Time-resolved amino acid uptake of Clostridium difficile 630Δerm and concomitant fermentation product and toxin formation

Abstract

BackgroundClostridium difficile is one of the major nosocomial threats causing severe gastrointestinal infections. Compared to the well documented clinical symptoms, little is known about the processes in the bacterial cell like the regulation and activity of metabolic pathways. In this study, we present time-resolved and global data of extracellular substrates and products. In a second part, we focus on the correlation of fermentation products and substrate uptake with toxin production.ResultsFormation of different fermentation products during growth in a comparison between the two different media in a global approach was studied using non-targeted gas chromatography–mass spectrometry (GC-MS) based analysis. During cultivation in a casamino acids medium and minimal medium, the clinical isolate C. difficile 630Δerm showed major differences in amino acid utilization: In casamino acids medium, C. difficile preferred proline, leucine and cysteine as carbon and energy sources while glutamate and lysine were not or hardly used. In contrast, proline and leucine were consumed at a significantly later stage in minimal medium. Due to the more complex substrate mixture more fermentation products were detectable in the casamino acids medium, accompanied by major changes in the ratios between oxidative and reductive Stickland products. Different glucose consumption dynamics were observed in presence of either casamino acids or the minimal set of amino acids, accompanied by major changes in butanoate formation. This was associated with a variation in both the toxin yield and a change in the ratio of toxin A to toxin B.ConclusionsSince in all media compositions, more than one substrate was available as a suitable carbon source, availability of different carbon sources and their metabolic fate appears to be the key factor for toxin formation.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-015-0614-2) contains supplementary material, which is available to authorized users.

Control of Clostridium difficile Physiopathology in Response to Cysteine Availability

“…Interestingly, it has recently been shown that when C. difficile grows in minimal medium with Casmino Acids, cysteine is consumed immediately and sulfide is produced (53). This finding is in complete agreement with our results.…”

“…In addition, for Clostridum sticklandii, using amino acids as the carbon and energy sources, cysteine is one of the six amino acids that is preferentially degraded, while valine, leucine, and isoleucine are used later, suggesting that certain amino acids regulate the metabolism of others (71). Cysteine is also one of the three amino acids that are preferentially used by C. difficile (53). Our results suggest that the presence of cysteine may delay the use of other amino acids, such as BCAAs, which are known to act as corepressors of CodY (17).…”

The protein inventory of Clostridium difficile grown in complex and minimal medium

“…In general, the preferred carbon and energy source as well as the nature of fermentation products vary depending on the medium C. difficile grows in, as shown in an in‐depth metabolomics analysis by Neumann‐Schaal et al. .…”

Oxidative Stickland reactions in an obligate aerobic organism – amino acid catabolism in the Crenarchaeon Sulfolobus solfataricus

“…Secretion of short chain‐organic acids was monitored by extraction, analysis and data processing procedures as described earlier . For calculation of the absolute concentrations, calibration curves ranging from 4 to 2000 μ m for each compound were measured.…”