13C NMR metabolomics: J-resolved STOCSY meets INADEQUATE.

Robust annotation of metabolites is a challenging task in metabolomics. Among available applications, 13C NMR experiment INADEQUATE determines direct 13C-13C connectivity unambiguously, offering indispensable information on molecular structure. Despite its great utility, it is not always practical to collect INADEQUATE data on every sample in a large metabolomics study because of its relatively long experiment time. Here, we propose an alternative approach that maintains the quality of information but saves experiment time. In this approach, individual samples in a study are first screened by 13C homonuclear J-resolved experiment (JRES). Next, JRES data are processed by statistical total correlation spectroscopy (STOCSY) to extract peaks that behave similarly among samples. Finally, INADEQUATE is collected on one internal pooled sample to select STOCSY peaks that originate from the same compound. We tested this concept using the 13C-labeled endometabolome of a model marine diatom strain incubated under various settings, intending to cover a range of metabolites produced under different external conditions. This scheme was able to extract known diatom metabolites proline, 2,3-dihydroxypropane-1-sulfonate (DHPS), ß-1,3-glucan, choline, and glutamate. This pipeline also detected unknown compounds with structural information, which is valuable in metabolomics where a priori knowledge of metabolites is not always available. The ability of this scheme was seen even in sugar regions, which are usually challenging in 1H NMR due to severe peak overlap. JRES and INADEQUATE were highly complementary; INADEQUATE provided directly-bonded 13C networks, whereas JRES linked INADEQUATE networks within the same compound but broken by nitrogen or sulfur atoms, highlighting the advantage of this integrated approach.

Bacterial transcriptional response to labile exometabolites from photosynthetic picoeukaryote Micromonas commoda.

Dissolved primary production released into seawater by marine phytoplankton is a major source of carbon fueling heterotrophic bacterial production in the ocean. The composition of the organic compounds released by healthy phytoplankton is poorly known and difficult to assess with existing chemical methods. Here, expression of transporter and catabolic genes by three model marine bacteria (Ruegeria pomeroyi DSS-3, Stenotrophomonas sp. SKA14, and Polaribacter dokdonensis MED152) was used as a biological sensor of metabolites released from the picoeukaryote Micromonas commoda RCC299. Bacterial expression responses indicated that the three species together recognized 38 picoeukaryote metabolites. This was consistent with the Micromonas expression of genes for starch metabolism and synthesis of peptidoglycan-like intermediates. A comparison of the hypothesized Micromonas exometabolite pool with that of the diatom Thalassiosira pseudonana CCMP1335, analyzed previously with the same biological sensor method, indicated that both phytoplankton released organic acids, nucleosides, and amino acids, but differed in polysaccharide and organic nitrogen release. Future ocean conditions are expected to favor picoeukaryotic phytoplankton over larger-celled microphytoplankton. Results from this study suggest that such a shift could alter the substrate pool available to heterotrophic bacterioplankton.

A suggested climate service for cyanobacteria blooms in the Baltic Sea - Comparing three monitoring methods.

