Cryptomonas pyrenoidifera organellar genomes and estimation of its ITS2 sequence diversity using lineage directed barcode primers

Cryptomonas is a common metropolitan freshwater microalgal genus that is easily recognizable due to its distinctive asymmetric cellular morphology and swimming pattern. There are currently 71 recognized Cryptomonas species but these can be difficult to identify by cellular morphology alone. This makes estimation of Cryptomonas diversity from environmental samples challenging as it requires culturing of strains followed by sequencing of genetic markers for clear identifications. In this paper we describe the isolation and culture of Cryptomonas pyrenoidifera from a small ephemeral pool in Southwestern Virginia, USA, with an annual recurrent population of this genus. Genomic resources from this isolate, including complete chloroplast and mitochondrial genomes, and nuclear and nucleomorph rRNA regions, were completed and archived. Metabarcoding, using lineage-directed primers designed to amplify the ITS2 region of Cryptomonas, was used to estimate an intra-specific variation of 99.3% for the C. pyrenoidifera ITS2 barcode from this natural environment. Metabarcoding also revealed that at least twelve phylogenetic species were present in this body of water. Our results provide new genomic resources for C. pyrenoidifera and suggest that metabarcoding with the ITS2 marker is a feasible approach to estimating Cryptomonas diversity and identification.