Figure 16.

Imaging the PG glycan backbone using MurNAc reporters. (A) E. coli ΔMurQ-KU was incubated in the presence of alkynyl MurNAc 42 and fosfomycin, fixed, subjected to CuAAC with azido-Cy5, and imaged by structured illuminated microscopy (SIM). Left, differential interference contrast (DIC); middle, SIM image; right, boxed area from middle image. (B) E. coli ΔMurQ-KU treated as in (A) and imaged by super-resolution 3D stochastic optical reconstruction microscopy (STORM). Red triangle indicates septal division plane. (C) E. coli ΔMurQ-KU were pre-treated with 42 and fosfomycin, then used to invade J774 macrophages. After fixation, CuAAC with azido-488, and nuclear staining with DAPI, invaded macrophages were imaged. White triangles indicate fluorescent fragments released from intracellular bacteria. Images in (A–C) are reproduced from ref ⁸⁹. (D) H. pylori HJH1 was incubated in the presence of alkynyl MurNAc 42 for a short pulse, fixed, subjected to CuAAC with azido-Alexa Fluor 555, counterstained with the fluorescent lectin WGA-Alexa Fluor 488, and imaged by 3D SIM. Image in (D) reproduced from ref ¹⁵⁴.