text:
Impact Factor 4.402 | CiteScore 7.8
More on impact ›
Frontiers in Plant Science
Toggle navigation Section
(current)Section
Why submit?
Contact editorial office
Suggest a Research Topic >
XML (NLM)
View Article Impact
Open Supplemental Data
Original Research ARTICLE
Front. Plant Sci., 13 February 2015 | https://doi.org/10.3389/fpls.2015.00065
Duration and intensity of shade differentially affects mycorrhizal growth- and phosphorus uptake responses of Medicago truncatula
Tereza Konvalinková1*, David Püschel1,2, Martina Janoušková1,2, Milan Gryndler1 and Jan Jansa1
1Laboratory of Fungal Biology, Institute of Microbiology, Academy of Sciences of the Czech Republic, Prague, Czech Republic
2Department of Mycorrhizal Symbioses, Institute of Botany, Academy of Sciences of the Czech Republic, Průhonice, Czech Republic
Plant and fungal partners in arbuscular mycorrhizal symbiosis trade mineral nutrients for carbon, with the outcome of this relationship for plant growth and nutrition being highly context-dependent and changing with the availability of resources as well as with the specific requirements of the different partners. Here we studied how the model legume Medicago truncatula, inoculated or not with a mycorrhizal fungus Rhizophagus irregularis, responded to a gradient of light intensities applied over different periods of time, in terms of growth, phosphorus nutrition and the levels of root colonization by the mycorrhizal fungus. Short-term (6 d) shading, depending on its intensity, resulted in a rapid decline of phosphorus uptake to the shoots of mycorrhizal plants and simultaneous accumulation of phosphorus in the roots (most likely in the fungal tissues), as compared to the non-mycorrhizal controls. There was, however, no significant change in the levels of mycorrhizal colonization of roots due to short-term shading. Long-term (38 d) shading, depending on its intensity, provoked a multitude of plant compensatory mechanisms, which were further boosted by the mycorrhizal symbiosis. Mycorrhizal growth- and phosphorus uptake benefits, however, vanished at 10% of the full light intensity applied over a long-term. Levels of root colonization by the mycorrhizal fungus were significantly reduced by long-term shading. Our results indicate that even short periods of shade could have important consequences for the functioning of mycorrhizal symbiosis in terms of phosphorus transfer between the fungus and the plants, without any apparent changes in root colonization parameters or mycorrhizal growth response, and call for more focused research on temporal dynamics of mycorrhizal functioning under changing environmental conditions.
The arbuscular mycorrhizal (AM) symbiosis is one of the most abundant symbioses on Earth, and it is important for the movement of nutrients through global ecosystems. Host plants exchange carbon (C) for mineral nutrients such as phosphorus (P), nitrogen (N) and zinc provided by AM fungi (Lekberg et al., 2010; Fitter et al., 2011).
Research in the last few decades has made a great progress in understanding P acquisition pathways of plants via AM fungi (Burleigh and Harrison, 1999; Rausch et al., 2001; Javot et al., 2007). Ecophysiological experiments using P-radioisotope labeling of soil patches accessible only to the AM hyphae have demonstrated the importance of the mycorrhizal P uptake pathway for plant nutrition. This pathway had previously been massively underestimated, but it can account for nearly all P uptake by a host plant under P-limiting soil conditions (Pearson and Jakobsen, 1993a; Smith et al., 2004).
Compared to P uptake, the C side of the symbiosis is much less understood. Initial estimates suggested that approximately 4% of the plant photosynthetic production is allocated to fungal symbionts (Paul and Kucey, 1981). Further studies showed that up to 20–30% of the C budget is being consumed by the fungal partners under some circumstances (Jakobsen and Rosendahl, 1990; Drigo et al., 2010). The average values for the different combinations of partners and different environmental conditions now stand somewhat below 10% of plant photosynthesis production (Grimoldi et al., 2006; Lendenmann et al., 2011). There is a great variability in these figures, however, depending on the identity of the fungal symbionts, with some fungal partners demanding more C than others (Pearson and Jakobsen, 1993b; Lendenmann et al., 2011). It has also been demonstrated that C allocation to the fungal partner is coupled with exchange of P and/or N (Hammer et al., 2011; Kiers et al., 2011; Fellbaum et al., 2012, 2014), meaning that nutrient availability and plant requirements can change C allocation patterns.
One approach to studying C allocation dynamics is to impose experimental shading on the host plant in order to manipulate the C source strength (Fellbaum et al., 2014; Knegt et al., 2015) and to determine how this changes the C allocation to the fungal partner. Previous research on the effects of light intensity on the AM symbiosis demonstrated a reduction of symbiotic benefits under reduced light intensities and a reduced fungal colonization rate in continuously or temporarily shaded plants (Son and Smith, 1988; Reinhard et al., 1994; Tuomi et al., 2001; Heinemeyer et al., 2003; Olsson et al., 2010). Other studies, however, found no reduction in symbiotic benefits and mycorrhizal colonization (Gehring, 2003; Korhonen et al., 2004; Millar and Ballhorn, 2013).
There is little doubt that the incoming light is the only source and light intensity thus being the primary determinant of metabolic energy availability for both the plant and the associated AM fungi. However, our understanding of the quantitative changes in symbiotic functioning as depends on the light intensity is still very limited. Further, we do not know what effects will the duration of exposure to different light intensities have on the functioning of the symbiosis on the gradient of plant responses ranging from positive to negative (Johnson et al., 1997, 2015; Johnson and Graham, 2013).
Here we study mycorrhizal functioning of the model plant Medicago truncatula Gaertn. across a gradient of light levels. We used a model mycorrhizal fungus, Rhizophagus irregularis (Blaszk., Wubet, Renker and Buscot) C. Walker and A. Schüßler as the fungal partner because it has previously been shown to provide major P uptake benefits to the plant, but also to consume significant amounts of plant C (Lendenmann et al., 2011). Our aim was to assess the effects of duration and intensity of shading on the symbiotic benefits and costs of our model plants.
