VOLUME 69 : 2021

Electrochemical Impedance Spectroscopy (EIS)-based measurement of the binding and lysis activity of immobilized bacteriophage UP87 on Aeromonas hydrophila

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Mariah Nicole D. Dujunco ,Danilo V. Barcelon Jr., Donna May Dela Cruz-Papa, Jose H. Bergantin, Jr.

ARTICLE DOI: https://doi.org/10.53603/actamanil.69.2021.brlg5871

In this study, a measurement system based on Electrochemical Impedance Spectroscopy (EIS) was assembled to determine whether bacteriophage UP87 immobilized on a gold electrode surface would exhibit binding and lytic activity towards Aeromonas hydrophila. Bacteriophage UP87, a lytic phage for A. hydrophila, was immobilized on gold screen-printed electrodes (Au-SPEs), which contained a self-assembled monolayer (SAM) of 11-mercaptoundecanoic acid (MUA) and dithiothreitol (DTT), via carbodiimide cross-linker chemistry. The phage solution incubation time on the Au SPE was optimized to ensure maximum phage coverage of the electrode. The optimum time for phage UP87 immobilization was 60 minutes. To confirm the successful immobilization of the phage on the electrode surface, the Au-SPEs were characterized using Fourier Transform Infrared Spectroscopy (FTIR). The study of the binding and lysis of A. hydrophila was done through bacterial solution incubation on the treated Au-SPEs for different durations. Binding of the bacterial cells was shown by the general increase in the charge transfer resistance (Rct) up to a maximum value obtained after 60 minutes of incubation. Lysis of the captured bacterial cells was indicated through the decrease in Rct value after 80 minutes of incubation. In addition, the phage-modified Au-SPE exhibited higher Rct values at 60 minutes of incubation period for A. hydrophila compared with the non-target bacteria Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus, thus showing satisfactory selectivity. This high selectivity was attributed to the pre-treatment of

the Au-SPE which blocked nonspecific interactions and highly favored the binding of A. hydrophila. The results of this study indicate that surface immobilization of phages can be used to concentrate them for binding and lysing bacteria in solution. The concentration-dependence of the bacteria’s binding to the phage-modified surface hints at the possibility of using the phage UP87 as a biorecognition element in a selective biosensor for A. hydrophila using the EIS technique.



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