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Salmonella enterica serovar Typhimurium must adapt to rapid environmental shifts, including those encountered upon entry and during replication to survive within macrophages during pathogenesis. Despite extensive RNA-seq-based investigations, questions remain regarding the range, timing and magnitude of response dynamics. Here we constructed a comprehensive GFP-reporter strain library representing 2,901 computationally identified Salmonella promoter regions to study time-resolved Salmonella transcriptional responses. Promoter activity was measured during in vitro growth and during intracellular infection of RAW 264.7 macrophages. Using bulk measurements and single-cell imaging, we uncovered condition-specific transcriptional regulation and population-level heterogeneity in SPI2-related promoter activity. We also discovered previously unidentified transcriptional activity from 234 promoters. These analyses revealed metabolic shifts including requirements for mntS expression to support manganese homeostasis and expression of Entner–Doudoroff pathway-associated genes to support growth within macrophages. Our library and datasets, made available through the online tool SalComKinetics, provide resources for systems-level interrogation of Salmonella transcriptional dynamics. 

Abstract Pandemic and endemic strains ofVibrio choleraearise from toxigenic conversion by the CTXφ bacteriophage, a process by which CTXφ infects nontoxigenic strains ofV. cholerae.CTXφ encodes the cholera toxin, an enterotoxin responsible for the watery diarrhea associated with cholera infections. Despite the critical role of CTXφ during infections, signals that affect CTXφ‐driven toxigenic conversion or expression of the CTXφ‐encoded cholera toxin remain poorly characterized, particularly in the context of the gut mucosa. Here, we identify mucin polymers as potent regulators of CTXφ‐driven pathogenicity inV. cholerae.Our results indicate that mucin‐associatedO‐glycans block toxigenic conversion by CTXφ and suppress the expression of CTXφ‐related virulence factors, including the toxin co‐regulated pilus and cholera toxin, by interfering with the TcpP/ToxR/ToxT virulence pathway. By synthesizing individual mucin glycan structuresde novo, we identify the Core 2 motif as the critical structure governing this virulence attenuation. Overall, our results highlight a novel mechanism by which mucins and their associatedO‐glycan structures affect CTXφ‐mediated evolution and pathogenicity ofV. cholerae, underscoring the potential regulatory power housed within mucus.