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A Novel, Rapid Phenotypic Assay for a Beta-Lactam Antibiotic Susceptibility and an Analysis of its Theoretical Limits


Liaw, Eric Jer-Jiun (2023) A Novel, Rapid Phenotypic Assay for a Beta-Lactam Antibiotic Susceptibility and an Analysis of its Theoretical Limits. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/qhvg-7q92.


Current management of bacterial infections is limited by the slow turnaround time of culture-based antibiotic susceptibility testing (AST). Culture-free phenotypic AST methods, though faster, are limited not only by analytical sensitivity but also by the low number, density, and purity of live pathogens present in clinical specimens before culturing. Separating and concentrating pathogens from clinical specimen matrices and improving the analytic sensitivity of phenotypic measurement technologies remain active areas of research. However, to date, the literature lacks consensus over what is a reasonable goal for the minimum number of pathogens in a clinical specimen needed to accurately perform phenotypic AST.

I describe "bulk filtration AST" and "digital filtration AST," two new filtration-based AST methods that improve an AST method previously published by others and myself. These methods use nucleic acid quantification to assess the activity of antibiotic classes (and only those classes) targeting peptidoglycan turnover, specifically the beta-lactams, which are the most frequently prescribed class of antibiotics. I use filtration AST to quantify the in vitro pharmacodynamics of beta-lactam antibiotics over time scales shorter than two hours, and I simultaneously validate the methods' accuracies on clinical isolates of Enterobacteriaceae. To analyze filtration AST results, either for fitting parameter values or for predicting susceptibility, I derive probabilistic models for the outcomes of each of the two filtration AST methods, then perform Bayesian parameter inference from my data.

I then propose a general mathematical framework for defining the concepts of the phenotypic assay and the ideal phenotypic assay. Within this framework, I calculate the ideal filtration AST performance as a function of the number of cells assayed, my fitted pharmacodynamic parameters, and other variables. Interestingly, the observed performance of my implementation of digital filtration AST is consistent with the implementation's approaching the ideal performance. I hope my demonstration of these new methods and my theoretical framework will help guide future research into rapid phenotypic AST.

Item Type:Thesis (Dissertation (Ph.D.))
Subject Keywords:antimicrobial susceptibility testing; rapid antimicrobial susceptibility testing; phenotypic antimicrobial susceptibility testing; digital assay; phenotypic assay; beta-lactam antibiotics; beta-lactam pharmacodynamics; Bayesian statistics; Markov birth-death process;
Degree Grantor:California Institute of Technology
Division:Biology and Biological Engineering
Major Option:Biology
Thesis Availability:Public (worldwide access)
Research Advisor(s):
  • Ismagilov, Rustem F.
Thesis Committee:
  • Murray, Richard M. (chair)
  • Ismagilov, Rustem F.
  • Newman, Dianne K.
  • Cai, Long
Defense Date:3 November 2022
Funding AgencyGrant Number
Defense Threat Reduction Agency (DTRA)MCDC-18-01-01-007
Burroughs Wellcome Fund Innovation in Regulatory Science AwardBWF.1014981
Center for Environmental Microbial Interactions (CEMI)UNSPECIFIED
Jacobs Institute for Molecular Engineering for MedicineUNSPECIFIED
Joan and Jerry Doren FellowshipUNSPECIFIED
NIHT32 GM008042
Record Number:CaltechTHESIS:05112023-130637882
Persistent URL:
Related URLs:
URLURL TypeDescription adapted for chapter 2
Liaw, Eric Jer-Jiun0000-0003-2244-8335
Default Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:15163
Deposited By: Eric Liaw
Deposited On:22 May 2023 19:50
Last Modified:05 Dec 2023 17:34

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