Validating tyrosinase homologue melA as a photoacoustic reporter gene for imaging Escherichia coli.
To understand the pathogenic processes for infectious bacteria, appropriate research tools are required for replicating and characterizing infections. Fluorescence and bioluminescence imaging have primarily been used to image infections in animal models, but optical scattering in tissue significantly limits imaging depth and resolution. Photoacoustic imaging, which has improved depth-to-resolution ratio compared to conventional optical imaging, could be useful for visualizing melA-expressing bacteria since melA is a bacterial tyrosinase homologue which produces melanin. Escherichia coli-expressing melA was visibly dark in liquid culture. When melA-expressing bacteria in tubes were imaged with a VisualSonics Vevo LAZR system, the signal-to-noise ratio of a 9×dilution sample was 55, suggesting that ∼20 bacteria cells could be detected with our system. Multispectral (680, 700, 750, 800, 850, and 900 nm) analysis of the photoacoustic signal allowed unmixing of melA-expressing bacteria from blood. To compare photoacoustic reporter gene melA (using Vevo system) with luminescent and fluorescent reporter gene Nano-lantern (using Bruker Xtreme In-Vivo system), tubes of bacteria expressing melA or Nano-lantern were submerged 10 mm in 1% Intralipid, spaced between <1 and 20 mm apart from each other, and imaged with the appropriate imaging modality. Photoacoustic imaging could resolve the two tubes of melA-expressing bacteria even when the tubes were less than 1 mm from each other, while bioluminescence and fluorescence imaging could not resolve the two tubes of Nano-lantern-expressing bacteria even when the tubes were spaced 10 mm from each other. After injecting 100-μL of melA-expressing bacteria in the back flank of a chicken embryo, photoacoustic imaging allowed visualization of melA-expressing bacteria up to 10-mm deep into the embryo. Photoacoustic signal from melA could also be separated from deoxy- and oxy-hemoglobin signal observed within the embryo and chorioallantoic membrane. Our results suggest that melA is a useful photoacoustic reporter gene for visualizing bacteria, and further work incorporating photoacoustic reporters into infectious bacterial strains is warranted.
University of Alberta, Department of Electrical and Computer Engineering, Donadeo Innovation Centre for Engineering, 9211-116 Street, Edmonton, Alberta T6G 1H9, CanadabUniversity of Alberta, Department of Oncology, Katz Group Centre, 114 Street & 87 Avenu.
Robert J
RJ
Paproski
University of Alberta, Department of Chemistry, 11227 Saskatchewan Drive, Edmonton, Alberta T6G 2G2, Canada.
Yan
Y
Li
University of Alberta, Department of Electrical and Computer Engineering, Donadeo Innovation Centre for Engineering, 9211-116 Street, Edmonton, Alberta T6G 1H9, Canada.
Quinn
Q
Barber
University of Alberta, Department of Oncology, Katz Group Centre, 114 Street & 87 Avenue, Edmonton, Alberta T6G 2E1, Canada.
John D
JD
Lewis
University of Alberta, Department of Chemistry, 11227 Saskatchewan Drive, Edmonton, Alberta T6G 2G2, Canada.
Robert E
RE
Campbell
University of Alberta, Department of Electrical and Computer Engineering, Donadeo Innovation Centre for Engineering, 9211-116 Street, Edmonton, Alberta T6G 1H9, Canada.
Roger
R
Zemp
D004867:Equipment Design; D019544:Equipment Failure Analysis; D004926:Escherichia coli / Q000166:cytology* / Q000201:enzymology*; D017930:Genes, Reporter / Q000502:physiology*; D008853:Microscopy / Q000379:methods*; D014442:Monophenol Monooxygenase / Q000378:metabolism*; D061088:Photoacoustic Techniques / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D014442:Monophenol Monooxygenase