Extended-spectrum β-lactamase-producing Enterobacteriaceae: risks during antibiotic treatment and potential solutions to cure carriage

Abstract

The abundant consumption and negligent use of antibiotics have resulted in the global emergence of antibiotic-resistant bacteria. This is largely due to the rapid spread of multi-resistance plasmids in bacterial communities via conjugation. The increased carriage of extended-spectrum β-lactamase (ESBL)-producing Enterobacteriaceae in the human gut increases the probability of conjugative ESBL plasmids spreading to new bacterial hosts. Therefore, identifying factors that affect the dispersal of plasmids is essential to control their spread. In this thesis, I demonstrate that bacteria-harbouring ESBL plasmids can evolutionarily rescue antibiotic-susceptible cells in a bacterial community via conjugation even under lethal β-lactam concentrations. Thus, antibiotic-sensitive pathogens may also become resistant after an apparently efficient treatment is initiated. In this thesis, a conjugative clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 plasmid system (i.e., midbiotics) was developed to eradicate sequence-specifically different ESBL-bacteria from bacterial community, such as gut microflora. Several genes can be targeted simultaneously with a single midbiotic plasmid. The dispersal of the midbiotic plasmids results in efficient resensitisation of the exposed strains to β-lactams. However, before introducing this system in vivo, the following concerns need to be resolved: the dissemination of unwanted genes in the flora, mutations that nullify CRISPR activity, and the spread of the conjugative plasmid without its ESBL-targeting plasmid partner. In addition to midbiotics, lytic phages, which infect and kill resistant bacterial pathogens, may provide a potential option to decrease ESBL carriage. In this thesis, it was demonstrated that phages can be isolated on-demand from environmental reservoirs to carry out personalised phage therapy against Enterobacteriaceae, which are frequently associated with ESBL infections. Keywords: Antibiotic resistance; horizontal gene transfer; conjugative plasmids; bacteriophages; phage therapy; CRISPR-Cas9.