Phage-based diagnostics have been implemented in various areas from biodefense to clinical settings. In this article, I highlight three FDA-approved or available products for phage-mediated detection of bacterial pathogens in the agricultural, clinical and biodefense environments.
Current methods used for microbial diagnostics
Currently, bacterial identification is still heavily culture-dependent. Though for many decades, phage typing was used to identify and distinguish different strains in a variety of sample types, the robustness of this assay is still lacking in that it requires meticulous maintenance of phage stocks and propagating strains. As they can be rapid and highly specific, more modern phage-based detection systems have the ability to bridge this gap.
What are examples of phage diagnostics products currently available?
How do they work?
Gamma Phage Assay: a traditional phage lysis assay
Initially developed in the 50s, the Gamma Phage Assay was modified in 2005 to become the first phage-based diagnostic test to gain FDA approval for human use within the CDC Laboratory Response Network for the detection of the biowarfare pathogen, Bacillus anthracis.
The traditional phage lysis assay is used alongside capsule detection to determine the presence and virulence of B. anthracis.
Bluephage®: a colorimetric assay
Coliphages have been researched extensively as possible indicators of fecal contamination. Bluephage® implements the use of somatic coliphages for fecal and viral detection and quantification in contaminated water. The technique provides detection of 1 somatic coliphage within 3.5 h, and is applicable to water, food and sludge samples.
The method utilizes a modified E. coli strain (knocked out uidB and uidC genes) incapable of intracellular glucuronic acid transportation. This subsequently overexpresses the uidA gene which encodes for the enzyme β-glucuronidase. Once the phages lyses the cell, the amassed enzyme within the bacterial cells is released into medium containing a substrate (chromogen) analogous to glucuronic acid, producing a color transition from yellow to blue to indicate a positive result.
Lumiphage: a bioluminescence assay
Guild BioSciences has engineered a reporter phage system that produces a signal upon infection of target bacteria. Bacterial (Vibrio harveyii) luciferase genes (luxA and luxB) were integrated into nonessential regions of a temperate phage genome to create a reporter phage capable of conferring bioluminescence.
The purified reporter phage is typically added directly to bacterial cultures, and as the phage is lysogenic and not lytic, bioluminescence increases exponentially with microbial growth.
Concluding remarks
Though there has been renewed interest in using phage as therapeutic tools and as an a potential answer to antibiotic resistance, the technologies highlighted here serve to remind us of the diversity in phage applications.
Beyond bacterial biocontrol and detection, phages have been and are used in areas including phage-derived enzymes, drug discovery and nanotechnology.
Further reading
Phage detection methods can be improved by combining them with techniques like real-time PCR and mass spectrometry to detect phage amplification. To learn more, have a look at these two examples:
- Sergueev, K. V., He, Y., Borschel, R. H., Nikolich, M. P., & Filippov, A. A. (2010). Rapid and sensitive detection of Yersinia pestis using amplification of plague diagnostic bacteriophages monitored by real-time PCR. PLoS One, 5(6), e11337. DOI: 10.1371/journal.pone.0011337
- Rees, J. C., & Barr, J. R. (2017). Detection of methicillin-resistant Staphylococcus aureus using phage amplification combined with matrix-assisted laser desorption/ionization mass spectrometry. Analytical and bioanalytical chemistry, 409(5), 1379-1386. DOI:
10.1007/s00216-016-0070-3
