An evaluation of bioassessment methodologies for macroinvertebrate classification in neotropical streams

Neotropical freshwater ecosystems are highly diverse, which makes characterizing them difficult. These ecosystems are also highly threatened by rapid land use change, extractive industry increase, and inadequate wastewater treatment. Accurate and efficient monitoring methods are increasingly needed for these ecosystems. Freshwater stream assessments have often relied on benthic macroinvertebrate samples for biomonitoring efforts. The classification of aquatic macroinvertebrates has traditionally been based on morpho-taxonomic determinations. This method, however, is time-consuming and requires expertise. Newer and more efficient DNA-based classification approaches, such as metabarcoding, could be considered for Neotropical streams. We took replicate samples of 11 stream sites along an altitudinal gradient in western Ecuador to compare morpho-taxonomic and metabarcoding methods for aquatic macroinvertebrate determination. Taxonomic resolution for the morpho-taxonomic method was to genus for insects and the lowest taxonomic level possible for non-insects. Taxonomic assignments for the metabarcoding approach were performed with BLAST sequence alignments and verified using BOLD-generated phylogenetic trees. The family and genus-level taxa overlap between morpho-taxonomic and metabarcoding determinations per site was generally low. We found that many Neotropical species lack reference DNA sequences, indicating the need for a comprehensive regional reference DNA database. Further research is needed to determine whether DNA-based biomonitoring assessments are comparable to those using conventional methods and to fully comprehend the implementation obstacles for this method in the region.