Hospital-acquired infections are a generally recognized problem for healthcare professionals. Clinical variants of Gram-negative and Gram-positive pathogens are characterized with enhanced antibiotic resistance and virulence due to mutations and the horizontal acquisition of respective genetic determinants. In this study, two Escherichia coli, two Klebsiella pneumoniae, three Pseudomonas aeruginosa, two Staphylococcus aureus, one Staphylococcus epidermidis and one Streptococcus pneumoniae showing broad spectra of antibiotic resistance were isolated from patients suffering from nosocomial infections in a local hospital in Almaty, Kazakhstan. The aim of the study was to compare general and species-specific pathways of the development of virulence and antibiotic resistance through opportunistic pathogens causing hospital-acquired infections. The whole-genome PacBio sequencing of the isolates allowed for the genotyping and identification of antibiotic resistance and virulence genetic determinants located in the chromosomes, plasmids and genomic islands. It was concluded that long-read sequencing is a useful tool for monitoring the epidemiological situation in hospitals. Marker antibiotic resistance mutations common for different microorganisms were identified, which were acquired due to antibiotic-selective pressure in the same clinical environment. The genotyping and identification of strain-specific DNA methylation motifs were found to be promising in estimating the risks associated with hospital infection outbreaks and monitoring the distribution and evolution of nosocomial pathogens.

Bacteria with similar genomes can exhibit different phenotypes due to alternative gene expression patterns. In this study, we analysed four antibiotic-resistant Staphylococcus aureus hospital isolates using transcriptomics, PacBio genome sequencing, and methylomics analyses. Transcriptomic data were obtained from cultures exposed to gentamicin, the iodine-alanine complex CC-196, and their combination. We observed strain-specific expression patterns of core and accessory genes that remained stable under antimicrobial stress – a phenomenon we term the Clonal Gene Expression Stability (CGES) that is the main discovery of the paper. An involvement of epigenetic mechanisms in stabilization of the CGES was hypothesized and statistically verified. Canonical methylation patterns controlled by type I restriction-modification systems accounted for ~ 10% of epigenetically modified adenine residues, whereas multiple non-canonically modified adenines were distributed sporadically due to imperfect DNA targeting by methyltransferases. Protein-coding sequences were characterized by a significantly lower frequency of modified nucleotides. Epigenetic modifications near transcription start codons showed a statistically significant negative association with gene expression levels. While the role of epigenetic modifications in gene regulation remains debatable, variations in non-canonical modification patterns may serve as markers of CGES.

1. Background: Iodine is a broad-spectrum antimicrobial disinfectant for topical application. Recent studies have shown promising results on the applicability of an iodine-containing complex, FS-1, against antibiotic-resistant pathogens. It was hypothesized that the antimicrobial activity of iodine-containing complexes may be modulated by the organic moiety of the complex, i.e., amino acids. 2. Methods: Gene regulation and metabolic alterations were studied in two model multidrug-resistant microorganisms, Staphylococcus aureus ATCC BAA-39, and Escherichia coli ATCC BAA-196, treated with three complexes containing iodine and three different amino acids: glycine, L-alanine, and L-isoleucine. The bacterial cultures were exposed to sub-lethal concentrations of the complexes in the lagging and logarithmic growth phases. Gene regulation was studied by total RNA sequencing and differential gene expression analysis. 3. Results: The central metabolism of the treated bacteria was affected. An analysis of the regulation of genes involved in stress responses suggested the disruption of cell wall integrity, DNA damage, and oxidative stress in the treated bacteria. 4. Conclusions: Previous studies showed that the application of iodine-containing complexes, such as FS-1, serves as a supplement to common antibiotics and can be a promising way to combat antibiotic-resistant pathogens. Current results shed light on possible mechanisms of this action by disrupting the cell wall barriers and imposing oxidative stress. It was also found that the effect of the complexes on metabolic pathways varied in the tested microorganisms depending on the organic moiety of the complexes and the growth phase when the complexes had been applied.

The problem of nosocomial infections is growing due to the introduction of new treatment regimens involving immunosuppressive drugs. The genomes of seven Gram-negative clinical isolates of Escherichia , Klebsiella , and Pseudomonas were sequenced and analyzed in this study to serve as model microorganisms to study drug-induced antibiotic resistance reversion.

Aim: Promising results on application of iodine-containing nano-micelles, FS-1, against antibiotic-resistant Escherichia coli was demonstrated. Materials & methods: RNA sequencing for transcriptomics and the complete genome sequencing by SMRT PacBio were followed by genome assembly and methylomics. Results & conclusion: FS-1-treated E. coli showed an increased susceptibility to antibiotics ampicillin and gentamicin. Cultivation with FS-1 caused gene expression alterations toward anaerobic respiration, increased anabolism and inhibition of many nutrient uptake systems. Main targets of iodine-containing particles were cell membrane structures causing oxidative, osmotic and acidic stresses. Identification of methylated nucleotides showed an altered pattern in the FS-1-treated culture. Possible role of transcriptional and epigenetic modifications in the observed increase in susceptibility to gentamicin and ampicillin were discussed.

Background: Recent studies showed promising results on application of iodine-containing nanomicelles, FS-1, against antibiotic resistant pathogens. The effect was studied on Escherichia coli ATCC BAA-196. Materials & methods: RNA sequencing for transcriptomics and the complete genome sequencing by SMRT PacBio RS II technology followed by genome assembly and methylomics study were performed. Results & conclusions: FS-1 treated E. coli showed an increased susceptibility to antibiotics ampicillin and gentamicin. The analysis of differential gene regulation showed that possible targets of iodine-containing particles are cell membrane fatty acids and proteins, particularly cytochromes, that leads to oxidative, osmotic and acidic stresses. Cultivation with FS-1 caused gene expression alterations towards anaerobic respiration, increased anabolism and inhibition of many nutrient uptake systems. Identification of methylated nucleotides showed an altered pattern in the FS-1 treated culture. Possible role of transcriptional and epigenetic modifications in the observed increase in susceptibility to gentamicin and ampicillin were discussed. Lay New approaches of combatting drug resistance infections are in demand as the development of new antibiotics is in a deep crisis. This study was set out to investigate molecular mechanisms of action of new iodine-containing nano-micelle drug FS-1, which potentially may improve the antibiotic therapy of drug resistant infections. Iodine is one of the oldest antimicrobials and until now there were no reports on development of resistance to iodine. Recent studies showed promising results on application of iodine-containing nano-micelles against antibiotic resistant pathogens as a supplement to antibiotic therapy. The mechanisms of action, however, remain unclear. The collection strain Escherichia coli ATCC BAA-196 showing an extended spectrum of resistance to beta-lactam and aminoglycoside antibiotics was used in this study as a model organism. Antibiotic resistance patterns, whole genomes and total RNA sequences of the FS-1 treated (FS) and negative control (NC) variants of E. coli BAA-196 were obtained and analyzed. FS culture showed an increased susceptibility to antibiotics associated with profound gene expression alterations switching the bacterial metabolism to anaerobic respiration, increased anabolism, osmotic stress response and inhibition of many nutrient uptake systems. Nucleotide methylation pattern were identified in FS and NC cultures. While the numbers of methylated sites in both genomes remained similar, some peculiar alterations were observed in their distribution along chromosomal and plasmid sequences.