DNA-sequence-based species delimitation for larval chironomid samples collected from Japanese farm ponds

Abstract

DNA-sequence-based species delimitation methods were applied to larval chironomid samples collected from 20 Japanese farm ponds. This is the first trial using DNA barcoding to reveal the species composition of chironomid communites in limnological waters. A total of 72 species were delimited from mitochondria DNA cytochrome c oxidase I sequences of the samples. Of these, the number of species delimited from the total sample, bottom sediment sample, and aquatic weed sample of each pond, were 0-21, 0-15 and 3-16, respectively. Greater numbers of collected larvae resulted in more delimited species. Scientific names were assigned for 31 species.

Pond chironomid communities revealed by molecular species delimitation reflect eutrophication

“…Twenty study ponds were selected for sampling chironomids from among farm ponds in the eastern part of the Banshu Plain (Takamura et al, 2017) to compare chironomid communities from different environmental conditions in the same year. Of these ponds, 17 ponds were ultimately included in this study (Figure 2), while the other three ponds were omitted due to the poor collection of chironomids.…”

“…The original sample collection was conducted from bottom sediments and aquatic weeds in May and September 2012 (Takamura et al, 2017). For benthic chironomids, a sampling researcher approached the central part of the pond on a fishing floater and took three cores of bottom sediment with an Ekman-Birge grab sampler (15 cm square in the mouth) in each pond.…”

“…The PCR products were purified using ExoSAP-IT (Affymetrix) and subjected to direct sequencing using a BigDye DNA Terminator v3.1 Cycle Sequencing Kit (Life Technologies) and an ABI 3730 Genetic Analyzer (Applied Biosystems). In the previous report (Takamura et al, 2017; May and September 2012), we collected 1,060 larval or pupal chironomids from bottom sediments and aquatic weeds, of which 786 individuals were successfully sequenced. In this report, our samples were 443 larvae from the bottom sediments in May 2012, of which 296 individuals were successfully sequenced.…”

“…Species delimited by these methods were 72 species for GMYC and 80 species for PTP in May and September for bottom sediments and aquatic weeds (Takamura et al, 2017; Tables S3 and S4, Figures 3 and 4). They were compared and lumped in the way that the resulting species number was the lowest.…”

“…However, many taxa that are not easily identified by morphological taxonomy have remained untouched in these researches. Researches on chironomids of bottom sediments and aquatic weeds in summer and fall, 2012 were made in these ponds based on the molecular species delimitation of larval chironomids, and only the species number of chironomids was compared between ponds and with other studies in Takamura et al, (2017). This report focuses on the effects of water quality, surrounding land use, and aquatic flora on the benthic chironomid communities of bottom sediments in the summer.…”