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Low light optical detection of whole-cell bioluminescent bacterial bioreporters for visualisation of surface decontamination using nonthermal plasma

Robinson, Gareth M.; Thorn, Robin; Hancock, C.; Reynolds, Darren M.

Authors

Gareth Robinson Gareth2.Robinson@uwe.ac.uk
Senior Lecturer in Molecular Microbiology

Robin Thorn Robin2.Thorn@uwe.ac.uk
Associate Professor Molecular Life Sciences

C. Hancock



Abstract

Introduction:
Maintenance of a clean, uncontaminated healthcare environment is instrumental in the delivery of modern healthcare. Nonthermal plasma is a technology with potential applications for sterilisation and disinfection in biological and medical fields. We describe the use of Salmonella enterica serotype Typhimurium DT104 modified with the Photorhabdus luminescens lux operon as a rapid in situ reporter of successful surface decontamination.

Method:
Plates of Salmonella Typhimurium DT104 pGLITE were treated a) immediately after inoculation or b) 24 hours post-inoculation with a HandiPlasma nonthermal plasma generator (Creo Medical, Chepstow, UK) and bioluminescence imaged in situ using an iXon EM+ DU-897 back-illuminated EMCCD camera (Andor, Belfast, UK). Alternatively, a 100 ┬ÁL volume of liquid suspension of bacteria was deposited onto a 3D cast of a human hand, treated for 200 seconds and imaged immediately after.

Results and Discussion
Nonthermal plasma treatment of mature bacterial growth (high bacterial load) on agar plates, resulted in reduction in optical detection of light output compared to argon treated plates. Against freshly inoculated plates (moderate bacterial load), prior to incubation, non-thermal plasma prevented bacterial growth on the agar surface in a 25 mm diameter zone of inhibition around the applicator. This was not observed for the argon control (Fig 1). Against liquid culture inoculated onto a model polyurethane hand, non-thermal plasma produced a clear reduction in light output at the point of application, which was not observed for the argon control. The in situ optical detection of whole cell bioluminescent bacterial reporters has proven to be a flexible method for visualising decontamination on three dimensional surfaces, allowing real-time analysis of bacterial killing, in addition to spatial data regarding the location of contaminating bacteria.

Citation

Robinson, G. M., Thorn, R., Hancock, C., & Reynolds, D. M. (2013, May). Low light optical detection of whole-cell bioluminescent bacterial bioreporters for visualisation of surface decontamination using nonthermal plasma. Paper presented at 3rd International Conference on Bio-Sensing Technology

Presentation Conference Type Conference Paper (unpublished)
Conference Name 3rd International Conference on Bio-Sensing Technology
Start Date May 12, 2013
End Date May 15, 2013
Publication Date May 12, 2013
Peer Reviewed Peer Reviewed
Keywords low light optical detection, whole-cell, bioluminescent bacterial bioreporters, visualisation, decontamination, nonthermal plasma
Related Public URLs http://www.nature.com/natureevents/science/events/16200-3rd_International_Conference_on_Bio_Sensing_Technology
Additional Information Title of Conference or Conference Proceedings : 3rd International Conference on Bio-Sensing Technology