SEQUENCING THE MICROBIAL COMMUNITY AT THE WESTERN CHANNEL OBSERVATORY / SEASONAL CHANGES IN DIVERSITY AND FUNCTION

The marine ecosystem is crucial for the functioning of the Earth system. It plays an essential part in modifying the climate, they account for half of the annual primary production on the planet and about two thirds of goods and services from our planet are provided by coastal and open ocean ecosystems. These environments are dominated by microorganisms, such as bacteria and archaea, which drive all global biogeochemical cycles and have a direct influence on the atmosphere by the production of chemically-active biogases. Therefore the biogeochemistry of the oceans is the consequence of microbial activity and understanding the Earth System requires a significant research effort on marine microbes. The enormous number of microbes and their vast metabolic diversity, suggests that the genetic diversity and function is far higher than that investigated so far. The huge extent of the microbial diversity in the marine environment was first demonstrated by the study of Craig Venter (Venter et al., 2004) who used Sanger sequence analysis of a shot-gun clone library produced from of a few hundred litres of the Sargasso Sea water. This extremely expensive sequence analysis found 1.2 million new genes within 1800 species of microbes, including 150 new species of bacteria. The sheer number of genes they found in the Sargasso is astounding and further supports the notion that the microbial life in the ocean is far more abundant and diverse than expected. In order to scale this level of diversity it is relevant to compare it with the number of genes in the human genome / currently thought to comprise 20,000 - 25,000 genes. This proposal is to extent the output from the PGP-funded project on Aquatic Microbial Metagenomics in that it utilises on the recently available novel sequencing methodology of pyrosequencing, which has been developed by 454 Life Sciences, Inc. (USA). A typical pyrosequencing run generates ca. 400,000 sequences from one sample. As it is an extention of the currently funded PGP project we do not request salary costs. We request the steering committee to consider providing 100% of the sequencing costs (currently NERC rules only allow 80%). We propose to use the novel high-throughput pyrosequencing approach to analyse both the genetic diversity and function of the bacteria and archaea at the NERC-funded Western Channel Observatory (WCO). We will use this approach to assess the changes in the composition and function that are characterised by seasonal cycles. The output of the proposed pyrosequencing approach will generate some 3.2 million sequence reads. Based on an average read length of 250 bp, this will produce approximately 800 million bp of genetic information comprising approximately 200 - 400 bacterial genome equivalents. In conclusion, it is important to understand that this project is novel in two points. Firstly, and for the first time on a large scale this project will analyse both the diversity and the activity of the microbes present in the same sample, as well as assessing the influence of different seasons on both. And secondly, the molecular approach that we will employ uses for the first time the new high-throughput pyrosequencing technique to analyse the gene expression of a whole microbial community.