Dense blooms of filamentous cyanobacteria are recurrent phenomena in the Baltic Sea, with occasional negative effects on the surrounding ecosystem, as well as on tourism, human health, aquaculture, and fisheries. Establishing a climate service is therefore suggested; including multi-method observations of cyanobacteria biomass, biodiversity, and biogeography, in correspondence to biotic and abiotic factors. Three different approaches were compared for determination of spatial and temporal variability and trends of the blooms; 1) microscopy-based long-term data, 2) satellite remote sensing, and 3) phycocyanin fluorescence mounted on a merchant vessel. Firstly, microscopy-based data on cyanobacteria biomass from the period 2000-2020 showed that the toxin producing genus Nodularia and non-toxic Aphanizomenon both had summer means of 15 µg C L-1, while Dolichospermum was less dominant with a mean of 8 µg C L-1. Some years also the Kattegat was affected by cyanobacteria blooms, likely transported here by ocean currents. Secondly, the satellite remote sensing time series for the period 2002-2020 indicated that near surface blooms were most frequent in the Northern Baltic Proper and that near surface blooms have increased in the Bothnian Sea, starting later in the season than in the Baltic Proper. The largest extents (i.e., total area covered) were observed in 2005, 2008, and 2018. Thirdly, phycocyanin fluorescence from a flow through sensor mounted on a merchant vessel was used as a proxy for cyanobacteria biomass and correlated to cyanobacteria biomass estimated by microscopy. However, the satellite remote sensing data on surface accumulations showed little resemblance to the data on cyanobacteria biomass based on water sampling and microscopy, interpreted as an effect of methods. Sensors on satellites mainly detect surface accumulations of cyanobacteria while the microscopy data was based on samples 0-10 m, thereby comprising a larger community. Data from satellite remote sensing of cyanobacteria was correlated to the phycocyanin fluorescence indicating that similar bio-optical properties are observed. Finally, results from a downscaled ocean climate model (NEMONordic) were used to produce future scenarios for temperature and salinity, which directly affects cyanobacteria blooms in the Baltic Sea, supposedly by increasing in abundance and change in species composition. Short-term forecasts can be used together with observations for early warning of cyanobacteria blooms, and we suggest an internationally coordinated cyanobacteria observation and warning system for the Baltic Sea area.

Covert therapeutic micro-processes in non-recovered eating disorders with childhood trauma: an interpersonal process recall study.

METHOD: To uncover therapeutic micro-processes from the perspectives of eating disorder (ED) treatment non-responders with childhood trauma (CT) late effects, we explored in-session experiences of poor long-term outcome patients. Female inpatients aged 28-59 (M = 40.2, SD = 5.0) from a randomised trial comparing Compassion Focused Therapy for EDs (n = 3) with Cognitive Behavioural Therapy for EDs (n = 3) were interviewed with video-assisted recall about a self-selected session. Data were analysed through Interpretative Phenomenological Analysis (IPA) with Grounded Theory (GT) elements. RESULTS: Covert patient strategies included self-effacement, regulating therapeutic distance to open up, and engaging with reflective rather than experiential interventions. First, self-effacement included submissive, passive or pretend responses to perceived criticising or violating therapist behaviours as well as other orientation and submission for approval. Second, some preferred a close patient-therapist alliance with therapist self-disclosure and reciprocity was a requirement for opening up; others required distance. Third, informants detached from experiential trauma work while engaging in joint reflection on post-trauma responses. CONCLUSION: Informants were preoccupied with calibrating the emotional-relational landscape in session; we hypothesized that psychological insecurity and affective intolerance from CT limit their freedom to explore own in-session experiences.

A patient perspective on individual ED treatment processes is scarce for difficult-to-treat eating disorders (EDs) with childhood trauma (CT). We therefore interviewed six poor long-term outcome inpatients through video-assisted recall about a self-selected therapy session. Patients' covert in-session strategies included self-effacing behaviours in relation to their therapist, their respective preferences for closeness or distance to their therapist to be able to open up in session, and being more prone to engage in therapists' reflective interventions rather than experiential. In sum, patients were preoccupied with calibrating the emotional­relational landscape between patient and therapist in session, which were strategies that went undetected. We hypothesized that a lack of psychological security and affective tolerance limit patients freedom to explore own experiences from the perspective of traumatic attachment and self-differentiation theory.

Temporal escape-adaptation to eutrophication by Skeletonema marinoi.

Diatoms commonly set off the spring-bloom in temperate coastal environments. However, their temporal offset may change in regions subject to nutrient enrichment, and by peaking earlier, such populations can maintain their position in the vernal plankton succession. We tested whether the marine keystone diatom Skeletonema marinoi can accomplish this through thermal evolutionary adaptation. Eight geographically separated subpopulations, representing hydromorphologically and climatologically similar inlets displaying a range of trophic states, were compared in a common-garden experiment. At early-spring temperatures, both doubling times and variation coefficients thereof, correlated negatively with the trophic state of the environment of origin, indicating selection for fast growth due to eutrophication. At mid-spring temperatures, the relationships were reversed, indicating selection in the opposite direction. At late-spring temperatures, no significant relationships were detected, suggesting relaxed selection. Subsequent field observations reflected these findings, where blooming temperatures decreased with trophic state. Natural selection thus moves along with eutrophication towards colder temperatures earlier in the spring, favouring genotypes with the capacity to grow fast. The thermal niche shift demonstrated herein may be an evolutionary mechanism essentially leading to trophic changes in the local ecosystem.

