Enhanced Delivery of 4-Thioureidoiminomethylpyridinium Perchlorate in Tuberculosis Models with IgG Functionalized Poly(Lactic Acid)-Based Particles.

The compound 4-thioureidoiminomethylpyridinium perchlorate (perchlozone©) is a novel anti-tuberculosis drug that is active in multiple drug resistance cases, but the compound is hepatotoxic. To decrease the systemic load and to achieve targeting, we encapsulated the drug into poly(lactic acid)-based micro- (1100 nm) and nanoparticles (170 nm) that were modified with single-chain camel immunoglobulin G (IgG) for targeting. Both micro- and nanoparticles formed stable suspensions in saline solution at particle concentrations of 10⁻50 mg/mL. The formulations were injected intraperitoneally and intravenously into the mice with experimental tuberculosis. The survival of control animals was compared to that of mice which were treated with daily oral drug solution, single intraperitoneal administration of drug-loaded particles, and those treated both intravenously and intraperitoneally by drug-loaded particles modified with polyclonal camel IgGs. The distribution of particles in the organs of mice was analyzed with immunofluorescence and liquid chromatography/mass spectrometry. Morphological changes related to tuberculosis and drug toxicity were registered. Phagocytic macrophages internalized particles and transported them to the foci of tuberculosis in inner organs. Nanoparticle-based drug formulations, especially those with IgG, resulted in better survival and lower degree of lung manifestations than the other modes of treatment.

Proteome analysis of the Mycobacterium tuberculosis Beijing B0/W148 cluster.

Beijing B0/W148, a "successful" clone of Mycobacterium tuberculosis, is widespread in the Russian Federation and some countries of the former Soviet Union. Here, we used label-free gel-LC-MS/MS shotgun proteomics to discover features of Beijing B0/W148 strains that could explain their success. Qualitative and quantitative proteome analyses of Beijing B0/W148 strains allowed us to identify 1,868 proteins, including 266 that were differentially abundant compared with the control strain H37Rv. To predict the biological effects of the observed differences in protein abundances, we performed Gene Ontology analysis together with analysis of protein-DNA interactions using a gene regulatory network. Our results demonstrate that Beijing B0/W148 strains have increased levels of enzymes responsible for long-chain fatty acid biosynthesis, along with a coincident decrease in the abundance of proteins responsible for their degradation. Together with high levels of HsaA (Rv3570c) protein, involved in steroid degradation, these findings provide a possible explanation for the increased transmissibility of Beijing B0/W148 strains and their survival in host macrophages. Among other, we confirmed a very low level of the SseA (Rv3283) protein in Beijing B0/W148 characteristic for all «modern¼ Beijing strains, which could lead to increased DNA oxidative damage, accumulation of mutations, and potentially facilitate the development of drug resistance.

Genome-wide Mycobacterium tuberculosis variation (GMTV) database: a new tool for integrating sequence variations and epidemiology.

BACKGROUND: Tuberculosis (TB) poses a worldwide threat due to advancing multidrug-resistant strains and deadly co-infections with Human immunodeficiency virus. Today large amounts of Mycobacterium tuberculosis whole genome sequencing data are being assessed broadly and yet there exists no comprehensive online resource that connects M. tuberculosis genome variants with geographic origin, with drug resistance or with clinical outcome. DESCRIPTION: Here we describe a broadly inclusive unifying Genome-wide Mycobacterium tuberculosis Variation (GMTV) database, (http://mtb.dobzhanskycenter.org) that catalogues genome variations of M. tuberculosis strains collected across Russia. GMTV contains a broad spectrum of data derived from different sources and related to M. tuberculosis molecular biology, epidemiology, TB clinical outcome, year and place of isolation, drug resistance profiles and displays the variants across the genome using a dedicated genome browser. GMTV database, which includes 1084 genomes and over 69,000 SNP or Indel variants, can be queried about M. tuberculosis genome variation and putative associations with drug resistance, geographical origin, and clinical stages and outcomes. CONCLUSIONS: Implementation of GMTV tracks the pattern of changes of M. tuberculosis strains in different geographical areas, facilitates disease gene discoveries associated with drug resistance or different clinical sequelae, and automates comparative genomic analyses among M. tuberculosis strains.

Unusual large-scale chromosomal rearrangements in Mycobacterium tuberculosis Beijing B0/W148 cluster isolates.

The Mycobacterium tuberculosis (MTB) Beijing family isolates are geographically widespread, and there are examples of Beijing isolates that are hypervirulent and associated with drug resistance. One-fourth of Beijing genotype isolates found in Russia belong to the B0/W148 group. The aim of the present study was to investigate features of these endemic strains on a genomic level. Four Russian clinical isolates of this group were sequenced, and the data obtained was compared with published sequences of various MTB strain genomes, including genome of strain W-148 of the same B0/W148 group. The comparison of the W-148 and H37Rv genomes revealed two independent inversions of large segments of the chromosome. The same inversions were found in one of the studied strains after deep sequencing using both the fragment and mate-paired libraries. Additionally, inversions were confirmed by RFLP hybridization analysis. The discovered rearrangements were verified by PCR in all four newly sequenced strains in the study and in four additional strains of the same Beijing B0/W148 group. The other 32 MTB strains from different phylogenetic lineages were tested and revealed no inversions. We suggest that the initial largest inversion changed the orientation of the three megabase (Mb) segment of the chromosome, and the second one occurred in the previously inverted region and partly restored the orientation of the 2.1 Mb inner segment of the region. This is another remarkable example of genomic rearrangements in the MTB in addition to the recently published of large-scale duplications. The described cases suggest that large-scale genomic rearrangements in the currently circulating MTB isolates may occur more frequently than previously considered, and we hope that further studies will help to determine the exact mechanism of such events.