Growth promotion of <i>Arabidopsis</i> seedling by VOCs emitted from <i>Paenibacillus polymyxa</i> E681.

<p>A) Illustration of plant growth promotion by VOCs produced by strain E681 in a 24-well microtitre system. The diameter of each well was 2 cm. The photograph was taken two weeks after inoculation with strain E681. Right plate = water control; Left plate = E681 treatment. The single arrows indicate inoculation site with strain E681 and water control. B) Plant growth at two weeks after exposure to VOCs released by <i>P. polymyxa</i> E681, <i>Bacillus subtilis</i> GB03, and water control in a microtitre system, as indicated by the differences in the total leaf surface areas. C) Plant growth at two weeks after exposure to VOCs released by <i>P. polymyxa</i> E681, <i>Bacillus subtilis</i> GB03, and water control in a microtitre system, as indicated by the differences in the foliar fresh weight in wild type Col-0 and its hormonal mutant lines, transgenic NahG (encodes salicylate hydroxylase and degrades salicylic acid (SA)), <i>ein2.5</i> (ET-insensitive), <i>coi1</i> (insensitive to jasmonic acid), and <i>gai2</i> (insensitive to gibberellic acid). Different letters indicate significant differences between treatments at 2, 4, and 6 cm away from bacteria inoculation in the microtitre system, according to least significant difference at <i>P</i> = 0.05.</p

Mitochondrial genome-based synthesis and timeline of Eurasian otter (<i>Lutra lutra</i>) phylogeography

Eurasian otters (Lutra lutra) have a broad distribution across Eurasia, but biogeographic data outside of western Europe is limited to disconnected pockets. Based on current subspecies designations, Asia appears to harbour a large proportion of the species’ diversity, with 10 of 12 Eurasian otter subspecies found in Asia. Here we provide a range-wide synthesis of mitochondrial data, inferring a timeline and pattern of phylogeographic signals. Whole mitochondrial genomes of 27 Eurasian otters across 4 subspecies are presented from newly generated data (n = 6; 4 from Korea, 1 from Hong Kong and 2 from UK), assembled from the Sequence Read Archive (n = 4), and sourced from GenBank (n = 17). We then combined whole mitochondrial genome results with cytochrome b data to increase the sample size and contextualise our results with prior studies. We identified five distinct lineages that were discordant with current subspecies classification. Phylogenetic dating revealed that the earliest diverging lineage was the Japanese lineage, with remaining lineages diverging ≥0.08 million years ago. Mitochondrial diversity calculated by sample locations seemed mainly driven by the presence of multiple lineages. When grouping samples by lineage, genetic diversity was highest in Lineage 1 (primarily found in China and Laos), followed by Lineage 2 (primarily found in Korea), and lowest diversity identified in Lineage 3 (primarily found in Europe). Our findings highlight previously undetected lineage diversity within Eurasian otters, but also the need for further taxonomic and genomic evaluation of the species in Asia. The identified unique, distinct lineages of Eurasian otters also warrant urgent conservation attention.</p

Induced Resistance by a Long-Chain Bacterial Volatile: Elicitation of Plant Systemic Defense by a C13 Volatile Produced by <em>Paenibacillus polymyxa</em>

<div><h3>Background</h3><p>Some strains of plant growth-promoting rhizobacteria (PGPR) elicit induced systemic resistance (ISR) by emission of volatile organic compounds (VOCs) including short chain alcohols, acetoin, and 2,3-butanediol. The objective of this study was to evaluate whether species-specific VOCs from PGPR strain <em>Paenibacillus polymyxa</em> E681 can promote growth and induce resistance in <em>Arabidopsis</em>.</p> <h3>Methodology/Principal Findings</h3><p>The efficacy of induction was strain-specific, with stronger protection against <em>Pseudomonas syringae</em> pv. maculicola ES4326 in plants exposed to VOCs from <em>P. polymyxa</em> E681 versus <em>Arabidopsis</em> plants exposed to VOCs from a reference strain <em>Bacillus subtilis</em> GB03, which was previously shown to elicit ISR and plant growth promotion. VOC emissions released from E681 primed transcriptional expression of the salicylic acid, jasmonic acid, and ethylene signaling marker genes <em>PR1</em>, <em>ChiB,</em> and <em>VSP2</em>, respectively. In addition, strain E681 produced more than thirty low molecular-weight VOCs, of which tridecane was only produced by E681 and not found in GB03 or IN937a volatile blends. These strain-specific VOCs induced <em>PR1</em> and <em>VSP2</em> genes.</p> <h3>Conclusions/Significance</h3><p>These results provide new insight into the existence of a long chain VOC signaling molecule produced by <em>P. polymyxa</em> that can serve as a bacterial trigger of induced systemic resistance <em>in planta</em>.</p> </div

