An Adapted Optical Density-Based Microplate Assay for Characterizing Actinobacteriophage Infection
Department
Biology
Document Type
Article
Publication Date
6-30-2023
Abstract
Bacteriophages are a key part of natural environments, and they have a powerful ability to shape bacterial populations. To understand how individual phages interact with slow-growing bacterial hosts such as actinomycetes, an easy and reliable method for quantifying long-term bacterial growth in the presence of phages is needed. Spectrophotometric microplate readers allow for high-throughput repeated measurements, but incubating a small volume for an extended time can present technical challenges. This procedure adapts a standard 96-well microplate to allow for the co-culturing of phages and bacteria without sub-sampling for 96 h, with the bacterial growth recorded every 8 h using spectrophotometric absorbance values. These optical density values are analyzed using R to yield infection metrics, including the percent growth inhibition, relative virulence, and the Stacy-Ceballos index. The methods outlined here provide an effective way to conduct and analyze extended-duration microplate growth curve experiments and includes modifications to reduce evaporation and lid condensation. These protocols facilitate microplate-based assays of interactions between slow-growing bacterial hosts and their bacteriophages.
Publication Title
Journal of Visualized Experiments
Publisher Statement
Copyright © 2023 MyJoVE Corporation
DOI
10.3791/65482
Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial-No Derivative Works 4.0 International License.
Recommended Citation
Christenson, Elijah I.; Zhang, Qingyang; and Plymale, Ruth, "An Adapted Optical Density-Based Microplate Assay for Characterizing Actinobacteriophage Infection" (2023). Articles. 377.
https://scholarlycommons.obu.edu/articles/377