Novel flow-through electrochemical biosensor for the detection of Listeria monocytogenes using oligonucleotides

Authors: Armstrong, Cheryl; Lee, Joseph - Joe; Gehring, Andrew; Capobianco, Joseph

Submitted to: Sensors
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/18/2021
Publication Date: 5/28/2021
Citation: Armstrong, C.M., Lee, J., Gehring, A.G., Capobianco Jr, J.A. 2021. Novel flow-through electrochemical biosensor for the detection of Listeria monocytogenes using oligonucleotides. Sensors. 21(11):3754. https://doi.org/10.3390/s21113754.
DOI: https://doi.org/10.3390/s21113754

Interpretive Summary: Food producers and regulators alike desire rapid in-line or near-line tests for identifying foods contaminated with bacteria that cause human illness. A novel method for detection of Listeria monocytogenes has been developed through the application of a previously developed biosensor platform, which was shown to identify low quantities of bacteria within unusually large volumes (up to 1 L) of foods samples. This previously developed sensor utilized antibodies to selectively identify target bacteria without the need for culture enrichment. However, antibodies that can reliably distinguish between closely related species of Listeria are not available. Therefore, to effectively apply this technology to Listeria, the shortcomings associated with antibodies were addressed by further expanding the sensors capabilities to accommodate genetic detection. Using this novel test, ~20,000 cells/mL could be detected within 2.5 hours of testing. Furthermore, the assay was capable of effectively distinguishing L. monocytogenes from the closely related species L. innocua; a current need identified by the Food Safety and Inspection Service.

Technical Abstract: Consumption of food contaminated by Listeria monocytogenes can result in Listeriosis, an illness with a hospitalization rate of 94% and a mortality rate of up to 30%. Due to the severity of the associated illness, regulatory agencies governing food safety in the United States retain zero tolerance policies for L. monocytogenes contamination in food products and processing environments. When pathogen detection is desired at such low concentrations, strategies such as increasing sample size or culture enrichment are often deployed to achieve these standards. Unfortunately, these tactics are not only time consuming but limit the ability for near-line detection. A novel flow-through electrochemical immunoelectrochemical biosensors was developed and has demonstrated promise for the rapid, sensitive detection of zero tolerance pathogens, such E.coli O157:H7, with 125 g samples in 1 L volumes without enrichment. In this work, the capabilities of this biosensor were further augmented to 1) include detection of L. monocytogenes and 2) accommodate genetic detection, which may address errors related to antibody specificity in phenotypic assays. Results of the present study demonstrated the sensors ability to specifically detect both synthesized DNA fragments as well as DNA from whole cell lysates. The sensor was able to distinguish L. monocytogenes from L. innocua with a limit of detection of ~20,000 cells/mL, which is in agreement with prior studies. As the protocol developed for this biosensor did not require a timely culture enrichment period, total assay time can be constrained to less than 2.5 hours. Furthermore, as the electrochemical detection assay can be performed with cost effective hand-held electronics, this sensor platform has the capability to be adopted for near-line product and/or environmental monitoring systems.