1985 — 2002 |
Levin, Bruce Richard |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Population Biology of Bacterial Viruses and Plasmids @ University of Massachusetts Amherst
The proposed investigations are to ascertain: i) the mechanisms responsible for the maintenance of plasmids in bacterial populations; ii) the nature and consequences of coevolution in bacteriophage and their hosts; iii) the ecological conditions and selection pressures responsible for the evolution and maintenance of restriction-modification systems; and iv) the ecological conditions and selection pressures responsible for the evolution and maintenance of temperate (as opposed to virulent) modes of bacteriophage replication. This investigation is to be performed at both a theoretical and empirical level. The former will involve the development and analysis of mathematical and computer simulation models of the population biology of bacteria with plasmids, virulent and temperate phage. The empirical portions of this investigation will be with naturally occuring, as well as laboratory strains of E. coli and its antibiotic resistance, R-plasmids, and virulent and temperate phage. Using experimental cultures of these organisms, estimates of the parameters of the models will be obtained and used for the numerical analysis. The validity and/or plausability of hypotheses generated from the analysis of these models and other sources will be tested with experimental (chemostat and serial transfer) populations of E. coli and its plasmids and phage and by the study of E. coli and phage from natural populations. --While this research is motivated primarily by the academic significance of these problems, the results obtained are anticipated to be of use for clinical epidemiological and other applied considerations of population biology of bacterial plasmids and viruses, e.g., the design of programs for the control of plasmid-borne multiple antibiotic resistance, the use of phage or plasmid-profile typing for bacterial clone idenfitication, and the design of programs for the use of bacteria and phage for biological control.
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1997 — 2008 |
Levin, Bruce Richard |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Population Genetics &Evolution of Antibiotic Resistance
DESCRIPTION (provided by applicant): Acquired antibiotic resistance is that which evolves in populations of susceptible commensal and pathogenic bacteria colonizing and infecting hosts that are under antibiotic treatment or prophylaxis. Acquired antibiotic resistance can result in treatment failure and contribute to transmissible or primary antibiotic resistance. The research proposed in this application will be devoted to understanding, in a quantitative and predictive way, the genetic, bacterial, host factors and population dynamic processes responsible for the evolution of acquired resistance in populations of bacteria infecting uncompromised mammals treated with single and multiple antibiotics. Towards this end, we will develop and analyze the properties of mathematical and computer simulation models of the within-host population dynamics of antibiotic treatment and the evolution of resistance and perform in vitro and in vivo (laboratory mouse) experiments with a capsulated E. coli (018:K1 :H7). In these experiments, we will estimate the parameters of these models, evaluate the reality of the assumptions behind their construction and test the validity of the predictions made from the analysis of their properties. The goals of this investigation are to; (1) Elucidate the conditions (dosage levels and treatment regimes) under which selection will favor the evolution of resistance in uncompromised mammals infected with antibiotic susceptible bacteria and treated with single antibiotics, multiple antibiotics and antibiotics for which clinical resistance requires multiple mutations. (2) Evaluate the contribution of post antibiotic effects (delays in the resumption of normal growth of antibiotic exposed bacteria after antibiotics are no longer at inhibitory concentrations) to the evolution of acquired resistance in populations of bacteria infecting antibiotic treated mammals. (3) Determine the contribution of elevated mutation rates to evolution acquired antibiotic resistance and the conditions under which antibiotic-mediated selection will result in the evolution of genes that augment mutation rates, mutator genes. The proposed research directed at these goals is in part, motivated by an academic interest in the mechanisms of adaptive evolution in bacteria. This research is also motivated by its direct utility to the health sciences and, in particular, to facilitate the design and evaluation of clinically effective antibiotic treatment protocols that minimize the likelihood of acquired antibiotic resistance evolving in the target population of bacteria.
