Comparative (meta-)genomics of marine polysaccharide-degrading bacteria with emphasis on CAZyme and PUL analyses in Bacteroidetes

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Subproject B1 focuses on comparative (meta-)genomics of polysaccharide-degrading bacteria. Marine phytoplankton blooms represent key events in global carbon cycling. They remove gigatons of atmospheric CO2 and trigger successive blooms of heterotrophic bacterioplankton clades that remineralize most of the fixed carbon. Prior to POMPU we studied such successions during four spring blooms off the North Sea island Helgoland (2009-2012). Using an integrated OMICS approach, we found that recurrently abundant bacterioplankton clades exhibited notably different substrate spectra. This was particularly evident with respect to gene frequencies and expression of carbohydrate-active enzymes (CAZymes) in Bacteroidota, indicating niche partitioning regarding algal polysaccharide degradation. In Bacteroidota the genes that encode the machinery for uptake and degradation of polysaccharides are usually organized in dedicated loci termed ‘polysaccharide utilization loci’ (PULs), and PUL analyses can potentially predict the targeted polysaccharide class.Our role in POMPU thus far was to study marine polysaccharide utilization by comparative genomic analyses of (i) isolates and (ii) of size-fractionated microbial communities. In the first project phase we analyzed (i) PUL repertoires of 53 sequenced North Sea Bacteroidota strains and (ii) a time series of 38 binned and taxonomically classified North Sea metagenomes in order to uncover the PUL repertoires of all abundant bacteria in the 0.2-3 μm (free-living) fraction, including yet uncultivated Bacteroidota. During the second project phase we shifted our focus towards (i) the ecological niches of Gammaproteobacteria during Helgoland spring blooms (ii) comparative genomics of free-living versus particle-associated Bacteroidota (3- 10 μm and >10 μm fractions) during the 2018 spring bloom, and (iii) gene expression analysis of distinct free-living bacterioplankton clades with substantially improved temporal resolution during the 2020 spring bloom using a combination of metagenomics, -transcriptomics, and - proteomics (A1, Z). The latter two approaches generated comprehensive datasets whose analyses will result in integrated ecological interpretations of two very contrasting blooms.During the POMPU extension we want to finalize our analyses of the 2018 and 2020 blooms and complement the 2020 study with data on particle-attached bacteria from the 3- 10 μm size fraction, in particular with respect to obtaining higher quality MAGs (metagenome-assembled genomes) and high temporal resolution metatranscriptome data. These data will also provide a basis for ongoing work packages in subprojects B3, B2 and A1. In addition, we will focus on detailed studies of as yet unexplored polysaccharide-degrading clades, integrate saccharide measurements (A4) with OMICS data and assist geneFISH experiments to visually assign selected PULs to free-living or particle-attached bacteria (B2).