In Situ Diazotroph Population Dynamics Under Different Resource Ratios in the North Pacific Subtropical Gyre.

Major advances in understanding the diversity, distribution, and activity of marine N2-fixing microorganisms (diazotrophs) have been made in the past decades, however, large gaps in knowledge remain about the environmental controls on growth and mortality rates. In order to measure diazotroph net growth rates and microzooplankton grazing rates on diazotrophs, nutrient perturbation experiments and dilution grazing experiments were conducted using free-floating in situ incubation arrays in the vicinity of Station ALOHA in March 2016. Net growth rates for targeted diazotroph taxa as well as Prochlorococcus, Synechococcus and photosynthetic picoeukaryotes were determined under high (H) and low (L) nitrate:phosphate (NP) ratio conditions at four depths in the photic zone (25, 45, 75, and 100 m) using quantitative PCR and flow cytometry. Changes in the prokaryote community composition in response to HNP and LNP treatments were characterized using 16S rRNA variable region tag sequencing. Microzooplankton grazing rates on diazotrophs were measured using a modified dilution technique at two depths in the photic zone (15 and 125 m). Net growth rates for most of the targeted diazotrophs after 48 h were not stimulated as expected by LNP conditions, rather enhanced growth rates were often measured in HNP treatments. Interestingly, net growth rates of the uncultivated prymnesiophyte symbiont UCYN-A1 were stimulated in HNP treatments at 75 and 100 m, suggesting that N used for growth was acquired through continuing to fix N2 in the presence of nitrate. Net growth rates for UCYN-A1, UCYN-C, Crocosphaera sp. (UCYN-B) and the diatom symbiont Richelia (associated with Rhizosolenia) were uniformly high at 45 m (up to 1.6 ± 0.5 d-1), implying that all were growing optimally at the onset of the experiment at that depth. Differences in microzooplankton grazing rates on UCYN-A1 and UCYN-C in 15 m waters indicate that the grazer assemblage preyed preferentially on UCYN-A1. Deeper in the water column (125 m), both diazotrophs were grazed at substantial rates, suggesting grazing pressure may increase with depth in the photic zone. Constraining in situ diazotroph growth and mortality rates are important steps for improving parameterization for diazotrophs in global ecosystem models.

Distinct ecological niches of marine symbiotic N2 -fixing cyanobacterium Candidatus Atelocyanobacterium thalassa sublineages.

A recently described symbiosis between the metabolically streamlined nitrogen-fixing cyanobacterium UCYN-A and a single-celled eukaryote prymnesiophyte alga is widely distributed throughout tropical and subtropical marine waters, and is thought to contribute significantly to nitrogen fixation in these regions. Several UCYN-A sublineages have been defined based on UCYN-A nitrogenase (nifH) sequences. Due to the low abundances of UCYN-A in the global oceans, currently existing molecular techniques are limited for detecting and quantifying these organisms. A targeted approach is needed to adequately characterize the diversity of this important marine cyanobacterium, and to advance understanding of its ecological importance. We present findings on the distribution of UCYN-A sublineages based on high throughput sequencing of UCYN-A nifH PCR amplicons from 78 samples distributed throughout many major oceanic provinces. These UCYN-A nifH fragments were used to define oligotypes, alternative taxonomic units defined by nucleotide positions with high variability. The data set was dominated by a single oligotype associated with the UCYN-A1 sublineage, consistent with previous observations of relatively high abundances in tropical and subtropical regions. However, this analysis also revealed for the first time the widespread distribution of the UCYN-A3 sublineage in oligotrophic waters. Furthermore, distinct assemblages of UCYN-A oligotypes were found in oligotrophic and coastally influenced waters. This unique data set provides a framework for determining the environmental controls on UCYN-A distributions and the ecological importance of the different sublineages.

Unusual marine unicellular symbiosis with the nitrogen-fixing cyanobacterium UCYN-A.