Growth-stage-related shifts in diatom endometabolome composition set the stage for bacterial heterotrophy.

Phytoplankton-derived metabolites fuel a large fraction of heterotrophic bacterial production in the global ocean, yet methodological challenges have limited our understanding of the organic molecules transferred between these microbial groups. In an experimental bloom study consisting of three heterotrophic marine bacteria growing together with the diatom Thalassiosira pseudonana, we concurrently measured diatom endometabolites (i.e., potential exometabolite supply) by nuclear magnetic resonance (NMR) spectroscopy and bacterial gene expression (i.e., potential exometabolite uptake) by metatranscriptomic sequencing. Twenty-two diatom endometabolites were annotated, with nine increasing in internal concentration in the late stage of the bloom, eight decreasing, and five showing no variation through the bloom progression. Some metabolite changes could be linked to shifts in diatom gene expression, as well as to shifts in bacterial community composition and their expression of substrate uptake and catabolism genes. Yet an overall low match indicated that endometabolome concentration was not a good predictor of exometabolite availability, and that complex physiological and ecological interactions underlie metabolite exchange. Six diatom endometabolites accumulated to higher concentrations in the bacterial co-cultures compared to axenic cultures, suggesting a bacterial influence on rates of synthesis or release of glutamate, arginine, leucine, 2,3-dihydroxypropane-1-sulfonate, glucose, and glycerol-3-phosphate. Better understanding of phytoplankton metabolite production, release, and transfer to assembled bacterial communities is key to untangling this nearly invisible yet pivotal step in ocean carbon cycling.

Diel investments in metabolite production and consumption in a model microbial system.

Organic carbon transfer between surface ocean photosynthetic and heterotrophic microbes is a central but poorly understood process in the global carbon cycle. In a model community in which diatom extracellular release of organic molecules sustained growth of a co-cultured bacterium, we determined quantitative changes in the diatom endometabolome and the bacterial uptake transcriptome over two diel cycles. Of the nuclear magnetic resonance (NMR) peaks in the diatom endometabolites, 38% had diel patterns with noon or mid-afternoon maxima; the remaining either increased (36%) or decreased (26%) through time. Of the genes in the bacterial uptake transcriptome, 94% had a diel pattern with a noon maximum; the remaining decreased over time (6%). Eight diatom endometabolites identified with high confidence were matched to the bacterial genes mediating their utilization. Modeling of these coupled inventories with only diffusion-based phytoplankton extracellular release could not reproduce all the patterns. Addition of active release mechanisms for physiological balance and bacterial recognition significantly improved model performance. Estimates of phytoplankton extracellular release range from only a few percent to nearly half of annual net primary production. Improved understanding of the factors that influence metabolite release and consumption by surface ocean microbes will better constrain this globally significant carbon flux.

Nitrogen fixation estimates for the Baltic Sea indicate high rates for the previously overlooked Bothnian Sea.

Dense blooms of diazotrophic filamentous cyanobacteria are formed every summer in the Baltic Sea. We estimated their contribution to nitrogen fixation by combining two decades of cyanobacterial biovolume monitoring data with recently measured genera-specific nitrogen fixation rates. In the Bothnian Sea, estimated nitrogen fixation rates were 80 kt N year-1, which has doubled during recent decades and now exceeds external loading from rivers and atmospheric deposition of 69 kt year-1. The estimated contribution to the Baltic Proper was 399 kt N year-1, which agrees well with previous estimates using other approaches and is greater than the external input of 374 kt N year-1. Our approach can potentially be applied to continuously estimate nitrogen loads via nitrogen fixation. Those estimates are crucial for ecosystem adaptive management since internal nitrogen loading may counteract the positive effects of decreased external nutrient loading.

