Plant- and stimulus-specific variations in remote-controlled sieve-tube occlusion

Phloem injury triggers local sieve-plate occlusion including callose-mediated constriction and protein plugging of sieve pores. In intact plants, reversible sieve-plate occlusion is induced by electric potential waves (EPWs)—accompanied by Ca²⁺-influx—as result of distant burning. Here, we present additional results which pertain to (a) the variability of EPW-profiles in relation to forisome conformation in intact Vicia faba plants and (b) the differential occlusion reactions to burning and cutting in various plant species. A correlation between stimulus perception and mode of phloem loading is discussed.Key words: callose, electrical potential waves, forisome, membrane potential, phloem transport, sieve-element occlusion, wound potentials     

Induction as well as suppression: How aphid saliva may exert opposite effects on plant defense

Aphids ingest from the sieve tubes and by doing so they are confronted with sieve-tube occlusion mechanisms, which are part of the plant defense system. Because aphids are able to feed over longer periods, they must be able to prevent occlusion of the sieve plates induced by stylet penetration. Occlusion probably depends upon Ca²⁺-influx into the sieve element (SE) lumen. Aphid behavior, biochemical tests and in vitro experiments demonstrated that aphid''s watery saliva, injected during initial phase of a stylet penetration into the SE lumen, contains proteins that are able to bind calcium and prevent calcium-induced SE occlusion. In this addendum, we speculate on the consequences of saliva secretion for plant resistance. (a) The release of elicitors (e.g., oligogalacturonides) due to cell wall digestion by gel saliva enzymes may increase the resistance of cortex, phloem parenchyma cells and companion cells (CC) around the puncture site. (b) Ca²⁺-binding by aphid watery saliva may suppress the local defense responses in the SEs. (c) Signaling cascades triggered in CCs may lead to systemic resistance.Key words: aphid saliva, calcium binding, elicitor, oligogalacturonides, local plant defense, systemic plant defense, phloem translocation, aphid/plant-interactionAfter having penetrated the sieve-element (SE) plasma membrane, aphids encounter unspecific wound-induced occlusion reactions to prevent sap leakage.^1–4 Occlusion mechanisms by callose, structural P-proteins and forisomes are likely induced by a sudden calcium influx into the sieve-tube lumen.⁵ Calcium possibly enters the sieve-tube lumen through the stylet wounding-site in the plasma membrane and/or stretch-activated calcium-channels.^6–8 After SE penetration, aphids secrete watery saliva that contains calcium-binding proteins presumed to sabotage sieve-plate occlusion.^9,10We demonstrated that Megoura viciae (Buckton) is most likely able to prevent or reduce sieve-tube occlusion in Vicia faba by secretion of watery saliva. By in vitro confrontation of isolated forisomes, protein bodies responsible for sieve-tube occlusion in Fabaceaen,⁵ and watery saliva concentrate, we were able to show that salivary proteins convey forisomes from a dispersed (+Ca²⁺) into a condensed (−Ca²⁺) state.¹⁰ The dispersed forisome functions in vivo as a plug, leading to stoppage of mass flow.⁵This in vitro evidence was corroborated by aphid behavior in response to leaf tip burning, which triggers an electrical potential wave (EPW) along the sieve tubes. Such an EPW induces Ca²⁺-influx and corresponding SE occlusion along the pathway.¹¹ The passage of the EPW is associated with a prolonged secretion of watery saliva of aphids. This is interpreted as an attempt to unplug the SEs by calcium binding.¹⁰ Similar behavioral changes in response to leaf-tip burning were observed in an extended set of aphid/plant species combinations, indicating that attempted sabotage of sieve-tube occlusion by aphid saliva is a widespread phenomenon (unpublished).Aphid feeding was reported to induce local (on the same leaf) and systemic (in distant leaves) reactions of the host plant. The local response led to enhanced feeding,^12–14 while the systemic response showed reduced ingestion and extended periods of watery saliva secretion in sieve tubes distant from previous feeding sites.^12–14 These contrasting observations were described to be independent of the aphid species.¹³ The question arises how aphids induce these seemingly opposite plant responses?The aphid stylet pushing forward through cortical and vascular tissue is surrounded by a sheath of gel saliva, secreted into the apoplast.^15,16 Gel saliva contains cellulase and pectinase that amongst others produce oligogalacturonides (OGs) along the stylet sheath by digestion of cell wall material.^17,18 Usually, OGs act as elicitors, triggering a variety of plant responses against pathogens and insects in which the activation of calcium channels is involved.^19,20 This seems to conflict with a suppression of resistance as observed for the impact of watery saliva in SEs.¹⁰ We will make an attempt to explain this paradoxon.OG induced defense responses may be triggered in all cell types adjacent to the salivary sheath (Fig. 1). Because watery saliva is only secreted briefly into these cells, which are punctured for orientation purposes (Hewer et al., unpublished), it seems unlikely that OG induced defense is suppressed here by saliva-mediated calcium binding.¹⁵ The diffusion range of OGs may be restricted to the close vicinity of the stylet sheath leading to an enhanced regional defense with a limited sphere of action (Fig. 1). Because the settling distance of aphids is restricted by their body size (1–10 mm),²¹ aphids feeding on the same leaf are probably hardly confronted with the regional defense induced by another aphid (Fig. 1). Otherwise, they would show an increased number of test probes before first phloem activity, as described for volatile mediated plant defense in cortex cells.¹³ Circumstantial support in favor of our hypothesis is provided by production of hydrogen peroxide in the apoplast,²² which is most likely associated with the action of OGs.