Mycobacterium tuberculosis is one of the most dangerous human pathogens, the causative agent of tuberculosis. While this pathogen is considered as extremely clonal and resistant to horizontal gene exchange, there are many facts supporting the hypothesis that on the early stages of evolution the development of pathogenicity of ancestral Mtb has started with a horizontal acquisition of virulence factors. Episodes of infections caused by non-tuberculosis Mycobacteria reported worldwide may suggest a potential for new pathogens to appear. If so, what is the role of horizontal gene transfer in this process?

Drug induced reversion of antibiotic resistance is a promising way to combat multidrug resistant infections. However, lacking knowledge of mechanisms of drug resistance reversion impedes employing this approach in medicinal therapies. Induction of antibiotic resistance reversion by a new anti-tuberculosis drug FS-1 has been reported. FS-1 was used in this work in combination with standard anti-tuberculosis antibiotics in an experiment on laboratory guinea pigs infected with an extensively drug resistant (XDR) strain M. tuberculosis SCAID 187.0. During the experimental trial, genetic changes in the population were analyzed by sequencing of M. tuberculosis isolates followed by variant calling. In total 11 isolates obtained from different groups of infected animals at different stages of disease development and treatment were sequenced. It was found that despite the selective pressure of antibiotics, FS-1 caused a counter-selection of drug resistant variants that speeded up the recovery of the infected animals from XDR tuberculosis. Drug resistance mutations reported in the genome of the initial strain remained intact in more sensitive isolates obtained in this experiment. Variant calling in the sequenced genomes revealed that the drug resistance reversion could be associated with a general increase in genetic heterogeneity of the population of M. tuberculosis. Accumulation of mutations in PpsA and PpsE subunits of phenolpthiocerol polyketide synthase was observed in the isolates treated with FS-1 that may indicate an increase of persisting variants in the population. It was hypothesized that FS-1 caused an active counter-selection of drug resistant variants from the population by aggravating the cumulated fitness cost of the drug resistance mutations. Action of FS-1 on drug resistant bacteria exemplified the theoretically predicted induced synergy mechanism of drug resistance reversion. An experimental model to study the drug resistance reversion phenomenon is hereby introduced.

Onosma roots are widely used in traditional medicine to treat various diseases throughout the world. In this study, for the first time, we investigated the component composition and biological activity of various extracts from the roots of Onosma gmelinii collected in the highlands of the Kakpakty Mountains of the Almaty region (Republic of Kazakhstan). Extracts were obtained by three different methods: percolation extraction, ultrasound-assisted extraction, and supercritical carbon dioxide extraction. The component composition of the extracts was determined by gas chromatography/mass spectrometry (GC/MS), naphthoquinones by thin-layer chromatography (TLC), and spectrophotometric method. In this study, the presence of shikonin and its derivatives in the extracts was confirmed. The concentration of naphthoquinones during CO₂ extraction was about 40, during ultrasonic extraction about 3, and during percolation extraction about 1.3. The GC-MS method identified 69 chemical compounds in the ultrasonic extract, 46 compounds in the CO₂ extract, and 51 compounds in the percolation extract. The extracts were tested on a panel of bacteria and viruses: two Gram-negative bacteria (Escherichia coli ATCC 8739, Pseudomonas aeruginosa ATCC 9027); nine Gram-positive bacteria (Staphylococcus aureus ATCC 6538-P, Staphylococcus aureus ATCC BAA-39, Staphylococcus epidermidis ATCC 51625, Staphylococcus epidermidis ATCC 12228, Streptococcus pyogenes ATCC 19615, Streptococcus pneumoniae ATCC BAA-660, Enterococcus hirae ATCC 10541, Enterococcus faecalis ATCC 51575, Enterococcus faecium ATCC 700221); and two fungal species (Candida albicans ATCC 10231, Candida albicans ATCC 2091); five subtypes of influenza virus A (A/FPV/Weybridge/78 (H7N7), A/Swine/Iowa/15/30 (H1N1), A/black-headed gull/Atyrau/743/04 (H13N6), A/FPV/Rostock/1934 (H7N1), A/Almaty/8/98 (H3N2)). The root extracts of Onosma gmelinii showed antibacterial activity in different degrees against all tested Gram-positive bacterial strains, while no inhibitory effect on Gram-negative bacteria was observed. The results indicated that the ultrasonic extract effectively inhibits the growth of the majority of tested Gram-positive bacteria (MBC from 18.3 to 293.0µg/mL). CO₂ extract had the greatest bactericidal activity (MBC from 0.1 to 24.4µg/mL). Percolation extract insignificantly inhibited bacterial growth (MBC from 2343.8 to 4687.5µg/mL). CO₂ extract and ultrasonic extract significantly reduced the activity of C. albicans. The results of the antiviral action showed that the ultrasonic extract has the greatest effectiveness against different subtypes of the influenza virus A, while other extracts did not show significant activity.

Application of supplementary drugs which increase susceptibility of pathogenic bacteria to antibiotics is a promising yet unexplored approach to overcome the global problem of multidrug-resistant infections. The discovery of a new drug, an iodine-containing nano-molecular complex FS-1, which has proven to improve susceptibility to antibiotics in various pathogens, including MRSA strain Staphylococcus aureus ATCC BAA-39^TM, allowed studying this phenomenon. Chromosomal DNA and total RNA samples extracted from the FS-1 treated strain (FS) and from the negative control (NC) cultures were sequenced by PacBio SMRT and Ion Torrent technologies, respectively. PacBio DNA reads were used to assemble chromosomal DNA of the NC and FS variants of S. aureus BAA-39 and to perform profiling of epigenetically modified nucleotides. Results of transcriptional profiling, variant calling and detection of epigenetic modifications in the FS variant were compared to the NC variant. Additionally, the genetic alterations caused by the treatment of S. aureus BAA-39 with FS-1 were compared to the results of a similar experiment conducted with another model organism, E. coli ATCC BAA-196. Several commonalities in responses of these phylogenetically distant microorganisms to the treatment with FS-1 were discovered, which included metabolic transition toward anaerobiosis and oxidative/osmotic stress response. S. aureus culture appeared to be more sensitive to FS-1 due to a higher penetrability of cells by iodine bound compounds, which caused carbonyl stress associated with nucleotide damaging by FS-1, abnormal epigenetic modifications and an increased rate of mutations. It was hypothesized that the disrupted pattern of adenine methylated loci within methicillin-resistance chromosome cassettes (SCCmec) may promote excision of this antibiotic resistance determinant from chromosomes while the altered pattern of cytosine methylation was behind the adaptive gene regulation in the culture FS. The selection against the antibiotic resistance in bacterial populations caused by abnormal epigenetic modifications exemplifies possible mechanisms of antibiotic resistance reversion induced by iodine-containing compounds. These finding will facilitate development of therapeutic agents against multidrug-resistant infections.