DNA Barcoding: a Quality Assurance Measure in Ecotoxicology

To date, ecotoxicological test species are usually identified based on morphological characters only, which may often lead to misidentifications. Since test species may differ in their response to toxic substances, accurate identification of test organisms is critical for the reliability and correct interpretation of ecotoxicological results.

A DNA-based service for accurate test species verification 

There is a growing interest in implementing DNA barcoding as a quality assurance measure in ecotoxicological testing. By sequencing and analysing reference genes from culture specimens on a regular basis, Ecotoxicology laboratories can guarantee the homogeneity of cultures and the species identification of test specimens. Therefore, two key issues, the accuracy and comparability of results, will be improved.

International standardisation organisations (e.g. ISO; OECD) are currently discussing the advantages of including the genetic characterisation of invertebrate and plant species in terrestrial ecotoxicological tests. As members of the Working Group 'Effects on soil fauna' of the ISO/TC 190/CS4 Committee, we are actively involved in a new ISO Project to standardise DNA barcoding as a quality control test in Ecotoxicology guidelines (ISO/NP 21286: Soil quality -- General guidance on the use of DNA barcoding in ecotoxicological testing).

Our verification of ecotoxicological test species service comprises two types of analyses:

  • Specimen identification, to determine the specific identity and, if possible, the genetic lineage of unknown specimens. This type of analysis is particularly useful when specimens collected in field studies require an accurate and cost-effective species-level identification.
  • Test of stock homogeneity, to verify that all the specimens in a laboratory stock belong to the same species, or even to the same genetic lineage. As a norm we DNA barcode 91 specimens per stock. This allows us to detect a contaminating species occurring at a minumum frequency of 5 % with a probability of 99 % in a cost-effective way. At the same time, you can precisely document the species or genetic strain being used in your tests. We strongly advise carrying out a test of stock homogeneity every time a new stock is acquired or used to initiate new cultures.

We specialise in DNA barcoding of terrestrial and aquatic test species from diverse taxonomic groups (click here for a complete list of species). If your target species is not in the list, please, do not hesitate to contact us and let us know about your specific testing requirements.

A curated reference database for Ecotoxicology test organisms

The efficiency of DNA barcoding depends on the availability of reliable reference sequences for comparison. We use reference DNA barcodes available in public international databases, as well as our own library of reference sequences. These sequences have been obtained from taxonomically validated specimens, so it is a solid standard for comparative analyses of samples from Ecotoxicology laboratories.

How we work

We provide sampling kits and sample collection guidelines within 2-4 working days once your order has been placed.

DNA barcoding projects involve DNA isolation, PCR amplification of a genome region (the DNA barcode) from the sample, and sequencing. Subsequently, electropherograms are analysed and the sequences obtained are compared to those in reference databases to find the matching species.

Deliverables include species identification or stock homogeneity certificates, the raw data generated (which will be delivered through our server), and a summary of the methods followed. We also provide a free archiving service for your own sequences. These sequences will be used as an additional reference database in your future orders.

The Eisenia barcoding initiative

The earthworms Eisenia fetida and E. andrei are two of the most widely used test species in Terrestrial Ecotoxicology. However, both species are difficult to identify using morphological characters alone. This may lead to frequent species misidentification, thus making comparisons of test results among ecotoxicology laboratories virtually impossible. Therefore, there is a wide interest in the Ecotoxicology community to study the species boundaries of earthworms being used by ecotoxicology laboratories. For this reason, the Biodiversität und Klima Forschungszentrum Frankfurt am Main, ECT Oekotoxikologie GmbH, and AllGenetics led an international initiative to DNA barcode Eisenia earthworms from more than 20 Ecotoxicology laboratories throughout the world. A ring test was carried out by five DNA barcoding labs in order to assess (1) the species homogeneity of Eisenia cultures, (2) the species boundaries between E. andrei and E. fetida, (3) the comparability of results among Ecotoxicology laboratories, and (4) the comparability of results among DNA barcoding laboratories.

The results of this project have been published in the following paper: Römbke et al. (2016). DNA barcoding of earthworms (Eisenia fetida/andrei complex) from 28 ecotoxicological test laboratories. Applied Soil Ecology 104:3-11.