Massively parallel transposon mutagenesis to identify relationships between biofilm formation and efflux activity in Enterobacteriaceae

Holden, Emma (2021) Massively parallel transposon mutagenesis to identify relationships between biofilm formation and efflux activity in Enterobacteriaceae. Doctoral thesis, University of East Anglia.

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Bacteria are usually found as part of structured, aggregated communities called biofilms. Progression through the biofilm life cycle requires temporally controlled gene expression to maximise fitness at each stage. Previous work identified that inhibition or deletion of efflux activity resulted in a severe reduction in biofilm formation, however the mechanism through which this occurs has not yet been described. In this work, I used TraDIS-Xpress; a massively parallel transposon mutagenesis approach to assay the impact of disruption or altered expression of all genes in the genome on biofilm formation and efflux activity in Escherichia coli and Salmonella Typhimurium. Pathways involved in biofilm formation in both species included fimbriae regulation and biosynthesis of flagella, nucleotides, curli and LPS. I identified genes with temporal contributions to biofilm fitness where their expression changed between being beneficial or detrimental depending on the stage at which they were expressed. Additionally, I characterised several genes in both E. coli and S. Typhimurium that had novel contributions to biofilm development. Efflux activity in both species was investigated in a similar way and identified genes involved in protein chaperoning, DNA housekeeping and signalling benefitted efflux in both species. Comparison of the genes and pathways involved in both biofilm development and efflux activity in both species revealed the importance of genes involved in DNA housekeeping, protein chaperoning, transcriptional regulation and stress responses. Overall, no one pathway was found to be the sole cause of the deficit in biofilm biomass seen in an efflux-deficient mutant. Therefore, it is most likely that disruption of efflux activity results in multiple pathways being altered, each of which impact biofilm matrix production to some degree. This work provides new insights into the requirements for successful biofilm formation through time and furthers our understanding of how biofilm development is affected by antimicrobial stress.

Item Type: Thesis (Doctoral)
Faculty \ School: Faculty of Medicine and Health Sciences > Norwich Medical School
Depositing User: Chris White
Date Deposited: 16 Mar 2022 16:12
Last Modified: 16 Mar 2022 16:12


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