SYTO16 labelling and flow cytometry of Mycobacterium avium

Mycobacterium avium cells were harvested from agar at different stages of their growth cycle, exposed to the minimum inhibitory concentration of isoniazid (INH) for 24 h and labelled with the fluorescent nucleic acid stain SYTO16. INH exposure led to a > 10-fold increase in the intensity of labelling in the majority of cells, and revealed discrete fluorescence peaks that were consistent with development of filamentous multinucleate cells during the growth cycle. Similar enhancement of labelling was observed in unfixed INH-treated cells viewed by fluorescence microscopy. INH appears to increase the permeability of Myco. avium cells to SYTO16. A combination of growth cycle-defined inocula, labelling with the new generation of fluorescent dyes and flow cytometry provides new opportunities to study the interrelationships between growth cycle events and antimicrobial susceptibility of mycobacteria.