Molecular insights into hidden diversity of the epilithic green algae in Chishui River, China

Epilithic green algae play crucial roles in aquatic ecosystems. While traditional morphological approaches have provided valuable insights into these algae, their limitations in resolving cryptic species and fully capturing the extent of diversity have impeded a comprehensive understanding. The diversity and phylogenetic relationships of these algae largely remain unexplored. In the present study, single molecular real-time (SMRT) sequencing of the full-length 18S rDNA was employed to investigate the molecular diversity and phylogenetic relationships of epilithic green algae in biodiversity conservation hotspots. A total of 79 709 algal reads were generated, yielding 169 amplicon sequence variants (ASVs) from 21 epilithic riverine green algal assemblages classified in the Ulvophyceae, Chlorophyceae and Trebouxiophyceae. The 10 families in which the greatest number of unique ASVs (diversity) were observed were the Ulvellaceae, Protosiphonaceae, Chlorococcaceae, Watanabeaceae, Chlorellaceae, Neochloridaceae, Chaetophoraceae, Hazeniaceae, Tupiellaceae and Scenedesmaceae. Unique clades formed by a substantial number of sequences of green algae were detected within the Ulvales/Ulotrichales (Ulvophyceae) and the Chlorococcaceae/Protosiphonaceae (Chlorophyceae). This study has unveiled hidden diversity within epilithic green algae, expanding our knowledge of their evolutionary relationships and providing fundamental data for taxonomic investigations within this group, as well as contributing to the assessment of algal diversity in biodiversity conservation hotspots. This study suggests that these epilithic green algal assemblages may be significantly more diverse than previously recognized, but we acknowledge the limited ecological and temporal scope of our sampling effort and the resulting effects on which taxa are identified, warranting further research. Molecular analysis shows high hidden diversity of epilithic green algaeThe families Ulvellaceae and Protosiphonaceae had the greatest number of unique ASVsMany unique clades within the Ulvales/Ulotrichales and Chlorococcaceae/Protosiphonaceae