Lead exposure is related to higher infection rate with the gapeworm in Norwegian house sparrows (Passer domesticus).

Anthropogenic pollution is identified as an important threat to bird and other wildlife populations. Many metals and toxic elements, along with poly- and perfluoroalkyl substances (PFASs) are known to induce immunomodulation and have previously been linked to increased pathogen prevalence and infectious disease severity. In this study, the house sparrow (Passer domesticus) was investigated at the coast of Helgeland in northern Norway. This population is commonly infected with the parasitic nematode "gapeworm" (Syngamus trachea), with a prevalence of 40-60 % during summer months. Gapeworm induces severe respiratory disease in birds and has been previously demonstrated to decrease survival and reproductive success in wild house sparrows. The aim of this study was to investigate whether a higher exposure to pollution with PFASs, metals and other elements influences gapeworm infection in wild house sparrows. We conducted PFASs and elemental analysis on whole blood from 52 house sparrows from Helgeland, including analyses of highly toxic metals such as lead (Pb), mercury (Hg) and arsenic (As). In addition, we studied gapeworm infection load by counting the parasite eggs in faeces from each individual. We also studied the expression of microRNA 155 (miR155) as a key regulator in the immune system. Elevated blood concentrations of Pb were found to be associated with an increased prevalence of gapeworm infection in the house sparrow. The expression of miR155 in the plasma of the house sparrow was only weakly associated with Pb. In contrast, we found relatively low PFASs concentrations in the house sparrow blood (∑ PFASs 0.00048-354 µg/L) and PFASs were not associated to miR155 nor infection rate. The current study highlights the potential threat posed by Pb as an immunotoxic pollutant in small songbirds.

IIb-RAD-sequencing coupled with random forest classification indicates regional population structuring and sex-specific differentiation in salmon lice (Lepeophtheirus salmonis).

The aquaculture industry has been dealing with salmon lice problems forming serious threats to salmonid farming. Several treatment approaches have been used to control the parasite. Treatment effectiveness must be optimized, and the systematic genetic differences between subpopulations must be studied to monitor louse species and enhance targeted control measures. We have used IIb-RAD sequencing in tandem with a random forest classification algorithm to detect the regional genetic structure of the Norwegian salmon lice and identify important markers for sex differentiation of this species. We identified 19,428 single nucleotide polymorphisms (SNPs) from 95 individuals of salmon lice. These SNPs, however, were not able to distinguish the differential structure of lice populations. Using the random forest algorithm, we selected 91 SNPs important for geographical classification and 14 SNPs important for sex classification. The geographically important SNP data substantially improved the genetic understanding of the population structure and classified regional demographic clusters along the Norwegian coast. We also uncovered SNP markers that could help determine the sex of the salmon louse. A large portion of the SNPs identified to be under directional selection was also ranked highly important by random forest. According to our findings, there is a regional population structure of salmon lice associated with the geographical location along the Norwegian coastline.

Simultaneous knockout of multiple LHCF genes using single sgRNAs and engineering of a high-fidelity Cas9 for precise genome editing in marine algae.

The CRISPR/Cas9 system is an RNA-guided sequence-specific genome editing tool, which has been adopted for single or multiple gene editing in a wide range of organisms. When working with gene families with functional redundancy, knocking out multiple genes within the same family may be required to generate a phenotype. In this study, we tested the possibility of exploiting the known tolerance of Cas9 for mismatches between the single-guide RNA (sgRNA) and target site to simultaneously introduce indels in multiple homologous genes in the marine diatom Phaeodactylum tricornutum. As a proof of concept, we designed two sgRNAs that could potentially target the same six light-harvesting complex (LHC) genes belonging to the LHCF subgroup. Mutations in up to five genes were achieved simultaneously using a previously established CRISPR/Cas9 system for P. tricornutum. A visible colour change was observed in knockout mutants with multiple LHCF lesions. A combination of pigment, LHCF protein and growth analyses was used to further investigate the phenotypic differences between the multiple LHCF mutants and WT. Furthermore, we used the two same sgRNAs in combination with a variant of the existing Cas9 where four amino acids substitutions had been introduced that previously have been shown to increase Cas9 specificity. A significant reduction of off-target editing events was observed, indicating that the altered Cas9 functioned as a high-fidelity (HiFi) Cas9 nuclease.

