Metabarcoding by capture using a single COI probe (MCSP) to identify and quantify fish species in ichthyoplankton swarms

Abstract

The ability to determine the composition and relative frequencies of fish species in large ichthyoplankton swarms could have extremely important ecological applications However, this task is currently hampered by methodological limitations. We proposed a new method for Amazonian species based on hybridization capture of the COI gene DNA from a distant species (Danio rerio), absent from our study area (the Amazon basin). The COI sequence of this species is approximately equidistant from all COI of Amazonian species available. By using this sequence as probe we successfully facilitated the simultaneous identification of fish larvae belonging to the order Siluriformes and to the Characiformes represented in our ichthyoplankton samples. Species relative frequencies, estimated by the number of reads, showed almost perfect correlations with true frequencies estimated by a Sanger approach, allowing the development of a quantitative approach. We also proposed a further improvement to a previous protocol, which enables lowering the sequencing effort by 40 times. This new Metabarcoding by Capture using a Single Probe (MCSP) methodology could have important implications for ecology, fisheries management and conservation in fish biodiversity hotspots worldwide. Our approach could easily be extended to other plant and animal taxa.

DNA metabarcoding of Neotropical ichthyoplankton: Enabling high accuracy with lower cost

“…This observation reinforces the idea that more sequences of fish species must be deposited in databases, especially in river basins with rich fish fauna as the Upper Paraná River. Another positive aspect is that, compared with DNA barcoding (Arroyave and Stiassny 2014, Becker et al 2015, Frantine-Silva et al 2015, our method can trigger significant advances in the quality of ecological studies in fish, as observed in some recent studies with metabarcoding (Guillerault et al 2017, Maggia et al 2017, Mariac et al 2018, Barbato et al 2019 and, mainly, to ensure significant reduction in labor time and cost per specimen reaching 80% of reduction (Shokralla et al 2015). Despite some differences in obtaining DNA samples, both environmental DNA (eDNA) and DNA metabarcoding from bulk samples are subject to false negatives and false positives (Ficetola Gentile et al 2015Gentile et al , 2016Divoll et al 2018).…”

“…Quali-and quantitative studies involving metabarcoding and eDNA have increased significantly in recent years (Hänfling et al 2016;Valentini et al 2016;Evans and Lamberti 2017;Yamamoto et al 2017;Nakagawa et al 2018;Mariac et al 2018). However, the reliable quantification of organisms remains a problem (Evans et al 2016;Kelly et al 2017;Hansen et al 2018;Mariac et al 2018). eDNA studies have shown a positive correlation between species-specific DNA concentrations in biomass and abundance in eDNA (Takahara et al 2012;Maruyama et al 2014;Doi et al 2015;Evans et al 2016).…”

“…The use of DNA barcodes in ichthyoplankton improved the quality of identifications, both for eggs and larvae, and emerged as an effective alternative for traditional identification methods (Becker et al 2015, Frantine-Silva et al 2015. Nevertheless, despite its high accuracy, when applied in extensive studies with thousands of eggs and larvae, DNA barcoding can be time-consum-ing and costly, because each organism has to be individually processed and sequenced (Evans et al 2016, Elbrecht and Leese 2017, Mariac et al 2018.…”

“…To solve this problem, DNA metabarcoding, based on next generation sequencing (NGS) could offer an alternative approach, allowing a reduction of labor time as well as costs. Although Neotropical ichthyofauna metabarcoding studies are scarce (Maggia et al 2017, Mariac et al 2018, Sales et al 2019, this technique has been utilized in several ecological studies on ichthyoplankton and also to identify gut contents (Cruaud et al 2017, Guillerault et al 2017, Maggia et al 2017, Divoll et al 2018, Barbato et al 2019).…”

Population‐level inferences from environmental DNA—Current status and future perspectives

“…Relatively small amounts of starting DNA material are required for this approach (Gnirke et al, ; Hodges et al, ), which has made it advantageous in studies where endogenous DNA content is low and fragment size is small, such as in ancient samples (Enk et al, ). Recently, target capture of mtDNA has been applied to both ancient eDNA from sediment (Slon et al, ) and modern eDNA from water samples (Mariac et al, ; Wilcox et al, ) for species detection. Although a reference genome is required to design capture probes for population genomic analyses, this genome may be sourced from a related species (even a distant one), since a perfect match between the probes and the target DNA is not a strict necessity (Enk et al, ).…”