Online first articles

Temporary rivers (TRs) are dynamic ecosystems that alternate between hydrological phases (i.e. flowing, disconnected pools, and dry). They are conservation refugia for aquatic species during dry seasons but are often neglected in bioassessment programs. To assess the biological quality of these ecosystems, morphology methods can be invasive, disrupting communities and diminishing their function as refugia. Environmental DNA (eDNA) metabarcoding provides a minimally invasive method, gathering community information from eDNA in water or sediment. We tested the effectiveness of eDNA methods alongside bulk DNA to characterize the macroinvertebrate communities and assess the biological quality of disconnected pools in TRs, comparing them with morphological methods. Additionally, we tested how the community patterns evolve over time using eDNA and how community composition shifts during disconnection. Biological quality was determined through macroinvertebrate indices widely used in Spain (i.e., IBMWP, family richness, and IASPT). eDNA samples were collected biweekly from three TRs in Catalonia, NE Spain. Macroinvertebrates were sampled during the three hydrological phases (connected, disconnecting, and disconnected pools). Macroinvertebrate samples were used to identify organisms using morphology and to sequence bulk DNA. eDNA and bulk DNA samples were sequenced through metabarcoding targeting the mitochondrial COI gene. Although communities determined by sediment eDNA did not detect variations in biotic indices (i.e., IBMWP and family richness), the method was useful to detect the replacement of EPT (Ephemeroptera, Plecoptera, Trichoptera) by OCH (Odonata, Coleoptera, Heteroptera). Additionally, sediment eDNA revealed significant impacts of hydrological changes on meiofauna (Ostracoda, Cladocera, Copepoda), a group often overlooked in stream assessments. These results indicate that sediment eDNA metabarcoding can serve as a valuable tool for the bioassessment of TRs, capturing the transitions between hydrological phases while preserving ecosystem integrity.