The Imaging of Guard Cells of thioglucosidase (tgg) Mutants of Arabidopsis Further Links Plant Chemical Defence Systems with Physical Defence Barriers.

The glucosinolate-myrosinase system is a well-known plant chemical defence system. Two functional myrosinase-encoding genes, THIOGLUCOSIDASE 1 (TGG1) and THIOGLUCOSIDASE 2 (TGG2), express in aerial tissues of Arabidopsis. TGG1 expresses in guard cells (GCs) and is also a highly abundant protein in GCs. Recently, by studying wild type (WT), tgg single, and double mutants, we showed a novel association between the glucosinolate-myrosinase system defence system, and a physical barrier, the cuticle. In the current study, using imaging techniques, we further analysed stomata and ultrastructure of GCs of WT, tgg1, tgg2 single, and tgg1 tgg2 double mutants. The tgg mutants showed distinctive features of GCs. The GCs of tgg1 and tgg1 tgg2 mutants showed vacuoles that had less electron-dense granular material. Both tgg single mutants had bigger stomata complexes. The WT and tgg mutants also showed variations for cell wall, chloroplasts, and starch grains of GCs. Abscisic acid (ABA)-treated stomata showed that the stomatal aperture was reduced in tgg1 single and tgg1 tgg2 double mutants. The data provides a basis to perform comprehensive further studies to find physiological and molecular mechanisms associated with ultrastructure differences in tgg mutants. We speculate that the absence of myrosinase alters the endogenous chemical composition, hence affecting the physical structure of plants and the plants' physical defence barriers.

Assessment of oxidative stress response genes in Avicennia marina exposed to oil contamination - Polyphenol oxidase (PPOA) as a biomarker.

Mangrove plants, which inhabit and form sensitive ecosystems in the intertidal zones of tropical and subtropical coastlines, though vulnerable to petroleum pollution, still maintain their growth under oil contamination. To elucidate the molecular response of mangrove plants to crude oil-sediment mixture, seeds of Avicennia marina were planted and grown on 0, 2.5, 5.0, 7.5 and10 % (w/w) oil-contaminated soil. Plant biomass was highly affected from 3.05 ± 0.28 (Control) to 0.50 ± .07 (10 %) and from 3.47 ± 0.12 to 1.88 ± 0.08 in 2 and 4 months old plants respectively. The expression analysis of 11genes belonging to detoxification pathways in the roots and leaves of 2 and 4 month-old plants was evaluated by qRT-PCR. Our results showed changes in expression levels of Fe-SOD, Mn-SOD, CAT, PRX, PPOs, GSTs, and NAP2 whose products are involved in reactive oxygen species (ROS) and xenobiotic detoxification. PPOA showed the highest expression induction of 43 ± 1.15, followed by CAT (12.61 ± 3.25) and PPOB (6.38 ± 1.34) in leaves of 2 months old seedlings grown on 7.5, 10 and 7.5 % oil contaminated soil respectively. PPOA (39.23 ± 2.1), PRX (32.13 ± 1.2) as well as PPOB (26.11 ± 1.3) showed the highest expression induction in leaves of 4 months old plants grown in 2.5 % oil contaminated soil. Our data indicated that PPOA can be a good biomarker candidate gene for long term exposure to oil contamination in A. marina.

Author Correction: The Seminavis robusta genome provides insights into the evolutionary adaptations of benthic diatoms.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The Seminavis robusta genome provides insights into the evolutionary adaptations of benthic diatoms.

