Kobari T, Tokumo Y, Sato I, Kume G, Hirai J.

Sci Rep. 2021 Dec 1;11(1):23265 doi:10.1038/s41598-021-02083-8

Metabarcoding analysis of trophic sources and linkages in the plankton community of the Kuroshio and neighboring waters.

eDNA analysis of “the diets of 22 higher taxonomic groups, 13 copepod families and 35 species, that dominate in the Kuroshio food web”. Larvacean DNA was more frequent in predator gut contents than in water samples. “The 18S rRNA V9 region was amplified using eukaryotic universal primers 1389F and 1510R.”

Stoeck T, Bass D, Nebel M, Christen R, Jones MD, Breiner HW, Richards TA

Mol Ecol. 2010 Mar;19 Suppl 1:21-31. doi:10.1111/j.1365-294X.2009.04480.x

Multiple marker parallel tag environmental DNA sequencing reveals a highly complex eukaryotic community in marine anoxic water.

The TAReuk454FWD1–TAReukREV3 pair (V4) detected more unique tags than the 1391F–EukB pair (V9).

Edgar RC.

Bioinformatics. 2018 Jul 15;34(14):2371-2375. doi:10.1093/bioinformatics/bty113

Updating the 97% identity threshold for 16S ribosomal RNA OTUs.

Following a benchmark on V4 regions of the 16S rRNA, proposes to raise the threshold to 99% or to just use sequence variants (also called ZOTUS, Zero-radius OTUs. Identity computed as the number of columns containing identical letters divided by the number of columns containing at least one letter.

Amaral-Zettler LA, McCliment EA, Ducklow HW, Huse SM.

PLoS One. 2009 Jul 27;4(7):e6372. doi:10.1371/journal.pone.0006372

A method for studying protistan diversity using massively parallel sequencing of V9 hypervariable regions of small-subunit ribosomal RNA genes.

Primers contain degenerate sequences and are either universal or eukaryote-based. Samples from Mount Hope Bay, Massachusetts and Palmer Station, Antarctica.

>18S_1380F eukaryotic 10.1371/journal.pone.0006372
CCCTGCCHTTTGTACACAC
>18S_1389F universal 10.1371/journal.pone.0006372
TTGTACACACCGCCC
>18S_1510R eukaryotic 10.1371/journal.pone.0006372
CCTTCYGCAGGTTCACCTAC

Tom O. Delmont, Morgan Gaia, Damien D. Hinsinger, Paul Fremont, Chiara Vanni, Antonio Fernandez Guerra, A. Murat Eren, Artem Kourlaiev, Leo d’Agata, Quentin Clayssen, Emilie Villar, Karine Labadie, Corinne Cruaud, Julie Poulain, Corinne Da Silva, Marc Wessner, Benjamin Noel, Jean-Marc Aury, Tara Oceans Coordinators, Colomban de Vargas, Chris Bowler, Eric Karsenti, Eric Pelletier, Patrick Wincker, Olivier Jaillon

bioRxiv 2020.10.15.341214; doi: https://doi.org/10.1101/2020.10.15.341214

Functional repertoire convergence of distantly related eukaryotic plankton lineages revealed by genome-resolved metagenomics

Assembled more than 700 eukaryotic metagenomes. 37 of them are Appendicularian. A phylogeny was made using DNA-dependent RNA polymerases.

Ann Bucklin, Katja T. C. A. Peijnenburg, Ksenia N. Kosobokova, Todd D. O’Brien, Leocadio Blanco-Bercial, Astrid Cornils, Tone Falkenhaug, Russell R. Hopcroft, Aino Hosia, Silke Laakmann, Chaolun Li, Luis Martell, Jennifer M. Questel, Deborah Wall-Palmer, Minxiao Wang, Peter H. Wiebe & Agata Weydmann-Zwolicka

Mar Biol 168, 78 (2021). doi:10.1007/s00227-021-03887-y

Toward a global reference database of COI barcodes for marine zooplankton.

MetaZooGene Barcode Atlas and Database (MZGdb). Database built on top of BOLD and GenBank. It includes species from which COI data is not yet available.