We used barrel medic (M. truncatula) that was inoculated or not with R. irregularis. The experiment encompassed both long- (38 d) and short-term (6 d) shading with three different intensities of shade (65, 35, or 10% of the incoming light intensity), as well as a full-light control treatment. Each treatment was replicated five times, resulting in a total of 70 pots.
Biological Materials
The seeds of M. truncatula J5 were surface-sterilized in concentrated (98%) sulfuric acid for 10 min and then rapidly washed with several liters of tap water to achieve synchronous germination. The seeds were germinated on a moist filter paper at 25°C for one day.
Inoculum of Sinorhizobium meliloti strain 1021 was grown in TY liquid medium (Somasegaran and Hoben, 1994) on a shaker at 24°C for 3 days. The bacteria were washed with 0.5% (w:v) MgSO4 solution and the suspension was adjusted to the optical density of 0.7 at 600 nm (which corresponded to approximately 2 × 109 cells mL-1).
Inoculum of the R. irregularis, The International Bank for the Glomeromycota (BEG) accession number 158, previously isolated from an arable field in Tänikon, Switzerland (Jansa, 2002), was produced in pot cultures with leek (Allium porrum L.) cv. Du Bouchet as a host plant in climate chambers (14 h photoperiod, 350 μmol photons m-2 s-1 provided by fluorescent tubes and sodium discharge lamps, 25/20°C day/night). The cultures were grown for 4 months prior to the experiment, resulting in 89% of the leek root length colonized by the AM hyphae, 59% of root length colonized by vesicles and hyphal length density in the substrate being 3.41 m g-1. The "mock-inoculum" for the non-mycorrhizal treatments was produced in pots with a non-mycorrhizal leek grown for four months under the same conditions as the mycorrhizal inocula. Shortly before the start of the experiment, the above-ground biomass of leek in the pots was removed, the roots were cut to pieces of about 1 cm in length and mixed back to the substrate, which was subsequently dried at room temperature and then used as the mycorrhizal or the control inoculum.
Cultivation of the Experimental Plants
Plants were grown in 2-L pots lined with plastic mesh (opening of 1 mm) at the bottom, sterilized with 96% ethanol and filled with substrate. The substrate consisted of autoclaved quartz sand (grain size < 3 mm), autoclaved zeolite MPZ 1-2.5 (Zeopol, http://www.zeolity.cz, grain size 1–2.5 mm) and γ-irradiated soil from Litoměřice, Czech Republic (pHH2O 7.88, 42% clay, 40% sand, total P 797 mg kg-1, water extractable P 3.3 mg kg-1, 2.26% C, 0.13% total N) mixed in a ratio 9:9:2 (v:v:v). The substrate was added with 2.5% (v:v) of the respective mycorrhizal or control inocula mixed thoroughly in the whole volume.
Rhizobial suspension (4 mL pot-1) and five germinated seeds were put in each pot about 1 cm below the surface. Surface of the substrate was kept moist with distilled water, and after 10 days, seedlings were thinned to two per pot. Plants were then watered daily with distilled water to approximately 85% water holding capacity of the substrate. Plants were grown during the summer period in the glasshouse of the Institute of Microbiology, Prague, with supplemental lighting (halogen lamps providing a photosynthetic flux density of 200 μmol m-2 s-1), extending the day length to 16 h. The plants were fertilized with a modified White mineral solution P2N3 (Gryndler et al., 1992) with reduced amount of P (5% of the original recipe), starting at two weeks after planting, and providing each pot with 50 mL of two-times concentrated solution pot-1 week-1.
Plants were exposed to shading treatments as described above. Full light corresponded to about 50% of the solar radiation outside the glasshouse. Plants were shaded by tents made of green shading fabrics used in horticulture. Pots were fully randomized in the glasshouse and they were rotated every week before application of the shade treatments, and then they were rotated within the tents.
13CO2 PULSE LABELING
Plants exposed to full light and 10% light intensity applied over long-term were pulse-labeled with 13CO2 3 days before the harvest. The pots were placed in an air-tight Plexiglas chamber (0.75 m3) equipped with a fan to mix the inner atmosphere. Temperature, humidity, and CO2 concentration inside the chamber were thoroughly monitored (using Testo 435-2 datalogger equipped with the IAQ probe, Testo AG, Lenzkirch, Germany). The air in chamber was enriched with 13CO2 by adding 40% phosphoric acid onto 1 g of 99% 13C-enriched calcium carbonate powder (Sigma–Aldrich, Buchs, Switzerland) placed in the labeling chamber to achieve a calculated1 CO2 concentration of 574 μmol mol-1, while the 13C enrichment inside the chamber reached 57 atom% 13C. The labeling took 1.5 h on a sunny day, with the light intensity at plant level reaching 891 μmol photons m-2 s-1.
Harvest and Sampling
Plants were harvested 7.5 weeks after planting. For all measurements and analyses, except the shoot architecture analysis, both plants from one pot were processed as one unit. The shoots were cut at the hypocotyl-root boundary, laid out between a PVC foil and a Plexiglas plate and photographed together with a ruler for shoot architectural analyses. Subsequently, the shoots were dried for 4 days at 65°C to determine the shoot dry weight. The roots were washed from the substrate with tap water, cut into 1-cm fragments and stirred in water. Then the roots were divided in two aliquots and the fresh weights of both were recorded. One aliquot was immersed in 10% KOH (w:v) to determine the AM fungal colonization, while the second aliquot was dried for 4 days at 65°C, weighed and the root dry weight of whole root system was calculated. The samples of substrate were stored at -80°C before being dried at 65°C for 4 days. Dried shoot, root, and substrate samples were milled to a fine powder using a ball mill (MM200, Retsch, Haan, Germany) before the elemental and isotopic analyses.
