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Title: Rapid nondestructive measurement of bacterial cultures with 3D interferometric imaging

Abstract

The agar culture plate has played a crucial role in bacteriology since the origins of the discipline. However, plating, inoculating, and waiting for microbes to develop colonies that are visible is time-consuming. In this work, we explored if white-light interferometry (WLI) would be a practical tool for reducing the wait time required before colonies can be identified and counted. High resolution surface profile imaging was used for non-destructive characterization and counting of bacterial colonies on agar before they became visible to the naked eye. The three-dimensional (3D) morphology of Gram-negative ( Pseudomonas fluorescens) and Gram-positive ( Bacillus thuringiensis) bacterial species were visualized over time by collecting surface profiles of colonies on agar plates with high vertical resolution (3-5 nanometers) and large field of view (3-5 mm). This unique combination of sensitive vertical resolution at low magnification enabled simultaneous detection of hundreds of microcolonies. The colonies were accurately counted within the first few hours after plating with results comparing favorably to counts made by traditional methods. Since the imaging is non-destructive it was used to track single cells multiplying into small colonies and the volumetric changes in the colony were used to accurately measure colony growth rates over time. Based onmore » the results herein, bioimaging with WLI was demonstrated as a powerful bacterial culture assay. Fast non-destructive counting of colony-forming units in a culture and simultaneous measurement of bacterial growth rates with this method suggest that it may lead to beneficial future applications in research, clinical, and other settings.« less

Authors:
ORCiD logo [1];  [1];  [1]; ORCiD logo [1]; ORCiD logo [1]; ORCiD logo [1]
  1. Pacific Northwest National Lab. (PNNL), Richland, WA (United States)
Publication Date:
Research Org.:
Pacific Northwest National Lab. (PNNL), Richland, WA (United States)
Sponsoring Org.:
USDOE
OSTI Identifier:
1525375
Report Number(s):
PNNL-SA-137620
Journal ID: ISSN 2045-2322
Grant/Contract Number:  
AC05-76RL01830
Resource Type:
Accepted Manuscript
Journal Name:
Scientific Reports
Additional Journal Information:
Journal Volume: 9; Journal Issue: 1; Journal ID: ISSN 2045-2322
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; bacterial colony; plate culture; counting; enumeration; CFU; white-light interferometry; bioimaging; optical imaging; surface profile; microscopy; growth rate; nondestructive

Citation Formats

Larimer, Curtis J., Brann, Michelle R., Powell, Joshua D., Marshall, Matthew J., Suter, Jonathan D., and Addleman, Raymond Shane. Rapid nondestructive measurement of bacterial cultures with 3D interferometric imaging. United States: N. p., 2019. Web. doi:10.1038/s41598-019-43839-7.
Larimer, Curtis J., Brann, Michelle R., Powell, Joshua D., Marshall, Matthew J., Suter, Jonathan D., & Addleman, Raymond Shane. Rapid nondestructive measurement of bacterial cultures with 3D interferometric imaging. United States. doi:10.1038/s41598-019-43839-7.
Larimer, Curtis J., Brann, Michelle R., Powell, Joshua D., Marshall, Matthew J., Suter, Jonathan D., and Addleman, Raymond Shane. Thu . "Rapid nondestructive measurement of bacterial cultures with 3D interferometric imaging". United States. doi:10.1038/s41598-019-43839-7. https://www.osti.gov/servlets/purl/1525375.
@article{osti_1525375,
title = {Rapid nondestructive measurement of bacterial cultures with 3D interferometric imaging},
author = {Larimer, Curtis J. and Brann, Michelle R. and Powell, Joshua D. and Marshall, Matthew J. and Suter, Jonathan D. and Addleman, Raymond Shane},
abstractNote = {The agar culture plate has played a crucial role in bacteriology since the origins of the discipline. However, plating, inoculating, and waiting for microbes to develop colonies that are visible is time-consuming. In this work, we explored if white-light interferometry (WLI) would be a practical tool for reducing the wait time required before colonies can be identified and counted. High resolution surface profile imaging was used for non-destructive characterization and counting of bacterial colonies on agar before they became visible to the naked eye. The three-dimensional (3D) morphology of Gram-negative (Pseudomonas fluorescens) and Gram-positive (Bacillus thuringiensis) bacterial species were visualized over time by collecting surface profiles of colonies on agar plates with high vertical resolution (3-5 nanometers) and large field of view (3-5 mm). This unique combination of sensitive vertical resolution at low magnification enabled simultaneous detection of hundreds of microcolonies. The colonies were accurately counted within the first few hours after plating with results comparing favorably to counts made by traditional methods. Since the imaging is non-destructive it was used to track single cells multiplying into small colonies and the volumetric changes in the colony were used to accurately measure colony growth rates over time. Based on the results herein, bioimaging with WLI was demonstrated as a powerful bacterial culture assay. Fast non-destructive counting of colony-forming units in a culture and simultaneous measurement of bacterial growth rates with this method suggest that it may lead to beneficial future applications in research, clinical, and other settings.},
doi = {10.1038/s41598-019-43839-7},
journal = {Scientific Reports},
number = 1,
volume = 9,
place = {United States},
year = {2019},
month = {5}
}

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