Fig 4

(A) Schematic of CRISPRi-based motility LOF switch. Lower left table summarizes switch activity and bacterial behaviors +/− aTc. Bent arrows denote promoters; “T” denotes transcriptional terminators. Solid lines represent constitutive interactions; dashed lines represent induced interactions. (B) Experimental timelines used to investigate in situ inactivation of swimming motility. (C) A maximum intensity projection acquired by LSFM of an animal colonized by Vibrio^motLOF at 6 hpi. Dashed line marks approximate intestinal boundaries. An arrowhead with a black stroke marks an area of swimming cells expressing only dTomato (magenta, “switch = OFF”). White tailed arrowheads mark aggregated cells (green, “switch = ON”). (D) Population center of mass over time for intestinal populations of wild-type Vibrio (gray) and Vibrio^motLOF (magenta/green). Lines are single bacterial populations within individual fish. Vertical dashed line marks time of aTc induction. (E) Abundances of Vibrio^motLOF at 24 and 48 hpi with aTc. Bars denote medians and interquartile ranges. Significant differences determined by Mann-Whitney. Underlying data plotted in panels D and E are provided in S1 Data. aTc, anhydrotetracycline; CRISPRi, CRISPR interference; hpi, hours post induction; LOF, loss-of-function; LSFM, light sheet fluorescence microscopy.