Saving our last-line antimicrobials

Antimicrobial resistance (AMR) is increasing globally, with low- and middle-income countries bearing the biggest burden of disease. Infections caused by two members of the Enterobacteriaceae family, Escherichia coli and Klebsiella pneumoniae, are particularly problematic due to their ability to harbour resistance mechanisms which result in the emergence of strains which are extensively resistant to antimicrobials of the B-lactam class. As this phenotype is predominantly driven by the acquisition of genes encoding enzymes which can hydrolyse the B-lactam ring the antimicrobial, the development of B-lactamase inhibitors (BLI)s has become an important strategy to mitigate the expression of the enzymes and restore the susceptibility of the bacteria to the antimicrobial. However, an increasing number of bacteria are able to undergo within-patient evolution and counter the BL + inhibitor combinations by inducing the hyperproduction of their B-lactamases, resulting again in AMR. Currently, there is limited research describing the mechanisms by which bacteria overcome the presence of BLIs and the overall changes within the genomic regulatory network that also occur in conjunction with resistance. There are even fewer studies focussing on bacteria originating from low-and middle-income countries, despite these areas being disproportionally affected by the burden of AMR. Additionally, studies that have been undertaken have focused on resistance to early-generation B-lactamase inhibitors (clavulanic-acid, tazobactam and sulbactam), rather than newer inhibitors (avibactam, relebactam and vaborbactam) or metallo-B-lactamase inhibitors - which are currently in the pre-clinical stage of development. Pre-emptively understanding how and why resistance may occur following the use of the last-line antimicrobials allows for mitigation strategies to be considered ahead of time, aiding the longevity of a given antimicrobial as a therapeutic option. The experimental approach taken will include both investigations into clinical isolates which are resistant to antimicrobials of interest; and the use of various culture-based approaches to generate resistance in vitro, which will then be further characterised using phenotypic assays. Long (ONT) and short-read (Illumina) whole genome sequencing will underpin these investigations and subsequent bioinformatic analyses will be used to elucidate the genomic basis of resistance.