Benthic diatoms are the main primary producers in shallow freshwater and coastal environments, fulfilling important ecological functions such as nutrient cycling and sediment stabilization. However, little is known about their evolutionary adaptations to these highly structured but heterogeneous environments. Here, we report a reference genome for the marine biofilm-forming diatom Seminavis robusta, showing that gene family expansions are responsible for a quarter of all 36,254 protein-coding genes. Tandem duplications play a key role in extending the repertoire of specific gene functions, including light and oxygen sensing, which are probably central for its adaptation to benthic habitats. Genes differentially expressed during interactions with bacteria are strongly conserved in other benthic diatoms while many species-specific genes are strongly upregulated during sexual reproduction. Combined with re-sequencing data from 48 strains, our results offer insights into the genetic diversity and gene functions in benthic diatoms.

The Role of a Glucosinolate-Derived Nitrile in Plant Immune Responses.

Glucosinolates are defense-related secondary metabolites found in Brassicaceae. When Brassicaceae come under attack, glucosinolates are hydrolyzed into different forms of glucosinolate hydrolysis products (GHPs). Among the GHPs, isothiocyanates are the most comprehensively characterized defensive compounds, whereas the functional study of nitriles, another group of GHP, is still limited. Therefore, this study investigates whether 3-butenenitrile (3BN), a nitrile, can trigger the signaling pathways involved in the regulation of defense responses in Arabidopsis thaliana against biotic stresses. Briefly, the methodology is divided into three stages, (i) evaluate the physiological and biochemical effects of exogenous 3BN treatment on Arabidopsis, (ii) determine the metabolites involved in 3BN-mediated defense responses in Arabidopsis, and (iii) assess whether a 3BN treatment can enhance the disease tolerance of Arabidopsis against necrotrophic pathogens. As a result, a 2.5 mM 3BN treatment caused lesion formation in Arabidopsis Columbia (Col-0) plants, a process found to be modulated by nitric oxide (NO). Metabolite profiling revealed an increased production of soluble sugars, Krebs cycle associated carboxylic acids and amino acids in Arabidopsis upon a 2.5 mM 3BN treatment, presumably via NO action. Primary metabolites such as sugars and amino acids are known to be crucial components in modulating plant defense responses. Furthermore, exposure to 2.0 mM 3BN treatment began to increase the production of salicylic acid (SA) and jasmonic acid (JA) phytohormones in Arabidopsis Col-0 plants in the absence of lesion formation. The production of SA and JA in nitrate reductase loss-of function mutant (nia1nia2) plants was also induced by the 3BN treatments, with a greater induction for JA. The SA concentration in nia1nia2 plants was lower than in Col-0 plants, confirming the previously reported role of NO in controlling SA production in Arabidopsis. A 2.0 mM 3BN treatment prior to pathogen assays effectively alleviated the leaf lesion symptom of Arabidopsis Col-0 plants caused by Pectobacterium carotovorum ssp. carotovorum and Botrytis cinerea and reduced the pathogen growth on leaves. The findings of this study demonstrate that 3BN can elicit defense response pathways in Arabidopsis, which potentially involves a coordinated crosstalk between NO and phytohormone signaling.

The Myb-like transcription factor phosphorus starvation response (PtPSR) controls conditional P acquisition and remodelling in marine microalgae.

Phosphorus (P) is one of the limiting macronutrients for algal growth in marine environments. Microalgae have developed adaptation mechanisms to P limitation that involve remodelling of internal phosphate resources and accumulation of lipids. Here, we used in silico analyses to identify the P-stress regulator PtPSR (Phaeodactylum tricornutum phosphorus starvation response) in the diatom P. tricornutum. ptpsr mutant lines were generated using gene editing and characterised by various molecular, genetics and biochemical tools. PtPSR belongs to a clade of Myb transcription factors that are conserved in stramenopiles and distantly related to plant P-stress regulators. PtPSR bound specifically to a conserved cis-regulatory element found in the regulatory region of P-stress-induced genes. ptpsr knockout mutants showed reduction in cell growth under P limitation. P-stress responses were impaired in ptpsr mutants compared with wild-type, including reduced induction of P-stress response genes, near to complete loss of alkaline phosphatase activity and reduced phospholipid degradation. We conclude that PtPSR is a key transcription factor influencing P scavenging, phospholipid remodelling and cell growth in adaptation to P stress in diatoms.

PAMP-INDUCED SECRETED PEPTIDE 3 modulates immunity in Arabidopsis.

