Proteomic Signatures of Clostridium difficile Stressed with Metronidazole, Vancomycin, or Fidaxomicin

Abstract

The anaerobic pathogen Clostridium difficile is of growing significance for the health care system due to its increasing incidence and mortality. As C. difficile infection is both supported and treated by antibiotics, a deeper knowledge on how antimicrobial agents affect the physiology of this important pathogen may help to understand and prevent the development and spreading of antibiotic resistant strains. As the proteomic response of a cell to stress aims at counteracting the harmful effects of this stress, it can be expected that the pattern of a pathogen’s responses to antibiotic treatment will be dependent on the antibiotic mechanism of action. Hence, every antibiotic treatment is expected to result in a specific proteomic signature characterizing its mode of action. In the study presented here, the proteomic response of C. difficile 630∆erm to vancomycin, metronidazole, and fidaxomicin stress was investigated on the level of protein abundance and protein synthesis based on 2D PAGE. The quantification of 425 proteins of C. difficile allowed the deduction of proteomic signatures specific for each drug treatment. Indeed, these proteomic signatures indicate very specific cellular responses to each antibiotic with only little overlap of the responses. Whereas signature proteins for vancomycin stress fulfil various cellular functions, the proteomic signature of metronidazole stress is characterized by alterations of proteins involved in protein biosynthesis and protein degradation as well as in DNA replication, recombination, and repair. In contrast, proteins differentially expressed after fidaxomicin treatment can be assigned to amino acid biosynthesis, transcription, cell motility, and the cell envelope functions. Notably, the data provided by this study hint also at so far unknown antibiotic detoxification mechanisms.

Proteomic Adaptation of Clostridioides difficile to Treatment with the Antimicrobial Peptide Nisin

“…GntR family members fulfil various biological functions in diverse bacterial groups. Amongst others, GntR-like proteins are involved in resistance to antimicrobial compounds including vancomycin and isoniazid, which inhibit cell wall synthesis [ 59 , 60 , 61 ]. In this study, we found the GntR-like protein CDIF630erm_01007 with significantly altered amounts after the addition of nisin to C. difficile .…”

“…Such divergent results could be potentially explained by differences in either, the applied antimicrobial concentration and the duration of treatment. Indeed, the existing literature points at different effects that can be observed when the same stressor is applied either in shock experiments (exponentially growing cells are treated with sub-lethal concentrations of the stressor) or in long-term stress experiment, where the bacterium is inoculated into a medium already containing the stressor and growth occurs in presence of this stressor for at least 16 h [ 53 , 61 , 65 , 66 ]. Moreover, the applicable concentration of a stressor seems to depend on the experimental setup followed.…”

“…Similarly, MurG accumulated up to 3.7-fold during adaptation of C. difficile to nisin, as observed in this study, supporting the assumption of a general role of MurG during adaptation and resistance development towards lipid II-targeting antimicrobials. On the other hand, it was found recently, that glucosamine-6-phosphate deaminase NagB (CDIF630erm_01147), glucose-6-phosphate isomerase Pgi (CDIF630erm_03585), both providing precursors for cell wall biosynthesis, and the peptidyl-prolyl cis-trans isomerase B PpiB (CDIF630erm_00459) involved in protein homeostasis of the cell envelope, exhibit a higher synthesis rate after vancomycin treatment in C. difficile [ 61 ]. In contrast, the analysis in this study revealed a lower amount of those proteins after nisin addition (up to 2.8-fold depletion), emphasizing unique features of each antimicrobial that are independent of the primary cellular target.…”

Impact of subinhibitory concentrations of metronidazole on proteome of Clostridioides difficile strains with different levels of susceptibility

“…These changes have been previously shown in a study on proteomic signatures of C . difficile stressed with sub lethal concentrations of MTZ, vancomycin, or fidaxomicin in exponential growth phase [ 56 ]. In addition to their roles in translation, some aminoacyl-tRNAs are also used in pathways directly implicated in antibiotic resistance.…”

Prevalence of infections with Clostridium difficile on potential pathology groups

“…As mentioned above, fidaxomicin is currently widely used in the treatment of Clostridium difficile infection, especially for recurrent cases. Fidaxomicin was approved in Germany and the US in 2011 for use in the treatment of Clostridium difficile infections in adult patients [17].…”