²² Observations of hydrogen peroxide production during aphid (Macrosiphum euphorbiae) infestation of tomato in a limited area along the leaf veins, the preferred feeding sites of this species, indicate a locally restricted defense response (Fig. 1 and ​and22).⁴ The question arises why the cell signals are not spread via plasmodesmata to adjacent cells to induce resistance in a more extended leaf area? Dissemination of the signals may be prevented by closure of plasmodesmata (Fig. 1) through callose deposition,^23,24 which is most likely directly coupled with calcium influx induced by OGs,²⁵ by apoplastic hydrogen peroxide and to a minor extent by stylet puncture (Fig. 2).^7,26Open in a separate windowFigure 1Hypothetical model on how stylet penetration induces and suppresses plant defense. Sheath saliva (light blue) that envelopes the stylet during propagation through the apoplast contains cellulase and pectinase,^17,18 enzymes producing elicitors (e.g., oligogalacturonides (oGs)) by local cell wall digestion.¹⁹ Parenchyma cells adjacent to the sheath may develop a defense response owing to signaling cascades triggered by oG-mediated Ca²⁺-influx.¹⁹ Together with a Ca²⁺-dependent transient closure of plasmodesmata by callose (black crosses),^23,24 the focused production of oGs may cause a defense response with a limited sphere of action (red—strong, brown—light, green—none). This restricted domain of defense may not be perceived by other aphids, since the settling distance is limited by the aphid body size. Nearby aphids do not show any sign of defense perception in their probing and feeding behavior.¹⁴ Signaling cascade compounds may be channeled from parenchyma cells to CCs (dashed yellow arrows), where they are subsequently released into the SEs. There they may act as long-distance systemic defense components (grey arrows). In contrast to the parenchyma domain (where only minor amounts of watery saliva are secreted), Ca²⁺-mediated reactions such as defense cascades and sieve-plate (SP) occlusion are suppressed in SEs by large amounts of watery saliva. The left aphid penetrates an SE and injects watery saliva (red cloud; ws) that inhibits local sieve-plate occlusion and,¹⁰ most likely, is transported by mass flow (black arrow) to adjacent SEs,²⁷ where occlusion is impeded as well. A short-distance systemic spread over a few centimeters may explain local suppression of plant defense resulting in a higher rate of colonization. Salivary proteins or their degradation products may serve as systemic defense signals as well (grey arrows), but may also diffuse via the PPUs into CCs where additional systemic signals are induced (yellow arrows).Open in a separate windowFigure 2Hypothetical involvement of Ca²⁺-channels in aphid-induced cell defense (detail). During probing with its stylet the aphid secretes gel saliva as a lubrication substance (light blue) into the apoplast.¹⁵ on the way to the sieve tubes, aphids briefly puncture most non-phloem cells (red) after which the puncturing sites are sealed with gel saliva.^7,16 Gel saliva also most likely prevents the influx of apoplastic calcium into pierced sieve elements (green) by sealing the penetration site.⁷ Watery saliva (red cloud), injected into the SE lumen,⁹ contains proteins which bind calcium ions (marked by X) that enter the SE via e.g., mechano sensitive Ca²⁺-channels activated by stylet penetration (blue tons).¹⁰ In this way, aphids suppress SE occlusion and activation of local defense cascades. In the parenchyma cells around the gel saliva sheath, a small cylindrical zone of defense may be induced by oligogalacturonides (oGs; brown triangles) produced by cell wall (grey) digestion.^17–19 Perceived by unknown receptor proteins (R; e.g., a receptor like protein kinase)³⁴ and kinase mediation (black dotted and dashed arrows), oGs lead to a Ca²⁺-influx through kinase activated calcium channels (orange tons).²⁵ Around the probing site, aphids apparently induce the production of superoxide by Ca²⁺-induced activation of the NADPH oxidase (violet box) and its following conversion to hydrogen peroxide (red spots) is mediated by superoxide dismutase (SoD).⁴ Hydrogen peroxide activates Ca²⁺-channels (violet tons) and diffuses through plasma membrane (curled arrows) therefore potentially acting as a intracellular signal.²⁶By contrast, Ca²⁺-influx into SEs, induced by presence of OGs or stylet insertion (Fig. 2), is not expected to trigger local defense given the abundant excretion of Ca²⁺-binding watery saliva.^7,10,25 Watery saliva may spread to down-stream and adjacent SEs through transverse and lateral sieve plates (Fig. 1).^7,27 Aphids puncturing nearby SEs may therefore encounter less severe sieve-plate occlusion which results in facilitated settling and thus in increased population growth. Aggregation of feeding aphids would self-amplify population growth until a certain density is attained. Farther from the colonization site, this effect may be lost due to dilution. Stimulation of aphid feeding by aphid infestation was observed locally on potato by Myzus persicae and M. euphorbiae, respectively, 96 h after infestation.¹³ However, a similar effect was not observed for M. persicae on Arabidopsis thaliana where aphids induced premature leaf senescence and resistance 12 h after infestation,²⁸ possibly induced by OGs.¹⁹As a speculation, OG induced Ca²⁺-influx into parenchyma cells adjacent to the salivary sheath activate Ca²⁺-induced signaling cascades via CaM,^26,29 CDPKs,^30,31 MAPKinases and reactive oxygen species (Fig. 2).³² Systemic resistance, induced by aphid infestation,^12–14 is mediated by unknown compounds such as, e.g., salivary proteins, their degradation products, signal cascade products or volatiles.¹³ Compounds produced in CCs first have to pass the PPUs, while SE signaling elements can be directly transported via mass flow (Fig. 1).The question arises if aphids profit from induced resistance on local (cortex and parenchyma cells) and systemic (distant plant organs) levels as holds for suppression of defense in SEs. Possibly settling and subsequent spread of competing pathogens/herbivores (e.g., fungi or other piercing-sucking insects) are suppressed by induced defense. In this context it is intriguing to understand how aphids cope with the self-induced systemic resistance, which probably lasts over weeks.³³     