Small post-translationally modified peptides are important signalling components of plant defence responses against phytopathogens, acting as both positive and negative modulators. PAMP-INDUCED SECRETED PEPTIDE (PIP) 1 and 2 have been shown to amplify plant immunity. Here we investigate the role of the related peptide PIP3 in the regulation of immune response in Arabidopsis. Treatment with synthetic PIP peptides led to similar transcriptome reprogramming, indicating an effect on innate immunity-related processes and phytohormones, including jasmonic acid (JA) biosynthesis and signalling. PIP3 overexpressing (OX) plants showed enhanced growth inhibition in response to flg22 exposure. In addition, flg22-induced production of reactive oxygen species and callose deposition was significantly reduced in PIP3-OX plants. Interestingly, PIP3-OX plants showed increased susceptibility toward both Botrytis cinerea and the biotrophic pathogen Pseudomonas syringae. Expression of both JA and salicylic acid (SA) biosynthesis and signalling genes was more induced during B. cinerea infection in PIP3-OX plants compared with wild-type plants. Promoter and ChIP-seq analyses indicated that the transcription factors WRKY18, WRKY33, and WRKY40 cooperatively act as repressors for PIP3. The results point to a fine-tuning role for PIP3 in modulation of immunity through the regulation of SA and JA biosynthesis and signalling pathways in Arabidopsis.

CRISPR/Cas9-mediated editing of Δ5 and Δ6 desaturases impairs Δ8-desaturation and docosahexaenoic acid synthesis in Atlantic salmon (Salmo salar L.).

The in vivo functions of Atlantic salmon fatty acyl desaturases (fads2), Δ6fads2-a, Δ6fads2-b, Δ6fads2-c and Δ5fads2 in long chain polyunsaturated fatty acid (LC-PUFA) synthesis in salmon and fish in general remains to be elucidated. Here, we investigate in vivo functions and in vivo functional redundancy of salmon fads2 using two CRISPR-mediated partial knockout salmon, Δ6abc/5Mt with mutations in Δ6fads2-a, Δ6fads2-b, Δ6fads2-c and Δ5fads2, and Δ6bcMt with mutations in Δ6fads2-b and Δ6fads2-c. F0 fish displaying high degree of gene editing (50-100%) were fed low LC-PUFA and high LC-PUFA diets, the former containing reduced levels of eicosapentaenoic (20:5n-3) and docosahexaenoic (22:6n-3) acids but higher content of linoleic (18:2n-6) and alpha-linolenic (18:3n-3) acids, and the latter containing high levels of 20:5n-3 and 22:6n-3 but reduced compositions of 18:2n-6 and 18:3n-3. The Δ6abc/5Mt showed reduced 22:6n-3 levels and accumulated Δ6-desaturation substrates (18:2n-6, 18:3n-3) and Δ5-desaturation substrate (20:4n-3), demonstrating impaired 22:6n-3 synthesis compared to wildtypes (WT). Δ6bcMt showed no effect on Δ6-desaturation compared to WT, suggesting Δ6 Fads2-a as having the predominant Δ6-desaturation activity in salmon, at least in the tissues analyzed. Both Δ6abc/5Mt and Δ6bcMt demonstrated significant accumulation of Δ8-desaturation substrates (20:2n-6, 20:3n-3) when fed low LC-PUFA diet. Additionally, Δ6abc/5Mt demonstrated significant upregulation of the lipogenic transcription regulator, sterol regulatory element binding protein-1 (srebp-1) in liver and pyloric caeca under reduced dietary LC-PUFA. Our data suggest a combined effect of endogenous LC-PUFA synthesis and dietary LC-PUFA levels on srebp-1 expression which ultimately affects LC-PUFA synthesis in salmon. Our data also suggest Δ8-desaturation activities for salmon Δ6 Fads2 enzymes.

bHLH-PAS protein RITMO1 regulates diel biological rhythms in the marine diatom Phaeodactylum tricornutum.

