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Discovering bacterial genes involved in the conjugative transfer of antibiotic resistance genes

by Laetitia Van Wonterghem, Matteo De Chiara, Gianni Liti, Jonas Warringer, Anne Farewell, Natalie Verstraeten, and Jan Michiels

By harnessing the power of genome-wide association studies, researchers at the KU Leuven Centre of Microbial and Plant Genetics and the VIB-KU Leuven Center for Microbiology have revealed novel host-encoded factors regulating horizontal gene transfer in bacteria.

Antibiotic resistance impedes our ability to treat common bacterial infections and it is considered one of the biggest threats to public health by the World Health Organization. Therefore, it is crucial to protect the efficacy of current and future antibiotics. In this respect, conjugative DNA elements are worrisome since they can serve as a vector for resistance genes, facilitating transfer between bacteria within a patient or between a patient and the hospital environment. As a consequence, conjugation inhibitors can improve the success of antibiotic treatment by halting the transfer of resistance genes. The aim of the current study was to identify bacterial genes that are involved in conjugation and therefore likely to be promising targets for the development of conjugation inhibitors.

Graphical abstract of the research
Graphical abstract of the research

To reach this aim, we quantified the efficiency by which 113 diverse Escherichia coli strains transfer a conjugative plasmid using flow cytometry, and we determined the genome sequence of all strains. Subsequently, we identified candidate conjugation genes in a hypothesis-free manner by associating the measured phenotype to the genotype in genome-wide association studies (GWAS). We used the computational power of the nodes in the VSC Genius cluster to summarize the genotype of all 113 E. coli strains (each containing 4.5 to 5.5 million base pairs) into 833 942 unique DNA words (unitigs) and to associate every DNA word with the conjugation efficiency. This approach allowed us to select fliF, fliK, kefB and ucpA as candidate conjugation genes and their role in horizontal gene transfer was subsequently experimentally confirmed using corresponding single-gene knockout mutants. The encoded gene products are involved in bacterial motility, energy supply, control of the intracellular pH or salinity, suggesting that these processes also play a role in conjugation.

"Thanks to the computational power and customer support provided by the VSC, we were able to run GWAS with unitigs and identify genes involved in conjugation." Laetitia Van Wonterghem

To conclude, thanks to the computational power and customer support provided by the VSC, we were able to run GWAS with unitigs and identify genes involved in conjugation. This bioinformatics technique can be applied to link any measurable discrete or continuous phenotype of interest to the genotype and therefore holds great potential for future research as sequence information becomes more and more available.

The full publication can be consulted below:

Van Wonterghem, L., De Chiara, M., Liti, G., Warringer, J., Farewell, A., Verstraeten, N., Michiels, J. (2022). Genome-Wide Association Study Reveals Host Factors Affecting Conjugation in Escherichia coli. Microorganisms, 10 (3), Art.No. 608. doi: 10.3390/microorganisms10030608


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