The State of the Art in Biodefense Related Bacterial Pathogen Detection Using Bacteriophages: How It Started and How It's Going

Abstract

Accurate pathogen detection and diagnosis is paramount in clinical success of treating patients. There are two general paradigms in pathogen detection: molecular and immuno-based, and phage-based detection is a third emerging paradigm due to its sensitivity and selectivity. Molecular detection methods look for genetic material specific for a given pathogen in a sample usually by polymerase chain reaction (PCR). Immuno-methods look at the pathogen components (antigens) by antibodies raised against that pathogen specific antigens. There are different variations and products based on these two paradigms with advantages and disadvantages. The third paradigm at least for bacterial pathogen detection entails bacteriophages specific for a given bacterium. Sensitivity and specificity are the two key parameters in any pathogen detection system. By their very nature, bacteriophages afford the best sensitivity for bacterial detection. Bacteria and bacteriophages form the predator-prey pair in the evolutionary arms race and has coevolved over time to acquire the exquisite specificity of the pair, in some instances at the strain level. This specificity has been exploited for diagnostic purposes of various pathogens of concern in clinical and other settings. Many recent reviews focus on phage-based detection and sensor technologies. In this review, we focus on a very special group of pathogens that are of concern in biodefense because of their potential misuse in bioterrorism and their extremely virulent nature and as such fall under the Centers for Disease and Prevention (CDC) Category A pathogen list. We describe the currently available phage methods that are based on the usual modalities of detection from culture, to molecular and immuno- and fluorescent methods. We further highlight the gaps and the needs for more modern technologies and sensors drawing from technologies existing for detection and surveillance of other pathogens of clinical relevance.

Keywords: bacteriophage; bioterrorism; biothreat; diagnostics; phage detection.

Figure 1

Summary of approaches used to detect biothreats using bacteriophage. (a) Methods dependent on the infection and metabolic activity or lysis of the bacteria cell, including reporter phages, molecular and immuno-detection for phage RNA/DNA and proteins, respectively, detection of phage proteins by mass spectrometry, and the traditional plaque assay or real-time, high throughput detection on the Omnilog™. (b) Use of phage components, including receptor binding proteins (RBP) reporter fusions to reporter molecules like florescent proteins, use of RBPs or lysins as capture agents on Qdots or gold nanoparticles, or use of lysins to release ATP that can be measured using a luminescent enzyme assay. (c) Engineered filamentous phages for spore detection and (d) as capture agents on plasmonic nanostructures coupled with surface enhanced Raman spectroscopy (SERS) for Brucella detection.