Diatoms and DNA barcoding: a pilot study on an environmental sample

Abstract

Diatoms live in all types of water bodies from fresh to marine habitats and also in soil and aero-terrestrial ecosystems. Because they are sensitive concerning pollution, acidification, and salinization, they are important bio-indicators and are often used for routine water quality assessments. Since morphological identification is time-consuming and demands specialized in-depth knowledge, diatoms are an ideal model group to establish DNA barcoding methods to provide an easy to use, standardized and fast organism identification tool. So far, little has been published on the topic DNA barcoding in diatoms (Evans et al. 2007; Kaczmarska et al. 2007). DNA barcoding in general demands a molecular locus, being variable enough to discriminate on species level for the organisms under study and a molecular reference database for comparison. The similarity or divergence of the molecular sequence of an unknown organism to a vouchered reference sequence in the database is used as quality indicator for species identification. DNA barcoding of environmental samples requires DNA extraction from an environmental pooled sample, PCR amplification of a target locus; cloning of the resulting PCR products, sequencing and the analysis. In a pilot study on a plankton water sample from Lake Tegel in Berlin, Germany, we wanted to test if identification of diatoms by classical identification method via morphology using light microscopy (LM) and by molecular means via DNA sequencing would result in comparable diversity assessments and test the potential for molecular based diatom identification. In the diatom flora of Berlin (Geissler & Kies 2003) 541 taxa are mentioned to occur in Lake Tegel (Geissler et al. 2006). We chose the 18S region to be amplified for our investigation as this marker has most often been used for phylogenetic analyses and therefore the number of comparable diatom sequences in reference databases was highest. 480 sequences were available for comparison (www.algaterra.org and GenBank: http://www.ncbi.nlm.nih.gov/). As the 480 used diatom sequences of the 18S region are from world wide occurrences, only a small number of individuals from Lake Tegel were expected to match precisely on the species level. By selecting this marker for our pilot investigation we have been aware, that it might not be the best for species discrimination but it was chosen to test the feasibility of the new methodology per se.