Plant, Fungal, and Substrate Analyses
To determine the P concentration in plant tissues, milled samples of shoots and roots (100 mg each) were incinerated in a muffle furnace (550°C) for 12 h, added with 1 mL of concentrated (69%) HNO3 and briefly heated to 250°C on a hot plate. Then they were transferred to volumetric flasks through a filter paper and made up to 50 mL with ultrapure water. Phosphorus concentration in the extracts was then measured using the malachite green method (Ohno and Zibilske, 1991).
The N and C concentrations and isotopic composition of N and C in shoots, roots, and substrate were measured in all pots subjected to 13C labeling, using an elemental analyzer (Flash EA 2000) coupled with an isotope ratio mass spectrometer (Delta V Advantage, ThermoFisher Scientific, Waltham, MA, USA). The isotopic compositions were expressed in notation (‰) relative to atmospheric air standard in case of 15N and to Vienna Pee Dee Belemnite (VPDB) standard in case of 13C (Eq. 1; Rs and Rst being the isotope ratios in the sample and the standard, respectively).
δ(‰)=(RsRst−1)×1000 (1)
Additionally, to calculate the amount of 13C in the samples which originated from the 13CO2 pulse (i.e., excess 13C), the δ values were converted to F-ratios (Eq. 2; where the stands for δ13C(‰) of the sample divided by 1000; 0.0111802 is the 13C/12C isotope ratio of the VPDB standard). The amounts of carbon in the roots, shoots or substrate (C, in moles) were calculated as in Eq. 3 (DW is the dry weight of the respective sample; B is the carbon concentration in the sample, g g-1; F is the 13C F-ratio in the sample). Finally, the amounts of 13C in the roots, the shoots or the substrate added by labeling (excess13C, in moles) were calculated (Eq. 4; Fs is 13C F-ratio in the sample; Fu is the average 13C F-ratio in the corresponding samples of five unlabeled plants).
F=(δ+1)×0.0111802(δ+1)×0.0111802+1 (2)
C=DW×B13×F+12×(1−F) (3)
excess13C=(Fs−Fu)×C (4)
Root samples for staining were first macerated in 10% KOH (30 min at 90°C), washed with tap water, neutralized in 1% HCl (1 h at room temperature followed by 15 min at 90°C) and stained with a mixture of Trypan and Methylene Blue (each 0.05% in lactic acid–glycerol–water, 1:1:1 v:v:v). The AM fungal colonization was assessed microscopically using the method of McGonigle et al. (1990).
Shoot architecture was assessed on each individual plant with image analyzing software (NIS-Elements AR, Nikon Instruments, Melville, NY, USA). The length of the shoot main axis and the number of branches were averaged for each pot using the values measured on the two individual plants. The mean surface of leaflets was calculated as an average of 18 measured leaflets per pot.
The data were analyzed with Statgraphics Plus for Windows version 3.1. The shoot and root weights, P and N contents, P concentrations, and the leaflet surface values were common log-transformed, whereas the root to shoot biomass ratios and excess13C values were square root-transformed. Fractional values of the mycorrhizal colonization of roots were arcsin (square root)-transformed before the analyses. Mycorrhizal responses were calculated as in Thonar et al. (2011). Data from long- and short-term shading treatments were analyzed separately.
Simple regression analyses using a linear model were performed to test the influence of light intensity on Rhizophagus-colonization of roots and on the mycorrhizal responses. General linear models (GLM) were employed to test the significances of the effects of inoculation treatment and light intensity on the plant dry weights, P contents and concentrations and plant architectural parameters, while two-way ANOVAs with inoculation treatment and light intensity as factors were carried out for the carbon and nitrogen data, because they were only collected for the full light and for the most intensive long-term shading treatments. The significance of individual factors and their interactions were judged by F-ratios. The effect was considered significant if p < 0.05.
Additionally, t-tests were performed to test the effect of mycorrhizal inoculation separately for the full light and for the 10% light intensity applied over long-term. Mann–Whitney test replaced the t-test comparisons if significant heteroscedasticity in the data was detected.
At full light, both shoot and root dry weights (Figure 1) were higher for the mycorrhizal than non-mycorrhizal plants (t8 = 3.43, p = 0.009 and t8 = 2.32, p = 0.048, respectively).
FIGURE 1. Shoot and root dry biomass of Medicago truncatula plants inoculated or not with Rhizophagus irregularis (black and white symbols, respectively), and subjected to different light regimes. Mean values ± SD (n = 5) are shown.
Under short-time shading, both shoot and root weights decreased significantly with the decrease of light intensity (F1,36 = 44.2, p < 0.001, and F1,36 = 39.2, p < 0.001, respectively). However, the differences between the inoculation treatments did not change significantly with light intensity as revealed by a non-significant interaction between the inoculation and the light factors (Supplementary Table S1).
Under long-term shading, the response of shoot weight to light intensity was not trivial – there was only a small difference between full light and 65% light intensity (Figure 1), while plant biomass production was strongly reduced under further shading levels. Therefore, there was a strong effect of light treatment on the shoot weight (F1,36 = 186, p < 0.001), but the effect of inoculation treatment and the interaction of the main factors were both not significant. In contrast to the full light treatment, non-mycorrhizal plants had higher shoot biomass than mycorrhizal plants at 10% light intensity applied over long time (t8 = 2.46, p = 0.039, Figure 1; Table 1). Root biomass also decreased significantly with the decrease of light intensity in the long-term shading (F1,36 = 296, p < 0.001). The differences in root biomass between the inoculation treatments were reversed at low light levels as compared to the full light, as indicated by a significant interaction between the inoculation and light factors (F1,36 = 5.58, p = 0.024). Indeed, at 10% light intensity applied over a long term, higher root weight was recorded for the non-mycorrhizal plants as compared to the mycorrhizal treatment (W = 0.00, p = 0.012).
