excitation energy. However, the increasing trend of the Zr-nat(gamma, xn)Zr-89,Zr-88 reaction cross-sections are sharper from the threshold value up to end-point bremsstrahlung energies of 17-22 MeV compared to the same for the Zr-nat(gamma, xn)Zr-87,Zr-86 reactions. This is due to the Giant Dipole Resonance (GDR) effect besides the role of excitation energy. After a certain value, the individual Zr-nat(gamma, xn) reaction cross-sections decrease with the increase of bremsstrahlung energy due to opening of other reactions, which indicates partitioning of energy in different channels.X11111sciescopu