Dear all,

We are pleased to announce that the 2nd edition of Eukaryotic Metabarcoding 
Workshop will take place from February 26th-March 2nd, 2018 in Berlin 


Dr Owen S. Wangensteen (University of Salford, UK)

Dr Vasco Elbrecht (University of Guelph, Canada)


Metabarcoding techniques are a set of novel genetic tools for qualitatively 
and quantitatively assessing biodiversity of natural communities. Their 
potential applications include (but are not limited to) accurate water 
quality, soil diversity assessment, trophic analyses of digestive contents, 
diagnosis of health status of fisheries, early detection of non-indigenous 
species, studies of global ecological patterns and biomonitoring of 
anthropogenic impacts. This workshop gives an overview of metabarcoding 
procedures with an emphasis on practical problem-solving and hands-on work 
using analysis pipelines on real datasets. After completing the workshop, 
students should be in a position to (1) understand the potential and 
capabilities of metabarcoding, (2) run complete analyses of metabarcoding 
pipelines and obtain diversity inventories and ecologically interpretable 
data from raw next-generation sequence data and (3) design their own 
metabarcoding projects, including bioinformatic data analysis and planning 
of laboratory work. All course materials (including copies of 
presentations, practical exercises, data files, and example scripts 
prepared by the instructing team) will be provided electronically to 

*Intended audience:*

This workshop is mainly aimed at researchers and technical workers with a 
background in ecology, biodiversity or community biology who want to use 
molecular tools for biodiversity research and at researchers in other areas 
of bioinformatics who want to learn ecological applications for 
biodiversity-assessment. In general, it is suitable for every researcher 
who wants to join the growing community of metabarcoders worldwide. This 
workshop will review mostly techniques and software useful for eukaryotic 
metabarcoding. Other workshops focused on procedures currently used in 
microbial metabarcoding will be available from Physalia-courses.

*Teaching format:*

The workshop is delivered over ten half-day sessions (see the detailed 
curriculum below). Each session consists of roughly a one hour lecture 
followed by two hours of practical exercises, with breaks at the 
organizer’s discretion.

*Assumed background:*

No programming or scripting experience is necessary, but some previous 
expertise using the Linux console and/or R will be most welcome. All 
examples will be run either in Linux or Mac environments, with some ssh 
connections to remote servers. For Windows users, a virtual box running 
Linux under Windows and/or the installation of an ssh client (e.g. PuTTY) 
will be needed. For MacOSX systems, installation of some additional Python 
packages might be needed for running the OBITools software suite. The 
syllabus has been planned for people which have some previous experience 
running simple commands from a terminal in Linux or Mac and using the R 
environment (preferently RStudio) for performing basic plots and 
statistical procedures. You will need to have a laptop with Python 2.7 
installed for running OBITools, but no experience with Python is necessary. 
If in doubt, take a look at the detailed session content below or send an 
email to us.


Monday 26th – Classes from 09:30 to 17:30

*Session 1*. Introduction to metabarcoding procedures. The metabarcoding 

In this session students will be introduced to the key concepts of 
metabarcoding and the different next-generation sequencing platforms 
currently available for implementing this technology. Some examples of 
results that can be obtained from metabarcoding projects are explained. We 
will outline the different steps of a typical metabarcoding pipeline and 
introduce some key concepts. We will also explain the format of the course. 
In this session, we will check that the computing infrastructure for the 
rest of the course is in place and all the needed software is installed. 
Core concepts introduced: high-throughput sequencing, multiplexing, NGS 
library, metabarcoding pipeline, metabarcoding marker, clustering 
algorithms, molecular operational taxonomic unit (MOTU), taxonomic 

*Session 2.* Molecular laboratory protocols. DNA extraction. Metabarcoding 
markers. Primer design. PCR and library preparation. Good laboratory 

In this session we will learn the basics about molecular laboratory 
procedures needed for metabarcoding. While there will be no hands-on 
laboratory practices, guidelines and best practices for all key laboratory 
steps will be discussed. We will explain sample collection techniques, 
including eDNA and bulk community samples, pretreatment and DNA extraction 
protocols. The diverse molecular markers available for different kinds of 
samples and target taxonomic groups will be discussed. The students will 
learn to design and test custom metabarcoding primers. They will know about 
sample tags, library tags, adapter sequences, PCR protocols and library 
preparation procedures. Core concepts introduced: good laboratory practice, 
proper sample collection, bulk (community DNA) and eDNA samples, DNA 
preservation, DNA extraction, PCR, clean up, metabarcoding marker, 
universality, specificity, taxonomic range, taxonomic resolution, primer 
bias, amplification errors, sequencing errors, DNA contaminations, in 
silico PCR, library generation, sequencing platforms, sample indexing, 
adapter sequences.

Tuesday 27th – Classes from 09:30 to 17:30

*Session 3&4*. The USEARCH pipeline.