TABLE 1. Medicago truncatula responses to mycorrhizal inoculation under the different light regimes.
Phosphorus Uptake
At full light, P contents in shoots and roots (Figure 2) were significantly higher in mycorrhizal than the non-mycorrhizal plants (t8 = 10.5, p < 0.001 and W = 20.0, p = 0.020, respectively). Phosphorus concentrations in both shoots and roots (Figure 3) were also higher in mycorrhizal as compared to the non-mycorrhizal plants (t8 = 15.4, p < 0.001 and t7 = 10.9, p < 0.001, respectively).
FIGURE 2. The amount of phosphorus (P) contained in the shoots and roots of M. truncatula plants inoculated or not with R. irregularis (black and white symbols, respectively), and subjected to different light regimes. Mean values ± SD (n = 5) are shown.
FIGURE 3. Phosphorus (P) concentrations in shoots and roots of the M. truncatula plants inoculated or not with R. irregularis (black and white symbols, respectively), and subjected to different light regimes. Mean values ± SD (n = 5) are shown.
Under short-term shading, the responses of shoot P content to light intensity strongly depended on the inoculation treatment, as revealed by a significant interaction between the inoculation and light factors (F1,36 = 15.8, p < 0.001). Whereas the shoot P content of mycorrhizal plants decreased with decreasing light intensity, the P content of shoots of the non-mycorrhizal plants remained unaffected by the shading (Figure 2), resulting in a significant decrease of the mycorrhizal shoot P content response with decreasing light intensity (F1,18=35.7, p<0.001, Table 1). The response of the root P content to the short-time shading was independent of the inoculation treatment, as revealed by a non-significant interaction between the inoculation and light factors and by a non-significant correlation of the mycorrhizal root P content response with light intensity (Supplementary Table S1; Table 1). Due to slowing biomass production of but continuing P uptake to the short-term shaded non-mycorrhizal plants, the P concentration in the shoots of non-mycorrhizal plants increased with increasing intensity of short-term shading, whereas the P concentration in the mycorrhizal plants stagnated or tended to decrease (Figure 3). These effects were significant as revealed by a significant interaction of the light and inoculation factors (F1,36 = 30.9, p < 0.001) and by a significant correlation of the mycorrhizal P concentration response with light intensity. Such a difference in response to shading between the inoculation treatments was not replicated in the root P concentrations (Figure 3), as revealed by a non-significant interaction between the light and inoculation treatments.
Under long-term shading, both shoot and root P contents were significantly reduced (F1,36 = 234, p < 0.001 and F1,35 = 251, p < 0.001, respectively). The decrease was most pronounced between 65 and 35% of the full light intensity and it was more rapid in mycorrhizal than in the non-mycorrhizal plants, as revealed by a significant interaction between the inoculation and light factors (F1,36 = 8.76, p = 0.005 and F1,35 = 9.88, p = 0.003, respectively) and by significant correlations between mycorrhizal P content responses in shoots and roots and the light intensity (F1,18 = 37.1, p < 0.001 and F1,18 = 16.5, p < 0.001, respectively). No significant differences were found between the inoculation treatments in the P content of either shoots or roots at 10% light intensity applied over a long-term. The P concentrations in both shoots and roots decreased significantly more along the shading gradient in the mycorrhizal as compared to the non-mycorrhizal plants, as documented by a significant interaction between the light and inoculation factors for both parameters (F1,36 = 41.1, p < 0.001, and F1,35 = 19.1, p < 0.001, respectively) and by a significant correlation of the mycorrhizal P concentration response for both shoots and roots with light intensity (F1,18 = 117, p < 0.001 and F1,18 = 19.0, p < 0.001, respectively).
Nitrogen Uptake
Shoot N content was not affected by the inoculation (Supplementary Table S1), but only by long-term shading (F1,16 = 773, p < 0.001). Nevertheless, δ15N values (Supplementary Figure S1), indicative of the origin of N in shoot biomass, differed between the inoculation treatments at full light (W = 0.00, p = 0.012). Mycorrhizal plants had lower δ15N value, indicating a higher proportion of symbiotically fixed nitrogen than in the non-mycorrhizal plants. This pattern was reversed in the plants exposed to 10% light intensity over a long term, as revealed by a significant interaction between the inoculation and light factors for the δ15N in the shoots (F1,16 = 51.2, p < 0.001). At 10% light intensity, mycorrhizal plants showed higher δ15N value than the non-mycorrhizal controls (t8 = 14.8, p < 0.001), indicating a lower proportion of symbiotically fixed N than in the non-mycorrhizal plants.
Plant Architecture
The root-to-shoot biomass ratio (Figure 4) did not differ between the inoculation treatments at full light and decreased with decreasing light intensity under both short- and long-term shading regimes (F1,36 = 16.7, p < 0.001 and F1,36 = 183, p < 0.001, respectively). The response to long-term shading was strongly affected by the inoculation treatment, as revealed by a significant interaction between the inoculation and light factors (F1,36 = 8.43, p = 0.006). There was a sharp drop of root-to-shoot biomass ratio between 65 and 35% light intensities in mycorrhizal plants, while the decrease was more gradual in the non-mycorrhizal plants, which consequently had a considerably higher root-to-shoot ratio than the former at 35% light intensity applied over a long-term (t8 = 5.98, p < 0.001). However, no significant difference among inoculation treatments was detected at 10% light intensity applied over a long-term.
FIGURE 4. Architecture of M. truncatula plants inoculated or not with R. irregularis (black and white symbols, respectively), and subjected to different light regimes. Mean values ± SD (n = 5) are shown.
Numbers of shoot branches (Figure 4) was higher in mycorrhizal than non-mycorrhizal plants at full light (F2,12 = 10.9, p = 0.002) and this parameter was unaffected by short-term shading. Under long-term shading, the number of branches decreased with decreasing light intensity (F1,36 = 177, p < 0.001), and this decrease was most pronounced between 65 and 35% light intensities. No plants had side branches at 10% light intensity applied over a long-term.
