# Trade-off between canonical and unusual recombination sites promotes diversity and stability of gene cassette arrays of mobile integrons

## TL;DR

This study explores how mobile integrons balance different DNA recombination sites to maintain genetic diversity and stability in antibiotic resistance genes.

## Contribution

The study identifies novel ΔattI sites and demonstrates their functionality in gene cassette recombination.

## Key findings

- Novel ΔattI-type gene cassettes confer resistance to four antibiotic families.
- Gene cassettes with identical recombination sites show high excision frequencies (over 97%).
- A balance between canonical and unusual recombination sites maintains cassette array diversity and stability.

## Abstract

Mobile integrons are the most efficient mechanism of Gram-negative bacteria to resist antimicrobial changing pressures in the nosocomial niche. Integron’s integrases mediate site-specific recombination of distinct DNA structures such as attI, attC and attG sites. Here, we identified novel ΔattI sites as part of 26 unusual ΔattI-type gene cassettes conferring resistance to four different antibiotic families. Since scarce data are found related to their functionality, we investigated site-specific recombination mediated by IntI1 of attI1-aadB-attI1, attI1-aadB-ΔattI1− 11, attCaadA1-aadB-ΔattI2− 238, and attCaadA1-ybeA-ΔattI2− 11 genetic architectures. All proved to be functional with some displaying excision and insertion rates equal to canonical gene cassettes. Interestingly, gene cassettes with the same recombination site upstream and downstream of the ORF, i.e. either two equal attI1 or two attC sites, had an excision frequency of more than 97%, which outlines a scenario in which the canonical gene cassette, once inserted, is almost instantaneously excised. These findings evidence that a trade-off between different recombination sites, including attI, attC and ΔattI sites in canonical and unusual ΔattI-type gene cassettes, is necessary to maintain a stable and diverse gene cassette array within the variable region of mobile integrons.

The online version contains supplementary material available at 10.1038/s41598-026-36353-0.