Metagenome of the Mediterranean deep chlorophyll maximum studied by direct and fosmid library 454 pyrosequencing

The deep chlorophyll maximum (DCM) is a zone of maximal photosynthetic activity, generally located toward the base of the photic zone in lakes and oceans. In the tropical waters, this is a permanent feature, but in the Mediterranean and other temperate waters, the DCM is a seasonal phenomenon. The metagenome from a single sample of a mature Mediterranean DCM community has been 454 pyrosequenced both directly and after cloning in fosmids. This study is the first to be carried out at this sequencing depth (ca. 600 Mb combining direct and fosmid sequencing) at any DCM. Our results indicate a microbial community massively dominated by the high-light-adapted Prochlorococcus marinus subsp. pastoris, Synechococcus sp., and the heterotroph Candidatus Pelagibacter. The sequences retrieved were remarkably similar to the existing genome of P. marinus subsp. pastoris with a nucleotide identity over 98%. Besides, we found a large number of cyanophages that could prey on this microbe, although sequence conservation was much lower. The high abundance of phage sequences in the cellular size fraction indicated a remarkably high proportion of cells suffering phage lytic attack. In addition, several fosmids clearly belonging to Group II Euryarchaeota were retrieved and recruited many fragments from the total direct DNA sequencing suggesting that this group might be quite abundant in this habitat. The comparison between the direct and fosmids sequencing revealed a bias in the fosmid libraries against low-GC DNA and specifically against the two most dominant members of the community, Candidatus Pelagibacter and P. marinus subsp. pastoris, thus unexpectedly providing a feasible method to obtain large genomic fragments from other less prevalent members of this community.