Antisense Blockade of Efflux Systems in Gram-Negative Pathogens

Antibiotic resistant bacteria, aka "super bugs", are a critical threat to public health worldwide, as the medical community is running out of effective antibiotics against a growing number of bacteria. One of the ways that bacteria develop resistance to antibiotics is by utilizing efflux systems that are used to pump the antibiotic out. A strategy that is currently being investigated is to restore the susceptibility of these bacteria to antibiotics by using peptide-conjugated phosphorodiamidate morpholino oligomers (PPMOs) to suppress genes within these bacteria that encode components of efflux pumps. This project studied the effectiveness of PPMOs that target the AcrAB-TolC efflux pump, which is a major component of the intrinsic antibiotic resistance mechanisms of E. coli and K. pneumoniae. Experiments tested for the effect of the PPMO targeting the acrA gene, specific sequences within the acrA gene, and the tolC gene. The effect of the PPMO was measured by a change in the minimum inhibitory concentration (MIC) of common antibiotics such as Piperacillin/Tazobactam (Pip/Tazo), Azithromycin, and Levofloxacin on strains of these two bacteria. The results show that PPMOs targeted to the acrA gene have a 4-8 fold effectiveness at lowering antibiotic MICs for the bacterial strains. PPMOs that targeted the tolC gene, on the other hand, have no synergistic effect in lowering antibiotic MICs for the bacterial strains. In addition, changing the sequence of the PPMOs targeting the acrA gene was shown to have an effect, albeit small, on susceptibility to antibiotics, which suggests that targeting specific regions of a gene of interest can induce more or less susceptibility in the bacteria to antibiotics.