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Published on: May 23, 2021
Minimalistic mycoplasmas harbor different functional toxin-antitoxin systems
Virginia Hill1,2, Hatice Akarsu1, Rubén Sánchez Barbarroja1
1Institute of Veterinary Bacteriology, University of Bern, Bern, Switzerland.
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.
Insights
This study identifies and confirms toxin-antitoxin systems in Mycoplasma mycoides subsp. capri, revealing their role in bacterial persistence and potential for a mycoplasma kill switch.
Area of Science:
- Microbiology
- Bacterial Genetics
- Molecular Biology
Background:
- Mycoplasmas are small bacteria causing chronic diseases and contributing to antimicrobial resistance.
- Virulence factors in mycoplasmas, particularly those promoting persistence, are poorly understood.
- Toxin-antitoxin (TA) systems are known bacterial persistence factors, but their presence in mycoplasmas is largely uncharacterized.
Purpose of the Study:
- To investigate the presence and function of candidate toxin-antitoxin (TA) systems in Mycoplasma mycoides subsp. capri.
- To assess the role of TA systems in mycoplasma persistence and explore their potential applications.
- To confirm the functionality of TA systems in other mycoplasma species.
Main Methods:
- Genome sequencing of 14 M. mycoides subsp. capri strains to identify TA systems.
- Transcriptomic and proteomic analyses to study TA system expression.
- Functional validation of candidate TA systems using heterologous expression in M. capricolum subsp. capricolum.
- Investigation of TA system homologs in other mycoplasma phylogenetic groups.
Main Results:
- Confirmed the presence of at least one TA system in all sequenced M. mycoides subsp. capri genomes, including novel AAA-ATPase/subtilisin-like protease and AbiE/Fic-RelB pairs.
- Identified signatures of horizontal gene transfer associated with TA systems in several mycoplasma species.
- Demonstrated the functionality of three candidate TA systems in M. capricolum subsp. capri, confirming their role in neutralization and persistence.
- Confirmed functional TA system homologs in Hominis and Pneumoniae mycoplasma groups.
- Showcased the potential of M. mycoides subsp. capri toxins, especially the subtilisin-like serine protease, for developing a mycoplasma kill switch.
Conclusions:
- Toxin-antitoxin systems are prevalent in Mycoplasma mycoides subsp. capri and likely contribute to bacterial persistence.
- Horizontal gene transfer plays a role in the distribution of TA systems among mycoplasma species.
- The identified TA systems are functional and can be validated in related mycoplasma hosts.
- Mycoplasma TA systems, particularly specific toxins, offer promising avenues for biotechnological applications like kill switches.
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