Urinary tract infections (UTIs) are one of the most common types of bacterial infection encountered by humans
throughout their lifespan and are a major health problem afflicting millions of people each year. Gram-negative
bacteria cause the majority of infections of which Escherichia coli is the primary pathogen and prone to
antibiotic resistance. The standard methods for species diagnosis and susceptibility determination are culture-
based protocols that take up to 48 h. As a result, UTIs are one of the most frequent reasons for antimicrobial
prescriptions in healthcare facilities, without the benefit of a confirmed diagnosis. Therefore, modern day rapid
diagnostic methods that promote rapid identification and antimicrobial stewardship are crucial.
The long-term objective of this R&D is to develop an easy to use platform for the rapid (=3 h)
identification and antimicrobial susceptibility testing (ID/AST) of UTI pathogens directly from urine. A
major goal is to develop a diagnostic that can bypass the need for bacterial amplification and isolation and thus
overcome the major time-limiting step of current diagnostics. The platform will consist of species-specific
bacteriophages (phages) engineered to express heterologous marker proteins, a lateral flow immunoassay,
and analysis by Sofia, a fluorescent assay reader produced and marketed by our Quidel collaborators. Upon
bacterial cell infection, the recombinant phages will produce large amounts of a foreign marker protein, which
following phage-mediated cell lysis, can be rapidly and sensitively detected by an immunoassay and
objectively read by Sofia. As the marker protein production is correlated to cell fitness, the platform termed
Phage-accelerated Test (PhACT), can rapidly determine an isolates sensitivity, or resistance to a particular
antibiotic, a key attribute in this 'post-antibiotic' era. The Phase I proof-of-principle studies will focus on
demonstrating feasibility for E. coli as this species is responsible for >70% of uncomplicated UTIs. Specific Aim
1 will generate a cocktail of genetically engineered E. coli phages harboring the foreign marker protein.
Specific Aim 2 will develop phage-mediated lateral-flow immunoassay and determine its performance
characteristics for direct ID/AST from urine samples. The research is significant as the technology enables
simultaneous detection and phenotypic drug susceptibility analysis without culture amplification and isolation,
thus bypassing the major time limiting step in diagnostics. This is key as decreasing length of time for
diagnosis and the administration of appropriate therapy positively impacts patient outcomes and promotes
antimicrobial stewardship.