Transcriptomics and methylomics study on the effect of iodine-containing drug FS-1 onEscherichia coliATCC BAA-196

Abstract

21 Recent studies have demonstrated that antibiotic resistance in pathogenic bacteria could be 22 reverted to the drug sensitive phenotype by the treatment with the newly introduced iodine 23 containing nanomolecular complex, FS-1. Antibiotic resistance reversion has been verified as a 24 promising therapeutic approach to combat multidrug resistant infections. The mechanisms of 25 action, however, remain unclear. A collection strain, Escherichia coli ATCC showing 26 an extended spectrum of resistance to beta-lactam and aminoglycoside antibiotics was used in 27 this study as a model organism. The FS-1 treated culture and the negative control variant were 28 sequenced by PacBio RS II System following the SMRTbell 20-kb library preparation protocol.29 Total RNA samples of these strains were sequenced by Ion Torrent. It was shown that the 30 treatment with FS-1 caused a profound gene expression alternation switching the bacterial 31 metabolism to anaerobic respiration, increased anabolism, oxidative/acidic stress response and an 32 inhibition of many nutrient uptake systems. All this leads to an increase in the susceptibility to 33 antibiotics even when FS-1 is removed from the medium. The later fact implies an involvement 34 of epigenetic mechanisms in gene regulation and antibiotic resistance reversion. This hypothesis 35 was investigated by base-call kinetic analysis in PacBio reads and DNA methylation profiling in 36 the sequenced genomes. Several DNA motifs of adenosine and cytosine methylation were 37 identified. While the numbers of methylated sites in chromosomes and plasmids of both 38 genomes, NC and FS, were similar, the distribution of the methylated sites was different. It may 39 explain the observed long lasting effect of the treatment of E. coli with FS-1 on antibiotic 40 susceptibility of this model microorganism.41 Author summary 42 The emergence of multidrug resistant bacteria is a great concern, since the misuse of antibiotics 43 have caused a strong selective pressure for these resistant bacteria and various treatment options 44 are becoming ineffective. Drug induced reversion of antibiotic resistant is considering a 45 promising approach to address this problem. This study was set out to investigate genetic 46 mechanisms of action of a new iodine-containing nano-micelle drug FS-1 that induces antibiotic 47 resistance reversion in bacteria. Escherichia coli ATCC BAA-196 was used as a model of 48 multidrug resistant microorganisms. With this purpose in mind, we have sequenced the genomic 49 DNA and total RNA samples of E. coli cultivated on medium containing FS-1, served as an 50 experimental (FS) variant, and bacteria cultivated on normal medium, served as negative control 51 (NC). RNA sequencing showed a differential gene expression in the FS-1 treated strain that may 52 explain the observed increase in susceptibility to gentamicin and ampicillin. Application of the 53 3 rd generation sequencing technology, SMRT PacBio, allowed an unambiguous whole genome 54 assembly of NC and FS variants, a...

The Effect of Iodine-Containing Nano-Micelles, FS-1, on Antibiotic Resistance, Gene Expression and Epigenetic Modifications in the Genome of Multidrug Resistant MRSA Strain Staphylococcus aureus ATCC BAA-39

“…Indeed, a drop of coverage of PacBio DNA reads generated from this region was observed in both, NC and FS cultures, suggesting an excision of the genomic islands at least in a fraction of cells; however, the coverage difference between NC and FS genomes was insignificant. A similar destabilization of the virulence plasmid in the FS-1 treated E. coli BAA-196 was reported in a previous publication (Korotetskiy et al, 2020b). The varying rate of excision of SCCmec cassettes from Staphylococcal populations was reported in another publication (Almebairik et al, 2020).…”

“…Moreover, cggR repressor of gluconeogenesis (Kohler et al, 2008;Chaffin et al, 2012) was activated by FS-1. Such activation of glycolysis is typical for S. aureus growing at anaerobic conditions (Kohler et al, 2008); however, it was opposite to what was observed in E. coli where FS-1 treatment caused upregulation of the genes involved in gluconeogenesis and downregulation of their glycolytic counterparts (Korotetskiy et al, 2020b). It may be explained by the fact that E. coli, in contrast to S. aureus, has several alternative glycolytic pathways bypassing glycolysis, which are Entner Doudoroff, Embden-Meyerhof-Parnas and oxidative pentose phosphate pathways (Hollinshead et al, 2016).…”

“…It suggests that the transmembrane flux of iodine-bound compounds into FS-1 treated E. coli cells was significantly lower even that in both cases FS-1 was applied in 1/2 MBC concentrations calculated for respective microorganisms (450 µg/ml for S. aureus and 500 µg/ml for E. coli). In contrast to S. aureus, no increase in the number of modified nucleotides and no increase in the rate of frameshift mutations was observed in E. coli treated with FS-1 (Korotetskiy et al, 2020b). In concordance with this was the detection of only traces of the radioactive iodine in the DNA extracted from the cells of E. coli treated with 131 I marked FS-1, while in the respective DNA sample extracted from the treated S. aureus cells, the residual radioactivity of DNA constituted 12.76% ± 9.04 of the total radioactivity of washed bacterial cells.…”

“…The residual radioactivity can be associated with both complexing of FS-1 with DNA and with a direct halogenation of nucleotides by iodine isotopes. In a similar study with E. coli ATCC BAA-196, the residual radioactivity of the extracted DNA constituted only 0.46% ± 0.15 of the residual radioactivity of the treated bacterial cells measured after washing the cells (Korotetskiy et al, 2020b).…”

“…Patterns of FS-1 Treated S. aureus ATCC BAA-39 and E. coli ATCC Recently, an analysis of the effect of FS-1 on gene expression in E. coli ATCC BAA-196 at condition NC − /FS + has been published (Korotetskiy et al, 2020b). In total, 725 homologous genes shared by the genomes S. aureus ATCC BAA-39 (CP033505.1) and E. coli ATCC BAA-196 (CP042865.1) were identified by the program GET_HOMOLOGUES.…”