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2009 — 2012 |
Levin, Bruce Richard |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Theoretical and Experimental Strudy of the Population and Evolutionary Dynamics O
DESCRIPTION (provided by applicant): Infections with resistant bacteria or resistance evolving during the course of treatment are major reasons antibiotic treatment fails. But this inherited resistance is not only reason treatment fails; patients remain ill for extensive periods or die due to infections with bacteria that are and remain fully susceptible to the antibiotics used for treatment. The goals of the proposed studies are to develop and evaluate antibiotic treatment protocols that are effective in rapidly clearing bacterial infections and, at the same time, minimize the likelihood of resistance evolving during the course of treatment. To achieve these goals, we will construct and analyze the properties of mathematical and computer simulation models that combine the pharmacodynamics of antibiotics and bacteria and the pharmacokinetics of the antibiotic treatment, with the population and evolutionary dynamics of bacteria in infected hosts. Using methicillin sensitive and resistant Staphylococcus aureus (MSSA and MSRA) and E. coli in invitro culture, we will estimate the parameters of these models and evaluate the validity of the assumptions behind their construction and test the predictions (hypotheses) generated from our analysis of their properties. Based on the results of these experiments, we will modify these models to make them more accurate and proposed the single and multi- drug treatment protocols to increase their efficacy in clearing bacterial infections and preventing the evolution of resistance. Of particular concern in these investigations are bacteria- and host-mediated processes that make genetically susceptible bacteria refractory to antibiotics. Included among these mechanisms of non-inherited resistance are subpopulations of non-growing bacteria (persistence) the physical structure of the infecting population (biofilms), the density of the infection, and physiological state of the bacteria (latent stages). PUBLIC HEALTH RELEVANCE: A theoretical and experimental study will be performed to improve the efficacy of antibiotic treatment and prevent the evolution of resistance. To achieve this end, we will use mathematical models, computer simulations and experiments with Staphylococcus aureus and E.coli. Particular consideration will be given to methicillin resistant S. aureus infections (MRSA).
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2014 |
Levin, Bruce Richard |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Diversity Supp: Population Genetics & Evolution of Antibiotic Resistant Bacteria
DESCRIPTION (provided by applicant): Infections with resistant bacteria or resistance evolving during the course of treatment are major reasons antibiotic treatment fails. But this inherited resistance is not only reason treatment fails; patients remain ill for extensive periods or die due to infections with bacteria that are and remain fully susceptible to the antibiotics used for treatment. The goals of the proposed studies are to develop and evaluate antibiotic treatment protocols that are effective in rapidly clearing bacterial infections and, at the same time, minimize the likelihood of resistance evolving during the course of treatment. To achieve these goals, we will construct and analyze the properties of mathematical and computer simulation models that combine the pharmacodynamics of antibiotics and bacteria and the pharmacokinetics of the antibiotic treatment, with the population and evolutionary dynamics of bacteria in infected hosts. Using methicillin sensitive and resistant Staphylococcus aureus (MSSA and MSRA) and E. coli in invitro culture, we will estimate the parameters of these models and evaluate the validity of the assumptions behind their construction and test the predictions (hypotheses) generated from our analysis of their properties. Based on the results of these experiments, we will modify these models to make them more accurate and proposed the single and multi- drug treatment protocols to increase their efficacy in clearing bacterial infections and preventing the evolution of resistance. Of particular concern in these investigations are bacteria- and host-mediated processes that make genetically susceptible bacteria refractory to antibiotics. Included among these mechanisms of non-inherited resistance are subpopulations of non-growing bacteria (persistence) the physical structure of the infecting population (biofilms), the density of the infection, and physiological state of the bacteria (latent stages). PUBLIC HEALTH RELEVANCE: A theoretical and experimental study will be performed to improve the efficacy of antibiotic treatment and prevent the evolution of resistance. To achieve this end, we will use mathematical models, computer simulations and experiments with Staphylococcus aureus and E.coli. Particular consideration will be given to methicillin resistant S. aureus infections (MRSA).
