EXPERIMENTAL EVOLUTION OF VIBRIO CHOLERAE SHOWS INTERPLAY BETWEEN MOTILITY AND GROWTH INDEPENDENT OF CHROMOSOMAL POSITION OF RIBOSOMAL GENES.

LETICIA INÉS LAROTONDA; ALFONSO SOLER BISTUE; DIEGO COMERCI; DIDIER MAZEL

Los Cocos

Congreso; XVII Congreso Argentino de Microbiología General; 2022

SAMIGE Sociedad Argentina de Microbiología

It is unclear how gene order within the chromosome influences bacterial evolution. The genomiclocation of genes encoding the flow of genetic information is biased towards the replication origin(oriC) in fast-growing bacteria. Under optimal conditions, fast-growing bacteria overlap replicationrounds, a process called multi-fork replication. Hence, genes close to the oriC benefit from arelatively higher dose during exponential growth. In previous work we aimed at studying the role ofchromosomal location of genes encoding the flow of genetic information on cell physiology. Werelocated the S10-spec-α locus (S10), the main ribosomal protein gene locus, and rplKAJL-rpoBC(rpoBC) locus, encoding the catalytic core of the RNA polymerase, to different chromosomalpositions in the fast-growing pathogen V. cholerae. We found that growth rate (GR) and fitnessinversely correlated the distance between each locus and oriC. To gain insight into the evolutionaryeffect of RP genomic position, we evolved strains bearing S10 at its current location or far from oriCfor 1000 generations. Although all populations increased their GR along the experiment thedifferences between the latter and the former persisted. Deep-sequencing showed that populationsfixed on average 1 mutation each 100 generations mainly at genes linked to flagellum biosynthesisregulation, lipopolysaccharide synthesis, chemotaxis, biofilm and quorum sensing. We selectedfast-growing clones displaying a ~10% GR increment finding that they harbored inactivatingmutations at, among other sites, flagellum master regulators flrAB. Using a novel multiple genomeediting technique called Multiplex Genome Editing by Natural Transformation we introduced thesesingle mutations into naïve V. cholerae strains. We observed a ~10% increase in growth ratedemonstrating the role of these mutations in GR increase. Therefore, along the evolution V. choleraeincreases its GR independently of the position of genes encoding ribosomal genes. The lack ofsuppressor mutations compensating S10 genomic location demonstrates that gene order influencesthe evolution of bacterial lineages in the long run.