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. 2020 Dec 2;6:58. doi: 10.1038/s41522-020-00166-4

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — The left panels show representative images of the BOAT assay performed with serial twofold dilutions (first six columns of the plate) of garvicin KS (a), micrococcin P1 (b) and both in combination (c) for the indicated strains. The concentration (in mg/ml) of the antimicrobials in the dilutions (dilution factors: D0–D7) is indicated on the far left of the images. At the same time, the assay was also performed using the control vehicles at their final concentrations (Ctrl, last six columns of each plate), see the “Methods” section for details on the composition. The development of red color indicates the retention of metabolic activity, and its quantification was performed by optical density readings at 492 nm (O.D. 492). The boxplots in the right panel show the trends of recovery of the bacterial metabolic activity as a function of the dilution factor for the different antimicrobials. Shown is the median distribution (thick line within boxes) and the degree of variability (amplitude of the box or interquartile region (IQR)) of the metabolic activities for the indicated strains measured at increasing dilution factors (D0–D7). Whiskers extending out the boxes mark the minimum and maximum observed values and the variability outside the middle 50% of values (whisker length). Outliers are represented as values that extend out of the whisker limit (1.5× IQR). Note that the combination treatment (GAK/MP1) led to a more delayed recovery of the bacterial metabolic activity compared to garvicin KS alone. d Boxplot showing the median distribution of logarithmic colony formation unit (Log₁₀CFU) values calculated after the BOAT assay for the indicated strains. The concentrations used were 5 mg/ml for garvicin KS and 0.1 mg/ml for micrococcin P1. The data represent the average values obtained from three independent experiments. Asterisks above the boxplots represent the statistical significance (p value) as determined by Welch’s t test by comparing the median of each group with that of the respective control (Ctrl) group. Asterisk representation of statistical significance: *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001; ****p ≤ 0.0001.