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2014 — 2017 |
Levin, Bruce Richard |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Population Dynamics and Evolution of Antibiotic Treatment and Resistance
DESCRIPTION (provided by applicant): Although pathogens with inherited resistance are an increasingly important problem, resistance is not the only reason antibiotic treatment fails and for some bacterial infections, like pneumococcal pneumonia, resistance is not a significant reason for treatment failure, at least not yet. One goal of this study is to develop and evaluate antibiotic treatment regimes that simultaneously maximize the rate of microbiological cure of infections with susceptible bacteria and minimize the likelihood of resistance emerging during the course of therapy. Another goal is to explore the potential efficacy of different antibiotics and pairs of antibiotics to treat infectons caused by widespread pathogenic bacteria that are designated non-susceptible (resistant) to the drugs currently employed for treatment. Towards these ends we will use a combination of mathematical and computer simulation models, parameter estimation, and pharmaco- population- and evolutionary- dynamic experiments to develop a framework for the design and interpretation of the results of different antibiotic choice and dosing regimens. These experiments will be done in vitro with antibiotic susceptible and resistant pathogenic strains of methicillin sensitive and resistant Staphylococcus aureus (MSSA and MRSA), Streptococcus pneumoniae, and Pseudomonas aeruginosa (including those from CF patients) each with antibiotics of four or more classes and pairs of these drugs. Particular consideration will be given to evaluating the (i) population dynamic and evolutionary consequences of exposure to low doses of antibiotics by these bacteria, (ii) the absolute and relative efficacy of different antibiotics and antibiotic pairs for treating infections of these bacteria within polysaccharide matrices known as biofilms or as colonies on the surfaces of tissues. Currently the criteria for susceptibility (resistance) and the rational design of antibiotic treatment are based on one and two parameters, respectively the Minimum Inhibitory Concentration (MIC) of the antibiotic estimated under conditions that are optimal for the action of the drug, and the MIC and one of three measures of the changes in the concentrations of the antibiotic in the plasma of patients, the peak concentration, the amount of time the concentration exceeds the MIC, or the area under the concentration time curve. As consequence of this parametric reductionism, are we not using antibiotics that could be effective? Are current antibiotic treatment regimes optimal for maximizing the rate of cure and minimizing the likelihood of resistance emerging and spreading during the course of therapy? This study is intended to answer these questions and identify measures to improve estimates of antibiotic susceptibility and the efficacy of antibiotic treatment.
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2020 — 2021 |
Levin, Bruce Richard |
R35Activity Code Description: To provide long term support to an experienced investigator with an outstanding record of research productivity. This support is intended to encourage investigators to embark on long-term projects of unusual potential. |
Theoretical and Experimental Studies of the Population and Evolutionary Dynamics of Bacteria and Bacteriophage.
ABSTRACT The viruses of bacteria and archaea, phage, are touted to be the most abundant organisms on Earth, and a great deal is known about their molecular biology, structure, and mechanisms of replication. Nevertheless there remain fundamental unanswered or incompletely answered questions about the population biology, ecological role, and evolution of these viruses. In this investigation we will address and provide answers to three of these fundamental questions: 1- What are the ecological, genetic and evolutionary conditions for lytic (virulent) phage to be maintained in and regulate the densities of bacterial populations? 2- Under what conditions will selection favor a temperate rather than a purely lytic mode of phage replication and transmission? 3- Under what conditions will selection by lytic and temperate phage favor the evolution and maintenance of CRISPR-Cas mediated adaptive immunity, rather than envelope or other constitutive resistance mechanisms? To address these questions, we will use mathematical and computer simulation models, the properties of which will be analyzed numerically with parameters estimated in the experimental systems that will be employed for population dynamic and evolutionary experiments. We will test the predictions (hypotheses) generated from our analysis of these models in populations of bacteria and phage maintained in liquid, surface, and semisolid culture. Based on the results of these experiments, we will modify our models to make them more realistic. The experiments will be done with Escherichia coli, Pseudomonas aeruginosa, Pseudomonas syringae, and Staphylococcus aureus and their lytic and/or temperate phage. In addition to their importance to academic ecology, population and evolutionary biology, the results of this basic science study may well have practical utility. This investigation will provide an empirically supported theory that could be used to facilitate the design and evaluation of programs to use phage for the treatment of bacterial infections in humans and domestic animals, and to control outbreaks of pathogenic bacteria in crops. !
