A Fluorescent Probe for Detecting and Identifying Genes Critical for Cell Entry.

The conventional method for quantitating () and relies on bacterial colony forming unit (CFU) enumeration on agar plates. Due to the slow growth rate of , it takes 3-6 weeks to observe visible colonies on agar plates. Imaging technologies that are capable of quickly quantitating both active and dormant tubercle bacilli and would accelerate research toward the development of anti-TB chemotherapies and vaccines. We have developed a fluorescent probe that can directly label the cell wall components. The fluorescent probe, designated as DLF-1, has a strong affinity to the D-Ala-D-Ala unit of the late peptidoglycan intermediates in the bacterial cell wall. We demonstrate that DLF-1 is capable of detecting in both the actively replicating and dormant states at 100 nM without inhibiting bacterial growth. The DLF-1 fluorescence signal correlated well with CFU of the labeled bacteria ( = 1 and 0.99 for actively replicating and dormant , respectively). DLF-1 can also quantitate labeled inside of cells. The utility of DLF-1 probe to quantitate was successfully applied to identify genes critical for cell invasion. In conclusion, this novel near infrared imaging probe provides a powerful new tool for enumerating with potential future use in bacterial virulence study.