The length of the main axis (Figure 4) depended strongly on the inoculation treatment under full light (t8 = 2.94, p = 0.019), with higher values observed in the mycorrhizal plants. The length of axis increased under both short- and long-term shading (F1,36 = 26.5, p < 0.001 and F1,36 = 66.6, p < 0.001, respectively). Furthermore, under long-term shading, a shortening of the longest branch was apparent between 35 and 10% light intensities, more pronouncedly in the mycorrhizal treatment, thus no difference among inoculation treatments was eventually observed at 10% light intensity.
The mean leaflet surface (Figure 4) did not differ among the inoculation treatments at full light. It was slightly but significantly increased by the short-term shading (F1,36 = 27.7, p = 0.001) and there was also a slight increase of the leaflet surface due to the mycorrhizal inoculation in shortly shaded plants (F1,36 = 4.29, p = 0.046). The response of leaflet surface to long-term shading was not monotonous, thus no significant effects were found there using the GLM analysis. Nevertheless, further GLM analysis with 100, 65 and 35% light intensities only applied over a long-term showed that leaflet surfaces strongly increased with decreasing light intensity (F1,26 = 97.1, p < 0.001) and this increase was intensified by mycorrhizal inoculation, as revealed by a significant interaction between the inoculation and the light factors (F1,26 = 7.45, p = 0.011). Leaflet surface at 10% light intensity was then approximately the same as at full light, and it did not differ between the inoculation treatments.
Mycorrhizal Colonization
The extent of root colonization by the mycorrhizal fungus in the Rhizophagus-inoculated treatment (Figure 5) varied between 34 and 96% of the root length colonized by AM fungal hyphae. Extent of root colonization by hyphae, vesicles, and arbuscules did not change significantly under short-term shading, but the first two decreased significantly under long-term shading (F1,18 = 25.1, p < 0.001 and F1,18 = 29.2, p < 0.001, respectively), whereas the decrease of the root occupancy by arbuscules was only marginally significant (p = 0.068). There were no AM fungal structures detected in the roots of non-mycorrhizal plants (data not shown).
FIGURE 5. The extent of root length colonized by R. irregularis in the inoculated M. truncatula plants subjected to different light regimes. Mean values ± SD (n = 5) are shown. H, colonization by mycorrhizal hyphae; A, colonization by arbuscules; V, colonization by vesicles.
Carbon Allocation
Both shoot and root 13C excess values (Supplementary Figure S2) showed a similar pattern as the shoot and root biomass data, although the differences between the two inoculation treatments were not statistically significant at any of the two light intensities applied over a long time. Both parameters were significantly higher at full light than at 10% light intensity applied over long-term (F1,16 = 510, p < 0.001 and F1,16 = 597, p < 0.001, respectively). Significant interaction between the light intensity and inoculation for the root 13C excess (F1,16 = 5.76, p = 0.029) indicates proportionally greater allocation of the 13C from the labeling pulse to the roots of unshaded mycorrhizal plants, while the reverse was observed in the shaded plants (Supplementary Figure S2). Simultaneously to root-to-shoot biomass ratios, the root-to-shoot ratios of 13C excess decreased with decreasing light intensity applied over a long-term (F1,16 = 338, p < 0.001) and this decrease was more pronounced in mycorrhizal than in the non-mycorrhizal plants, as revealed by a significant interaction of the inoculation and light treatments (F1,16 = 5.75, p = 0.029). The excess 13C in the substrate did not depend on any of the tested factors (data not shown).
This research highlighted several important features in the responses of plant-AM fungal symbiosis to shading with different intensities and durations. These results improve our understanding of phosphorus and carbon economy of mycorrhizal legumes along a gradient of light availabilities, and generate testable hypotheses for further studies. Here is the summary of the most important observations:
(1) In mycorrhizal plants subjected to a sudden light deprivation, P acquisition to the shoots rapidly declined, and the decline was proportional to the intensity of shade. Such a decline was not observed in the non-mycorrhizal plants. In contrast, there was no indication of a decline of the P flow to the roots of mycorrhizal plants via AM hyphal networks as compared with the non-mycorrhizal plants, with the gradually raising P concentration in the roots of the mycorrhizal plants indicating P accumulation in the intraradical fungal tissues.
(2) Long-term shading, particularly if more than 50% of the light was cut off, resulted in a significant decline of plant biomass production and massive rearrangements of plant architecture, indicating a strong C limitation. These morphological adaptations were particularly boosted by the presence of the AM fungus, but only to a certain level of light deprivation. At the most extreme shading intensity applied over a long-term, positive effects of the mycorrhizal symbiosis on the plant growth, nutrition, as well as architectural adaptations completely vanished. Root colonization levels by R. irregularis also declined with increasing intensity of shade applied over a long-term.
(3) Isotopic composition of shoot N in the plants inoculated with R. irregularis indicated an important redirection of C fluxes toward the root symbionts when the host plant was intensively shaded over a long period of time. In comparison with the non-mycorrhizal plants, Rhizophagus-inoculated plants seemed to have benefited from rhizobial symbiosis more at full light than when shaded. The N-isotopic data thus provide indirect evidence that the C flux from the shoots has been preferentially diverted to the mycosymbiont at the expense of the rhizobia when the plant was shaded.
