Integrons are bacterial recombination systems that allow adaptation to environmental stresses and are largely responsible for the emergence and rise of antibiotic multiresistance in Gram-negative bacteria. These versatile systems function by capturing, stockpiling, excising and reordering mobile genetic elements known as cassettes (1). The stable platform of the integron contains an integrase gene (intI), and a cassette promoter (Pc) driving the expression of genes encoded in the variable cassette array located downstream of the integration point, the attI recombination site. Cassettes consist of promoterless coding DNA sequences (CDSs), associated with a recombination site (attC) (2). In this study, we developed a method to access cassette excision dynamics within the model SCI of V. cholerae (3) at a population level, using long-read sequencing technology. By taking advantage of the Oxford Nanopore's Read Until technology (4), we achieved a 30,000-fold enrichment of sequencing data precisely targeting the cassettes DNA within the SCI. This high-resolution approach enabled precise measurement of cassette excision rates and facilitated the detection of cassette recruitment events into a synthetic mobile integron. Our dataset will serve as the basis for the development of predictive models, elucidating parameters important for cassette excisions such as the attC site sequence, the cassette position in the array, and the size of the cassette. These models will also enable us to predict the likelihood of each cassette being recruited into mobile integrons.

References

1. Stokes,H.W. and Hall,R.M. (1989) A novel family of potentially mobile DNA elements encoding site-specific gene-integration functions: integrons. Mol. Microbiol., 3,

2. Escudero,J.A., Loot,C., Nivina,A. and Mazel,D. (2015) The integron: adaptation on demand. Microbiol. Spectr., 3,

3. Mazel,D., Dychinco,B., Webb,V.A. and Davies,J. (1998) A distinctive class of integron in the Vibrio cholerae genome. Science, 280,

4. Loose,M., Malla,S. and Stout,M. (2016) Real-time selective sequencing using nanopore technology. Nat. Methods, 13,