The alkaline single-cell gel electrophoresis (comet) assay was used to investigate the genotoxicity of fecal water (FW) isolated from 47 Irish subjects using Caco-2 colonocytes as target cells. Two methods of comet assay analysis were compared to determine the extent of DNA damage and to categorize the samples as having no, low-to-moderate, or high genotoxicity. FW was isolated from stool samples by centrifugation and tested for its ability to induce DNA damage in Caco-2 cells. DNA damage was assessed using the comet assay by measuring the extent of DNA migration from the nucleus (microns, tail length) or by classifying the nuclei into five different categories depending on their morphology. Data collected from the two methods were used to categorize the FW samples on the basis of their genotoxic activity. Both methods showed good agreement. There was an approximately 50:50 split, with half the samples having some level of genotoxic activity and half having no genotoxicity. About one-third of the samples were considered to be highly genotoxic. There was a trend for low pH of the FW to be associated with increased DNA damage, but this was not significant. The results presented in this report show a relatively high incidence of genotoxic FW in samples derived from a free-living Irish population. Our data demonstrate the suitability of classifying nuclei on the basis of their morphology as a means of determining DNA damage. This procedure is very rapid and, therefore, advantageous in analyzing a large number of slides in the absence of an image analysis system.
Woods, J. A., Dunne, C., Collins, J. K., Shanahan, F., & O'Brien, N. M. (2002). Genotoxicity of fecal water in a free-living Irish population. Nutrition and Cancer, 42(1), 62-9. https://doi.org/10.1207/S15327914NC421_9