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Net growth of microbial populations, i.e., changes in abundances over time, can be studied using 16S rRNA fluorescence in situ hybridization (FISH). However, this approach does not differentiate between mortality and cell division rates. We used FISH-based image cytometry in combination with dilution culture experiments to study net growth, cell division, and mortality rates of four bacterial taxa over two distinct phytoplankton blooms: the oligotrophs SAR11 and SAR86, the copiotrophic phylum Bacteroidetes, and its genus Aurantivirga. Cell volumes, ribosome content, and frequency of dividing cells (FDC) co-varied over time. Among the three, FDC was the most suitable predictor to calculate the cell division rates for the selected taxa. The FDC-derived cell division rates for SAR86 of up to 0.8 d-1 and Aurantivirga of up to 1.9 d-1 differed, as expected for oligotrophs and copiotrophs. Surprisingly, SAR11 also reached high cell division rates of up to 1.9 d-1, even before the onset of phytoplankton blooms. For all four taxonomic groups, the abundance-derived net growth (-0.6 to 0.5 d-1) was about an order of magnitude lower than the cell division rates. Consequently, mortality rates were comparably high to cell division rates, indicating that about 90% of bacterial production is recycled without apparent time lag within one day. Our study shows that determining taxon-specific cell division rates complements omics-based tools and provides unprecedented clues on individual bacterial growth strategies including bottom-up and top-down controls.

Additional file 2: Compilation of supplementary figures. Figure S1. a) Diversities of macroalgae, seawater and sediment samples as assessed by Shannon and Simpson indices as well as Good’s coverage of 16S rRNA ASVs. Statistical significance was assessed using a pairwise Wilcoxon test with Holm p-value adjustment for multiple comparisons (*, p < 0.05; **, p < 0.01; ***, p < 0.001). b) Rarefaction curves of the top 200 ASVs for all six samples and all four seasons. Figure S2. The most abundant taxa as assessed by 16S rRNA gene amplicon data. Figure S3. Phycosphere composition as assessed by 16S rRNA gene amplicon data as a function of host species and season. Figure S4. Phylogenies and abundances of the 86 most abundant families as assessed by 16S rRNA gene amplicon sequencing. Figure S5. 16S rRNA phylogenetic tree reconstruction for 202 genera that were represented by at least three cultured strains. Figure S6. Compositional differences of strains depending on sample source and season. Figure S7. Numbers of colony forming units (CFUs) per gram of sample depending on habitat and season. Figure S8. Workflow for translating GTDB taxonomic classifications to SILVA taxonomic classifications. Figure S9. Proportions of genes within 965 metagenome-assembled genomes (MAGs) and 1,618 draft genomes (DGs) with EggNOG, COG (2020), Pfam, UniProtKB, and KEGG annotations, as well as the percentage of genes lacking any functional annotation. Figure S10. CAZymes in metagenome-assembled genomes (MAGs) and draft genomes (DGs) of different phyla. Figure S11. CAZymes versus sulfatase gene frequencies in prominent phyla and families as assessed in 1,294 metagenome-assembled genomes (MAGs) and 963 draft genomes (DGs) from all six sample sources. Figure S12. Categories of loci used to find putative PULs in this study. Figure S13. Histograms of the lengths of the four loci described in Fig. S12 in metagenome-assembled genomes (MAGs) and draft genomes (DGs). Figure S14. Tree of all 159 clusters derived from 3,769 PUL-associated SusC-like protein sequences from Bacteroidota metagenome-assembled genomes (MAGs) and draft genomes (DGs). Figure S15. Basic quality metrics of the 1,619 metagenome-assembled genomes (MAGs) and 965 draft genomes (DGs). Box-plots (A-E) show the minimum value, first quartile, median, third quartile and maximum value. Figure S16. Biosynthetic gene cluster (BGC) sizes in genomes from distinct phyla. Figure S17. Clustering of biosynthetic gene clusters (BGCs) according to sample type and phylogeny. Figure S18. Sizes of PULs and PUL-like loci in genomes from distinct Bacteroidota families (categories: hybrid susCD, single susCD, tandem repeat susCD, and tandem repeat plus hybrid susCD PULs).