Reduction of Ca_v1.3 channels in dorsal hippocampus impairs the development of dentate gyrus newborn neurons and hippocampal-dependent memory tasks

<div><p>Ca_v1.3 has been suggested to mediate hippocampal neurogenesis of adult mice and contribute to hippocampal-dependent learning and memory processes. However, the mechanism of Ca_v1.3 contribution in these processes is unclear. Here, roles of Ca_v1.3 of mouse dorsal hippocampus during newborn cell development were examined. We find that knock-out (KO) of Ca_v1.3 resulted in the reduction of survival of newborn neurons at 28 days old after mitosis. The retroviral eGFP expression showed that both dendritic complexity and the number and length of mossy fiber bouton (MFB) filopodia of newborn neurons at ≥ 14 days old were significantly reduced in KO mice. Both contextual fear conditioning (CFC) and object-location recognition tasks were impaired in recent (1 day) memory test while passive avoidance task was impaired only in remote (≥ 20 days) memory in KO mice. Results using adeno-associated virus (AAV)-mediated Ca_v1.3 knock-down (KD) or retrovirus-mediated KD in dorsal hippocampal DG area showed that the recent memory of CFC was impaired in both KD mice but the remote memory was impaired only in AAV KD mice, suggesting that Ca_v1.3 of mature neurons play important roles in both recent and remote CFC memory while Ca_v1.3 in newborn neurons is selectively involved in the recent CFC memory process. Meanwhile, AAV KD of Ca_v1.3 in ventral hippocampal area has no effect on the recent CFC memory. In conclusion, the results suggest that Ca_v1.3 in newborn neurons of dorsal hippocampus is involved in the survival of newborn neurons while mediating developments of dendritic and axonal processes of newborn cells and plays a role in the memory process differentially depending on the stage of maturation and the type of learning task.</p></div

Induced systemic resistance and priming of defense-related genes by tridecane in <i>Arabidopsis</i>.

<p>A) Effect of tridecane on <i>Arabidopsis</i> growth. Plants were exposed to 10 mM and 100 µM tridecane, strain E681, and water for 2 weeks in presence (black bar) and absence of Ba(OH)₂ (white bar). The photograph shows effect of CO₂ on <i>Arabidopsis</i> growth treatments with 10 mM and 100 µM tridecane, strain E681, and water by the addition of Ba(OH)₂ for trapping CO₂ that precipitated as BaCO₃. B) Induced systemic resistance against <i>P. syringae</i> pv. maculicola ES4326 elicited by strain E681 and 10 mM and 100 µM tridecane using the I-plate system. Disease severity (0 = no symptom, 5 = severe chlorosis) was recorded seven days after pathogen challenge at 10⁸ cfu/ml. Inset picture indicates chemical structure of tridecane. Gene expression levels of <i>PR1</i> for salicylic acid signaling (C and F), <i>ChiB</i> for ET signaling (D and G), and <i>VSP2</i> for jasmonic acid signaling (E and H), as determined by quantitative reverse transcriptase (qRT)-PCR after tridecane emission (C, D, and E) at 0 and 6 hour post-inoculation (hpi) and after pathogen challenge for detecting defense priming (F, G, and H). The expression ratio (C–H), a ratio of the expression in the strain E681 or tridecane-inoculated treatment relative to expression of <i>Actin</i> gene, is shown as the mean ± SEM. Different letters indicate significant differences between treatments (A and B) according to least significant difference at <i>P</i> = 0.05.</p

Induction of resistance against <i>Pseudomonas syringae</i> pv. maculicola ES4326 in <i>Arabidopsis</i> exposed to bacterial VOCs.

<p>Induced systemic resistance against <i>P. syringae</i> pv. maculicola ES4326 elicited by VOCs of <i>P. polymyxa</i> E681 and a water control, using the microtitre system. Disease severity (0 = no symptom, 10 = severe chlorosis) was recorded seven days after pathogen challenge. Different letters indicate significant differences between treatments, according to least significant difference at <i>P</i> = 0.05. The error bars indicate SEM.</p

Induced resistance and priming of defense-related genes by long chain VOCs.

