A novel route to antibiotic resistance in Acinetobacter baumannii clinical isolates via IS26-mediated gene amplification
Christopher J. Harmer 1, Francois Lebreton 2, Jason Stam 2, Patrick T. McGann 2 and Ruth M. Hall 1
1 School of Life and Environmental Sciences, The University of Sydney, NSW, Australia.
2 Multidrug-Resistant Organism Repository and Surveillance Network (MRSN), Walter Reed Army Institute of Research, MD USA.
IS26 plays a major role in the mobilisation and accumulation of antibiotic resistance determinants in Gram negative bacteria. To date the focus has been on the ability of IS26 to create complex mosaic resistance regions containing several different antibiotic resistance determinants by employing an arsenal of different movement mechanisms. However, the role that IS26 plays in amplifying individual resistance genes had not been carefully examined.
An extensively resistant GC1 Acinetobacter baumannii isolate MRSN56 was recovered in 2010 from the wounds of a soldier injured in Afghanistan and repatriated to the USA for treatment. Treatment with tobramycin, one of the few remaining options, led to the emergence of tobramycin resistance, though no known tobramycin resistance genes were present. Subsequent examination showed that substantial amplification of Tn6020, which carries the aphA1 kanamycin and neomycin resistance gene, had led to aphA1 conferring an unexpected tobramycin resistance phenotype. As only short-read data were available for MRSN56 and the resistant isolates, the ability to examine the mechanics of amplification was limited and they were sequenced on an Oxford Nanopore MinION platform to generate complete genomes. They revealed surprising new capabilities of IS26. In each resistant isolate, IS26 was involved in the amplification of aphA1. In two instances, different IS26-mediated adjacent deletions which created a circular translocatable unit (TU) were followed by IS26-mediated targeted conservative cointegration to generate 7-65 copies of the TU containing Tn6020 in a tandem array. In another, homologous recombination between the two copies of IS26 in Tn6020 generated a circular TU followed by IS26-mediated copy-in cointegration which generated a tandem array of 77 copies of the TU.
Amplification of resistance genes is an important and under-appreciated route to the emergence of novel resistance phenotypes. The role that IS26 plays in gene amplification is likely to be substantial and requires further examination.