Quantification of the dark fungal taxon Cryptomycota using qPCR.

Fungi are present in a wide variety of natural environments, and in the last years, various studies have shown that they are quite abundant in aquatic ecosystems. In addition, a whole new highly diverse phylum, the Cryptomycota, was discovered. Nevertheless, research on aquatic fungi and a detailed evaluation of their functions and distribution are still sparse. One of the main reasons is a limitation in reliable identification and quantification methods. To bridge part of the research gap, this study aims to implement a quantitative PCR method to detect and quantify the newly discovered phylum. We developed and validated a Cryptomycota-specific qPCR primer pair targeting the 5.8S region that detects the majority of Cryptomycota, but Microsporidia. The resulting amplicon is 102 bp long. We used different environmental samples to evaluate the primer pair, various fungal sequences as negative control and positive control sequences. Obtained amplicons were sequenced using Illumina, and the obtained ASVs were all classified as Cryptomycota. The qPCR method works reliably and specifically for the quantification of Cryptomycota in environmental samples.