Does growth efficiency determine the inoculum effect?

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Project Summary/Abstract 2.8 million Americans are infected with antibiotic resistant bacteria annually. This leads to increased deaths and hospital costs. Moreover, there are few new antibiotics being developed. There is a dire need to extend the usefulness of existing antibiotics by understanding how bacteria tolerate antibiotic treatment. One way that bacteria tolerate antibiotics is the inoculum effect (IE), where the initial density of a bacterial population determines the concentration of antibiotic required to kill the population: a population of bacteria with a higher density will require more antibiotics to kill. IE has been observed in nearly all bacteria and antibiotics, and in the clinical setting. Surprisingly, we currently do not have treatment approaches that reduce IE, which would increase the ability to treat infections. It has been recently discovered that bacterial metabolic rate can determine the effectiveness of antibiotics. A higher metabolic rate reduces the amount of antibiotic required to kill a population of bacteria. This finding is particularly relevant in the context of IE, as bacterial density, metabolism, and growth are interrelated. Highly dense populations have a short period of growth before they exhaust their food source. This small window of growth leads to an overall low metabolic rate. Lower density populations have a longer period of growth before exhausting their food source. This long period of growth leads to an overall high metabolic rate. Thus, due its overall higher metabolic rate, the population with low density would require less antibiotic to kill relative to the high density population. This relationship between growth rate and metabolism can be described as growth efficiency. It is possible that changes in growth efficiency can determine IE, but we have not yet studied this. This is an important question to address as there is growing interest in using metabolite adjuvants with antibiotic treatment to increase antibiotic efficacy. However, without knowing how metabolism, or growth efficiency, affect IE, we risk enhancing the ability of bacteria tolerate antibiotics, which will complicate infections. The goal of this research is to investigate how growth efficiency determines IE. This will help us achieve our long term goal of altering bacterial metabolism to make bacteria easier to kill using antibiotics. To address our goal, we will first examine how growth efficiency determines IE in Pseudomonas aeruginosa and Staphylococcus aureus with a goal of determining growth conditions that can reduce, or even eliminate, IE. Next, we will discover what genes and biochemical pathways determine growth efficiency and IE towards developing a molecular mechanism. Our work may lead to the development of new antibiotics that reduce, or eliminate, IE. We will use a combination of experiments that measure growth, metabolism and the ability of antibiotics to kill bacteria. We will couple these experiments with mathematical modeling, computer simulations and bioinformatics. The proposed work will involve graduate and undergraduate students in interdisciplinary research with a focus on understanding antibiotic tolerance.