First Viruses Infecting the Marine Diatom Guinardia delicatula

The marine diatom Guinardia delicatula is a cosmopolitan species that dominates seasonal blooms in the English Channel and the North Sea. Several eukaryotic parasites are known to induce the mortality of this species. Here, we report the isolation and characterization of the first viruses that infect G. delicatula. Viruses were isolated from the Western English Channel (SOMLIT-Astan station) during the late summer bloom decline of G. delicatula. A combination of laboratory approaches revealed that these lytic viruses (GdelRNAV) are small tailless particles of 35–38 nm in diameter that replicate in the host cytoplasm where both unordered particles and crystalline arrays are formed. GdelRNAV display a linear single-stranded RNA genome of ~9 kb, including two open reading frames encoding for replication and structural polyproteins. Phylogenetic relationships based on the RNA-dependent-RNA-polymerase gene marker showed that GdelRNAV are new members of the Bacillarnavirus, a monophyletic genus belonging to the order Picornavirales. GdelRNAV are specific to several strains of G. delicatula. They were rapidly and largely produced (<12 h, 9.34 × 104 virions per host cell). Our analysis points out the host's variable viral susceptibilities during the early exponential growth phase. Interestingly, we consistently failed to isolate viruses during spring and early summer while G. delicatula developed important blooms. While our study suggests that viruses do contribute to the decline of G. delicatula's late summer bloom, they may not be the primary mortality agents during the remaining blooms at SOMLIT-Astan. Future studies should focus on the relative contribution of the viral and eukaryotic pathogens to the control of Guinardia's blooms to understand the fate of these prominent organisms in marine systems

Impact de la température sur les interactions entre la microalgue marine Micromonas et les virus qui l’infectent

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An abscisic acid-independent oxylipin pathway controls stomatal closure and immune defense in <em>Arabidopsis</em>

International audiencePlant stomata function in innate immunity against bacterial invasion and abscisic acid (ABA) has been suggested to regulate this process. Using genetic, biochemical, and pharmacological approaches, we demonstrate that (i) the Arabidopsis thaliana nine-specific-lipoxygenase encoding gene, LOX1, which is expressed in guard cells, is required to trigger stomatal closure in response to both bacteria and the pathogen-associated molecular pattern flagellin peptide flg22; (ii) LOX1 participates in stomatal defense; (iii) polyunsaturated fatty acids, the LOX substrates, trigger stomatal closure; (iv) the LOX products, fatty acid hydroperoxides, or reactive electrophile oxylipins induce stomatal closure; and (v) the flg22-mediated stomatal closure is conveyed by both LOX1 and the mitogen-activated protein kinases MPK3 and MPK6 and involves salicylic acid whereas the ABA-induced process depends on the protein kinases OST1, MPK9, or MPK12. Finally, we show that the oxylipin and the ABA pathways converge at the level of the anion channel SLAC1 to regulate stomatal closure. Collectively, our results demonstrate that early biotic signaling in guard cells is an ABA-independent process revealing a novel function of LOX1-dependent stomatal pathway in plant immunity

Eradications as scientific experiments: first attempt to eradicate two major invasive taxa, <em>Rattus rattus</em> and <em>Carpobrotus spp</em>. from a Mediterranean island

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Involvement of OST1 and MAPKs in biotic- and ABA-mediated stomatal closure.

<p>(A) Dose response effects of ABA, flg22, and 13-KODE were performed on WT (Ler) and <i>ost1-2</i> mutant plants according to the experimental design described in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001513#pbio-1001513-g002" target="_blank">Figure 2</a>, and data represent means ± SD of three independent experiments. (B) OST1 activity in Col-0 suspension cells was analyzed at 10 min by in-gel kinase assay after the treatments: ABA (30 µM), flg22 (100 nM), and 9- and 13-KODE (100 nM). 0.1% ethanol in water (mock). Kinase activity was estimated on proteins immunoprecipitated with anti-SnRK2-6 (OST1) antibody. The gel is typical of three independent experiments. (C) Stomatal aperture measurements were performed on WT (Col-0) and <i>mpk9-1/12-1</i> mutant plants after incubation with ABA (10 µM) and flg22 (5 µM). (D) Protein kinase activities of Col-0 suspension cells were analyzed at 10 min by in-gel kinase assay after the treatments: ABA (30 µM), flg22 (100 nM), and 9- and 13-KODE (100 nM). Kinase activity was estimated on 20 µg of crude protein extracts, and the gel is typical of three independent experiments. (E) Typical immunoblot of three independent experiments performed with total proteins extracted from 2-wk-old plants (Col-0) either treated with ABA (100 µM) or flg22 (1 µM). Mock treatments were 0.1% ethanol in water for ABA and water for flg22. MAPKs were detected (upper panel) with the rabbit anti-phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) monoclonal antibody. Coomassie staining of proteins is shown for loading control (lower panel). (F) Stomatal aperture measurements were performed on WT (Col-0), <i>mpk3</i>, and <i>mpk6</i> mutant plants after incubation with ABA (10 µM), flg22 (5 µM), linoleic acid (18∶2, 100 nM), and 9-KODE (1 nM).</p

Stomatal closure responses to fatty acids and hydroperoxides.