Additional file 1. Compilation of supplementary results, supplementary methods and of software tools used in this study.

Additional file 1: Figure S1. Average coverage and sequence diversity for SR and LR metagenomic samples. a. The estimated abundance-weighted average community coverage as determined in Nonpareil for each SR and LR metagenomic sample. For LR metagenomes, we first selected ~250 bp fragments from each LR and then used them to generate a model. Coverage was predicted from generated models and the original sequencing effort b. Sequence diversity (total diversity; Nd) as defined in Nonpareil. Figure S2. Read overlapping between short- and long-read metagenomic samples. The bars show the mapping of SR on LR for each time point. The dark shades indicate the mapped fractions, and the light shades show unmapped SR and LR. Figure S3. Assembly statistics for contigs generated using short- and long-read metagenomic samples. a. N statistics for contigs generated using SR and LR metagenomic samples. b. Distribution of sequencing depth (x-axis) for contigs generated using SR and LR metagenomic samples. Figure S4. Distribution of predicted protein lengths using different gene prediction tools vs. best hit match in UniProt TrEMBL. a Only predicted proteins >= 100 amino acids were used for all comparisons. The boxplots show the quotients between the length of predicted proteins using FragGeneScan (FGS), MetaGenemark, and Prodigal and the best match in UniProt TrEMBL for the unassembled long-reads of the 2020-03-10 sample. b. Distribution of predicted protein lengths from contigs vs. best match in UniProt TrEMBL for the 2020.03.10 LR sample. Figure S5. Statistics for pairs of MAGs recovered from SR and LR metagenomes. a. Difference between the number of predicted genes in MAG pairs recovered in SR and LR metagenomic samples. The boxplot in the lower right corner summarizes the comparison of predicted genes for MAG pairs. b. GC content for pairs of SR and LR MAGs colored according to their class taxonomic affiliation. The right side of the plot shows histograms for the distribution of GC content values of MAGs belonging to Bacteroidia and Gammaproteobacteria class levels. The thick black line depicts the median value of the distribution. c,d. Relative abundance for all MAGs recovered (c) and pairs of 99% ANI MAGs (d). Colors represent the taxonomic affiliation of MAGs according to GTDB-tk. Figure S6. Relationship between relative abundances of MAG pairs in SR and LR metagenomes for each time point. Figure S7. Comparison of the breadth of the coverage vs. sequencing depth for cross-mapping of reads. The figures summarize the mapping of (a) short-reads on long-read derived MAGs and (b) long-reads on short-read MAGs. Uniquely detected species are colored according to the inferred taxonomy. The rest of the points represent MAG species detected in both technologies. The dotted purple line represents the expected breadth of coverage for a given level of sequencing depth, according to Lander and Waterman (1998). The letters indicate the novelty taxonomic level for each of the MAGs (p=phylum, c=class, o=order, f=family, g=genus, and the species level is omitted for clarity).

Bacterial diversity and function across time and space in the Arctic Ocean, including the Polar Night, remain virtually unknown. In the FRAM Observatory, we study microbial composition and genetic potential in ice-covered and ice-free regions of the Fram Strait, the major gateway between the Arctic and Atlantic Oceans. A continuous amplicon time-series, derived from moored autonomous samplers, revealed marked taxonomic and functional seasonality among bacterial communities in the ice-free West Spitsbergen Current, with distinct succession of taxonomic modules. PacBio long-read metagenomes showed peaks of proteorhodopsin- and DMSP-utilizing genes in late summer, whereas winter mixing of the water column covaried with ammonia- and nitrite-metabolizing bacterial genes. In the ice-covered East Greenland Current, taxonomic and functional diversity varied less with seasons, with prominent influence of ice cover and polar water masses. For instance, high-ice conditions coincided with higher number of peptidoglycan-utilizing genes. Continuous observations were contextualized with five-year amplicon data from summertime samples collected across Fram Strait, integrating seasonal and interannual patterns of bacterial community dynamics. This fundamental baseline information helps understanding ecological and biogeochemical processes in a marine region severely affected by climate change.