Nitrogen fixation - the reduction of dinitrogen (N2) gas to biologically available nitrogen (N) - is an important source of N for terrestrial and aquatic ecosystems. In terrestrial environments, N2-fixing symbioses involve multicellular plants, but in the marine environment these symbioses occur with unicellular planktonic algae. An unusual symbiosis between an uncultivated unicellular cyanobacterium (UCYN-A) and a haptophyte picoplankton alga was recently discovered in oligotrophic oceans. UCYN-A has a highly reduced genome, and exchanges fixed N for fixed carbon with its host. This symbiosis bears some resemblance to symbioses found in freshwater ecosystems. UCYN-A shares many core genes with the 'spheroid bodies' of Epithemia turgida and the endosymbionts of the amoeba Paulinella chromatophora. UCYN-A is widely distributed, and has diversified into a number of sublineages that could be ecotypes. Many questions remain regarding the physical and genetic mechanisms of the association, but UCYN-A is an intriguing model for contemplating the evolution of N2-fixing organelles.

Unscrambling Cyanobacteria Community Dynamics Related to Environmental Factors.

Future climate scenarios in the Baltic Sea project an increase of cyanobacterial bloom frequency and duration, attributed to eutrophication and climate change. Some cyanobacteria can be toxic and their impact on ecosystem services is relevant for a sustainable sea. Yet, there is limited understanding of the mechanisms regulating cyanobacterial diversity and biogeography. Here we unravel successional patterns and changes in cyanobacterial community structure using a 2-year monthly time- series during the productive season in a 100 km coastal-offshore transect using microscopy and high-throughput sequencing of 16S rRNA gene fragments. A total of 565 cyanobacterial OTUs were found, of which 231 where filamentous/colonial and 334 picocyanobacterial. Spatial differences in community structure between coastal and offshore waters were minor. An "epidemic population structure" (dominance of asingle cluster) was found for Aphanizomenon/Dolichospermum within the filamentous/colonial cyanobacterial community. In summer, this clusters imultaneously occurred with opportunistic clusters/OTUs, e.g., Nodularia spumigena and Pseudanabaena. Picocyanobacteria, Synechococcus/Cyanobium, formeda consistent but highly diverse group. Overall, the potential drivers structuring summer cyanobacterial communities were temperature and salinity. However, the different responses to environmental factors among and within genera suggest high niche specificity for individual OTUs. The recruitment and occurrence of potentially toxic filamentous/colonial clusters was likely related to disturbance such as mixing events and short-term shifts in salinity, and not solely dependent on increasing temperature and nitrogen-limiting conditions. Nutrients did not explain further the changes in cyanobacterial community composition. Novel occurrence patterns were identified as a strong seasonal succession revealing a tight coupling between the emergence of opportunistic picocynobacteria and the bloom offilamentous/colonialclusters. These findings highlight that if environmental conditions can partially explain the presence of opportunistic picocyanobacteria, microbial and trophic interactions with filamentous/colonial cyanobacteria should also be considered as potential shaping factors for single-celled communities. Regional climate change scenarios in the Baltic Sea predict environmental shifts leading to higher temperature and lower salinity; conditions identified here as favorable for opportunistic filamentous/colonial cyanobacteria. Altogether, the diversity and complexity of cyanobacterial communities reported here is far greater than previously known, emphasizing the importance of microbial interactions between filamentous and picocyanobacteria in the context of environmental disturbances.

Identification of Associations between Bacterioplankton and Photosynthetic Picoeukaryotes in Coastal Waters.

Photosynthetic picoeukaryotes are significant contributors to marine primary productivity. Associations between marine bacterioplankton and picoeukaryotes frequently occur and can have large biogeochemical impacts. We used flow cytometry to sort cells from seawater to identify non-eukaryotic phylotypes that are associated with photosynthetic picoeukaryotes. Samples were collected at the Santa Cruz wharf on Monterey Bay, CA, USA during summer and fall, 2014. The phylogeny of associated microbes was assessed through 16S rRNA gene amplicon clone and Illumina MiSeq libraries. The most frequently detected bacterioplankton phyla within the photosynthetic picoeukaryote sorts were Proteobacteria (Alphaproteobacteria and Gammaproteobacteria) and Bacteroidetes. Intriguingly, the presence of free-living bacterial genera in the photosynthetic picoeukaryote sorts could suggest that some of the photosynthetic picoeukaryotes were mixotrophs. However, the occurrence of bacterial sequences, which were not prevalent in the corresponding bulk seawater samples, indicates that there was also a selection for specific OTUs in association with photosynthetic picoeukaryotes suggesting specific functional associations. The results show that diverse bacterial phylotypes are found in association with photosynthetic picoeukaryotes. Taxonomic identification of these associations is a prerequisite for further characterizing and to elucidate their metabolic pathways and ecological functions.