Volatile organic compounds as metabolic markers in bacterial and phage systems

Clostridium difficile and Staphylococcus aureus are dangerous known to cause major illness and mortality globally. Bacteriophages, or phages for short, have been used to treat bacterial infections such as the C. difficile strains R20291 and ATT. Metals such as copper, arsenic and cadmium can also be antimicrobial because these metals become toxic in excessive amounts. Volatile organic compounds (VOCs) are produced by bacteria during their various metabolic processes. These VOCs can potentially be used as biomarkers for the detection and diagnosis of bacterial infections. Environmental stress, such as exposure to phages and metals, can modify the metabolic profiles of C. difficile and S. aureus, hence altering their VOC production. In terms of VOC production, the effects of phages and metals on the metabolic pathways of C. difficile and S. aureus have not yet been investigated, as far as we know. Therefore, the purpose of this thesis is to investigate the metabolic profiles of C. difficile strains, the effect of certain phages on these strains, and the effect of the metals Cu, As, and Cd on S. aureus in terms of changing VOC production. Proton transfer reaction time-of-flight mass spectrometry (PTR-TOF-MS) was used to determine the metabolic pathway via detection of the VOCs released from the C. difficile strains, R20291, which leads to higher morbidity and mortality, and the ATT strain during their growth states. Furthermore, the effect of different phages on the alteration of metabolic pathways of the C. difficile strains R20291 and ATT in terms of VOC production was additionally examined. Solid phase microextraction-gas chromatography mass spectrometry (SPEM-GC-MS) is used to investigate the changes in metabolic pathways in S. aureus cultures with and without subinhibitory concentrations of Cu, As, and Cd via measuring VOC abundances.

Overall, the results indicate that the two C. difficile strains, R20291 and ATT, have distinct metabolic pathways based on the abundance of VOCs, which was determined to be greater in R20291, and this divergence may be attributed to genetic variation resulting in distinct active metabolic pathways. The results of the study show that the different phages have different effects on the alteration of metabolic pathways through the reduction of VOC abundance in infected C. difficile cultures. In comparison to a control, S. aureus treated with subinhibitory concentrations of copper, arsenic, and cadmium was shown to have lower abundances of nine, ten, and fourteen VOCs, respectively, where copper has the least impact on bacterial growth and cadmium has the greatest. In conclusion, the study has provided evidence that phages and metals can lead to alterations in the metabolic pathways of bacteria that, in general, result in a reduction in VOC abundances. Amino acids catabolism, TCA cycle pathways, and pyruvate pathways are three potential pathways that are impacted by phages and metals.