Submitted to: Journal of Food Science
Publication Type: Peer reviewed journal
Publication Acceptance Date: 10/20/2003
Publication Date: 12/20/2003
Citation: Chantarapanont, W., Berrang, M.E., Frank, J.F. 2003. Direct microscopic observation and visualization of campylobacter jejuni viability on chicken skin. Journal of Food Science. 66(12):2222-2230. Interpretive Summary: Campylobacter is a human bacterial pathogen that has been associated with poultry. Campylobacter can be found on the skin of broilers during and after processing. It has been unclear where on the skin Campylobacter attaches and survives the best. We developed a method to visualize Campylobacter on chicken skin. By using a confocal laser scanning microscope this can be done on fully hydrated skin without artificially modifying the surface. A strain of Campylobacter that fluoresces with a green color when exposed to certain wavelengths of light was used to inoculate chicken skin thus avoiding confusion with other bacteria present. Following rinsing, special dyes were applied to allow the discrimination between live and dead cells of Campylobacter. Campylobacter cells remaining on the skin after rinsing were mostly located in crevices, feather follicles or entrapped in water on the surface. Most living Campylobacter cells were in crevices and feather follicles. These small diameter openings provide a microenvironment in which Campylobacter can become firmly associated with the skin and survive. These findings and this method will allow further study of the ecology of Campylobacter on poultry skin, and may help in the effort to find ways to interfere with its attachment
Technical Abstract: The objective of this study was to develop a method to identify specific sites on chicken skin, which allow Campylobacter jejuni survival. This method employs confocal laser scanning microscopy (CSLM) visualization of Campylobacter jejuni transformed with Pcgfp plasmid (GFP-Campylobacter) exposed to 5-cyano-2,3-ditolyl tetrazolium choride (CTC), a redox dye which is taken up, reduced to CTC-formazan, and accumulated intra-cellularly in respiratory active cells. An Ar/Kr laser (excitation wavelength = 488 nm) was used to excite both GFP-Campylobacter and CTC. The emission wavelength of 483-495 nm as grey color for chicken skin-reflected light, 495-540 nm as green color fro GFP-Campylobacter image, and 600-670 nm as red color for CTC image. After 1 h inoculation with 2 ml of 108-109 cfu/ml GFP-Campylobacter suspension, 105-106 cfu of C. jejuni remained on 1 cm2 of chicken breast skin after rinsing. Green fluorescence of all C. jejuni colonies as well as the CTC-formazan in viable Campylobacter was clearly visible on chicken skin. The data indicated that GFP-Campylobacter remaining on the chicken skin surface after rinsing were mostly located in crevices, entrapped inside feather follicles with water and also entrapped in the surface water layer. Most of viable cells were entrapped with water in the skin crevices and feather follicles. These sites provide suitable microenvironment for GFP-Campylobacter to survive.