Understanding Multidrug-Resistant Pathogen Infections and their Treatment with Antibiotics and Bacteriophages - PROJECT SUMMARY Antibiotic resistance is a global public health problem that has been aggravated in the last decades by the emergence and spread of multidrug-resistant microorganisms. Each year, it is estimated that resistant pathogens are responsible for more than 99,000 deaths in the United States. Particularly problematic is the ESKAPE group (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp), bacteria capable of ‘escaping’ the action of antibiotics and which represent new paradigms in pathogenesis, transmission, and resistance. Together with Escherichia coli, the ESKAPE group are responsible for most life-threatening bacterial infections in health care facilities. These bacteria exhibit an intrinsic low susceptibility to multiple antibiotics, as well as an overwhelming capacity to develop resistance via mutations. The ability to predict the mechanisms and genetic basis of antibiotic resistance is paramount to establishing new treatment protocols to fight these bacterial infections. To elucidate the evolutionary trajectories to antibiotic resistance in S. aureus, P. aeruginosa, and E. coli, and how the immune system changes these trajectories, experimental evolution assays will be performed in vivo using the larvae of Galleria mellonella (a lepidopteran model used for the study of infections). These Adaptive Laboratory Evolution experiments will be conducted in the presence of the antibiotics used clinically to treat infections caused by these bacteria. At multiple time-points and at the end of the experiment, the genetic basis of resistance and the order of appearance of these genetic variants will be characterized to determine the contribution of each mutation to the observed resistance. Furthermore, the fitness cost and the cross-resistance and collateral susceptibility of these mutations will be identified (Aim 1). In view of treatment failures arising from antibiotic resistance, novel strategies are needed to deal with these infections. Hence, there has been a resurgence of interest in the use of lytic bacteriophages for treating bacterial infections: phage therapy. Currently, phage therapy is only used as a last line therapy for patients for which no other treatment has been successful. Often, these patients are being treated with multiple antibiotics both before and during phage treatment. For phage therapy to become commonly employed, these bacterial viruses must be used in conjunction with antibiotics; and, moreover, we must know how these viruses, antibiotics, and the host’s immune system interact. To design and evaluate protocols based on the combination of antibiotics and lytic phages to treat bacterial infections, the research proposed will develop and analyze the properties of mathematical and computer-simulation models, estimate the parameters of these models in vitro and in vivo, analyze the properties of these models using G. mellonella, to ultimately determine the optimal combinations of antibiotics and lytic phages for the treatment of infections with S. aureus (Aim 2).