The functional diversity of microbes along the seasonal extremes in the Arctic Ocean including the Polar Night are virtually unknown. Here, using PacBio long-read metagenomes derived from automated samplers over an annual cycle, we elucidate functional microbial seasonality in the Fram Strait in the context of a high-resolution amplicon time-series. In the ice-free West Spitsbergen Current, the transition from the phototrophy-dominated spring and summer ecosystem states to the dark winter was evident in bacterial genomes. Proteorhodopsin- and DMSP-utilizing genes peaked in late summer, marking a transition phase. Winter mixing of the water column covaried with microbial taxa encoding ammonia- and urea-metabolizing genes, with probable implications for nitrogen recycling and the following phytoplankton bloom. In the ice-covered East Greenland Current, functional diversity varied with the extent of ice cover and polar water masses. During intermittent low-ice conditions in winter, the metagenomic repertoire resembled that during summer, indicating rapidly (i.e. within weeks) shifting ecosystem states with ice cover. Overall, we provide a baseline to understand ecological and biogeochemical processes in a region severely affected by climate change, with implications for the present and future Arctic Ocean.

Additional file 2. Supplementary materials

Additional file 1. Supplementary figures

Anaerobic microbial hydrocarbon degradation is a major biogeochemical process at marine seeps. Here we studied the response of the microbial community to petroleum seepage simulated for 190 days in a sediment core from the Caspian Sea using a sediment-oil-flow-through (SOFT) system. Untreated (without simulated petroleum seepage) and SOFT sediment microbial communities shared 43% bacterial genus-level 16S rRNA-based operational taxonomic units (OTU0.945) but shared only 23% archaeal OTU0.945. The community differed significantly between sediment layers. The detection of fourfold higher deltaproteobacterial cell numbers in SOFT than in untreated sediment at depths characterized by highest sulfate reduction rates and strongest decrease of gaseous and mid-chain alkane concentrations indicated a specific response of hydrocarbon-degrading Deltaproteobacteria. Based on an increase in specific CARD-FISH cell numbers, we suggest the following groups of sulfate-reducing bacteria to be likely responsible for the observed decrease in aliphatic and aromatic hydrocarbon concentration in SOFT sediments: clade SCA1 for propane and butane degradation, clade LCA2 for mid- to long-chain alkane degradation, clade Cyhx for cycloalkanes, pentane and hexane degradation, and relatives of Desulfobacula for toluene degradation. Highest numbers of archaea of the genus Methanosarcina were found in the methanogenic zone of the SOFT core where we detected preferential degradation of long-chain hydrocarbons. Sequencing of masD, a marker gene for alkane degradation encoding (1-methylalkyl)succinate synthase, revealed a low diversity in SOFT sediment with two abundant species-level MasD OTU0.96.

Gammaproteobacterial Reinekea spp. were detected during North Sea spring algae blooms in the years 2009-2012, with relative abundances of up to 16% in the bacterioplankton. Here, we explore the ecophysiology of 'R. forsetii' strain Hel1_31_D35 that was isolated during the 2010 spring bloom using (i) its manually annotated, high-quality closed genome, (ii) re-analysis of in situ data from the 2009-2012 blooms and (iii) physiological tests. High resolution analysis of 16S rRNA gene sequences suggested that 'R. forsetii' dominated Reinekea populations during these blooms. This was corroborated by retrieval of almost complete Hel1_31_D35 genomes from 2009 and 2010 bacterioplankton metagenomes. Strain Hel1_31_D35 can use numerous low-molecular weight substrates including diverse sugar monomers, and few but relevant algal polysaccharides such as mannan, α-glucans, and likely bacterial peptidoglycan. It oxidizes thiosulfate to sulfate, and ferments under anoxic conditions. The strain can attach to algae and thrives at low phosphate concentrations as they occur during blooms. Its genome encodes RTX toxin and secretion proteins, and in cultivation experiments Hel1_31_D35 crude cell extracts inhibited growth of a North Sea Polaribacter strain. Our data suggest that the combination of these traits make strain Hel1_31_D35 a versatile opportunist that is particularly competitive during spring phytoplankton blooms.

