Pathogenic E. coli contamination in unpasteurized apple juice or cider is thought to originate from animal feces, and fecal contamination of apples has been recognized by the FDA as an important health issue. In a companion article, reflectance imaging techniques were shown inadequate for the detection of thin smears of feces applied to apples. The objective of this study was to evaluate the use of fluorescence imaging techniques to detect fecal contamination on apple surfaces. A hyperspectral imaging system based on a spectrograph, camera, and UV light source was used to obtain hyperspectral images of Red Delicious, Fuji Golden Delicious, and Gala apples. Fresh dairy feces were applied to each apple as both a thick patch and as a thin smear Results indicate that multispectral fluorescence techniques can be used to effectively detect fecal contamination on apple surfaces. Both principal component analysis and examination of emission maxima identified the same four multispectral bands (450, 530, 685, and 735 nm) as being the optimal bands to allow discrimination of contaminated apple surfaces. Furthermore, the simple two-band ratio (e.g., 685 to 450 nm) reduced the variation in normal apple surfaces while accentuating differences between contaminated and uncontaminated areas. Because of the limited sample size, delineation of an optimal detection scheme is beyond the scope of the current study. However, the results suggest that use of multispectral fluorescence techniques for detection of fecal contamination on apples in a commercial setting may be feasible.