Cell preparation method with trypsin digestion for counting of colony forming units inCandida albicans-infected mucosal tissues

Abstract

Colony forming units (CFU) of Candida albicans in cell suspensions or homogenates prepared from C. albicans-infected tissues are not always accurate indicators of the severity of infection in mucosal tissues. In order to improve the reliability of CFU counts in the murine oral candidiasis model, we developed a new cell preparation method in which a dispersal process involving trypsin digestion was included in the processing of Candida albicans-infected tongue tissues. Trypsin digestion, which was added to the conventional method for preparing Candida suspension from tongue homogenates, improved the recovery yield as evidenced by an increase in Candida CFU's. This method also increased the number of planktonic Candida cells which could pass through a mesh filter, perhaps because the trypsin contributed to the break up of the complex mass of tissue-debris and C.albicans cells. Using this trypsin digestion technique, we confirmed the protective action of farnesol by a relative decrease in the number of viable Candida cells in homogenates of infected tongues, which was correlated with improvement of the symptoms of oral candidiasis. These results indicate that our new method of trypsin digestion is valuable in that it reflects the protective activity of some bioactive substances against mucosal candidiasis.