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2021 |
Levin, Bruce Richard Weiss, David S [⬀] |
U19Activity Code Description: To support a research program of multiple projects directed toward a specific major objective, basic theme or program goal, requiring a broadly based, multidisciplinary and often long-term approach. A cooperative agreement research program generally involves the organized efforts of large groups, members of which are conducting research projects designed to elucidate the various aspects of a specific objective. Substantial Federal programmatic staff involvement is intended to assist investigators during performance of the research activities, as defined in the terms and conditions of award. The investigators have primary authorities and responsibilities to define research objectives and approaches, and to plan, conduct, analyze, and publish results, interpretations and conclusions of their studies. Each research project is usually under the leadership of an established investigator in an area representing his/her special interest and competencies. Each project supported through this mechanism should contribute to or be directly related to the common theme of the total research effort. The award can provide support for certain basic shared resources, including clinical components, which facilitate the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence. |
Heteroresistance Interdisciplinary Research Unit
ABSTRACT Antibiotic resistance is one of the most serious medical challenges of our time. This crisis puts patients at risk of untreatable bacterial infections and threatens major advances of modern medicine that rely on antibiotics (transplants, chemotherapy, etc). There are at least 2.8 million antibiotic resistant infections each year in the US, leading to over 35,000 deaths [1]. Without significant action, worldwide annual mortality due to these infections is predicted to reach 10 million by 2050, surpassing that predicted for cancer [2]. Understanding resistance mechanisms is critical to designing novel approaches and therapeutics to combat resistant bacteria. Heteroresistance (HR) is an enigmatic form of antibiotic resistance in which a bacterial isolate harbors a resistant subpopulation that can rapidly replicate in the presence of an antibiotic, while a susceptible subpopulation is killed [3, 4]. Not only do many species of bacteria exhibit this form of phenotypic resistance, but it has been reported against nearly all classes of antibiotics [3, 5, 6]. Unfortunately, our understanding of HR is extremely limited and its relevance during infection has been unclear. We recently demonstrated that HR to diverse antibiotics, including the last-line antibiotic colistin, can cause treatment failure in an in vivo model [4, 5, 7]. Furthermore, when the frequency of the resistant subpopulation is very low (<1 in 10,000 cells) HR is misclassified as susceptible by clinical diagnostic tests, yet is still able to mediate treatment failure [4]. Our surveillance data reveal that HR to diverse classes of antibiotics is widespread even among highly resistant carbapenem-resistant Enterobacteriaceae (CRE) and Acinetobacter baumannii (CRAB). Further, we recently discovered that targeting pan-resistant bacteria with two antibiotics to which a strain exhibits HR reliably leads to effective combination therapy, highlighting that knowledge of HR can be used to guide effective therapies [5]. Taken together, these data highlight a largely unappreciated and undetected epidemic of HR in the clinic that may cause unexplained antibiotic treatment failure but can also be exploited therapeutically. The Heteroresistance Interdisciplinary Research Unit (HR-IRU) brings together an interdisciplinary team of experts in an unprecedented effort to understand the mechanisms, dynamics, and prevalence of HR. The proposed projects, supported by Clinical Isolate and Single-Cell Analysis Cores, will use a combination of genetics, single cell microscopy, dynamic flow and in vivo infection studies, modeling, and epidemiological analyses to make foundational insights into HR. At a basic level, this work will significantly broaden our understanding of how traits exhibited by subpopulations of cells can impact bacterial physiology. At a translational level, this effort will be a critical step in our fight against antibiotic resistant bacteria and lay the foundation for the discovery of novel therapeutics, diagnostics, and approaches to alleviate human suffering.
