Grant: Travel Awards - Institute Community Support

Genetically identical bacteria growing in the same environment exhibit highly individualistic phenotypes and behaviors, referred to as "phenotypic heterogeneity". These differences are attributed to variation in growth rates, gene expression, stress resistance, and other quantifiable phenotypes. Phenotypic heterogeneity can have a significant impact in the context of infection with a pathogen such as Mycobacterium tuberculosis, which is able to survive and establish a persistent infection in the host. Recent studies have highlighted the heterogeneous and divergent fates of individual lesions within a single host; however, these approaches so far have provided limited insight into phenotypic heterogeneity at the single-cell level. Here we report the development and characterization of fluorescent reporter strains of M. tuberculosis to measure various metabolic activities at single-cell resolution. We use a combination of microfluidic cultures and time-lapse microscopy to analyze bacterial phenotypes and growth behavior at the single-cell level. The new reporter strains were also used in murine models of tuberculosis to evaluate phenotypic heterogeneity during survival and persistence in the infected host. By culturing our reporter strains in microfluidic devices and documenting their growth and behavior by time-lapse microscopy, we were able to capture the remarkable phenotypic heterogeneity exhibited by mycobacterial populations during growth and how the metabolic states diversify when exposed to various environmental stresses, such as starvation or antibiotic treatment. Therefore, these strains may serve as a "phenotyping toolkit" to study diverse phenotypic states at the single-cell level. We are also carrying out infection studies in the murine TB model to capture this phenotypic heterogeneity in vivo during infection.