Periodic light-dark cycles govern the timing of basic biological processes in organisms inhabiting land as well as the sea, where life evolved. Although prominent marine phytoplanktonic organisms such as diatoms show robust diel rhythms, the mechanisms regulating these processes are still obscure. By characterizing a Phaeodactylum tricornutum bHLH-PAS nuclear protein, hereby named RITMO1, we shed light on the regulation of the daily life of diatoms. Alteration of RITMO1 expression levels and timing by ectopic overexpression results in lines with deregulated diurnal gene expression profiles compared with the wild-type cells. Reduced gene expression oscillations are also observed in these lines in continuous darkness, showing that the regulation of rhythmicity by RITMO1 is not directly dependent on light inputs. We also describe strong diurnal rhythms of cellular fluorescence in wild-type cells, which persist in continuous light conditions, indicating the existence of an endogenous circadian clock in diatoms. The altered rhythmicity observed in RITMO1 overexpression lines in continuous light supports the involvement of this protein in circadian rhythm regulation. Phylogenetic analysis reveals a wide distribution of RITMO1-like proteins in the genomes of diatoms as well as in other marine algae, which may indicate a common function in these phototrophs. This study adds elements to our understanding of diatom biology and offers perspectives to elucidate timekeeping mechanisms in marine organisms belonging to a major, but under-investigated, branch of the tree of life.

Dietary fatty acid source has little effect on the development of the immune system in the pyloric caeca of Atlantic salmon fry.

The quality and relative amounts of dietary lipids may affect the health and growth of cultured Atlantic salmon. So far, little is known about their effects on the performance of the fish immune system during early life stages and, in particular their importance in the transition from endogenous nutrition (yolk) in the alevin stage to exogenous nutrition in the later fry stage. We investigated the immunomodulatory effects of fish oil, vegetable oil and phospholipid-rich oil in feeds for farmed Atlantic salmon using a transcriptomic approach. The experiment allowed a fine-scale monitoring of gene expression profiles in two tissues, the pyloric caeca of the intestine and the liver, in a 94 days-long first feeding experiment. The analysis of transcriptional profiles revealed that first feeding induced a strong immunomodulation in the pyloric caeca after 48 days of feeding, lasting up to day 94 and possibly beyond. On the other hand, the differential effect of the three dietary regimes was negligible. We interpret this upregulation, undetectable in liver, as a potentiation of the immune system upon the first contact of the digestive system with exogenous feed. This process involved a complex network of gene products involved in both cellular and humoral immunity. We identified the classical pathway of the complement system, acting at the crossroads between innate and adaptive immunity, as a key process modulated in response to the switch from endogenous to exogenous nutrition.

Unique photosynthetic electron transport tuning and excitation distribution in heterokont algae.

Heterokont algae are significant contributors to marine primary productivity. These algae have a photosynthetic machinery that shares many common features with that of Viridiplantae (green algae and land plants). Here we demonstrate, however, that the photosynthetic machinery of heterokont algae responds to light fundamentally differently than that of Viridiplantae. While exposure to high light leads to electron accumulation within the photosynthetic electron transport chain in Viridiplantae, this is not the case in heterokont algae. We use this insight to manipulate the photosynthetic electron transport chain and demonstrate that heterokont algae can dynamically distribute excitation energy between the two types of photosystems. We suggest that the reported electron transport and excitation distribution features are adaptations to the marine light environment.

Accumulation of Ag(I) by Saccharomyces cerevisiae Cells Expressing Plant Metallothioneins.