In this session, we will work with the USEARCH and VSEARCH software suites, 
using a real sequence dataset as example for testing our metabarcoding 
pipeline. We will outline the steps needed to start analysing raw data from 
high-throughput sequencers. The students will learn about key 
bioinformatics workflows and they will perform quality control, sample 
demultiplexing, paired-end merging, sequence filtering, removal of chimeric 
sequences, format conversion, dereplication of unique sequences, sequence 
clustering as well as taxonomy assignment using reference databases. We 
will run most commands in an R environment using a user friendly modular 
wrapper script, with specific focus on when and why each module is 
necessary. Core concepts introduced: fastq and fasta formats, Phred quality 
score, paired-end alignment, demultiplexing, sequence filtering, chimeras, 
dereplication, unique sequences, reads, singleton sequences, abundance 
recalculation, OTU clustering, sequence repositories, identity assignment, 
BLAST, GenBank, Barcode Of Life Datasystems (BOLD).


Wednesday 28th – Classes from 09:30 to 17:30

*Session 5*. The OBITools pipeline I. Workflow, first steps and quality 
control. Clustering algorithms with variable thresholds.

In this session, we will work with the OBITools software suite, using the 
same dataset we used in USEARCH for testing some alternative metabarcoding 
pipelines from a Linux terminal environment. We will also introduce 
different algorithms for clustering sequences into MOTUs, such as CROP and 
SWARM. We will learn the differences between constant and variable identity 
threshold for delineating the MOTUS. Core concepts introduced: reference 
clustering, de novo clustering, unsupervised-learning clustering, Bayesian 
clustering, step aggregation methods, hard identity threshold, flexible 
identity threshold.

*Session 6.* The OBITools pipeline II. Taxonomic assignment using ecotag.

In this session we will continue with the OBITools pipeline. We will learn 
about phylogenetic algorithms for taxonomic assigment. The ecotag algorithm 
will be used for adding taxonomic information to the MOTUs in our example 
dataset and the results will be compared to those from other assignment 
software. The students will learn how to build local reference databases 
from the information available in public sequence repositories and how to 
add new custom sequences to these local reference databases. They will also 
learn how sequence databases interact with taxonomy databases for 
retrieving the phylogenetic information for the assignment algorithms. Core 
concepts introduced: local reference database, phylogenetic assignment, 
best match, assignment of higher taxa, ecoPCR and ecoPCR format, taxonomic 
database, taxonomic identifier (taxid).

Thursday 1st – Classes from 09:30 to 17:30

*Session 7.* Comparing the results from different pipelines. Refining the 
final datasets. Collapsing, renormalising and blank correction. 
Visualization of results.

In this session, students will learn about procedures for refining and 
curating the final datasets obtained from the previous pipelines. They will 
learn about blank correction, renormalization procedures for deleting false 
positive results, and taxonomy collapsing of related MOTUs for obtaining 
enhanced final datasets. We will compare the results from the different 
pipelines tested and we will discuss how to interpret them in order to 
obtain ecologically relevant information. Core concepts introduced: 
renormalization, taxonomy collapsing, blank correction.

*Session 8.* Presenting the final results. α- and ß- diversity patterns.

In this session we will continue with the presentation of final results. 
Students will learn how to plot taxonomic summaries from their datasets, 
including krona plots, a graphic representation showing relative abundances 
of reads at different taxonomic levels. Resampling and rarefaction 
procedures for assessing biodiversity patterns will be introduced. 
Qualitative and quantitative indices for assessing dissimilarity between 
samples will be explained. We will introduce the UniFrac dissimilarity 
distance between samples, an index taking in account not only abundances of 
the different MOTUs but also their taxonomic affinities. Core concepts 
introduced: taxonomic summary, krona plots, α-diversity, ß-diversity, 
rarefaction, MOTU richness, UniFrac distances, ordination techniques, 
multidimensional scaling (MDS).

Friday 2nd – Classes from 09:30 to 17:30

*Session 9.* Experimental design. Customization.

In this session we will learn how to design a successful metabarcoding 
project and how to customize it in function of the specific needs. We will 
discuss the best strategies for obtaining good results by optimizing time, 
money and computing resources. The idea is to make this session as 
interactive and useful as possible. We will present some current and future 
projects in the format of an open discussion and we will try to propose the 
best solutions for every potential problem in a collaborative way. The rest 
of the session will be dedicated to introduce current research and possible 
future developments of metabarcoding / metagenomics techniques and to 
provide a list of useful resources for further learning, continuous 
training and future research opportunities. Core concepts discussed: 
optimal multiplexing, ecological replication, technical replication, 
sequencing depth, price per sample.

*Session 10.* Hands-on project brainstorming

In small groups the participants will have the opportunity to plan and 
develop metabarcoding projects based on their research questions and 
taxonomic groups of interest. Project proposals will be presented and 
discussed in the form of 5-minute presentations, and they will be evaluated 
and improved by interacting with workshop participants. We will finish the 
workshop with an interactive open questions session. Core Concepts: 
Experimental planning, developing successful research projects and 
proposals, concept evaluation and improvement by peer review, using 
metabarcoding as a tool to answer exciting research questions.
For more information please visit our website:

MORPHMET may be accessed via its webpage at
You received this message because you are subscribed to the Google Groups 
"MORPHMET" group.
To unsubscribe from this group and stop receiving emails from it, send an email 

Reply via email to