Póster presentado en la Annual Conference 2013 of the Association for General and Applied Microbiology (VAAM) in collaboration with the Royal Netherlands Socity for Microbiology (KNVM), celebrada del 10 al 13 de marzo de 2013 en Bremen (Alemania), High quality 16S ribosomal RNA (rRNA) gene sequences from the type strains of all species with validly published names as defined by the International Code of Nomenclature of Bacteria is a prerequisite for their accurate affiliations within the global genealogical classification and for the recognition of potentially new species. During the last few years the Living Tree Project (LTP, http://www.arb-silva.de/projects/living-tree) has taken care to create a high quality, aligned 16S and 23S rRNA gene sequence database of all type strains. The manual curation of the sequence dataset and type strain information revealed that in total 552 “orphan” species (about 5.7% of the currently classified species) had to be excluded from the reference trees. Among them, 322 type strains were not represented by a SSU entry in the public sequence repositories. The remaining 230 type strains had to be discarded due to bad sequence quality. Since 2010, the LTP team coordinated a network of researchers and culture collections to improve the situation by (re)-sequencing the type strains of these “orphan” species. Here we report 351 16S rRNA gene sequences of type strains. 201 species could not be sequenced because either cultivable type strains were not available (121), or the cultures had either been lost or were never deposited in first place (66), or due to other constraints (14). The International Code of Nomenclature of Bacteria provides a number of mechanisms to deal with the problem of missing type strains and we recommend that due consideration be given to the appropriate mechanisms to help solve some of these issues

Capítulo en el libro, Krieg, Noel R., et al. (eds.). Bergey's Manual of Systematic Bacteriology. Volume 4: The Bacteroidetes, Spirochaetes, Tenericutes (Mollicutes), Acidobacteria, Fibrobacteres, Fusobacteria, Dictyoglomi, Gemmatimonadetes, Lentisphaerae, Verrucomicrobia, Chlamydiae, and Planctomycetes. 2nd ed. New York: Springer ; [Athens, Ga.]: Bergey's Manual Trust, 2010(1984)

Hypersaline environments, such as crystallizer ponds of solar salterns, show one of the highest concentration of viruses reported for aquatic systems. All the halophages characterized so far are isolates obtained by cultivation from described haloarchaeal species that have only low abundance in the environment. We employed a culture-independent metagenomic approach to analyse for the first time complete genomes in the halophage community and explored the in situ diversity by transmission electron microscopy and pulsed-field gel electrophoresis. We report the genomic sequence of a not yet isolated halophage (named as environmental halophage 1 'EHP-1′) whose DNA was obtained from crystallizer samples with a salinity of 31%. The sequenced genome has a size of 35 kb and a G + C content around 51%. The G + C content is lower than that of previously characterized halophages. However, G + C content and codon usage in EHP-1 are similar to the recently cultivated and sequenced Haloquadratum walsbyi, the major prokaryotic component in solar salterns around the world. Forty open reading frames have been predicted, including genes that putatively code for proteins involved in DNA replication (ribonucleotide reductases, thymidylate kinase) normally found in lytic viruses. © 2007 The Authors., This work was funded by the FP6-EU Network of Excellence Marine Genomics (Contract Nb 505403), the Max Planck Society, the Spanish Ministry of Science and Education Grants BOS2003-05198-C02-01 and -02 and the Generalitat Valenciana (Grant ACOMP06/097)