<p>A)Induced systemic resistance against <i>P. syringae</i> pv. maculicola ES4326 elicited by strain E681 and 10 mM and 100 µM of decane, undecane, and dodecane using the I-plate system. Disease severity (0 = no symptom, 5 = severe chlorosis) was recorded seven days after pathogen challenge at 10⁸ cfu/ml. Gene expression levels of <i>PR1</i> for salicylic acid signaling (B and E), <i>ChiB</i> for ET signaling (C and F), and <i>VSP2</i> for jasmonic acid signaling (D and G), as determined by quantitative reverse transcriptase (qRT)-PCR after tridecane emission (B, C, and D) at 0 and 6 hour post-inoculation (hpi) and after pathogen challenge for detecting defense priming (E, F, and G). The expression ratio (B–G), a ratio of the expression in the strain E681 or tridecane treatment relative to water-treated control, is shown as the mean ± SEM. Different letters indicate significant differences between treatments (A) according to least significant difference at <i>P</i> = 0.05.</p

Impairments of hippocampus-dependent memory tasks in Ca_v1.3 KO mice.

<p>(A) Scheme of CFC learning and memory tests. Both recent and remote CFC memories were assessed in the same chamber at Days 0, 1, 2 and 23. (B) Freezing responses of CFC memory tasks. (Day 0, WT, 0 ± 0, n = 5, KO, 0 ± 0, n = 5; Day 1, WT, 39.80 ± 3.33%, n = 15, KO, 8.39 ± 1.40%, n = 11, <i>p</i> < 0.00001; Day 2, WT, 59.59 ± 5.60%, n = 13, KO, 47.18 ± 2.74%, n = 9, <i>p</i> = 0.098; Day 23, WT, 56.75 ± 4.58%, n = 10, KO, 38.74 ± 14.30%, n = 6, p = 0.305). *, **, *** indicate <i>p</i> < 0.05, <i>p</i> < 0.01, <i>p</i> < 0.001, respectively, unless otherwise mentioned. Two-way ANOVA, F_G = 18.24, <i>p</i> = 0.000; F_T = 17.88, <i>p</i> = 0.000; F_G+T = 2.31, <i>p</i> = 0.106. (C) Scheme of PA tasks. (D) Latency of entrance to dark room of PA tasks. (Day 0, WT, 13.13 ± 2.88 s, n = 24, KO, 13.12 ± 1.75 s, n = 24, <i>p</i> = 0.990; Day 1, WT, 268.89 ± 16.18 s, n = 24, KO, 239.34 ± 20.72 s, n = 24, <i>p</i> = 0.267; Day 21, WT, 241.38 ± 28.90 s, n = 13, KO, 294.95 ± 5.05 s, n = 13, <i>p</i> = 0.080; Day 42, WT, 232.67 ± 34.10 s, n = 9, KO, 107.22 ± 27.86 s, n = 12, <i>p</i> = 0.010; Day 63, WT, 224.59 ± 34.76 s, n = 7, KO, 46.29 ± 10.71 s, n = 10, <i>p</i> = 0.000). Two-way ANOVA, F_G = 2.45, <i>p</i> = 0.12; F_T = 17.47, <i>p</i> = 0.00; F_G+T = 2.33, <i>p</i> = 0.08. (E) Schemes of OR and OLR tasks. (F) Preference index measurement of OR/OLR tasks. (OR task: WT, 76.41 ± 1.66%, n = 11, KO, 72.54 ± 4.0%, n = 9, <i>p</i> = 0.339; OLR task: WT, 55.19 ± 4.04%, n = 11, KO, 42.89 ± 4.13%, n = 9, <i>p</i> = 0.048).</p

Expression of Ca_v1.3 in adult hippocampal area.

