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

Sandra Maaß,Dörte Becher,Dirk Albrecht,Anke Trautwein-Schult,Andreas Otto,Katharina Riedel

doi:10.3390/cells7110213

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.

Highlights

As a spore-forming anaerobic bacterium that primarily causes antibiotic-associated diarrhea, Clostridium difficilerepresents an enormous financial burden for the health care system [2]
Comparative 2D PAGE was applied for the first time to detect, visualize, and quantify protein amounts and protein synthesis rates during stress adaptation in C. difficile
Adaptation to antibiotics was monitored after adding sublethal concentrations of vancomycin, metronidazole, or fidaxomicin to exponentially growing cells, resulting in reduced growth rates compared to an untreated control culture (Figure 1)

Summary

Introduction

Represents an enormous financial burden for the health care system [2] This is even more important as the incidence in hospitals is increasing, both in frequency and severity, resulting in considerable morbidity and mortality. In clinical practice most patients experiencing a first episode of Clostridium-associated diarrhea are cured by treatment with metronidazole or vancomycin, the former preferred for mild or moderate episodes and the latter for severe episodes [3,4,5]. Both antibiotics exhibit different chemical structures resulting in varying mechanisms of action. Today vancomycin is no longer considered a first-line treatment due to the increasing incidence of vancomycin-resistant enterococci [7,8], which continue to be a clinical challenge despite the availability of new therapeutic agents [9]

Results

Discussion

Conclusion