High-Throughput Sequencing for Life-History Sorting and for Bridging Reference Sequences in Marine Gerromorpha (Insecta: Heteroptera)

Accurate identification and association of larval specimens with adults is a major challenge in insect taxonomy. Fortunately, it is now possible for nonexperts to sort collections of bulk samples with DNA barcodes rapidly and cost-effectively. We demonstrate this process using nanopore barcoding of 757 marine insects (Insecta: Gerromorpha), of which 81% were nymphs and many samples did not have co-occurring adult males for specific identification. We successfully associated 738 specimens (97%) to nine gerromorphan species, which would have been impossible to identify using morphological characters alone. This improved ability to incorporate information from all life-history stages has led to greater precision of species distributional ranges-knowledge that will be crucial for a more complete understanding of marine insects. We also highlighted two distinct, nonoverlapping Gerromorpha COI sequence databases on GenBank-a consequence of using two different primer sets to amplify different regions of COI. This issue inevitably hinders species identification with DNA-based methods, particularly for poorly represented groups such as marine insects. We bridged these databases by analyzing full-length COI sequences. We believe this will inspire future studies to incorporate DNA-based methods for more adult-larval association studies and for enhancing existing genetic resources, especially in understudied groups.