Gut microbiomes, such as the microbial community that colonizes the rumen, have vast catabolic potential and play a vital role in host health and nutrition. By expanding our understanding of metabolic pathways in these ecosystems, we will garner foundational information for manipulating microbiome structure and function to influence host physiology. Currently, our knowledge of metabolic pathways relies heavily on inferences derived from metagenomics or culturing bacteria in vitro. However, novel approaches targeting specific cell physiologies can illuminate the functional potential encoded within microbial (meta)genomes to provide accurate assessments of metabolic abilities. Using fluorescently labeled polysaccharides, we visualized carbohydrate metabolism performed by single bacterial cells in a complex rumen sample, enabling a rapid assessment of their metabolic phenotype. Specifically, we identified bovine-adapted strains ofBacteroides thetaiotaomicronthat metabolized yeast mannan in the rumen microbiome ex vivo and discerned the mechanistic differences between two distinct carbohydrate foraging behaviors, referred to as “medium grower” and “high grower.” Using comparative whole-genome sequencing, RNA-seq, and carbohydrate-active enzyme fingerprinting, we could elucidate the strain-level variability in carbohydrate utilization systems of the two foraging behaviors to helppredict individual strategies of nutrient acquisition. Here, we present a multi-faceted study using complimentary next-generation physiology and “omics” approaches to characterize microbial adaptation to a prebiotic in the rumen ecosystem.

Tesis doctoral.-- Doctoral Programme of Environmental and Biomedical Microbiology., The use of ribosomal markers in bacterial systematics intimately linked the Cytophaga-Flavobacteria-Bacteroides groups even though phenotypic studies classified them apart. With regard to phylogenetic evidence, they were circumscribed as the phylum Bacteroidetes but they stay principally studied as independent clades due lack of phenotypic coherence. The phenotypes foremost attributed to Bacteroidetes are the ability to decompose complex organic matter, gliding motility and pigmentation with flexirubins and carotenoids. The recent outburst of culture-independent methodologies and development of polynucleotide sequencing techniques has unveiled Bacteroidetes’ prevalence in many relevant ecosystems. For example, some saline environments seem to be selective of these bacteria out-competing other phyla. The goals of this doctoral thesis targets are (1) reviewing the taxonomy of the Bacteroidetes, (2) finding coding genes that could translate into an identifying phenotype and (3) finding what of their genes are selected in saline environments.For the taxonomic review of the phylum not only their 16S rDNA gene phylogeny has been updated, but also that of the 23S rDNA, signature nucleotides on their 16S rRNA sequence, insertion/deletion patterns in two of their proteins and a phylogeny based on the multilocus sequence analysis of 29 genes. This new taxonomy, on top of naming a new phylum, three classes, three orders, eight families and a genus (all of them with standing in bacterial nomenclature), paveda precise phylogenomic analysis of the Bacteroidetes, which on earlier studies included members ofthe new phylum Rhodothermaeota. However, it curtailed probabilities of finding salinity-relatedgenes, since most extreme halophiles belong to the new phylum.Through the compared genomics of 89 meticulously selected genomes we found that the most conserved genes in the phylum (in 90% of the genomes), not housekeeping, are those coding thetype-IX secretion system, hitherto only seen in the Bacteroidetes. This secretion system is involvedin the gliding motility of some Bacteroidetes, but it is also an excreting route for hydrolyticenzymes. Next, were genes coding the respiratory super-complex Alternative Complex III –cytochrome caa3 oxydoreductase at 83% of the genomes, all of aerobic species. The anaerobic lineage of the Bacteroidales can fulfill the aerobic respiration thanks to a cytochrome bdoxidoreductase with high oxygen affinity that helps detoxifying their surroundings. Genes of the Complex II, Succinate dehydrogenase, of the respiratory chain are single-copy through all Bacteroidetes’ genomes, and at the Bacteroidales other vestigial genes of the tricarboxylic acidcycle denote their aerobic past. Last, the genes of the respiratory Complex I are not arranged phylogenomically in the Bacteroidetes. These genes are substituted by an alternative complex I thatpumps sodium into the inter-membrane space instead of protons, the sodium pumping NADH: quinone oxydoreductase (Na+-NQR), only on marine species or gut microbiota species. Moreover, we found evidences of this complex being invented by the Bacteroidetes and its horizontal transmission within saline environments.The simultaneous expression of the type-IX secretion system and the respiratory chain of the Bacteroidetes, which is adapted to the environments they live at, is likely responsible for the adaptive radiation of the phylum. Some of these genes are horizontally transferred, but all together they predict the original bacteroidete was a heterotroph aerobic decomposer of complex organicmatter, likely living in freshwater., This research was funded by the Max Planck Society, the Spanish Ministry of Economy and Competitivity projects CGL2012-39627-C03-03, CLG2015-66686-C3-1-P and PGC2018-096956-B-C41 that also supported the work with European Regional Development Fund (FEDER) funds,the preparatory phase of the Microbial Resource Research Infrastructure (MIRRI) funded by the EU (grant number 312251), and the financial support of Deep Blue Sea Enterprise S.L.

