The value of acridine orange and of electron microscopy in determining the viability of Mycobacterium leprae murium

Abstract 1. 1) An investigation has been made of the possibility of determining whether M. leprae murium were alive or dead by applying the acridine orange technique—living organisms have been said to fluoresce green and dead organisms to fluoresce red when treated with acridine orange. 2. 2) Suspensions of M. leprae murium were prepared from rat lepromata, subjected to various treatments, and then treated with acridine orange. It was not possible to differentiate living and dead organisms by differences in fluorescent colour. In general, the organisms in a fresh suspension fluoresced very faintly and the only treatment which resulted in all the organisms fluorescing red was heating the suspension in a water bath. 3. 3) It was shown that exposure to blue light of organisms which were fluorescing red resulted in a change in the fluorescent colour from red to green. This was the result of a change in the acridine orange itself due possibly to photo-dimerization of the dye. 4. 4) The morphology of the organisms as observed by electron microscopy appears to be of value in assessing viability. Two forms of the organism were seen, one of which is believed to be the normal form and the other a degenerate form which is non-viable. Determination of the percentage of degenerate forms gives an indication of the viability of a suspension.