The detection of Escherichia coli biofilm in green oak with spectroscopic imaging techniques based on confocal laser scanning microscope

Suranaree University of Technology, Institute of Engineering, School of Agricultural Engineering 30000. Nakhonratchasima, THIALAND

^* Corresponding author: LSRYTW@g.sut.c.th

Abstract

Microbial recontamination in post- harvest products, specifically for fresh cut fruits and vegetables, often raises concern of consumer health safety since pathogen bacteria readily forms sticky biofilm for their protection that often render ineffective of cleaning or disinfection process. Therefore, tracking biofilm rather than bacteria cells may also be used for indication of microbial recontamination. This research focuses on acquisition of spectroscopic imaging and information for E. coli biofilm adherent on green oak leaf at microscopic level for visible bandwidth. A confocal laser scanning microscope (CLSM) was applied to investigate for the evident of biofilm formation. In order to enhance spectroscopic signal, staining surface sample with propidium iodide, commonly used for staining dead cell, was specifically investigated to a potential use for determination of E. colis biofilm as an evident of recontamination occurrence. Green oak leaf samples were inoculated with a small drop of viable E. coli of 2.7x10⁹ CFU/ml covering around 7x7 mm² and all samples were spectroscopic imaged every day for 7 days. Absorbance spectrum acquired by CLSM for the E. coli inoculated samples presented three identity peaks at wavelengths of 620, 670, and 690 nm but the latter identity wavelength appeared to include common autofluorescence of leaf surface physiology so arguable interpretation is unavoidable for 690 nm. In opposite, the identity peaks at 620 and 670 nm presented strong detection of E. coli ‘s biofilm for storage time beyond day 5 at a significant level of confident 99%. However, the lowest minimum detection limit in term of E. coli concentration required more precisely experimental validation.