Optical multi-channel interrogation instrument for bacterial colony characterization

Authors: DOH, IYLL-JOON - Purdue University; KIM, HUISUNG - Purdue University; STURGIS, JENNIFER - Purdue University; RAJWA, BARTEK - Purdue University; ROBINSON, J - Purdue University; BAE, EUIWON - Purdue University

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/11/2021
Publication Date: 2/25/2021
Citation: Doh, I., Kim, H., Sturgis, J., Rajwa, R., Robinson, J.P., Bae, E. 2021. Optical multi-channel interrogation instrument for bacterial colony characterization. PLoS ONE. doi.org/10.1371/journal.pone.0247721. https://doi.org/10.1371/journal.pone.0247721.
DOI: https://doi.org/10.1371/journal.pone.0247721

Interpretive Summary: Rapid detection and identification of human pathogens present in food samples remains a major issue in food safety. The use of elastic light scattering (ELS) technology has provided one rapid method of identification, although it has some limitations. ELS technology involves shining a laser on a bacterial colony resulting in a unique scatter image that can be used to identify the bacteria by comparison to a reference library of scatter images. This study describes a single instrument that includes multiple optical chRapid detection and identification of human pathogens present in food samples remains a major issue in food safety. The use of elastic light scattering (ELS) technology has provided one rapid method of identification, although it has some limitations. ELS technology involves shining a laser on a bacterial colony resulting in a unique scatter image that can be used to identify the bacteria by comparison to a reference library of scatter images. This study describes a single instrument that includes multiple optical channels to simultaneously measure various optical and associated biophysical characteristics of a bacterial colony. The multi-channel device can provide five distinct optical features without the need to transfer the sample to multiple locations or instruments. The available measurement channels are bright-field light microscopy, 3-D colony-morphology, 2-D spatial optical-density distribution, spectral ELS pattern, and spectral optical density. The series of multiple morphological interrogations is beneficial in understanding the bio-optical features of a bacterial colony and the correlations among them, resulting in an enhanced power of phenotypic bacterial discrimination. This new instrument will enhance the ability of ELS technology to accurately identify human pathogens in contaminated foods and to simultaneously measure various optical and associated biophysical characteristics of a bacterial colony. The multi-channel device can provide five distinct optical features without the need to transfer the sample to multiple locations or instruments. The available measurement channels are bright-field light microscopy, 3-D colony-morphology, 2-D spatial optical-density distribution, spectral ELS pattern, and spectral optical density. The series of multiple morphological interrogations is beneficial in understanding the bio-optical features of a bacterial colony and the correlations among them, resulting in an enhanced power of phenotypic bacterial discrimination. This new instrument will enhance the ability of ELS technology to accurately identify human pathogens in contaminated foods.

Technical Abstract: A single instrument that includes multiple optical channels was developed to simultaneously measure various optical and associated biophysical characteristics of a bacterial colony. The multi-channel device can provide five distinct optical features without the need to transfer the sample to multiple locations or instruments. The available measurement channels are bright-field light microscopy, 3-D colony-morphology map, 2-D spatial optical-density distribution, spectral forward-scattering pattern, and spectral optical density. The series of multiple morphological interrogations is beneficial in understanding the bio-optical features of a bacterial colony and the correlations among them, resulting in an enhanced power of phenotypic bacterial discrimination. To enable a one-shot interrogation, a confocal laser scanning module was built as an add-on to an upright microscope. Three different-wavelength diode lasers were used for the spectral analysis, and high-speed pin photodiodes and CMOS sensors were utilized as detectors to measure the spectral OD and light-scatter pattern. The proposed instrument and algorithms were evaluated with four bacterial genera, Escherichia coli, Listeria innocua, Salmonella Typhimurium, and Staphylococcus aureus; their resulting data provided a more complete picture of the optical characterization of bacterial colonies.