Effects of Environmental and Clinical Interferents on the Host Capture Efficiency of Immobilized Bacteriophages

Abstract

The concept of using bacteriophages to control populations of pathogenic bacteria is gaining momentum, driven mainly by the growing global crisis over antibiotic resistance in both the natural environment and healthcare settings. Bacteriophages (phages) are natural predators of bacteria and are innocuous to humans, animals or plants.Functionalizing surfaces with phage offers the promise of designing devices that can actively capture and deactivate bacteria such as water filters, wound dressings or antibacterial coatings. Our laboratory has previously proven the feasibility of this idea in a clean water matrix, demonstrating that phage-functionalized surfaces are promising candidates for selective capture and inactivation of bacterial pathogens. However, the complex composition of many natural samples (e.g., surface waters, waste water, blood, etc.) can potentially interfere with the interaction of phage and its bacterial host, leading to a decline in the efficiency of the phage-functionalized surface. In this study, the bacterial capture efficiency of phage-functionalized surfaces was assessed in the presence of potential environmental and biomedical interferents. Two phage-bacteria systems were used in this study, namely PRD1 with Salmonella Typhimurium and T4 with Escherichia coli. The potential environmental interferents tested were humic and fulvic acids, colloidal latex microspheres (as a model for environmental colloids), extracellular polymeric substances (EPS), as well as a natural unfiltered groundwater. Albumin, fibrinogen, and blood serum were also tested as representative interferents of interest for biomedical applications such as wound dressings. The inactivation of the selected phages by the potential interferents was first evaluated for each phage suspended in aqueous media containing each interferent. Next, the bacterial (host) capture efficiency of a iii phage-functionalized substrate was evaluated in the presence of each interferent.Interestingly, humic and fulvic acids reduced the capture efficiency of T4-functionalized surfaces by over 60%, even though they did not lead to inactivation of the suspended virions. Neither humics nor fulvics affected the capture efficiency of PRD1. EPS and human serum decreased the host capture efficiency for immobilized PRD1 and T4 by over 70%, and also impaired the infectivity of the non-immobilized (planktonic) phage, although to a much lower extent (less than 50%). The fundamental mechanisms leading to the observed decrease in performance of the phage-functionalized surfaces in the presence of selected interferents is discussed in detail in the thesis. These findings demonstrate the inadequacy of traditional phage selection methods (i.e., infectivity of suspended phage towards its host in clean buffer) for designing antimicrobial surfaces

Strategies for Surface Immobilization of Whole Bacteriophages: A Review

“…For immobilization on silicate-based materials, a preferred method is silanization of the substrate using alkoxysilanes which act as hetero-bifunctional crosslinkers between inorganic mineral surfaces and organic ligands 37,50,51,60,62,67,79,80,95,141,142,[144][145][146]173,174 .…”

“…However, a clean surface is also susceptible to non-specific binding of proteins in aqueous solutions. Non-specific binding of interferents can hinder specific biofunctionalization, obscure the transduction of specific interaction events and decrease the signal-to-noise ratio (SNR) in the case of biosensors, and generally compromise reproducibility, even when phage infectivity is not impacted 51 .…”

Optimizing Bacteriophage Surface Densities for Bacterial Capture and Sensing in Quartz Crystal Microbalance with Dissipation Monitoring

“…In many cases, these daughter phages eventually escape the host bacterium by lysis . Given these properties, surface immobilized bacteriophages can be used as a biorecognition element on acoustic, electrical, or optical bacterial biosensor platforms or even as a host-specific antimicrobial surface modification. − …”

Prevention of encrustation and blockage of urinary catheters by Proteus mirabilis via pH-triggered release of bacteriophage

“…Notable also is the stability of the phage within the coating, which remained viable throughout the entire experimental procedure, including encapsulation within the hydrogel reservoir, coating with an organic polymeric mixture, storage at 4 1C, and exposure to the challenging conditions of the bladder (involving both neutral and alkaline pHs, as well as the high ionic strength of the urine media). Phage stability with respect to clinical application is considered to be a barrier to their eventual use within the clinical setting, 50 hence the proof of stability within a hydrogel matrix, as well as the maintenance of infectivity in the face of more representative physiological conditions presented in this work signify the successfully maintained viability of the viruses despite the removal of the phage from its natural conditions and exposure to experimental interferents.…”