Cotylorhiza tuberculata is an important scyphozoan jellyfish producing population blooms in the Mediterranean probably due to pelagic ecosystem's decay. Its gastric cavity can serve as a simple model of microbial–animal digestive associations, yet poorly characterized. Using state-of-the-art metagenomic population binning and catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH), we show that only four novel clonal phylotypes were consistently associated with multiple jellyfish adults. Two affiliated close to Spiroplasma and Mycoplasma genera, one to chlamydial ‘Candidatus Syngnamydia’, and one to bacteroidetal Tenacibaculum, and were at least one order of magnitude more abundant than any other bacteria detected. Metabolic modelling predicted an aerobic heterotrophic lifestyle for the chlamydia, which were found intracellularly in Onychodromopsis-like ciliates. The Spiroplasma-like organism was predicted to be an anaerobic fermenter associated to some jellyfish cells, whereas the Tenacibaculum-like as free-living aerobic heterotroph, densely colonizing the mesogleal axis inside the gastric filaments. The association between the jellyfish and its reduced microbiome was close and temporally stable, and possibly related to food digestion and protection from pathogens. Based on the genomic and microscopic data, we propose three candidate taxa: ‘Candidatus Syngnamydia medusae’, ‘Candidatus Medusoplasma mediterranei’ and ‘Candidatus Tenacibaculum medusae’. This research from RRM’s group was partially supported by the Spanish Ministry of Economy projects CGL2012-39627-C03-03 and CLG2015_66686-C3-1-P, which were also supported with European Regional Development Fund (FEDER) funds. KTK’s research was supported, in part, by the U.S. National Science Foundation (Award No. 1241046). RRM acknowledges the economic support of grant PR2015-00008 included in the program Salvador de Madariaga of the Ministry of Education, Culture and Sports in order to undertake a research stay at the MPI-MM in Bremen. TVP acknowledges the predoctoral fellowship of the Ministerio de Economía y Competitividad of the Spanish Government for the FPI fellowship (Nr BES-2013-064420) supporting his research activities.

