Pentafluorosulfanyl-containing Triclocarban Analogs with Potent Antimicrobial Activity

Concerns have been raised about the long-term accumulating effects of triclocarban, a polychlorinated diarylurea widely used as an antibacterial soap additive, in the environment and in human beings. Indeed, the Food and Drug Administration has recently banned it from personal care products. Herein, we report the synthesis, antibacterial activity and cytotoxicity of novel N,N′-diarylureas as triclocarban analogs, designed by reducing one or more chlorine atoms of the former and/or replacing them by the novel pentafluorosulfanyl group, a new bioisostere of the trifluoromethyl group, with growing importance in drug discovery. Interestingly, some of these pentafluorosulfanyl-bearing ureas exhibited high potency, broad spectrum of antimicrobial activity against Gram-positive bacterial pathogens, and high selectivity index, while displaying a lower spontaneous mutation frequency than triclocarban. Some lines of evidence suggest a bactericidal mode of action for this family of compounds.

Determination and Therapeutic Exploitation of Ebola Virus Spontaneous Mutation Frequency

ABSTRACTEbola virus (EBOV) is an RNA virus that can cause hemorrhagic fever with high fatality rates, and there are no approved vaccines or therapies. Typically, RNA viruses have high spontaneous mutation rates, which permit rapid adaptation to selection pressures and have other important biological consequences. However, it is unknown if filoviruses exhibit high mutation frequencies. Ultradeep sequencing and a recombinant EBOV that carries the gene encoding green fluorescent protein were used to determine the spontaneous mutation frequency of EBOV. The effects of the guanosine analogue ribavirin during EBOV infections were also assessed. Ultradeep sequencing revealed that the mutation frequency for EBOV was high and similar to those of other RNA viruses. Interestingly, significant genetic diversity was not observed in viable viruses, implying that changes were not well tolerated. We hypothesized that this could be exploited therapeutically.In vitro, the presence of ribavirin increased the error rate, and the 50% inhibitory concentration (IC50) was 27 μM. In a mouse model of ribavirin therapy given pre-EBOV exposure, ribavirin treatment corresponded with a significant delay in time to death and up to 75% survival. In mouse and monkey models of therapy given post-EBOV exposure, ribavirin treatment also delayed the time to death and increased survival. These results demonstrate that EBOV has a spontaneous mutation frequency similar to those of other RNA viruses. These data also suggest a potential for therapeutic use of ribavirin for human EBOV infections.IMPORTANCEEbola virus (EBOV) causes a severe hemorrhagic disease with high case fatality rates; there are no approved vaccines or therapies. We determined the spontaneous mutation frequency of EBOV, which is relevant to understanding the potential for the virus to adapt. The frequency was similar to those of other RNA viruses. Significant genetic diversity was not observed in viable viruses, implying that changes were not well tolerated. We hypothesized that this could be exploited therapeutically. Ribavirin is a viral mutagen approved for treatment of several virus infections; it is also cheap and readily available. In cell culture, we showed that ribavirin was effective at reducing production of infectious EBOV. In mouse and monkey models of therapy given post-EBOV exposure, ribavirin treatment delayed the time to death and increased survival. These data provide a better understanding of EBOV spontaneous mutation and suggest that ribavirin may have great value in the context of human disease.

GENETIC AND MORPHOTYPIC HETEROGENEITY OF SWIMMING POOL BACTERIAL POPULATIONS

The populations of resident bacterial species of the swimming pool community such as Pseudomonas aeruginosa, Bacillus sp., Acinetobacter lwoffii and Pseudomonas alcaligenes were analyzed. All these species showed stable in time heterogeneity by spontaneous mutation frequency and biofilm forming ability. There was notably high occurrence of mutators in all investigated populations. Our results show high level of genetic plasticity and adaptivity under conditions of starvation and exposure to biocides.

An orphan DNA (cytosine-5-)-methyltransferase in Vibrio cholerae

5-Methyl cytosine (m5C) was detected in genomic DNA of the enteric pathogen Vibrio cholerae by HPLC analysis and immunoblotting with m5C-specific antibody. Although cleavage with the restriction endonuclease EcoRII revealed the absence of a Dcm homologue in V. cholerae, analysis of the genome sequence indicated the presence of a gene, designated in this study as vchM, which encodes a DNA (cytosine-5-)-methyltransferase (m5C-MTase) designated M.Vch. M.Vch is not associated with a restriction endonuclease or a mismatch very short patch repair (Vsr)-like endonuclease and is hence an ‘orphan’ or solitary MTase, although analysis of a phylogenetic tree indicated that related cytosine MTases are all components of restriction-modification systems. M.Vch recognizes and methylates the first 5′ C in the degenerate sequence 5′-RCCGGY-3′. RT-PCR analysis suggested that vchM gene expression is increased during the stationary phase of growth. During stationary phase, the spontaneous mutation frequency in the V. cholerae wild-type strain was significantly higher than in the corresponding vchM mutant strain, suggesting that the presence of M.Vch and the absence of a very short patch (VSP) repair-like system imposes upon V. cholerae a mutator phenotype.

YtkD and MutT Protect Vegetative Cells but Not Spores of Bacillus subtilis from Oxidative Stress

ABSTRACT ytkD and mutT of Bacillus subtilis encode potential 8-oxo-dGTPases that can prevent the mutagenic effects of 8-oxo-dGTP. Loss of YtkD but not of MutT increased the spontaneous mutation frequency of growing cells. However, cells lacking both YtkD and MutT had a higher spontaneous mutation frequency than cells lacking YtkD. Loss of either YtkD or MutT sensitized growing cells to hydrogen peroxide (H2O2) and t-butylhydroperoxide (t-BHP), and the lack of both proteins sensitized growing cells to these agents even more. In contrast, B. subtilis spores lacking YtkD and MutT were not sensitized to H2O2, t-BHP, or heat. These results suggest (i) that YtkD and MutT play an antimutator role and protect growing cells of B. subtilis against oxidizing agents, and (ii) that neither YtkD nor MutT protects spores against potential DNA damage induced by oxidative stress or heat.