Mahsa Mousavi‐Derazmahalleh, Audrey Stott, Rose Lines, Georgia Peverley, Georgia Nester, Tiffany Simpson, Michal Zawierta, Marco De La Pierre, Michael Bunce, Claus T. Christophersen

Molecular Ecology Resources. 2021

eDNAFlow, an automated, reproducible and scalable workflow for analysis of environmental DNA (eDNA) sequences exploiting Nextflow and Singularity

DSL1

Miya M, Sato Y, Fukunaga T, Sado T, Poulsen JY, Sato K, Minamoto T, Yamamoto S, Yamanaka H, Araki H, Kondoh M, Iwasaki W.

MiFish, a set of universal PCR primers for metabarcoding environmental DNA from fishes: detection of more than 230 subtropical marine species.

R Soc Open Sci. 2015 Jul 22;2(7):150088. doi:10.1098/rsos.150088

“Out of the 180 marine fish species contained in the four tanks with reference sequences in a custom database, we detected 168 species (93.3%) distributed across 59 families and 123 genera.”

“considering the unconventional base pairing in the T/G bond, the designed primers use G rather than A when the template is variably C or T, and T rather than C when the template is A or G;”

“2 l lots of seawater from the 10 l samples were vacuum-filtered onto 47 mm diameter glass-fibre filters [and then] stored in −20°C before eDNA extraction.” “Two litres of Milli-Q water was used as the negative control.” ”Lysis using proteinase K [at] 56°C [...] for 30 min.” “Six random hexamers (N) are used to enhance cluster separation on the flowcells during [basecall]”

“35 cycles of a 12 μl reaction volume containing 6.0 μl 2× KAPA HiFi HotStart ReadyMix (including DNA polymerase, reaction buffer, dNTPs and MgCl2 (at a final concentration of 2.5 mM)) (KAPA Biosystems, Wilmington, MA, USA), 0.7 μl of each primer (5 μM), 2.6 μl sterile distilled H2O and 2.0 μl template. [...] The thermal cycle profile after an initial 3 min denaturation at 95°C was as follows: denaturation at 98°C for 20 s; annealing at 65°C for 15 s; and extension at 72°C for 15 s with the final extension at the same temperature for 5 min.” “The first PCR product was diluted 10 times using Milli-Q water and used as a template for the second PCR.”

“The pre-processed reads from the above custom pipeline were dereplicated using a ‘derep_fulllength’ command in UCLUST”

“Optimal experimental conditions for the first PCR with these primers were achieved through trial and error, and we found that choice of a PCR kit (KAPA HiFi HotStart ReadyMix) and associated high-annealing temperatures (65–67°C) in the first PCR are the two most important factors contributing to successful amplifications showing distinct single PCR bands on the agarose gel.”

“MiFish-U/E primers also amplified eDNA from a [...] spotted dolphin”

“The occasional detection of [non-tank species] in the negative controls strongly suggests cross contamination in the laboratory, which seems unavoidable in eDNA studies using PCR amplifications.”

Vorobev A, Dupouy M, Carradec Q, Delmont TO, Annamalé A, Wincker P, Pelletier E.

Genome Res. 2020 Apr;30(4):647-659. doi:10.1101/gr.253070.119

Transcriptome reconstruction and functional analysis of eukaryotic marine plankton communities via high-throughput metagenomics and metatranscriptomics.

Metagenomics-based transcriptomes (MGTs): metatranscriptomic-based unigenes of the MATOU-v1 catalog detected by metagenomic reads mapping in at least three different Tara Oceans samples and displaying no more than 90% of their total genomic occurrence signal in a single sample, grouped based on the covariation of their genomic abundance across the samples, with a minimum of 500 unigenes per group. Oikopleuridae detected (Figure 1).

Djurhuus A, Closek CJ, Kelly RP, Pitz KJ, Michisaki RP, Starks HA, Walz KR, Andruszkiewicz EA, Olesin E, Hubbard K, Montes E, Otis D, Muller-Karger FE, Chavez FP, Boehm AB, Breitbart M.

Nat Commun. 2020 Jan 14;11(1):254. doi: 10.1038/s41467-019-14105-1.

Environmental DNA reveals seasonal shifts and potential interactions in a marine community.

Found Oikopleura 18S sequences in MiSeq run SRR8473276.

PLoS Genet. 2019 Feb 8;15(2):e1007943. doi:10.1371/journal.pgen.1007943

Berry TE, Saunders BJ, Coghlan ML, Stat M, Jarman S, Richardson AJ, Davies CH, Berry O, Harvey ES, Bunce M.

Marine environmental DNA biomonitoring reveals seasonal patterns in biodiversity and identifies ecosystem responses to anomalous climatic events.

Oikopleura detected in the 18S data.