Concentrations and ecological risks of metals in surface sediments of some coastal creeks in the Niger Delta,Nigeria

The concentrations of nine metals were measured by atomic absorption spectrophotometry in surface sediments of three coastal creeks, namely, the Ifie, Egbokodo and Ubeji creeks, in the Niger Delta of Nigeria, from August 2012 to January 2013. The aim of the study was to provide information on the spatial and seasonal distribution patterns, degree of contamination, and ecological risks of metals in these sediments. The mean concentrations of the nine metals in these creek sediments ranged from 0.30 to 3.20?mg kg^?1 Cd; 10.7 to 24.7?mg kg^?1 Pb, 125 to 466?mg kg^?1 Cr; 3.1.10 to 14.9?mg kg^?1 Cu; 4.7 to 14.3?mg kg^?1 Co; 61.1 to 115?mg kg^?1 Ni; 106 to 183?mg kg^?1 Mn; 52.0 to 170?mg kg^?1 Zn and 5 469 to 20 639?mg kg^?1 Fe. In general, the metal concentrations were higher in the dry season than the wet season, except for Cr. The concentrations of Cd, Cr, Ni and Zn were above their regulatory control limits in sediment as specified by the Nigerian Regulatory Authority and Cd was identified as the main ecological risk factor. The enrichment factors for the studied metals followed the order: Cd > Cr > Ni > Zn > Pb > Co > Mn > Cu. The average multiple pollution index values indicated that these sediments were severely polluted with significant inputs from Cd, Ni and Cr.     

DNA-DNA hybridization evidence of the rapid rate of muroid rodent DNA evolution

Single-copy nuclear DNAs (scnDNAs) of eight species of arvicoline and six species of murine rodents were compared using DNA-DNA hybridization. The branching pattern derived from the DNA comparisons is congruent with the fossil evidence and supported by comparative biochemical, chromosomal, and morphological studies. The recently improved fossil record for these lineages provides seven approximate divergence dates, which were used to calibrate the DNA-hybridization data. The average rate of scnDNA divergence was estimated as 2.5%/Myr. This is approximately 10 times the rate in the hominoid primates. These results agree with previous reports of accelerated DNA evolution in muroid rodents and extend the DNA-DNA hybridization data set of Brownell.      

