# A viable but nonculturable state of Mycobacterium avium in response to macrolide antibiotics: a recipe for relapses?

## TL;DR

This study explores how Mycobacterium avium becomes drug-tolerant by entering a nonculturable state when exposed to macrolide antibiotics, which may explain treatment failures and relapses.

## Contribution

The study provides evidence that macrolides induce a viable but nonculturable state in M. avium, offering a new perspective on drug tolerance mechanisms.

## Key findings

- Clarithromycin induces a viable but nonculturable state in M. avium.
- Bacteria in this state show lipid accumulation and reduced acid-fastness.
- Resuscitation-promoting factors can revive these nonculturable bacteria.

## Abstract

Mycobacterium avium complex disease is difficult to treat, with high failure and recurrence rates despite multidrug, macrolide-based treatments. The bacterial mechanisms involved in this drug tolerance and persistence are incompletely understood. Recent evidence has suggested persistence through metabolic adaptations indicative of the viable but nonculturable state, including a decreased respiratory rate and a switch to lipid accumulation and metabolism. To assess the contribution of switching to viable but nonculturable state to macrolide tolerance, we performed time–kill kinetics assays for clarithromycin against M. avium. In these experiments, we performed Auramine-O (for acid-fastness, representing active mycobacteria) and Nile red (for lipid accumulation) staining and stimulation using resuscitation-promoting factors of Mycobacterium tuberculosis. Loss of auramine staining, increased Nile red staining and increased population sizes after stimulation with resuscitation-promoting factors support the hypothesis that clarithromycin induces a viable but nonculturable state in M. avium. Induction of a viable but nonculturable state is one of the mechanisms of macrolide tolerance in M. avium. It might be one of the drivers of the high failure and recurrence rates of macrolide-based treatments. Antimicrobials active against viable but nonculturable M. avium may improve treatment outcomes.