|Complete Diversity and Relative Abundance of Active vs. Holocene Black Sea Microbiota as Revealed by 454 Pyrosequencing of rRNA Genes and Reverse Transcribed rRNA. Funded by the Mellow Foundation (WHOI Independent Studies) and ICoMM.
Microfossils or fossil organic compounds (lipid biomarkers) provide important clues about past environmental conditions and climate-change but most planktonic species are soft-bodied and do not fossilize. Lipid biomarkers are generally not very specific. However, during the last few years we and others have shown that several Holocene planktonic species, including those that do not leave fossils, can be identified based on their preserved DNA. Due to early diagenesis, fossil DNA of ancient plankton members generally comprises less than 0.1 % of the total DNA pool and a few bacterial species make up the bulk of the sedimentary DNA. At WHOI and at the Royal Netherlands Institute for Sea Research, we developed molecular biological strategies to screen sediment records for preserved planktonic DNA but it was thus far impossible to generate a complete picture of the ancient species-composition. However, recently it was shown that the pyrosequencing-based 454 platform (454 Life Sciences, Branford, CT), which can sequence millions of base pairs in a matter of hours, enabled the identification of thousands of low-abundance populations within deep-sea water samples among a small number of most abundant populations without the need for cloning. This technique most likely also enables an unprecedented fast and detailed screening of the near to complete diversity and richness of past planktonic community members based on traces of their preserved rRNA genes among a small number of dominant, possibly living microbiota.
In order to test this new technique, we selected particulate organic matter samples from the chlorophyll/plankton maximum, as well as the suboxic and anaerobic part of the photic zone for subsequent “total community 454 pyrosequencing”. This part of the Black Sea water column harbors the microbiota that, upon death or grazing, sediment out and become buried within the sediment record. Four samples will be selected from a 7,500-year-old sediment record of the Black Sea including the fluffy surface layer with recently deposited organic matter. From the same 8 samples, reverse transcribed RNA will also be subjected to 454 sequencing to study the complete active/living community structure and their relative abundance. The latter will reveal important clues about the true age of sedimentary biomarkers (both lipids and DNA) and whether they are reliable proxies for past microbial communities.