Observation of Cytotoxicity of Phosphonium Derivatives Is Explained: Metabolism Inhibition and Adhesion Alteration.

The search for new antibiotics, substances that kill prokaryotic cells and do not kill eukaryotic cells, is an urgent need for modern medicine. Among the most promising are derivatives of triphenylphosphonium, which can protect the infected organs of mammals and heal damaged cells as mitochondria-targeted antioxidants. In addition to the antioxidant action, triphenylphosphonium derivatives exhibit antibacterial activity. It has recently been reported that triphenylphosphonium derivatives cause either cytotoxic effects or inhibition of cellular metabolism at submicromolar concentrations. In this work, we analyzed the MTT data using microscopy and compared them with data on changes in the luminescence of bacteria. We have shown that, at submicromolar concentrations, only metabolism is inhibited, while an increase in alkyltriphenylphosphonium (CnTPP) concentration leads to adhesion alteration. Thus, our data on eukaryotic and prokaryotic cells confirm a decrease in the metabolic activity of cells by CnTPPs but do not confirm a cytocidal effect of TPPs at submicromolar concentrations. This allows us to consider CnTPP as a non-toxic antibacterial drug at low concentrations and a relatively safe vector for delivering other antibacterial substances into bacterial cells.

Constructing of Bacillus subtilis-Based Lux-Biosensors with the Use of Stress-Inducible Promoters.

Here, we present a new lux-biosensor based on Bacillus subtilis for detecting of DNA-tropic and oxidative stress-causing agents. Hybrid plasmids pNK-DinC, pNK-AlkA, and pNK-MrgA have been constructed, in which the Photorhabdus luminescens reporter genes luxABCDE are transcribed from the stress-inducible promoters of B. subtilis: the SOS promoter PdinC, the methylation-specific response promoter PalkA, and the oxidative stress promoter PmrgA. The luminescence of B. subtilis-based biosensors specifically increases in response to the appearance in the environment of such common toxicants as mitomycin C, methyl methanesulfonate, and H2O2. Comparison with Escherichia coli-based lux-biosensors, where the promoters PdinI, PalkA, and Pdps were used, showed generally similar characteristics. However, for B. subtilis PdinC, a higher response amplitude was observed, and for B. subtilis PalkA, on the contrary, both the amplitude and the range of detectable toxicant concentrations were decreased. B. subtilis PdinC and B. subtilis PmrgA showed increased sensitivity to the genotoxic effects of the 2,2'-bis(bicyclo [2.2.1] heptane) compound, which is a promising propellant, compared to E. coli-based lux-biosensors. The obtained biosensors are applicable for detection of toxicants introduced into soil. Such bacillary biosensors can be used to study the differences in the mechanisms of toxicity against Gram-positive and Gram-negative bacteria.

ArdB Protective Activity for Unmodified λ Phage Against EcoKI Restriction Decreases in UV-Treated Escherichia coli.

Anti-restriction proteins ArdB/KlcA specifically inhibit restriction (endonuclease) activity of restriction-modification (RM) type I systems. Molecular mechanisms of ArdB/KlcA-based anti-restriction remain unknown. In this study, we quantitate effects of ArdB on protection of unmodified λ phage DNA from EcoKI restriction. After UV irradiations, which produce significant amounts of unmodified chromosomal DNA in Escherichia coli K12 cells, the protective activity of ArdB decreases. Unlike ArdB, DNA-mimicking protein Ocr retains its ability to protect the unmodified λ phage regardless of UV dose. We hypothesize that the observed decrease in ArdB protective activity in UV-treated cells is due to its binding to unmodified chromosomal DNA, which decreases effective concentrations of free ArdB molecules available for λ phage protection against type I restriction enzymes.

Seasonal changes in luminescent intestinal microflora of the fish inhabiting the Bering and Okhotsk seas.

Here, we present a study of luminescent intestinal microflora of the fish inhabiting Bering and Okhotsk seas in summer and winter seasons. Sampling of intestinal luminescent microflora was carried for several years, with all recovered species belonging to psychrophilic bacteria of either Aliivibrio logei or Photobacterium phosphoreum species. A seasonal change in fish intestinal luminescent microflora detected include an increase in prevalence of P. phosphoreum bacteria in summer and an increase in prevalence of A. logei bacteria in winter seasons. In fact, 90% of all luminescent bacteria isolated in winter period (January-March) were A. logei, while 88% of luminescent isolates recovered in summer period (July-September) were that of P. phosphoreum species. Seasonal changes were similar across all six sampling expeditions, three in winter and three in summer seasons, evenly spread through 2010-2018 period.

A combination of luxR1 and luxR2 genes activates Pr-promoters of psychrophilic Aliivibrio logei lux-operon independently of chaperonin GroEL/ES and protease Lon at high concentrations of autoinducer.

UNLABELLED: Lux-operon of psychrophilic bacteria Aliivibrio logei contains two copies of luxR and is regulated by Type I quorum sensing (QS). Activation of lux-operon of psychrophilic bacteria A. logei by LuxR1 requires about 100 times higher concentrations of autoinducer (AI) than the activation by LuxR2. On the other hand, LuxR1 does not require GroEL/ES chaperonin for its folding and cannot be degraded by protease Lon, while LuxR2 sensitive to Lon and requires GroEL/ES. Here we show that at 10(-5) - 10(-4)М concentrations of AI a combination of luxR1 and luxR2 products is capable of activating the Pr-promoters of A. logei lux-operon in Escherichia coli independently of GroEL/ES and protease Lon. The presence of LuxR1 assists LuxR2 in gro(-) cells when AI was added at high concentration, while at low concentration of AI in a cell LuxR1 decreases the LuxR2 activity. These observations may be explained by the formation of LuxR1/LuxR2 heterodimers that act in complex with AI independently from GroEL/ES and protease Lon. IMPORTANCE: This study expands current understanding of QS regulation in A. logei as it implies cooperative regulation of lux-operon by LuxR1 and LuxR2 proteins.

Lux-operon of the marine psychrophilic bacterium Aliivibrio logei: a comparative analysis of the LuxR1/LuxR2 regulatory activity in Escherichia coli cells.

The lux-operon of the psychrophilic bioluminescent bacterium Aliivibrio logei is regulated by quorum sensing (QS). The key components of this system are LuxI, which catalyses synthesis of the autoinducer (AI), and LuxR, which activates transcription of the entire lux-operon. The lux-operon of A. logei contains two copies of the luxR gene: luxR1 and luxR2. In the present study, lux-operon sequence analysis from 16 strains of A. logei, isolated from cold habitats of the White, Baltic, Okhotsk and Bering seas, was carried out. Phylogenetic analysis showed that all isolated strains of A. logei have both copies of luxR genes which are homologous to luxR genes of the related Aliivibrio salmonicida. Evaluation of LuxR1 and LuxR2 activity showed that LuxR2 remains active at significantly lower concentrations of AI (10- 9 M) than LuxR1, which is active only at high AI concentrations (10- 6 M). As the QS response is already prominent at AI concentrations as low as 10- 8 to 10- 9 M, we conclude that LuxR2 is the main activator of the lux-operon of A. logei. The thermolabilities of LuxR1 and LuxR2 are similar and exceed that of LuxR of the mesophilic bacterium Aliivibrio fischeri. In contrast to LuxR2, LuxR1 is not a substrate of Lon protease and does not require the chaperonin GroEL/ES for its folding. This study expands our current understanding of QS regulation in A. logei as it implies differential regulation by LuxR1 and LuxR2 proteins.