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2021 |
Levin, Bruce Richard |
U19Activity Code Description: To support a research program of multiple projects directed toward a specific major objective, basic theme or program goal, requiring a broadly based, multidisciplinary and often long-term approach. A cooperative agreement research program generally involves the organized efforts of large groups, members of which are conducting research projects designed to elucidate the various aspects of a specific objective. Substantial Federal programmatic staff involvement is intended to assist investigators during performance of the research activities, as defined in the terms and conditions of award. The investigators have primary authorities and responsibilities to define research objectives and approaches, and to plan, conduct, analyze, and publish results, interpretations and conclusions of their studies. Each research project is usually under the leadership of an established investigator in an area representing his/her special interest and competencies. Each project supported through this mechanism should contribute to or be directly related to the common theme of the total research effort. The award can provide support for certain basic shared resources, including clinical components, which facilitate the total research effort. These scientifically meritorious projects should demonstrate an essential element of unity and interdependence. |
Heteroresistance Interdisciplinary Research Unit (Project 3)
ABSTRACT Heteroresistance (HR) is a phenomenon in which minority populations of antibiotic resistant bacteria are maintained in populations dominated by cells susceptible to that antibiotic. We have shown that HR is often undetected by clinical diagnostics, but that the subpopulations of resistant cells present in HR can lead to in vivo treatment failure. It is critical to gain a thorough understanding of HR in order to design more effective and sensitive diagnostics for its detection, and to guide clinical treatment. One major unaddressed area of investigation is which parameters control the dynamics of the resistant subpopulations in HR. The goal of Project 3 is to elucidate the dynamics of resistant subpopulations through experimental testing, supported by quantitative modeling of the pharmaco-, population- and evolutionary dynamics of heteroresistant bacteria. Importantly, this will include studies of both their dynamics upon antibiotic treatment as well as the reversion of the population toward susceptibility upon removal of the drug. Toward this end we will develop and analyze the properties of mathematical models of HR based on the mechanisms of heteroresistance derived from molecular, genetic and single-cell microfluidic studies in Projects 1 and 2. The parameters used for the numerical analyses of the properties of these models will be estimated with clinical isolates of HR Enterobacteriaceae (Enterobacter, Escherichia, Klebsiella) and Acinetobacter baumannii obtained from Core B and studied in depth in Projects 1 and 2. For each subpopulation we will estimate: (i) parameters of comprehensive pharmacodynamic functions, (ii) the rates of transition between susceptible and resistant states, and (iii) the fitness costs of these resistant states. Using Hollow Fiber Bioreactors, batch culture, and microfluidics, we will evaluate how well the models, with independent estimates of their parameters, fit the pharmacodynamic of HR-bacteria confronted with antibiotics and, with continuous culture devices, how well these models account for the dynamics of drug treatment of heteroresistant infections. In serial transfer culture we will estimate the rates of transition to baseline susceptible states following the removal of the antibiotics. Based on the results of these experiments, and in an iterative process, the models will be modified to make them more accurate and predictive analogs of the pharmacodynamics of HR. For each HR isolate studied, we will also perform experiments to determine if the frequency of the resistant subpopulations change as a consequence of antibiotic-mediated selection (i.e. if the baseline frequency of the resistant cells increases), and elucidate if there are conditions under which HR will be replaced by permanent resistance. The results from this research will for the first time, provide a broad and detailed understanding of the dynamics of HR, facilitating an understanding of the parameters that control the frequency of the resistant subpopulations. These studies will have a major impact on the development of diagnostic procedures to detect HR and the design of protocols for treating infections with bacteria that exhibit HR to the treating antibiotic.
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