The various applications of Ag(I) generated the necessity to obtain Ag(I)-accumulating organisms for the removal of surplus Ag(I) from contaminated sites or for the concentration of Ag(I) from Ag(I)-poor environments. In this study we obtained Ag(I)-accumulating cells by expressing plant metallothioneins (MTs) in the model Saccharomyces cerevisiae. The cDNAs of seven Arabidopsis thaliana MTs (AtMT1a, AtMT1c, AtMT2a, AtMT2b, AtMT3, AtMT4a and AtMT4b) and four Noccaea caerulescens MTs (NcMT1, NcMT2a, NcMT2b and NcMT3) fused to myrGFP displaying an N-terminal myristoylation sequence for plasma membrane targeting were expressed in S. cerevisiae and checked for Ag(I)-related phenotype. The transgenic yeast cells were grown in copper-deficient media to ensure the expression of the plasma membrane high-affinity Cu(I) transporter Ctr1, and also to elude the copper-related inhibition of Ag(I) transport into the cell. All plant MTs expressed in S. cerevisiae conferred Ag(I) tolerance to the yeast cells. Among them, myrGFP-NcMT3 afforded Ag(I) accumulation under high concentration (10⁻50 µM), while myrGFP-AtMT1a conferred increased accumulation capacity under low (1 µM) or even trace Ag(I) (0.02⁻0.05 µM). The ability to tolerate high concentrations of Ag(I) coupled with accumulative characteristics and robust growth showed by some of the transgenic yeasts highlighted the potential of these strains for biotechnology applications.

Genome editing in diatoms: achievements and goals.

Diatoms are major components of phytoplankton and play a key role in the ecology of aquatic ecosystems. These algae are of great scientific importance for a wide variety of research areas, ranging from marine ecology and oceanography to biotechnology. During the last 20 years, the availability of genomic information on selected diatom species and a substantial progress in genetic manipulation, strongly contributed to establishing diatoms as molecular model organisms for marine biology research. Recently, tailored TALEN endonucleases and the CRISPR/Cas9 system were utilized in diatoms, allowing targeted genetic modifications and the generation of knockout strains. These approaches are extremely valuable for diatom research because breeding, forward genetic screens by random insertion, and chemical mutagenesis are not applicable to the available model species Phaeodactylum tricornutum and Thalassiosira pseudonana, which do not cross sexually in the lab. Here, we provide an overview of the genetic toolbox that is currently available for performing stable genetic modifications in diatoms. We also discuss novel challenges that need to be addressed to fully exploit the potential of these technologies for the characterization of diatom biology and for metabolic engineering.

Benzyl Cyanide Leads to Auxin-Like Effects Through the Action of Nitrilases in Arabidopsis thaliana.

Plants within the Brassicales order generate glucosinolate hydrolysis products that can exert different biological effects on several organisms. Here, we evaluated the physiological effects of one of these compounds, benzyl cyanide (phenylacetonitrile), when exogenously applied on Arabidopsis thaliana. Treatment with benzyl cyanide led to a dose-dependent reduction of primary root length and total biomass. Further morphological changes like elongated hypocotyls, epinastic cotyledons, and increased formation of adventitious roots resembled a severe auxin-overproducer phenotype. The elevated auxin response was confirmed by histochemical staining and gene expression analysis of auxin-responsive genes. Nitriles are converted by specific enzymes, nitrilases (NIT1-3), to their corresponding carboxylic acids. The nitrilase mutants nit1 and nit2 tolerated benzyl cyanide treatments better than the wild type, with nit2 being less resistant than nit1. A NIT2RNAi line suppressing several nitrilases was resistant to all tested benzyl cyanide concentrations. When exposed to phenylacetic acid (PAA) - the corresponding carboxylic acid of benzyl cyanide - wild type and mutant seedlings were, however, equally susceptible and showed a more severe auxin phenotype than upon cyanide treatment. Here, we demonstrate that the auxin-like effects triggered by benzyl cyanide on Arabidopsis are due to its nitrilase-mediated conversion to the natural auxin PAA.

Arabidopsis mutants impaired in glutathione biosynthesis exhibit higher sensitivity towards the glucosinolate hydrolysis product allyl-isothiocyanate.

