Characterization of the Mobility and Evolution of Non-Transmissible Plasmids

Project Information

EvoPlas

Grant agreement ID: 101062386

DOI

10.3030/101062386

Project closed

EC signature date 10 June 2022

Start date 1 July 2022

End date 30 June 2024

Funded under

• Marie Skłodowska-Curie Actions (MSCA)

Total cost

No data

EU contribution

€ 195 914,88

Coordinated by

INSTITUT PASTEUR
France

Periodic Reporting for period 1 - EvoPlas (Characterization of the Mobility and Evolution of Non-Transmissible Plasmids)

Reporting period: 2022-07-01 to 2024-06-30

Summary of the context and overall objectives of the project

Bacteria are extraordinarily adaptive organisms spread all over the globe and making up about half of the cells in our bodies. Their main source of novel functions is horizontal gene transfer, usually prompted by mobile genetic elements such as plasmids, that provide them with a wide range of traits. Plasmid conjugation is the main mechanism of dissemination of antimicrobial resistance (AMR), an increasingly important health problem. However, more than half of the plasmids found in bacterial genomes lack genes encoding conjugation functions and it is not known how they are transferred between cells. Since extensive data shows that many of these plasmids are transferred, it is important and urgent to study how this takes place to understand and predict their epidemiological patterns.

EvoPlas has two main objectives. First, to unravel the mechanisms by which the putatively non-transmissible plasmids are transferred between cells, thereby showing the actual ability of plasmids to spread within populations. Second, to identify the evolutionary origins of non-transmissible plasmids and understand their stability in bacterial lineages. EvoPlas will use bioinformatics and experimental biology to leverage massive genomic data in order to unravel key aspects of bacterial evolution by horizontal gene transfer. It will additionally provide cutting-edge knowledge on the evolution of bacterial conjugation, novel insights into integrative mobile genetic elements and provide major advances on the understanding of how AMR spreads. Hence, EvoPlas has the potential to uncover new insights into bacterial and plasmid evolution and provide novel approaches to fight against the spread of AMR.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

The project EvoPlas had three main objectives. The three objectives proposed were the following:

Objective 1: To characterize the diversity and distribution of oriTs.
To fulfil this objective, we collected and characterized the largest database of experimentally validated oriTs so far. During the procedure of this WP, we run into a risk previously foreseen in the section 3.1 of the DoA (Risk and Contingency table, WP1, Medium Risk): most of the oriTs are unknown, limiting our study to a few species. To overcome this limitation: (1) we focused the first characterization on the model species in which most oriTs are already described; and (2) we developed a novel methodology to ab initio identify oriTs in any bacterial species without the need of having already described cases.

Objective 2: To unveil the mechanisms of mobility of the NT plasmids.
To answer this question, we helped ourselves with the outcome of the previous objective. We used the collection of oriTs, and remaining data on non-canonical mechanisms of mobility, to characterize the mobility of those plasmids that were previously considered non-mobile. This objective was key to answer the first biological question highlighted in the section 1.1 of the DoA: “Do ‘non-transmissible’ plasmids move between bacteria more than envisioned?”.

Objective 3: To identify the evolutionary origin of NT plasmids.
This objective was an ambitious section of the project, whose accomplishment would answer the second and last biological question proposed in the DoA: “What is the evolutionary origin and fate of ‘non-transmissible’ plasmids?”. To reach this objective, we used the data obtained from the previous two objectives to: (1) understand the evolutionary origin of these plasmids from the degradation of previously conjugative replicons; and (2) to propose the theory that some could have evolved de novo from non-mobile elements.

All these results have led to three different scientific publications:
Ares-Arroyo M, Coluzzi C, Rocha EPC. Origins of transfer establish networks of functional dependencies for plasmid transfer by conjugation. (2023) Nucleic Acids Res. Apr 24;51(7):3001-3016. doi: 10.1093/nar/gkac1079. PMID: 36442505; PMCID: PMC10123127.
Ares-Arroyo M, Nucci A, Rocha EPC. (2024). Identification of novel origins of transfer across bacterial plasmids. Doi: 10.1101/2024.01.30.577996 [Under review in Nature Microbiology]
Ares-Arroyo M, Coluzzi C, Moura de Sousa J, Rocha EPC. (2024). EcoEvoRxiv. Hijackers, hitchhikers, or co-drivers? The mysteries of microbial mobilizable genetic elements. Doi: 10.32942/X2R89M [Under review in PLOS Biology]

Additionally, this data gave rise to scientific collaboration:
Benz F, Camara-Wilpert S, Russel J, Wandera KG, Čepaitė R, Ares-Arroyo M, Gomes-Filho JV, Englert F, Kuehn JA, Gloor S, Mestre MR, Cuénod A, Aguilà-Sans M, Maccario L, Egli A, Randau L, Pausch P, Rocha EPC, Beisel CL, Madsen JS, Bikard D, Hall AR, Sørensen SJ, Pinilla-Redondo R. (2024) Type IV-A3 CRISPR-Cas systems drive inter-plasmid conflicts by acquiring spacers in trans. Cell Host Microbe. Jun 12;32(6):875-886.e9. doi: 10.1016/j.chom.2024.04.016. Epub 2024 May 15. PMID: 38754416.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

Scientific impact:
The main goal of EvoPlas is to better comprehend the basis of plasmid mobility and evolution. As mentioned in the DoA of the project, plasmid mobility is the major cause of the dissemination of antimicrobial resistance. So, this project unlocks the study of molecular epidemiology and inference of spread of antimicrobial resistance in critical environments such as hospitals and nursing homes. Indeed, the results of this project goes in line with the first two objectives of the WHO/FAO/OIE Global Action Plan on Antimicrobial Resistance and in line with the EU Action Plan on Antimicrobial Resistance. In the end, in this project we have been able to identify major mechanisms of mobility that have been ignored for the past decades. We can ensure with confidence that the vast majority of plasmids are able to transfer between bacteria (which was under debate until now), and we can understand the patterns of plasmid mobility (most plasmids require other replicons to move).
Its scientific impact has been immediate, with a first publication as breakthrough article and cover of the Issue in Nucleic Acids Research (Vol 57, Issue 51), and two more publications being currently under review in Nature Microbiology and PLoS Biology, respectively. Notably, the project has been awarded by international conferences such as the ones organized by the International Society of Plasmid Biology, or the International Society for Microbial Ecology.

Industrial Impact:
Even more, some of the results have been focused on developing a method to infer ab initio the origins of transfer of plasmids and other conjugative elements. This is the first method ever described trying to identify this short non-coding regions without further information needed. The applications of this tool for biotechnological purposes are enormous, e.g. the delivery of plasmids as pharmaceutical tools, or bacterial manipulation in the laboratory. This tool is currently being under review for publication, being its potential usability groundbreaking in the future.

Societal Impact:
Regarding the societal impact, beyond the one directly produced by the scientific advances, we also find the one of communication. This key aspect is included in the AMR EU Action Plan, in which research, education and communication are essential pillars for the fight to tackle antimicrobial resistance.