Large seasonal and spatial variation in nano- and microphytoplankton diversity along a Baltic Sea-North Sea salinity gradient.

Aquatic phytoplankton experience large fluctuations in environmental conditions during seasonal succession and across salinity gradients, but the impact of this variation on their diversity is poorly understood. We examined spatio-temporal variation in nano- and microphytoplankton (> 2 µm) community structure using almost two decades of light-microscope based monitoring data. The dataset encompasses 19 stations that span a salinity gradient from 2.8 to 35 along the Swedish coastline. Spatially, both regional and local phytoplankton diversity increased with broad-scale salinity variation. Diatoms dominated at high salinity and the proportion of cyanobacteria increased with decreasing salinity. Temporally, cell abundance peaked in winter-spring at high salinity but in summer at low salinity. This was likely due to large filamentous cyanobacteria blooms that occur in summer in low salinity areas, but which are absent in higher salinities. In contrast, phytoplankton local diversity peaked in spring at low salinity but in fall and winter at high salinity. Whilst differences in seasonal variation in cell abundance were reasonably well-explained by variation in salinity and nutrient availability, variation in local-scale phytoplankton diversity was poorly predicted by environmental variables. Overall, we provide insights into the causes of spatio-temporal variation in coastal phytoplankton community structure while also identifying knowledge gaps.

Basin-specific changes in filamentous cyanobacteria community composition across four decades in the Baltic Sea.

Almost every summer, dense blooms of filamentous cyanobacteria are formed in the Baltic Sea. These blooms may cause problems for tourism and ecosystem services, where surface accumulations and beach fouling are commonly occurring. Future changes in environmental drivers, including climate change and other anthropogenic disturbances, may further enhance these problems. By compiling monitoring data from countries adjacent to the Baltic Sea, we present spatial and temporal genus-specific distribution of diazotrophic filamentous cyanobacteria (Nostocales) during four decades (1979-2017). While the summer surface salinity decreased with a half up to one unit, the surface temperature in July-August increased with 2-3 °C in most sub-basins of the Baltic Sea, during the time period. The biovolumes of the toxic Nodularia spumigena did not change in any of the sub-basins during the period. On the other hand, the biovolume of the non-toxic Aphanizomenon sp. and the potentially toxic Dolichospermum spp. increased in the northern parts of the Baltic Sea, along with the decreased salinity and elevated temperatures, but Aphanizomenon sp. decreased in the southern parts despite decreased salinity and increased temperatures. These contradictory changes in biovolume of Aphanizomenon sp. between the northern and southern parts of the Baltic Sea may be due to basin-specific effects of the changed environmental conditions, or can be related to local adaptation by sub-populations of the genera. Overall, this comprehensive dataset presents insights to genus-specific bloom dynamics by potentially harmful diazotrophic filamentous cyanobacteria in the Baltic Sea.

Resting Stages of Skeletonema marinoi Assimilate Nitrogen From the Ambient Environment Under Dark, Anoxic Conditions.

The planktonic marine diatom Skeletonema marinoi forms resting stages, which can survive for decades buried in aphotic, anoxic sediments and resume growth when re-exposed to light, oxygen, and nutrients. The mechanisms by which they maintain cell viability during dormancy are poorly known. Here, we investigated cell-specific nitrogen (N) and carbon (C) assimilation and survival rate in resting stages of three S. marinoi strains. Resting stages were incubated with stable isotopes of dissolved inorganic N (DIN), in the form of 15 N-ammonium (NH4+ ) or -nitrate (NO3- ) and dissolved inorganic C (DIC) as 13 C-bicarbonate (HCO3- ) under dark and anoxic conditions for 2 months. Particulate C and N concentration remained close to the Redfield ratio (6.6) during the experiment, indicating viable diatoms. However, survival varied between <0.1% and 47.6% among the three different S. marinoi strains, and overall survival was higher when NO3- was available. One strain did not survive in the NH4+ treatment. Using secondary ion mass spectrometry (SIMS), we quantified assimilation of labeled DIC and DIN from the ambient environment within the resting stages. Dark fixation of DIC was insignificant across all strains. Significant assimilation of 15 N-NO3- and 15 N-NH4+ occurred in all S. marinoi strains at rates that would double the nitrogenous biomass over 77-380 years depending on strain and treatment. Hence, resting stages of S. marinoi assimilate N from the ambient environment at slow rates during darkness and anoxia. This activity may explain their well-documented long survival and swift resumption of vegetative growth after dormancy in dark and anoxic sediments.

