Generation and maintenance of the circularized multimeric IS26-associated translocatable unit encoding multidrug...

Area of Science:

• Microbiology
• Molecular Biology
• Genetics

Background:

• IS26 elements are frequently found in multidrug resistance (MDR) regions of gram-negative bacteria.
• IS26 forms pseudocompound transposons (PCTn) capable of tandem amplification.
• A circular intermediate, the translocatable unit (TU), is generated but previously only detected via PCR.

Purpose of the Study:

• To investigate the generation and amplification of TUs from PCTn in a Klebsiella pneumoniae MDR clone.
• To understand the dynamics of TU formation and amplification under antimicrobial selection.
• To elucidate the role of RecA in the amplification processes.

Main Methods:

• Utilized a Klebsiella pneumoniae MDR clone containing an MDR plasmid with IS26 elements.
• Applied antimicrobial selection pressure to induce and study amplification.
• Analyzed mono- and multimeric forms of the TU and their amplification states.
• Investigated the RecA dependency of TU generation and amplification.

Main Results:

• Mono- and multimeric forms of the TU were generated from the PCTn in the MDR plasmid.
• The inserted TU form also underwent tandem amplification within the plasmid.
• Amplification was induced by antimicrobial selection and maintained without antibiotics.
• Circularized TUs were generated in a RecA-dependent manner from tandemly amplified TUs.
• Tandemly amplified TUs could be generated via RecA-dependent and independent pathways.

Conclusions:

• Demonstrated direct generation of mono- and multimeric TUs from PCTn in a K. pneumoniae MDR clone.
• Showcased that IS26-mediated amplification can be stable, persisting post-selection.
• Highlighted the dual role of RecA in TU formation and amplification, providing novel insights into bacterial genome plasticity.