Characterisation of nongrowing Salmonella during macrophage infection

Abstract

Persisters are antibiotic tolerant cells thought to be responsible for the recalcitrance and the recurrence of many bacterial infections. Salmonella has been shown to form persisters during macrophage infection, the source of which is a reservoir of nongrowing bacteria. This thesis describes the use of dual RNA-seq, combined with fluorescence dilution and FACS, to analyse the transcriptomes of nongrowing and growing Salmonella as well as the transcriptomes of their respective host macrophages during infection. Interestingly, nongrowing Salmonella were found to express the SPI-2 type three secretion system (T3SS) and infected macrophages were found to be polarised towards an M2 phenotype compared to uninfected bystander cells. Further activity in nongrowing Salmonella was uncovered using fluorescence dilution combined with DAPI-staining, which provided evidence of DNA synthesis during infection. In addition, the toxin-antitoxin module yhfG-fic was investigated in Salmonella after a deletion mutant in this module was found to have a defect during infection, however this was discovered to be due to a polar effect of the deletion on the expression of the downstream gene pabA. Heterogeneity in the lag distributions of growing and nongrowing Salmonella post-infection was also investigated in this study, using the ScanLag technique. Deletion of TA modules, which have previously been linked to persister formation, had a minimal effect of the lag distributions of nongrowing Salmonella, however the lag times of growing bacteria were increased. The SOS response was found to affect lag times and is important for Salmonella recovery post-infection, regardless of growth state.