Protocol for detecting Yersinia pseudotuberculosis nitric oxide exposure during in vitro growth

Yersinia pseudotuberculosis (Yptb) is a bacterial pathogen that causes foodborne illness. Defense against the host antimicrobial gas, nitric oxide (NO), by the bacterial NO-detoxifying gene, hmp, promotes Yptb replication in mouse models of infection. Here, we detail the use of fluorescent signals as readouts for NO exposure within individual cells and subsequent detection of heterogeneity within a population, using single-cell imaging and analysis. This protocol quantifies NO exposure in culture, without capturing the full complexity of the host environment. For complete details on the use and execution of this protocol, please refer to Patel et al. (2021).