Marine primary production accounts for half of the Earth' s carbon fixation and therefore has a significant impact on the global carbon cycle. However, although marine primary producers fix such a significant fraction of carbon, their biomass makes up only a small fraction of the total organic carbon pool. This is due to the extremely high turnover of phytoplankton and phytoplankton-derived organic matter within the oceans. This turnover is mediated by marine heterotrophic microorganisms (Bacteria and Archaea). Marine microorganisms therefore significantly affect the global carbon cycle. The objectives of this thesis were to investigate how the taxonomic and functional diversity of marine microorganisms affects the bacterially mediated carbon turnover in the Atlantic Ocean. In chapter 1 and 2 I developed methods which enable a shipboard high-throughput analysis of microbial diversity and abundance. These methods were developed to overcome the time delay between sampling and results and enable a comprehensive interpretation of the microbial community composition even in remote sampling sites. In chapter 3 I explored the biogeographical distribution patterns of the free-living (FL) and particle-associated (PA) bacterial communities across different provinces of the Atlantic Ocean. The FL and PA bacterial community compositions were more similar under copiotrophic condition and more dissimilar under oligotrophic conditions. I could associate these results to the relative age of the available particles as well as the availability of organic matter. In Chapter 4 I investigated alternative substrate uptake mechanisms in marine bacteria. Using fluorescently labelled polysaccharides (FLA-PS) and super-resolution structured illumination microscopy I could show that a significant fraction of marine bacteria use a a selfisha substrate utilisation mechanism. I combined this analysis with fluorescence in situ hybridization (FISH) to taxonomically identify the organisms as belonging to the Bacteroidetes, Planctomycetes and Gammaproteobacteria. The discovery of a widespread alternative substrate utilisation mechanism significantly affects our global estimates of carbon turnover by marine bacteria. Finally, in Chapter 5 I investigated the extracellular hydrolysis rates and bacterial community dynamics within fluorescently labelled polysaccharide incubations. These analyses lead to the identification of the dominant microorganisms associated with the hydrolysis of polysaccharides in the marine environment. The work done in this thesis has furthered our understanding of the activity and distribution patterns of bacterial polysaccharide utilisation mechanisms across the Atlantic Ocean. Additionally, the activity of the individual mechanisms could be associated with specific microbial groups, thereby linking the taxonomy and function of the dominant marine polysaccharide degrading organisms. This study will enable us to make better predictions of the impact which marine microorganisms have on global biogeochemical cycles.

Oceans cover ~70% of the Earths surface and are the second largest global carbon reservoir. Major processes in marine carbon cycling are summarized in the biological carbon pump. Within the biological carbon pump, pivotal stages are the microbial loop and aggregate related processes. Within the surface layer and on aggregates, carbon is channelled within a complex food web based on microbial processes. These processes counteract the biological carbon pump, showing the importance of the microbial loop for carbon sequestration. During the iron fertilization experiment LOHAFEX, a phytoplankton bloom of nano- and picoplankton was induced in the South Atlantic. We used catalyzed reporter deposition fluorescence in situ hybridization (CARD FISH) and 454 tag pyrosequencing to investigate the bacterial and archaeal community response to this bloom. The bacterial and archaeal community was stable over the course of the experiment and only members of the SAR11 and SAR86 clades showed elevated cell numbers. This led to the hypothesis of a top-down control exerted by a community of nano- and picoplankton grazers. Consequently, we used the same techniques to investigate the nano- and picoplankton community during the LOHAFEX experiment. We discovered a stable community with high but constant abundance of Phaeocystis, the major bloom forming organism, and a short peak of Micromonas and Pelagophyceae after the second iron fertilization. This again led to the hypothesis of a strong top-down control and a tight coupling of the microbial loop. We investigated the bacterial community on aggregates at different depth from the Canary Current Upwelling system. A free drifting sediment trap was used to sample aggregates in situ at 100 m and 400 m depth. We used a three dimensional FISH approach to quantify the bacterial community. Synechococcus dominated the bacterial community on marine snow at both depths, while Bacteroidetes and Alteromonas abundance significantly decreased with depth. We hypothesize a change in the bacterial community due to a combined effect of changes in nutrient quality due to degradation processes, grazing, decreasing temperature and increasing pressure. In summary, a strong top-down control was exerted on the bacterial and archaeal, and the nano- and picoplankton community, indicating a tight coupling of the microbial loop during the iron fertilizing experiment LOHAFEX. Marine snow investigations off Cape Blanc showed that Bacteroidetes and Alteromonas dominated the bacterial community but decreased with depth, indicating a nutrient quality, grazer, pressure and temperature dependent community composition.