Phosphorus Fluxes
Our model plant (M. truncatula) is known to rely strongly on the mycorrhizal P uptake pathway for its P acquisition (Smith et al., 2004). In concert with this fact, concentrations of P in the mycorrhizal plants, especially in the roots, highly exceeded those in non-mycorrhizal plants (Figure 3). The discrepancy of P responses in roots and shoots in plants exposed to a short-term shading provides strong evidence that sudden light deprivation will markedly slow down the flow of P from the fungus to the plant: As the shade intensifies, the concentration of P in the roots does not decrease or even tends to increase, strengthening the mycorrhizal P concentration response for the roots. On the other hand, a significant decrease of the mycorrhizal P concentration response was observed in the shoots with increasing shade intensity (Table 1). It is well known that once in the plant tissues, P is rapidly redistributed within the entire plant body, effectively counteracting local P accumulation (Jeschke et al., 1997; Huve et al., 2007). Thus, the selective accumulation of P in the roots can only be explained by P contained in the AM fungal tissues, presumably as long-chain polyphosphates (Viereck et al., 2004). As the P uptake in our mycorrhizal plants was to some extent in the luxurious range, i.e., P acquisition was not the primary and immediate limitation of plant growth, rapid reduction of P transfer from the fungus to the plant did not immediately affect the growth of mycorrhizal vs. control plants. However, further shading may have had negative impact on plant growth as the P concentrations in the shoots were almost equalized between the mycorrhizal and non-mycorrhizal plants after just one week of shading.
Shading applied over a long-term not only dramatically reduced plant growth, but it also markedly decreased P concentration in the shoots and roots of the mycorrhizal plants, whereas the decrease in P concentration with increasing shade was virtually not existent in the non-mycorrhizal plants (Figure 3). This can be explained by reduced efficiency of the mycorrhizal P uptake pathway upon long-term C-limitation. Mycorrhizal plants thus may suffer, under intensive and long-term shading, not only from energy shortage (which is likely to be more severe in mycorrhizal as compared to the non-mycorrhizal plants) but also from reduced P uptake rates, since the plant usually down-regulate the direct P uptake pathway in response to colonization with AM fungi (Burleigh and Bechmann, 2002). However, since the P uptake per unit of biomass remained higher in the mycorrhizal than in the non-mycorrhizal plants at all levels of shading, there are still net P benefits of the mycorrhizal symbiosis detectable at all shading levels applied in our study. This means that the plant C investments into the fungus were always rewarded by an enhanced P uptake. To completely shut down the operation of mycorrhizal exchange of P for C, one may thus need to apply even greater intensities of shade than reported here.
To gain deeper and more direct insights into the P fluxes between soil, mycorrhizal fungi and the plants, radioisotope labeling would offer more specific tool to separate and quantify the P flows via the direct (root) and the mycorrhizal uptake pathways (Frossard et al., 2011). Although we did not use this direct tracing in our current study, we believe that the magnitude of the effects presented here and the clear and consistent influence of the light intensity gradient on the mycorrhizal P uptake and P concentration responses justify the above conclusions. We also hope that our results would stimulate further studies aiming at understanding the responses of mycorrhizal plants to energy deprivation at different temporal scales and advocate using both P and C isotope labeling to disentangle the rearrangement of the respective fluxes between the fungi and the plant in response to sudden environmental changes.
Growth, Carbon, and Nitrogen Fixation
Shading applied over a long-term greatly impacted the growth and architecture of the plants, with strong compensatory mechanisms (plant elongation and increasing leaf surface) being activated upon intermediate shading levels and further stimulated by the mycorrhizal symbiosis. However, below a certain threshold (35% light intensity), the carbon costs of the symbiosis outweighed any biomass or plant architectural benefits and caused a significant growth depression of the mycorrhizal plants. The extreme shading levels tested here (10% light intensity) were still ecosystem relevant as they corresponded to the light received by understory plants under a closed (temperate to boreal) forest canopy as compared to an open landscape (Martens et al., 2000), or to extended periods of cloudy weather, or events of atmospheric smoke or volcanic ash accumulation, as compared to open skies (McCormick et al., 1995; Cirino et al., 2014).
In response to C limitation imposed by long-term shading, the plants obviously down-regulated the extent of root colonization by the AM fungus, most probably due to high C costs of the symbiosis (Son et al., 1988; Reinhard et al., 1994; Grman, 2012). Interestingly though, the colonization of roots was by far not eliminated, as also shown in other recent reports (Fellbaum et al., 2014; Knegt et al., 2015). Why and how plants still maintain relatively high colonization levels of their roots even when they are strongly C-limited and not connected to neighboring plants via common mycorrhizal networks (Lekberg et al., 2010; Walder et al., 2012; Fellbaum et al., 2014; Knegt et al., 2015) thus remains to be explained – either this is due to only a limited capacity of the host to repel the AM fungi from its roots or the fungus is retained as an insurance for a sudden change of environmental conditions.
Partly, the lack of a mycorrhizal growth depression at intermediate shading levels could possibly be attributed to the intensification of plant photosynthesis rates due to AM symbiosis establishment, a phenomenon described in a handful of experiments carried mainly under artificially lit chambers with only a fraction of a full sunlight intensity (Wright et al., 1998; Kaschuk et al., 2009, 2010). This is certainly a topic worth further investigations, advocating different light levels and temporal patterns of shading to be carefully incorporated into the experimental designs rather than scrutinizing interactions between mycorrhiza and photosynthesis up-regulation at a single light intensity as done previously. This is because some effects may only appear at certain light intensities or at a certain light regime and not others, as we also demonstrate here.
Another plant adaptation for higher C demand of the mycosymbiont under light-limiting conditions may be an increase of photosynthetically active area. Root-to-shoot biomass ratio of Medicago declined with decreasing intensity of light while mean leaflets area increased (except at the lowest light intensity applied over a long-time) and these morphological adaptations were more pronounced in the mycorrhizal plants at the 35% light intensity as compared to their non-mycorrhizal counterparts (Figure 4). This might be a reason why the plant still benefited from the symbiosis (in terms of biomass production and improved P acquisition) at this light-level despite the high costs of the symbiosis. To our knowledge, our observation is the first documented evidence of a clear morphological adaptation to light deprivation boosted by the symbiotic fungi.
