Stress-induced formation of cell wall-deficient cells in filamentous actinomycetes

Abstract

The cell wall is a shape-defining structure that envelopes almost all bacteria. One of its main functions is to serve as a protection barrier to environmental stresses. Bacteria can be forced in a cell wall-deficient state under highly specialized conditions, which are invariably aimed at interrupting cell wall synthesis. Therefore, the relevance of such cells has remained obscure. Here we show that many filamentous actinomycetes have a natural ability to generate a new, cell wall-deficient cell type in response to hyperosmotic stress, which we call S-cells. This wall-deficient state is transient, as S-cells are able to switch to the canonical mycelial mode-of-growth. Remarkably, prolonged exposure of S-cells to hyperosmotic stress yielded variants that are able to proliferate indefinitely without their cell wall. This is the first report that demonstrates the formation of wall-deficient cells as a natural adaptation strategy and their potential transition into stable wall-less forms solely caused by prolonged exposure to osmotic stress. Given that actinomycetes are potent antibiotic producers, our work also provides important insights into how biosynthetic gene clusters and resistance determinants may disseminate into the environment.

Genome rearrangements and megaplasmid loss in the filamentous bacterium Kitasatospora viridifaciens are associated with protoplast formation and regeneration

“…The activation of transposases are typically stimulated by stressful conditions, such as radiation, oxidative stress, temperature or inhibitory concentrations of metals and antibiotics (Vandecraen et al 2017). It was recently demonstrated that elevated levels of osmolytes induces hyperosmotic stress (Fuchino et al 2017;Ramijan et al 2018), which are conditions that are also used during preparation of protoplasts. This stress could also stimulate transposition events and consequent chromosomal rearrangements.…”

“…For Pulsed-Field Gel Electrophoresis (PFGE), 10 6 spores ml -1 of K. viridifaciens were inoculated in 25 ml of TSBS with 0.5% glycine or LPB (Ramijan et al 2018). Cultures were grown at 30°C agitating at 200 rpm for 16 and 40 h, respectively.…”

Genome rearrangements and megaplasmid loss in the filamentous bacteriumKitasatospora viridifaciensare associated with protoplast formation and regeneration

“…The activation of transposases are typically stimulated by stressful conditions, such as radiation, oxidative stress, temperature or inhibitory concentrations of metals and antibiotics [38]. It was recently demonstrated that elevated levels of osmolytes induces hyperosmotic stress [25, 39], which are conditions that are also used during preparation of protoplasts. This stress could also stimulate transposition events and consequent chromosomal rearrangements.…”

“…Consistent with this idea is that other cell wall-deficient cells, called L-forms, which have likewise been exposed to osmotic stress conditions, carry chromosomal lesions. In this context it is interesting to note that in three independent L-form lineages of K. viridifaciens lesions in the right chromosomal arms were found in addition to loss of KVP1 [25]. These three strains retained a similar region of KVP1 in their genomes, with a size of 164,773 bp for alpha and M1, and 164,642 bp for M2.…”

“…For Pulsed-Field Gel Electrophoresis (PFGE), 10 6 spores⋅ml −1 of K. viridifaciens were inoculated in 25 ml of TSBS with 0.5% glycine or LPB [25]. Cultures were grown at 30°C agitating at 200 rpm for 16 and 40 hours, respectively.…”