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Topic

Breaking the Culture Barrier: Shaping the Future of Diagnostics with Direct-from-Specimen Omics

Department of Biochemistry and Microbiology

Date & location

• Friday, March 20, 2026
• 10:00 A.M.
• Clearihue Building, Room B017

Examining Committee

Supervisory Committee

• Dr. David Goodlett, Department of Biochemistry and Microbiology, ̽��ϵ�� (Co-Supervisor)
• Dr. Michael Chen, Department of Biochemistry and Microbiology, UVic (Co-Supervisor)
• Dr. Caroline Cameron, Department of Biochemistry and Microbiology, UVic (Member)
• Dr. Lisa Reynolds, Department of Biochemistry and Microbiology, UVic (Member)
• Dr. Fraser Hof, Department of Chemistry, UVic (Outside Member)

External Examiner

• Dr. Young Ah Goo, Department of Biochemistry and Molecular Biophysics, Washington University in St. Louis

Chair of Oral Examination

• Dr. Sang Nam, Gustavson School of Business, UVic

Abstract

Microbial analysis directly from specimen offers a powerful alternative to traditional culture-based diagnostics by enabling microbial detection and characterization in their native host environment. In the context of urinary tract infections (UTIs), reliance on urine culture remains a major limitation: culture is slow (24-48 hrs), often discordant with urinalysis (urine dipstick) results, and can obscure clinically relevant microbial features through ex vivo growth. This dissertation breaks the culture barrier by developing and applying direct from urine omics approaches that improve diagnostic speed while preserving biologically meaningful pathogen signatures.

A central focus of this work is the development of a lipidomics-based Fast Lipid Analysis Technique (FLAT) for culture-free detection of uropathogens using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). In a clinical cohort of 402 outpatient urine samples, FLAT correctly identified common uropathogens and showed 99% agreement with urine culture for negative samples, substantially outperforming urinalysis, which demonstrated only 37% agreement with culture. Using this approach, negative UTIs were rapidly ruled out in 77% of cases, reducing the need for culture and accelerating clinical decision making.

While early implementations of FLAT showed strong performance for Gram-negative bacteria, detection of Gram-positive uropathogens was limited by inefficient cardiolipin release. To address this, a lysozyme pretreatment step was introduced to disrupt the Gram-positive peptidoglycan layer prior to lipid extraction. Optimization using contrived urine samples resulted in a 100-fold improvement in the limit of detection. Validation in a clinical cohort of 76 culture-confirmed Gram-positive urine samples yielded a 95% detection rate, while Gram-negative detection remained uncompromised, enabling reliable identification of both organism classes within a single workflow.

Beyond diagnostics, this dissertation demonstrates the biological value of direct from specimen analysis for therapeutic target discovery. Proteomic profiling of Escherichia coli obtained directly from urine of UTI-positive patients was compared to the same isolates after a single laboratory passage. This analysis revealed substantial host-specific adaptation, with 37 proteins consistently present in patient-derived samples but absent after culture. These included outer membrane transporters, virulence-associated proteins, stress-response enzymes, and essential metabolic factors, many of which represent potential diagnostic markers or therapeutic targets that are missed by culture-based approaches.

Together, these findings establish direct from specimen lipidomics and proteomics as practical and informative alternatives to traditional culture. By combining rapid diagnosis with biologically relevant pathogen profiling, this work demonstrates how breaking the culture barrier can improve infectious disease diagnostics and expand our understanding of microbial behavior in the host environment.