Demographic Data of a Population of Insured Swedish Dogs Measured in a Questionnaire Study

Dogs, in the age range 1–3 years old, were randomly selected from the largest animal insurance database in Sweden for inclusion in the study. The study was performed in 1997, and a total of 680 dog owners were selected for the study. A total of 461 dog owners completed the survey, at an overall response rate of 68%. Data was compared to a recent gallup performed on a sample of all dogs in Sweden. The demographic statistics of the insured dog population were in many aspects similar to the total dog population of Sweden. Typical for both insured dogs and the total population of dogs were a low proportion of neutered dogs, that many dogs were bought at an early age, that many dogs were in contact with a "breeder" when sold, and a similar profile of health status. However, "dog breeders" seemed to have their dogs insured to a higher extent than the general dog owner. It was concluded that as the populations were alike in many respects, it is reasonable to use the insurance database for epidemiological studies on diet and exercise in Swedish dogs.     

Rise of oceanographic barriers in continuous populations of a cetacean: the genetic structure of harbour porpoises in Old World waters

Background

Understanding the role of seascape in shaping genetic and demographic population structure is highly challenging for marine pelagic species such as cetaceans for which there is generally little evidence of what could effectively restrict their dispersal. In the present work, we applied a combination of recent individual-based landscape genetic approaches to investigate the population genetic structure of a highly mobile extensive range cetacean, the harbour porpoise in the eastern North Atlantic, with regards to oceanographic characteristics that could constrain its dispersal.

Results

Analyses of 10 microsatellite loci for 752 individuals revealed that most of the sampled range in the eastern North Atlantic behaves as a 'continuous' population that widely extends over thousands of kilometres with significant isolation by distance (IBD). However, strong barriers to gene flow were detected in the south-eastern part of the range. These barriers coincided with profound changes in environmental characteristics and isolated, on a relatively small scale, porpoises from Iberian waters and on a larger scale porpoises from the Black Sea.

Conclusion

The presence of these barriers to gene flow that coincide with profound changes in oceanographic features, together with the spatial variation in IBD strength, provide for the first time strong evidence that physical processes have a major impact on the demographic and genetic structure of a cetacean. This genetic pattern further suggests habitat-related fragmentation of the porpoise range that is likely to intensify with predicted surface ocean warming.     

Contractile behavior of the forelimb digital flexors during steady-state locomotion in horses (Equus caballus): an initial test of muscle architectural hypotheses about in vivo function

The forelimb digital flexors of the horse display remarkable diversity in muscle architecture despite each muscle-tendon unit having a similar mechanical advantage across the fetlock joint. We focus on two distinct muscles of the digital flexor system: short compartment deep digital flexor (DDF(sc)) and the superficial digital flexor (SDF). The objectives were to investigate force-length behavior and work performance of these two muscles in vivo during locomotion, and to determine how muscle architecture contributes to in vivo function in this system. We directly recorded muscle force (via tendon strain gauges) and muscle fascicle length (via sonomicrometry crystals) as horses walked (1.7 m s(-1)), trotted (4.1 m s(-1)) and cantered (7.0 m s(-1)) on a motorized treadmill. Over the range of gaits and speeds, DDF(sc) fascicles shortened while producing relatively low force, generating modest positive net work. In contrast, SDF fascicles initially shortened, then lengthened while producing high force, resulting in substantial negative net work. These findings suggest the long fibered, unipennate DDF(sc) supplements mechanical work during running, whereas the short fibered, multipennate SDF is specialized for economical high force and enhanced elastic energy storage. Apparent in vivo functions match well with the distinct architectural features of each muscle.     

Latitudinal clines in body size,but not in thermal tolerance or heat‐shock cognate 70 (HSC70), in the highly‐dispersing intertidal gastropod Littorina keenae (Gastropoda: Littorinidae)

Natural populations of widely‐distributed animals often exhibit clinal variation in phenotypic traits or in allele frequencies of a particular gene over their geographical range. A planktotrophic intertidal snail, Littorina keenae is broadly distributed along the north‐eastern Pacific coast through a large latitudinal range (24°50′N–43°18′N). We tested for latitudinal clines in two complex phenotypic traits – thermal tolerance and body size – and one single locus trait – heat shock cognate 70 (HSC70) – in L. keenae along almost its entire geographical range. We found only weak evidence for a latitudinal cline in the thermal tolerance and no evidence for a cline in allele frequencies at HSC70. However, as predicted by Bergmann's rule, we detected a strong latitudinal cline that accounted for 60% of the variance in body size (R² = 0.598; P < 0.001). In contrast, body size did not significantly affect thermal tolerance. HSC70 showed no genetic differentiation among the populations, supporting our previous mitochondrial gene‐based estimate of high gene flow during this snail's free‐swimming larval stage. Given that L. keenae experiences panmixia along its species range, the observed size cline may be partially or entirely caused by a phenotypically plastic response to local thermal environments rather than by genetic divergence in body size among populations in response to locally optimizing natural selection. © 2010 The Linnean Society of London, Biological Journal of the Linnean Society, 2010, 100 , 494–505.