Change processes related to long-term outcomes in eating disorders with childhood trauma: An explorative qualitative study.

To date, eating disorder (ED) treatments are lacking for ED patients with psychiatric sequelae of childhood trauma, and successful outcomes are scarce. Therefore, the aim of this study was to explore therapeutic change processes from a patient perspective in relation to good versus poor long-term ED outcome at 1-year follow-up. Outcome categories were based on clinician assessment of ED behavior and diagnoses, body mass index, and Eating Disorder Examination Questionnaire scores. Eleven White, cisgendered female ED patients with childhood trauma were interviewed after a 3-month inpatient treatment: data was analyzed with elements from grounded theory and interpretative phenomenological analysis. The qualitative analysis rendered change-related descriptions (9 subcategories) and obstacles to change (6 subcategories), and 3 process-related domains differentiated good from poor long-term outcome: trauma exposure (4 subcategories), patient agency (6 subcategories), and patient-therapist dynamics (3 subcategories). First, sensory and emotional trauma exposure in good outcome informants was contrasted with avoiding or not addressing trauma and body in poor outcome informants. Second, promotion of patient agency while receiving support in the good outcome group was contrasted to an orientation toward others' needs, distrust in own abilities, and difficulties showing vulnerability in the poor outcome group. Third, poor outcome informants described either a distanced or immersed/idealizing relationship to their therapist, as opposed to more balanced between self-assertion and vulnerability in good outcome informants. Our findings raise new hypotheses that trauma work, fostering patient agency, and focusing on relational dynamics in patient-therapist dyad may be important in producing enduring ED outcomes for these patients. (PsycINFO Database Record (c) 2019 APA, all rights reserved).

Nitrate and ammonium fluxes to diatoms and dinoflagellates at a single cell level in mixed field communities in the sea.

Growth of large phytoplankton is considered to be diffusion limited at low nutrient concentrations, yet their constraints and contributions to carbon (C) and nitrogen fluxes in field plankton communities are poorly quantified under this condition. Using secondary ion mass spectrometry (SIMS), we quantified cell-specific assimilation rates of C, nitrate, and ammonium in summer communities of large phytoplankton when dissolved inorganic nitrogen concentrations are low in temperate coastal regions. Chain-forming diatoms composed 6% of total particulate organic carbon, but contributed 20% of C assimilation, 54% of nitrate assimilation and 32% of ammonium assimilation within the plankton community. In contrast, large dinoflagellates composed 11% of total POC, and contributed 14% of the C assimilation, 4% of ammonium and 9% of nitrate assimilation within the plankton community. Measured cell-specific C and nitrate assimilation rate match the Redfield ratio and the maximal nitrate assimilation in Chaetoceros spp. predicted by mass transfer theory. However, average ammonium assimilation rates were 30 and 340% higher than predicted by mass transfer theory in Tripos/Ceratium and Chaetoceros, respectively, suggesting that microbial interactions in the phycosphere may facilitate substantial luxury ammonium uptake by Chaetoceros in environments with fluctuating nitrate concentrations.

High single-cell diversity in carbon and nitrogen assimilations by a chain-forming diatom across a century.

Almost a century ago Redfield discovered a relatively constant ratio between carbon, nitrogen and phosphorus in particulate organic matter and nitrogen and phosphorus of dissolved nutrients in seawater. Since then, the riverine export of nitrogen to the ocean has increased 20 fold. High abundance of resting stages in sediment layers dated more than a century back indicate that the common planktonic diatom Skeletonema marinoi has endured this eutrophication. We germinated unique genotypes from resting stages originating from isotope-dated sediment layers (15 and 80 years old) in a eutrophied fjord. Using secondary ion mass spectrometry (SIMS) combined with stable isotopic tracers, we show that the cell-specific carbon and nitrogen assimilation rates vary by an order of magnitude on a single-cell level but are significantly correlated during the exponential growth phase, resulting in constant assimilation quota in cells with identical genotypes. The assimilation quota varies largely between different clones independent of age. We hypothesize that the success of S. marinoi in coastal waters may be explained by its high diversity of nutrient demand not only at a clone-specific level but also at the single-cell level, whereby the population can sustain and adapt to dynamic nutrient conditions in the environment.

