Rapid Colorimetric Detection of Bacterial Species through the Capture of Gold Nanoparticles by Chimeric Phages

Abstract

Rapid, inexpensive, and sensitive detection of bacterial pathogens is an important goal for several aspects of human health and safety. We present a simple strategy for detecting a variety of bacterial species based on the interaction between bacterial cells and the viruses that infect them (phages). We engineer phage M13 to display the receptor-binding protein from a phage that naturally targets the desired bacteria. Thiolation of the engineered phages allows the binding of gold nanoparticles, which aggregate on the phages and act as a signal amplifier, resulting in a visible color change due to alteration of surface plasmon resonance properties. We demonstrate the detection of two strains of Escherichia coli , the human pathogens Pseudomonas aeruginosa and Vibrio cholerae , and two strains of the plant pathogen Xanthomonas campestris . The assay can detect ∼100 cells with no cross-reactivity found among the Gram-negative bacterial species tested here. The assay can be performed in less than an hour and is robust to different media, including seawater and human serum. This strategy combines highly evolved biological materials with the optical properties of gold nanoparticles to achieve the simple, sensitive, and specific detection of bacterial species.

Controlled phage therapy by photothermal ablation of specific bacterial species using gold nanorods targeted by chimeric phages

“…Detection of Specific Bacterial Species by Phage-AuNRs. We previously reported a detection method for specific bacterial species using thiolated phages to target aggregation of gold nanospheres (AuNPs), which causes a red shift of LSPR peaks in the UV-vis spectrum (31). In this prior work, abundant thiol groups were incorporated on carboxylates of the phage capsid (with three or more solvent-accessible residues on each g8p protein) to induce aggregation of gold nanoparticles, and removal of free thiolated phage was required to remove background signal.…”

“…Five chimeric phages recognizing other Gram-negative bacterial strains (I + E. coli, P. aeruginosa, V. cholerae, and two strains of X. campestris) were propagated in E. coli cells, as previously described (31), and functionalized with AuNRs, as described above. As seen with M13KE-AuNRs targeting E. coli (SI Appendix, Fig.…”

“…To verify whether NIR irradiation destroyed the infectious potential of the phages, M13KE-AuNRs were irradiated for 10 min and then used to infect E. coli for phage propagation. Putative viral DNA was extracted and assayed by qPCR (31). No DNA was detected from propagation of the treated sample (SI Appendix, Fig.…”

“…First, greater delivery of nanoparticles per bacterial receptor could be achieved using phages due to the comparatively large surface area of phage, which may accommodate multiple nanoparticles; this property could be useful if bacterial receptors are in low abundance. A related benefit is that the aggregation of nanoparticles with phages on bacteria produces a visible shift in the LSPR spectrum (31), and one might therefore envision applications that combine treatment and detection of bacteria. Second, in addition to the targeting mechanisms evolved by whole phages as described above, chimeric phages can be rationally designed to achieve specificity against different bacterial hosts (8).…”

“…In this scheme, the phage confers specific targeting while the AuNRs achieve the desired effect. Using chimeric phages we previously engineered (31) to target several bacterial pathogens (two strains of Escherichia coli, Pseudomonas aeruginosa, Vibrio cholerae, and two strains of the plant pathogen Xanthomonas campestris), we first show that the phage-AuNRs can be used to detect specific bacteria through phage-mediated aggregation of AuNRs on the bacterial surface, which causes a red shift of the longitudinal LSPR peak. Next, we use NIR irradiation to induce death of the targeted bacteria via photothermal heating, both in solution and in a P. aeruginosa biofilm grown on a substrate of mammalian epithelial cells.…”