Molecular methods for characterizing communities of marine animals

We are all blind men trying to describe a monstrous elephant of ecological and evolutionary diversity” (Nanney, 2004) is one of my favorite quotes. It describes well one of the main challenges that scientists face: most eukaryotic species in marine samples are small, difficult to identify and often undescribed.  The ability to delineate and detect species using DNA and RNA markers has revolutionized the way we can characterize spatial and temporal patterns of diversity (i.e., the way communities respond to ongoing environmental changes). We no longer have to rely on indicator species or focus on conspicuous taxonomic groups. We can now study entire communities of microscopic metazoan, protists and microbes that have been largely overlooked.


            

Part of the work I have conducted with colleagues and students has been to facilitate the use of these molecular approaches for characterizing metazoan communities. Since Paul Hebert’ influential work, millions of DNA barcodes for 100s of thousands of species have been made public online (e.g., GenBank). DNA barcodes are small fragments of DNA that can be used as species identifiers (for example in the absence of well defined morphological characters). We have been interested in finding ways to make use of this incredible resource in ecological and evolutionary studies.

 

 

Projects we have worked on include the design of broad-range PCR primers to allow the simultaneous detection of hundreds of species in ecological samples. We tested and implemented molecular approaches that allow the description of feeding behaviors with unprecedented levels of taxonomic resolution. We published protocols for sample preparation and data analysis. More recently, I have been working with my colleague Ryuji Machida at Academia Sinica (Taiwan) and others on the development of a curated and pre-formatted database of mitochondrial sequences for the taxonomic assignment of sequences isolated from the environment (link to MIDORI server). 

Environmental DNA isolated from seawater samples reveals nearby animal, plant and microbial communities (photo: Sean Mattson)

The MIDORI server a user-friendly web platform that uses a curated reference dataset, MIDORI, for high throughput taxonomic classification of unknown metazoan mitochondrial encoded gene sequences. Currently three methods of taxonomic assignments: RDP Classifier, SPINGO and SINTAX, are implemented. Link to publication.

Small invertebrates that live on and between sand grains are so numerous and diverse that DNA sequencing is often needed to understand who is there and why (photo: Sean Mattson).

SELECTED PUBLICATIONS ON THIS TOPIC

Leray M, Ho S-L, Lin J, Machida R. 2018. MIDORI server: a webserver for taxonomic assignment of unknown metazoan mitochondrial-encoded sequences using a curated database. Bioinformatics bty454

Leray M, Knowlton N. 2017. Random sampling causes the low reproducibility of rare eukaryotic OTUs in Illumina COI metabarcoding. PeerJ5,e3006

Machida RJ, Leray M, Ho S-L, Knowlton N. 2017. Metazoan mitochondrial gene sequence reference datasets for taxonomic assignment of environmental samples. Scientific Data4,170027

Leray M, Knowlton N. 2016. Visualizing patterns of marine eukaryotic diversity from metabarcoding data using QIIME. In, Bourlat S.J. (eds) Methods in Molecular Biology, Marine Genomics Methods and Protocols. Springer Protocols, pp 219-235

Bourlat SJ, Haenel Q, Finnman J, Leray M. 2016. Preparation of amplicon libraries for metabarcoding of marine eukaryotes using Illumina MiSeq, The dual-PCR method. In, Bourlat S.J. (eds) Methods in Molecular Biology, Marine Genomics Methods and Protocols. Springer Protocols, pp 197-207

Leray M, Haenel Q, Bourlat SJ. 2016. Preparation of amplicon libraries for metabarcoding of marine eukaryotes using Illumina MiSeq, The adapter ligation method. In, Bourlat S.J. (eds) Methods in Molecular Biology, Marine Genomics Methods and Protocols. Springer Protocols, pp 209-218

​Leray M, Yang JY, Meyer CP, Mills SC, Agudelo N, Ranwez V, Boehm JT, Machida RJ. 2013. A new versatile primer set targeting a short fragment of the mitochondrial COI region for metabarcoding metazoan diversity, application for characterizing coral reef fish gut contents. Frontiers in Zoology 10,34

​Leray M, Agudelo N, Mills SC, Meyer CP. 2013. Effectiveness of annealing blocking primers versus restriction enzymes for characterization of generalist diets, unexpected prey revealed in the gut contents of two coral reef fish species. PLoS One8,e58076

​Leray M, Boehm JT, Mills SC, Meyer CP. 2012. Moorea BIOCODE barcode library as a tool for understanding predator-prey interactions, insights into the diet of common predatory coral reef fishes. Coral Reefs31,383-388

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