<p>Measurements were performed on 2 h light-preincubated epidermal peels from 4–5-wk-old Arabidopsis plants after 2.5 h incubation, and data represent means ± SD of three independent experiments. (A) Dose response effects of linoleic acid (18∶2) on Col-0 plants. Student's <i>t</i> test analysis of means indicates that the effect of 18∶2 becomes significant at 10 nM. (B) Effects of oleic (18∶1), linoleic (18∶2), and linolenic (18∶3) acids at a concentration of 100 nM. Values marked with asterisks were statistically different from the corresponding light controls. (C) Dose response effects of FAHs (9- and 13-HPODE) H₂O₂ and t-BuOOH on Col-0 plants. Student's <i>t</i> test analysis of means indicates that the effects of 9- and 13-HPODE become significant at 0.1 nM. (D) NAC (1 mM) was added to epidermal peels 30 min prior to addition of hydroperoxides. Values marked with asterisks were significantly different from those of the corresponding sample without NAC pre-incubation, and means marked with identical letters were not statistically different (a: <i>F</i> = 0.737, <i>df</i> = 11, <i>p</i> = 0.559; b: <i>F</i> = 1.792, <i>df</i> = 11, <i>p</i> = 0.189).</p

Stomatal closure responses to thiol-reagents.

<p>Measurements were performed on Col-0 plants according to the experimental design described in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1001513#pbio-1001513-g002" target="_blank">Figure 2</a>, and data represent means ± SD of three independent experiments. (A) Dose response effects of RES oxylipins (9- and 13-KODE). Student's <i>t</i> test analysis of means indicates that the effects of 9- and 13-KODE become significant at 0.1 nM. (B) Comparison of the effects of 9- and 13-KODE to their corresponding alcohols 9- and 13-HODE. Values marked with asterisks were significantly different from those of the corresponding alcohols. (C) Structures and reactivity of compounds with thiols. (D) NAC (1 mM) was added to epidermal peels 30 min prior to addition of thiol-reagents or ABA. Marked values were significantly different from those of samples without NAC pre-incubation.</p

Stomatal defense and stomatal closure responses of <i>lox1</i> mutant lines.

<p>(A) Growth of <i>Pst</i> DC3000 was assessed 3 days after spray inoculation at 5×10⁷ cfu/mL. Bars represent means of five replicates ± SE. Similar results were obtained in three independent experiments. (B) Stomatal apertures were measured on 2 h light-preincubated epidermal peels, incubated for 1 h with bacterial suspensions (10⁸ cfu/mL) or 2.5 h with flg22 (5 µM) or (C) with ABA. Data represent means ± SD of three independent experiments, and values marked with asterisks were statistically different from those of the corresponding light controls. In (C), means marked with identical letters were not statistically different (a: <i>F</i> = 0.421, <i>df</i> = 11, <i>p</i> = 0.743; b: <i>F</i> = 3.683, <i>df</i> = 11, <i>p</i> = 0.062; c: <i>F</i> = 3.726, <i>df</i> = 11, <i>p</i> = 0.061).</p

Effects of NAC and 13-HODE on different stimuli inducing stomatal closure.

<p>Stomatal aperture measurements were determined on leaf peels from 4–5-wk-old Arabidopsis plants on 2 h light-preincubated epidermal peels after 2.5 h incubation with the following inducers, (A and B) 13-KODE, PCMB, sulforaphane, ABA, or (C) flg22 (5 µM), or after 1 h incubation with the bacterial suspension of <i>Pst</i> DC3000 (10⁸ cfu/mL). Pre-treatments with NAC (1 mM) or with 13-HODE (1 nM or 1 µM for ABA treatments) were performed 30 min prior to addition of stomatal closure inducers. Data represent means ± SD of three independent experiments. Asterisks represent significant differences from controls (in (A) samples pre-treated with 13-HODE, in (C) light). In (B) letters represent values not statistically different (a: <i>F</i> = 0.0127, <i>df</i> = 8, <i>p</i> = 0.987; b: <i>F</i> = 0.310, <i>df</i> = 14, <i>p</i> = 0.865).</p

Eradications as scientific experiments : progress in simultaneous eradications of two major invasive taxa from a Mediterranean island

BACKGROUNDBlack rats, Rattus rattus, and mat-forming iceplants, Carpobrotus aff. acinaciformis and Carpobrotus edulis, are pervasive pests on Mediterranean islands. Their cumulative impacts on native biotas alter the functioning of island ecosystems and threaten biodiversity. A report is given here of the first attempt to eradicate both taxa from a protected nature reserve in south-eastern France (Bagaud Island). In order to minimise unwanted hazardous outcomes and produce scientific knowledge, the operations were embedded in a four-step strategy including initial site assessment, planning, restoration and monitoring. RESULTSTrapping, which resulted in the removal of 1923 rats in 21045 trap-nights, made it possible to eliminate a substantial proportion of the resident rat population and to reduce the amount of rodenticide delivered in the second stage of the operation. Forty tons of Carpobrotus spp. were manually uprooted from a total area of 18000 m(2); yet careful monitoring over a decade is still required to prevent germinations from the seed bank. CONCLUSIONTwo years after the beginning of the interventions, both eradication operations are still ongoing. Biosecurity measures have been implemented to reduce reinvasion risks of both taxa. With the long-term monitoring of various native plants and animals, Bagaud Island will become a reference study site for scientific purposes