The relative increase of costs of the AM symbiosis for the host plant upon decreasing light intensity could have also been buffered (partly or fully) by a reduction of symbiotic nitrogen fixation, which may be more C costly than the AM symbiosis (Paul and Kucey, 1981; Voisin et al., 2013). Our results provide indirect support for such a scenario, using the 15N natural abundance approach (George et al., 1993). Partitioning of C between the N- and P-supplying symbionts is likely to be regulated by the demand for these macronutrients (Mortimer et al., 2009; Wang et al., 2011). Thus, under conditions of suboptimal P-supply – here induced by lower efficiency of the mycorrhizal P-uptake pathway due to imbalance of symbiotic reward – the C is preferentially allocated to the fungus, resulting in increasing the share of N originating from the soil and/or N fertilizer rather than the N derived from the symbiotic N fixation (Voisin et al., 2013). Multitude of the different fluxes and dependencies make the situation rather complex (Supplementary Figure S3) and further studies on C partitioning between different root symbionts and its dynamics under changing environmental conditions are certainly warranted.
In this study, we detected surprisingly weak change in allocation of recent (13C labeled) photosynthates belowground (Root-to-shoot excess 13C) due to the mycorrhizal symbiosis along the light intensity gradient. This may be, on the one hand, due to the rather small share of C allocated to the fungus (Paul and Kucey, 1981; but Lendenmann et al., 2011) or, more likely, due to some important component missing from the C budget. Although we measured the 13C excess in the cultivation substrate next to our plants, it did not show any dependence on the mycorrhizal status of the plants. Given the short duration of labeling, it is unlikely that measuring C specifically allocated to the extraradical mycelium by means of signature fatty acids (Olsson et al., 2010) will bring much different view. However, we missed a potentially important fraction of C in this experiment, which is the one rapidly respired from the rhizosphere within a few days after labeling (Grimoldi et al., 2006; Lendenmann et al., 2011). Therefore, it seems that the delay of three days between labeling and harvest may be not the most appropriate timing to measure the mycorrhizal C drain belowground without accounting for belowground respiration. Alternative explanation would be that the amount of C allocated to the fungus was relatively small as compared to the C stored in plants. As there is still only a limited knowledge on the absolute numbers of C allocation to the fungus in mycorrhizal symbiosis, this scenario cannot be ruled out yet, but certainly deserves a dedicated attention in the future.
Ecosystem Relevance of the Results
The most surprising observation of this study was that just a few days of intense but still realistic levels of shading obviously interfered with the exchange of P for C between the plant and the AM fungus, although the P continued to be taken up from the soil by the AM fungus and transported toward the roots via the hyphae. This effectively confirms the observations made previously in a simplified in vitro systems (Lekberg et al., 2010; Hammer et al., 2011), but with a more realistic whole-plant model. It also indicates that the fungus has sufficient energy reserves to feed the P uptake and transport machinery over a period of several days independently from the C input from the host plant, because in our system there was no other (better quality) host plant available to feed the mycorrhizal network as in the other recent studies (Fellbaum et al., 2014; Knegt et al., 2015).
What do these results mean in a real-world? Possibly, a few cloudy days in a row or temporary shading due to industrial smoke, smog, or volcanic ash may significantly slow-down the exchange of goods at the symbiotic interface, without swiftly affecting the root colonization levels and with possibly both partners remaining prepared to restore the functioning of the interaction. For the conditions of long-term shading, frequently encountered in many ecological situations, we show that mycorrhizal fungi may be able to maintain significant levels of root colonization and that the plants are likely to possess mechanisms to compensate for the C demands of the mycosymbionts, at least up to a certain degree. This appears, again, as an important pre-requisite for an adequate reaction to changing environmental conditions: The mycorrhizal plant is thus not particularly disadvantaged in comparison with a potential non-mycorrhizal competitor and can quickly start profiting from already established mycorrhizal symbiosis – provided the conditions, especially the energy inputs, are favorable for expression of the symbiotic benefits.
Given the above thoughts and lack of specific knowledge, our results call for more detailed studies on the dynamics of mycorrhizal functioning, especially the carbon fluxing from plants to its microbial symbionts, in rapidly changing (micro-) climatic conditions – and this is what is very rarely done in glasshouse experiments (Olsson et al., 2010). Further we showed that the responses of plants to mycorrhizal symbiosis across a light intensity gradient are often not trivial, advocating specific efforts to be dedicated to manipulation of light intensity levels and shading durations in future studies on mycorrhizal ecophysiology. This appears important in order to dissect the mechanisms and quantify the volumes of carbon-for-phosphorus exchanges between the partners in mycorrhizal symbiosis as well as their control mechanisms.
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
This research was carried out in the joint working group of the Institute of Microbiology and the Institute of Botany. It was financially supported by the Czech Ministry of Education, Youth and Sports (project LK11224) and the J. E. Purkyně fellowship to Jan Jansa, as well as by the long-term development programmes RVO 61388971 and RVO 67985939. The assistance of Jiří Machač with plant architectural analyses is gratefully acknowledged.
The Supplementary Material for this article can be found online at: http://www.frontiersin.org/journal/10.3389/fpls.2015.00065/abstract
^ Infrared CO2 sensor is differentially sensitive to 13CO2 and 12CO2. Therefore, total CO2 concentration of highly 13C enriched atmosphere cannot be measured directly, but we rather summed the CO2 concentration measured just before the labeling with calculated surplus of 13CO2 released from the 13C-carbonate.
Burleigh, S. H., and Bechmann, I. E. (2002). Plant nutrient transporter regulation in arbuscular mycorrhizas. Plant Soil 244, 247–251. doi: 10.1023/A:1020227232140
CrossRef Full Text | Google Scholar
Burleigh, S. H., and Harrison, M. J. (1999). "Mt4, a phosphorus starvation-inducible cDNA from Medicago truncatula, which is down-regulated both by phosphorus fertilization and arbuscular-mycorrhizal colonization," in Phosphorus in Plant Biology: Regulatory Roles in Molecular, Cellular, Organismic, and Ecosystem Processes, eds J. P. Lynch and J. Deikman (Rockville, MD: American Society of Plant Biologists), 359–360.
