# Dynamics of a large multidrug-resistant plasmid encoding New Delhi metallo-β-lactamase-1 and oxacillinase-58 carbapenemases in Acinetobacter baumannii clinical isolates from a tertiary hospital in Malaysia

## TL;DR

This study tracks the spread of a large plasmid in Acinetobacter baumannii that carries genes for antibiotic resistance, including NDM-1 and OXA-58, in a Malaysian hospital over several years.

## Contribution

The study reveals the ongoing horizontal transmission and structural variability of a multidrug-resistant plasmid in unrelated A. baumannii isolates.

## Key findings

- The plasmid carrying blaNDM-1 and blaOXA-58 was found in four new MDR A. baumannii isolates from 2018–2020.
- Structural variability in the plasmid was driven by insertion sequences and xrs modules, with some isolates lacking blaOXA-58.
- An 11.1 kb plasmid with tetracycline resistance genes was identified in one isolate, highlighting the role of xrs modules in AMR dissemination.

## Abstract

Acinetobacter baumannii carrying the blaNDM-1 gene, which encodes New Delhi metallo-β-lactamase-1 (NDM-1), exhibits resistance to nearly all β-lactams and is not affected by β-lactamase inhibitors, limiting treatment options. The blaNDM-1 gene is often associated with other antimicrobial resistance (AMR) genes, resulting in multidrug-resistant (MDR) phenotypes. We previously reported a large, circa (ca.) 170 kb plasmid co-harbouring blaNDM-1- and blaOXA-58-encoded carbapenemases in clinical MDR Acinetobacter nosocomialis and A. baumannii isolates from a tertiary hospital in Terengganu, Malaysia, in 2015 and 2016, respectively. In this study, we identified four additional MDR A. baumannii isolates from the same hospital (2018–2020) carrying blaNDM-1 on a similar plasmid. Complete genome sequences were obtained using a hybrid assembly of short-read DNA Nanoball Sequencing(DNBSeq) and long-read (Oxford Nanopore Technologies) data. All isolates belonged to distinct, unrelated clonal lineages, indicating the ongoing horizontal transmission of this plasmid. Comparative analysis revealed substantial structural variability among the plasmids, largely driven by insertion sequence elements and mobile xrs (also known as pdif) modules. In two A. baumannii isolates (i.e. AC1932 and AC2014), blaOXA-58 and several adjacent xrs modules were absent. Despite this, carbapenem minimum inhibitory concentrations remained comparable across all isolates, strongly indicating that blaNDM-1 is the primary carbapenem resistance determinant. Notably, in A. baumannii AC1839, we identified a 15,434 bp presumptive transposon carrying a type III cyclic oligonucleotide-based anti-phage signalling system, inserted within the xrs-rich region of the plasmid. AC1839 also carried a separate 11.1 kp plasmid carrying the tetracycline resistance genes tetA(39)-tetR within an xrs module. Identical xrs modules were identified in other unrelated Acinetobacter plasmids. These results underscore the mobility and potential roles of xrs modules in AMR gene dissemination and, more generally, in shaping Acinetobacter plasmid evolution.