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theses

p-Cresol is an aromatic organic compound that is widely used in the manufacture of chemical products such as disinfectants and preservatives. It is a toxic compound and considered as pollutant in the environment. Studies have discovered anaerobic microbial degradation of p-cresol under nitrate-reducing, sulfate-reducing, iron-reducing, and methanogenic environments (ref). p-Cresol is found naturally in the human gut system as a product of tyrosine and phenylalanine metabolism. Clostridium difficile infection (CDI) is a disease affecting the gut system with symptoms ranging from diarrhea to inflammation of colon. The infection usually occurs following a disruption of healthy microbial community such as after antibiotic treatment. The organism colonizes by producing toxins that cause death on the epithelial cells. C. difficile produces p-cresol and is able to tolerate higher concentration of p-cresol that would be too toxic for other microorganisms. It is one of the strategies for C. difficile to suppress the growth of other microorganisms and to maintain its dominance in the gut system during infection. Antibiotic treatment is effective for CDI but has 20-40% chance of recurrence. Fecal microbiota transplantation (FMT) has become an alternative treatment that is able to resolve recurrence of infection. It restores the microbial community in the gut back to its normal healthy state, which is hard to achieve with the antibiotic treatment. p-Cresol degradation has been found in various anaerobic environments and this study aims to investigate if there are any microorganisms in the human gut that are able to degrade p-cresol that can potentially be used to develop a more targeted FMT treatment for CDI. Anaerobic gut cultures were set up from stool samples of two donors under sulfate-reducing and methanogenic conditions with p-cresol amendment as carbon source. Four batches were set up throughout the experiment with different adjustments on the method to get a better representation of the gut culture. p-Cresol degradation was not observed in any of the gut cultures that were set up. On the other hand, p-cresol was produced in most of the gut cultures, both in the active cultures and the background controls. As a comparison to the gut cultures, two different environmental cultures were set up under methanogenic condition from freshwater sediments and anaerobic digestate from a waste water treatment plant and were able to degrade 0.5 mM p-cresol within 12 days of incubation. It confirmed p-cresol degradation that has been found in various environmental cultures in previous studies. High-throughput Illumina sequencing of 16S rRNA gene showed the change of microbial community in the gut cultures after incubation. It confirmed the presence of five phyla that has been reported to be found in the human gut, which are Firmicutes, Bacterioidetes, Verrucomicrobia, Proteobacteria, and Actinobacteria. The gut cultures showed a pattern of a decrease in Firmicutes and an increase in Bacteroidetes except for one batch from a post-antibiotic treatment stool sample. This batch had a different microbial composition on day 0 compared to other batches from healthy stool samples suggesting that antibiotic treatment can affect microbial community in the human gut as other studies have reported. The fourth batch were set up with different stool sample handling method and composition of media and showed a more balanced ratio of Firmicutes and Bacteroidetes before and after incubation, suggesting that the methods used in the gut culture set up could affect the microbial community composition in the culture.

ETD_8487

M.S.

Includes bibliographical references

by Ruzka Taruc

doi:10.7282/T3Q81H73

ETD graduate

The author owns the copyright to this work.