<p>(A) Ca_v1.3 expression in dorsal hippocampal area. Ca_v1.3 is shown in red and DAPI, a nuclear maker, is shown in blue. <i>Scale bars</i>, 200 μm (10x) and 50 μm (40x). (B) Images of developmental profiling of Ca_v1.3 expression in adult hippocampal newborn neurons. Confocal images of adult hippocampal newborn neurons, infected with GFP-retrovirus and stained with Ca_v1.3 antibody (red), were taken at 3, 7, 14 and 28 days after infection. White arrows indicate newborn cells infected with retrovirus. <i>Scale bars</i>, 50μm (40x) and 10 μm (40x/6x-zoom). (C) Ca_v1.3 antibody fluorescent intensity of newborn neurons (GFP (+), filled bar) and control mature neurons (GFP (-), open bar) of dorsal hippocampus shown at (B). A.U. indicates arbitrary unit. (Day 3, GFP(+), 658.10 ± 41.58, n = 9, GFP(-), 1302.51 ± 40.98, n = 50; Day 7, GFP(+), 558.19 ± 61.26, n = 9, GFP(-), 1149.03 ± 126.35, n = 50; Day 14, GFP(+), 950.79 ± 83.09, n = 7, GFP(-), 1264.75 ± 97.98, n = 50; Day 28, GFP(+), 1217.75 ± 55.34, n = 13, GFP(-), 1470.64 ± 115.84, n = 50; <i>p</i>(Day 3) < 0.000, <i>p</i>(Day 7) = 0.000, <i>p</i>(Day 14) = 0.035, <i>p</i>(Day 28) = 0.041). Two-way ANOVA, F_G = 66.17, <i>p</i> = 0.000; F_T = 15.22, <i>p</i> = 0.000; F_G+T = 3.20, <i>p</i> = 0.031. (D) Normalized Ca_v1.3 antibody fluorescent intensity of newborn neurons to that of mature neurons. (Day 3, 49.52 ± 3.61%, n = 9; Day 7, 48.26 ± 3.08%, n = 9; Day 14, 73.42 ± 5.94%, n = 7; Day 28, 83.76 ± 3.58%, n = 13; <i>p</i>(Day 3–7) = 0.795, <i>p</i>(Day 7–14) = 0.001, <i>p</i>(Day 14–28) = 0.138). One-way ANOVA, F = 20.913, <i>p</i> = 0.000. (E) Comparison of Ca_v1.3 expression among DG, CA1 and CA3 regions of dorsal hippocampus shown at (A) (each, n = 10). (DG, 1851.50 ± 54.44, n = 10; CA1, 2072.08 ± 38.63, n = 10; CA3, 2298.10 ± 115.40, n = 10; <i>p</i>(DG-CA1) = 0.004, <i>p</i>(CA1-CA3) = 0.080, <i>p</i>(DG-CA3) = 0.003). One-way ANOVA, F = 8.42, <i>p</i> = 0.001. *, **, *** indicate <i>p</i> < 0.05, <i>p</i> < 0.01, <i>p</i> < 0.001, respectively.</p

Proliferation and survival of DG newborn cells of dorsal hippocampus in Ca_v1.3 KO mouse.

<p>(A) Confocal images of BrdU (+) cells (red) and NeuN (+) cells (green) in Ca_v1.3 KO and WT mouse. Images are acquired at 1, 14 and 28 days after BrdU injection. <i>Scale bar</i>, 50 μm. (B) Number of BrdU (+) cells. (Day 1, WT, 394.667 ± 30.78 cells, n = 8; KO, 387 ± 49.05, n = 6, <i>p</i> = 0.660; Day 14, WT, 376.6 ± 45.85 cells, n = 6; KO, 35.8 ± 13.22 cells, n = 6, <i>p</i> = 0.472; Day 28, WT, 219 ± 13.61 cells, n = 7; KO, 159.83 ± 23.70 cells, n = 6, <i>p</i> = 0.046). * indicates <i>p</i> < 0.05. Two-way ANOVA, F_G = 3.80, <i>p</i> = 0.061; F_T = 59.12, <i>p</i> = 0.000; F_G+T = 0.84, <i>p</i> = 0.444. (C) Number of BrdU (+) cells of KO mice normalized to that of WT mice at given day. (Day 1, WT, 100 ± 7.80%, n = 8, KO, 98.06 ± 12.43%, n = 10; Day 14, WT, 100 ± 12.17%, n = 6, KO, 93.95 ± 3.50%, n = 6; Day 28, WT, 100 ± 6.73%, n = 7, KO, 72.98 ± 11.85%, n = 6, <i>p</i> = 0.046). Two-way ANOVA, F_G = 4.61, <i>p</i> = 0.040; F_T = 1.82, <i>p</i> = 0.179; F_G+T = 1.90, <i>p</i> = 0.168. (D) Number of BrdU (+) cells per DG area at Day 28. (WT, 21.24 ± 1.22 cells/mm², n = 7; KO, 15.47 ± 2.12, n = 6, <i>p</i> = 0.032). (E) <i>Left</i>, example images for area measurements of DG (white dot line) and GCL (yellow line). <i>Right</i>, NeuN (+) cells (green) of DG in Ca_v1.3 KO and WT mice. <i>Scale bars</i>, 100 um (10x), 50 μm (40x), 10 μm (<i>insets</i>, 40x/5x-zoom). (F) DG area (WT, 9.92 ± 0.19 mm², n = 6, KO, 9.58 ± 0.18 mm², n = 6, <i>p</i> = 0.833), (G) GCL area (WT, 1.85 ± 0.063 mm², n = 6, KO, 1.91 ± 0.05 mm², n = 7, <i>p</i> = 0.445) and (H) Density of NeuN (+) cells in GCL (WT, 6071 ± 691.88 cells/mm², n = 11, KO, 6304.71 ± 339.34 cells/mm², n = 12, <i>p</i> = 0.897).</p