Upon tissue damage the plant secondary metabolites glucosinolates can generate various hydrolysis products, including isothiocyanates (ITCs). Their role in plant defence against insects and pest and their potential health benefits have been well documented, but our knowledge regarding the endogenous molecular mechanisms of their effect in plants is limited. Here we investigated the effect of allyl-isothiocyanate (AITC) on Arabidopsis thaliana mutants impaired in homeostasis of the low-molecular weight thiol glutathione. We show that glutathione is important for the AITC-induced physiological responses, since mutants deficient in glutathione biosynthesis displayed a lower biomass and higher root growth inhibition than WT seedlings. These mutants were also more susceptible than WT to another ITC, sulforaphane. Sulforaphane was however more potent in inhibiting root growth than AITC. Combining AITC with the glutathione biosynthesis inhibitor L-buthionine-sulfoximine (BSO) led to an even stronger phenotype than observed for the single treatments. Furthermore, transgenic plants expressing the redox-sensitive fluorescent biomarker roGFP2 indicated more oxidative conditions during AITC treatment. Taken together, we provide genetic evidence that glutathione plays an important role in AITC-induced growth inhibition, although further studies need to be conducted to reveal the underlying mechanisms.

Rearing Water Treatment Induces Microbial Selection Influencing the Microbiota and Pathogen Associated Transcripts of Cod (Gadus morhua) Larvae.

We have previously shown that K-selection and microbial stability in the rearing water increases survival and growth of Atlantic cod (Gadus morhua) larvae, and that recirculating aquaculture systems (RAS) are compatible with this. Here, we have assessed how water treatment influenced the larval microbiota and host responses at the gene expression level. Cod larvae were reared with two different rearing water systems: a RAS and a flow-through system (FTS). The water microbiota was examined using a 16S rDNA PCR/DGGE strategy. RNA extracted from larvae at 8, 13, and 17 days post hatching was used for microbiota and microarray gene expression analysis. Bacterial cDNA was synthesized and used for 16S rRNA amplicon 454 pyrosequencing of larval microbiota. Both water and larval microbiota differed significantly between the systems, and the larval microbiota appeared to become more dissimilar between systems with time. In total 4 phyla were identified for all larvae: Actinobacteria, Bacteroidetes, Firmicutes, and Proteobacteria. The most profound difference in larval microbiota was a high abundance of Arcobacter (Epsilonproteobacteria) in FTS larvae (34 ± 9% of total reads). Arcobacter includes several species that are known pathogens for humans and animals. Cod larval transcriptome responses were investigated using an oligonucleotide gene expression microarray covering approximately 24,000 genes. Interestingly, FTS larvae transcriptional profiles revealed an overrepresentation of upregulated transcripts associated with responses to pathogens and infections, such as c1ql3-like, pglyrp-2-like and zg16, compared to RAS larvae. In conclusion, distinct water treatment systems induced differences in the larval microbiota. FTS larvae showed up-regulation of transcripts associated with responses to microbial stress. These results are consistent with the hypothesis that RAS promotes K-selection and microbial stability by maintaining a microbial load close to the carrying capacity of the system, and ensuring long retention times for both bacteria and water in the system.

Molecular adaptations to phosphorus deprivation and comparison with nitrogen deprivation responses in the diatom Phaeodactylum tricornutum.

Phosphorus, an essential element for all living organisms, is a limiting nutrient in many regions of the ocean due to its fast recycling. Changes in phosphate (Pi) availability in aquatic systems affect diatom growth and productivity. We investigated the early adaptive mechanisms in the marine diatom Phaeodactylum tricornutum to P deprivation using a combination of transcriptomics, metabolomics, physiological and biochemical experiments. Our analysis revealed strong induction of gene expression for proteins involved in phosphate acquisition and scavenging, and down-regulation of processes such as photosynthesis, nitrogen assimilation and nucleic acid and ribosome biosynthesis. P deprivation resulted in alterations of carbon allocation through the induction of the pentose phosphate pathway and cytosolic gluconeogenesis, along with repression of the Calvin cycle. Reorganization of cellular lipids was indicated by coordinated induced expression of phospholipases, sulfolipid biosynthesis enzymes and a putative betaine lipid biosynthesis enzyme. A comparative analysis of nitrogen- and phosphorus-deprived P. tricornutum revealed both common and distinct regulation patterns in response to phosphate and nitrate stress. Regulation of central carbon metabolism and amino acid metabolism was similar, whereas unique responses were found in nitrogen assimilation and phosphorus scavenging in nitrogen-deprived and phosphorus-deprived cells, respectively.