Ocean acidification and desalination: climate-driven change in a Baltic Sea summer microplanktonic community.

Helcom scenario modelling suggests that the Baltic Sea, one of the largest brackish-water bodies in the world, could expect increased precipitation (decreased salinity) and increased concentration of atmospheric CO2 over the next 100 years. These changes are expected to affect the microplanktonic food web, and thereby nutrient and carbon cycling, in a complex and possibly synergistic manner. In the Baltic Proper, the extensive summer blooms dominated by the filamentous cyanobacteria Aphanizomenon sp., Dolichospermum spp. and the toxic Nodularia spumigena contribute up to 30% of the yearly new nitrogen and carbon exported to the sediment. In a 12 days outdoor microcosm experiment, we tested the combined effects of decreased salinity (from 6 to 3) and elevated CO2 concentrations (380 and 960 µatm) on a natural summer microplanktonic community, focusing on diazotrophic filamentous cyanobacteria. Elevated pCO2 had no significant effects on the natural microplanktonic community except for higher biovolume of Dolichospermum spp. and lower biomass of heterotrophic bacteria. At the end of the experimental period, heterotrophic bacterial abundance was correlated to the biovolume of N. spumigena. Lower salinity significantly affected cyanobacteria together with biovolumes of dinoflagellates, diatoms, ciliates and heterotrophic bacteria, with higher biovolume of Dolichospermum spp. and lower biovolume of N. spumigena, dinoflagellates, diatoms, ciliates and heterotrophic bacteria in reduced salinity. Although the salinity effects on diatoms were apparent, they could not clearly be separated from the influence of inorganic nutrients. We found a clear diurnal cycle in photosynthetic activity and pH, but without significant treatment effects. The same diurnal pattern was also observed in situ (pCO2, pH). Thus, considering the Baltic Proper, we do not expect any dramatic effects of increased pCO2 in combination with decreased salinity on the microplanktonic food web. However, long-term effects of the experimental treatments need to be further studied, and indirect effects of the lower salinity treatments could not be ruled out. Our study adds one piece to the complicated puzzle to reveal the combined effects of increased pCO2 and reduced salinity levels on the Baltic microplanktonic community.

A psychometric evaluation of the Swedish version of the Responses to Positive Affect questionnaire.

BACKGROUND: Previous research mainly focused on responses to negative affect in relation to depression, and less on responses to positive affect. Cognitive responses to positive affect are interesting in the context of emotion regulation and emotion disorders: positive rumination is associated to hypomania risk and bipolar disorder. There is to date no questionnaire in Swedish that captures the phenomena of cognitive response styles. AIMS: The aim of this study was to investigate the replicability of the Responses to Positive Affect questionnaire (RPA) in a newly translated Swedish version and to test its psychometric properties. METHODS: Swedish undergraduates (n = 111) completed a set of self-report questionnaires in a fixed order. RESULTS: The hypothesized three-factor model was largely replicated in the subscales Self-focused positive rumination, Emotion-focused positive rumination and Dampening. The two positive rumination subscales were strongly associated with each other and current positive affect. The subscales showed acceptable convergent and incremental validity with concurrent measures of depression, hypomania, anxiety, repetitive negative thinking, and positive and negative affect. The model explained 25% of the variance in hypomania, but fell short in the explanation of depression. CONCLUSIONS: The Swedish version of the RPA shows satisfactory reliability and initial findings from a student sample indicate that it is a valid measure comparable with the original RPA questionnaire. RESULTS give emphasis to the importance of further exploration of cognitive response styles in relation to psychopathology.