Visible surface films, termed slicks, can extensively cover freshwater and marine ecosystems, with coastal regions being particularly susceptible to their presence. The sea-surface microlayer (SML), the upper 1-mm at the air-water interface in slicks (herein slick SML) harbors a distinctive bacterial community, but generally little is known about SML viruses. Using flow cytometry, metagenomics, and cultivation, we characterized viruses and bacteria in a brackish slick SML in comparison to non-slick SML as well as seawater below slick and non-slick areas (subsurface water = SSW). Size-fractionated filtration of all samples distinguished viral attachment to hosts and particles. The slick SML contained higher abundances of virus-like particles, prokaryotic cells, and dissolved organic carbon compared to non-slick SML and SSW. The community of 428 viral operational taxonomic units (vOTUs), 426 predicted as lytic, distinctly differed across all size fractions in the slick SML compared to non-slick SML and SSW. Specific metabolic profiles of bacterial metagenome-assembled genomes and isolates in the slick SML included a prevalence of genes encoding motility and carbohydrate-active enzymes (CAZymes). Several vOTUs were enriched in slick SML, and many virus variants were associated with particles. Nine vOTUs were only found in slick SML, six of them being targeted by slick SML-specific clustered-regularly interspaced short palindromic repeats (CRISPR) spacers likely originating from Gammaproteobacteria. Moreover, isolation of three previously unknown lytic phages for Alishewanella sp. and Pseudoalteromonas tunicata, abundant and actively replicating slick SML bacteria, suggests that viral activity in slicks contributes to biogeochemical cycling in coastal ecosystems.

Phages modulate bacterial metabolism during infection by regulating gene expression, which influences aquatic nutrient cycling. However, the effects of shifting nutrient regimes are less understood. Here, we analyzed transcriptomes of an ecologically relevant Gammaproteobacterium and its lytic phage in high (HNM) and low (LNM) nutrient medium. Despite different infection characteristics, including reduced burst size and longer latent period in LNM, the phage had a fixed expression profile. Bacterial transcription was instead different depending on nutrient regime, with HNM bacteria focusing on growth while LNM bacteria focused on motility and membrane transport. Additionally, phage infection had a larger effect on bacterial gene expression in LNM compared to HNM, e.g. suppressing increased iron uptake and altering expression of phosphorus uptake genes. Overall, phage infection influenced host metabolism more in LNM, which was more similar to natural conditions, emphasizing the importance of considering natural conditions to understand phage and host ecology.

Phage predation constitutes a major mortality factor for bacteria in aquatic ecosystems, and thus, directly impacts nutrient cycling and microbial community dynamics. Yet, the population dynamics of specific phages across time scales from days to months remain largely unexplored, which limits our understanding of their influence on microbial succession. To investigate temporal changes in diversity and abundance of phages infecting particular host strains, we isolated 121 phage strains that infected three bacterial hosts during a Baltic Sea mesocosm experiment. Genome analysis revealed a novel Flavobacterium phage genus harboring gene sets putatively coding for synthesis of modified nucleotides and glycosylation of bacterial cell surface components. Another novel phage genus revealed a microdiversity of phage species that was largely maintained during the experiment and across mesocosms amended with different nutrients. In contrast to the newly described Flavobacterium phages, phages isolated from a Rheinheimera strain were highly similar to previously isolated genotypes, pointing to genomic consistency in this population. In the mesocosm experiment, the investigated phages were mainly detected after a phytoplankton bloom peak. This concurred with recurrent detection of the phages in the Baltic Proper during summer months, suggesting an influence on the succession of heterotrophic bacteria associated with phytoplankton blooms.

Polynucleobacter asymbioticusstrain QLW-P1DMWA-1(T)represents a group of highly successful heterotrophic ultramicrobacteria that is frequently very abundant (up to 70% of total bacterioplankton) in freshwater habitats across all seven continents. This strain was originally isolated from a shallow Alpine pond characterized by rapid changes in water temperature and elevated UV radiation due to its location at an altitude of 1300 m. To elucidate the strain's adjustment to fluctuating environmental conditions, we recorded changes occurring in its transcriptomic and proteomic profiles under contrasting experimental conditions by simulating thermal conditions in winter and summer as well as high UV irradiation. To analyze the potential connection between gene expression and regulation via methyl group modification of the genome, we also analyzed its methylome. The methylation pattern differed between the three treatments, pointing to its potential role in differential gene expression. An adaptive process due to evolutionary pressure in the genus was deduced by calculating the ratios of non-synonymous to synonymous substitution rates for 20Polynucleobacterspp. genomes obtained from geographically diverse isolates. The results indicate purifying selection.

Polynucleobacter (Burkholderiaceae, Betaproteobacteria) and Limnohabitans (Comamonadaceae, Betaproteobacteria) are abundant freshwater bacteria comprising large genetic and taxonomic diversities, with species adapted to physico-chemically distinct types of freshwater systems. The relative importance of environmental drivers, i.e., physico-chemistry, presence of microeukaryotes and geographic position for the diversity and prevalence has not been investigated for both taxa before. Here, we present the first pan-European study on this topic, comprising 255 freshwater lakes. We investigated Limnohabitans and Polynucleobacter using an amplicon sequencing approach of partial 16S rRNA genes along environmental gradients. We show that physico-chemical factors had the greatest impact on both genera. Analyses on environmental gradients revealed an exceptionally broad ecological spectrum of operational taxonomic units (OTUs). Despite the coarse resolution of the genetic marker, we found OTUs with contrasting environmental preferences within Polynucleobacter and Limnohabitans subclusters. Such an ecological differentiation has been characterized for PnecC and LimC before but was so far unknown for less well studied subclusters such as PnecA and PnecB. Richness and abundance of OTUs are geographically clustered, suggesting that geographic diversity patterns are attributable to region-specific physico-chemical characteristics (e.g., pH and temperature) rather than latitudinal gradients or lake sizes.