Abstract
Mycoplasmas are minute bacteria controlled by very small genomes ranging from 0.6 to 1.4 Mbp. They encompass several important medical and veterinary pathogens that are often associated with a wide range of chronic diseases. The long persistence of mycoplasma cells in their hosts can exacerbate the spread of antimicrobial resistance observed for many species. However, the nature of the virulence factors driving this phenomenon in mycoplasmas is still unclear. Toxin-antitoxin systems (TA systems) are genetic elements widespread in many bacteria that were historically associated with bacterial persistence. Their presence on mycoplasma genomes has never been carefully assessed, especially for pathogenic species. Here we investigated three candidate TA systems in M. mycoides subsp. capri encoding a (i) novel AAA-ATPase/subtilisin-like serine protease module, (ii) a putative AbiEii/AbiEi pair and (iii) a putative Fic/RelB pair. We sequence analyzed fourteen genomes of M. mycoides subsp. capri and confirmed the presence of at least one TA module in each of them. Interestingly, horizontal gene transfer signatures were also found in several genomic loci containing TA systems for several mycoplasma species. Transcriptomic and proteomic data confirmed differential expression profiles of these TA systems during mycoplasma growth in vitro. While the use of heterologous expression systems based on E. coli and B. subtilis showed clear limitations, the functionality and neutralization capacities of all three candidate TA systems were successfully confirmed using M. capricolum subsp. capricolum as a host. Additionally, M. capricolum subsp. capricolum was used to confirm the presence of functional TA system homologs in mycoplasmas of the Hominis and Pneumoniae phylogenetic groups. Finally, we showed that several of these M. mycoides subsp. capri toxins tested in this study, and particularly the subtilisin-like serine protease, could be used to establish a kill switch in mycoplasmas for industrial applications.
Highlights
Mycoplasmas are minute bacteria that evolved through regressive evolution from a Gram-positive ancestor with low G+C content related to the genus Clostridia [1]
They were thought to lack many of the common bacterial virulence traits including classical exotoxins and toxinantitoxin (TA) systems
We confirmed the presence of different functional TA systems in several isolates of the caprine pathogen Mycoplasma mycoides subsp. capri
Summary
Mycoplasmas are minute bacteria that evolved through regressive evolution from a Gram-positive ancestor with low G+C content related to the genus Clostridia [1] This extensive loss of genes affected many metabolic pathways including the biosynthesis of the peptidoglycan cell wall and even their capacity to synthesize essential cellular building blocks and nutrients, which make them strictly dependent on their hosts [2]. Bacterial persistence is generally associated with a state of growth arrest resulting in non-dividing cells that can survive environmental stresses such as temperature, pH, nutrient starvation [15] This phenomenon can be triggered in response to external factors such as phage infections or antibiotic exposure [16]. The continuous rise of antibiotic resistance observed for several mycoplasma species [17] could be the result of an increased selection of persister cells as it was recently shown for several natural and laboratory strains of E. coli [18]
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