A food color based colorimetric assay for Cryptococcus neoformans laccase activity

Cryptococcus neoformans is a fungal pathogen that causes cryptococcosis mostly in immune compromised patients, such as those with HIV/AIDS. One survival mechanism of C. neoformans during infection is melanin production, which catalyzed by laccase, and protects fungal cells against immune attack. Hence comparative assessment of laccase activity is useful for characterizing cryptococcal strains. We serendipitously observed that culturing C. neoformans with food coloring resulted in the degradation of some dyes with phenolic structures. Consequently, we investigated the color changes for the food dyes metabolized by C. neoformans laccase and explored using this effect for the development of a colorimetric assay to measure laccase activity. We developed several versions of a food dye based colorimetric laccase assay that can be used to compare the relative laccase activities between different C. neoformans strains. We found that phenolic color degradation was glucose dependent, which may reflect changes in the reduction properties of the media. Our food color based colorimetric assay has several advantages over the commonly used 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) assay for laccase activity, including lower cost, irreversibility, and does not require constant monitoring. This method has potential applications to bioremediation of water pollutants in addition to its use in determining laccase virulence factor expression.