Evaluating Species Delimitation Methods in Chloroidium (Trebouxiophyceae, Chlorophyta): Efficacy of DNA Barcodes and Description of Chloroidium pseudoellipsoideum sp. nov. from Arctic Soils.

Despite extensive research into green microalgae belonging to the genus Chloroidium, their species diversity and biotechnological potential remain poorly characterized. The strain VKM Al-418, the subject of this study, was isolated from the soil of Duvannyi Yar (Russian Federation). The independent species status of this strain is supported by distinct morphological characteristics, robust phylogenetic placement based on the 18S-ITS1-5.8S-ITS2 fragment, and unique features in the secondary structures of both ITS1 and ITS2, including one compensatory base change (CBC) in the highly conserved helix III of ITS2. Additionally, the species delimitation was also confirmed using five independent algorithmic approaches analyzing four different DNA barcodes. The concatenated ITS1-5.8S-ITS2 fragment is more reliable for species discrimination than the individual ITS1 or ITS2 barcodes. Of the species delimitation methods evaluated, ASAP (Assemble Species by Automatic Partitioning) and GMYC (Generalized Mixed Yule Coalescent) performed best in distinguishing Chloroidium species across multiple barcode regions in our analysis. The fatty acid profile of strain VKM Al-418 was analyzed at 9 °C, 22 °C, and 27 °C and exhibited high plasticity in response to temperature, indicative of an adaptive strategy to its harsh environment. Using this integrative taxonomic approach, we describe Chloroidium pseudoellipsoideum sp. nov., a new species with a distinct phylogenetic positioning and promising biotechnological properties.