Cirino, G. G., Souza, R. A. F., Adams, D. K., and Artaxo, P. (2014). The effect of atmospheric aerosol particles and clouds on net ecosystem exchange in the Amazon. Atmos. Chem. Phys. 14, 6523–6543. doi: 10.5194/acp-14-6523-2014
Drigo, B., Pijl, A. S., Duyts, H., Kielak, A., Gamper, H. A., Houtekamer, M. J.,et al. (2010). Shifting carbon flow from roots into associated microbial communities in response to elevated atmospheric CO2. Proc. Natl. Acad. Sci. U.S.A. 107, 10938–10942. doi: 10.1073/pnas.0912421107
Pubmed Abstract | Pubmed Full Text | CrossRef Full Text | Google Scholar
Fellbaum, C. R., Gachomo, E. W., Beesetty, Y., Choudhari, S., Strahan, G. D., Pfeffer, P. E.,et al. (2012). Carbon availability triggers fungal nitrogen uptake and transport in arbuscular mycorrhizal symbiosis. Proc. Natl. Acad. Sci. U.S.A. 109, 2666–2671. doi: 10.1073/pnas.1118650109
Fellbaum, C. R., Mensah, J. A., Cloos, A. J., Strahan, G. E., Pfeffer, P. E., Kiers, E. T.,et al. (2014). Fungal nutrient allocation in common mycorrhizal networks is regulated by the carbon source strength of individual host plants. New Phytol. 203, 646–656. doi: 10.1111/nph.12827
Fitter, A. H., Helgason, T., and Hodge, A. (2011). Nutritional exchanges in the arbuscular mycorrhizal symbiosis: implications for sustainable agriculture. Fungal Biol. Rev. 25, 68–72. doi: 10.1016/j.fbr.2011.01.002
Frossard, E., Achat, D. L., Bernasconi, S. M., Bunemann, E. K., Fardeau, J. C., Jansa, J.,et al. (2011). "The use of tracers to investigate phosphate cycling in soil-plant systems," in Phosphorus in Action: Biological Processes in Soil Phosphorus Cycling, eds E. Bünemann, A. Oberson, and A. Frossard (Berlin: Springer), 59–91. doi: 10.1007/978-3-642-15271-9_3
Gehring, C. A. (2003). Growth responses to arbuscular mycorrhizae by rain forest seedlings vary with light intensity and tree species. Plant Ecol. 167, 127–139. doi: 10.1023/A:1023989610773
George, T., Singleton, P. W., and Vankessel, C. (1993). The use of 15N natural abundance and nitrogen yield of nonnodulating isolines to estimate nitrogen fixation by soybeans (Glycine max L) across 3 elevations. Biol. Fertil. Soils 15, 81–86. doi: 10.1007/BF00336422
Grimoldi, A. A., Kavanova, M., Lattanzi, F. A., Schaufele, R., and Schnyder, H. (2006). Arbuscular mycorrhizal colonization on carbon economy in perennial ryegrass: quantification by 13CO2/12CO2 steady-state labelling and gas exchange. New Phytol. 172, 544–553. doi: 10.1111/j.1469-8137.2006.01853.x
Grman, E. (2012). Plant species differ in their ability to reduce allocation to non-beneficial arbuscular mycorrhizal fungi. Ecology 93, 711–718. doi: 10.1890/11-1358.1
Gryndler, M., Vejsadová, H., and Vančura, V. (1992). The effect of magnesium-ions on the vesicular arbuscular mycorrhizal infection of maize roots. New Phytol. 122, 455–460. doi: 10.1111/j.1469-8137.1992.tb00073.x
Hammer, E. C., Pallon, J., Wallander, H., and Olsson, P. A. (2011). Tit for tat? A mycorrhizal fungus accumulates phosphorus under low plant carbon availability. FEMS Microbiol. Ecol. 76, 236–244. doi: 10.1111/j.1574-6941.2011.01043.x
Heinemeyer, A., Ridgway, K. P., Edwards, E. J., Benham, D. G., Young, J. P. W., and Fitter, A. H. (2003). Impact of soil warming and shading on colonization and community structure of arbuscular mycorrhizal fungi in roots of a native grassland community. Glob. Chang. Biol. 10, 52–64. doi: 10.1111/j.1365-2486.2003.00713.x
Huve, K., Merbach, W., Remus, R., Luttschwager, D., Wittenmayer, L., Hertel, K.,et al. (2007). Transport of phosphorus in leaf veins of Vicia faba L. J. Plant Nutr. Soil Sci. 170, 14–23. doi: 10.1002/jpin.200625057
Jakobsen, I., and Rosendahl, L. (1990). Carbon flow into soil and external hyphae from roots of mycorrhizal cucumber plants. New Phytol. 115, 77–83. doi: 10.1111/j.1469-8137.1990.tb00924.x
Jansa, J. (2002). Effect of Soil Tillage on Arbuscular Mycorrhizal Fungi and on their Role in Nutrient Uptake by Crops. Ph.D. thesis, Swiss Federal Institute of Technology (ETH), Zürich.
Javot, H., Penmetsa, R. V., Terzaghi, N., Cook, D. R., and Harrison, M. J. (2007). A Medicago truncatula phosphate transporter indispensable for the arbuscular mycorrhizal symbiosis. Proc. Natl. Acad. Sci. U.S.A. 104, 1720–1725. doi: 10.1073/pnas.0608136104
Jeschke, W. D., Kirkby, E. A., Peuke, A. D., Pate, J. S., and Hartung, W. (1997). Effects of P deficiency on assimilation and transport of nitrate and phosphate in intact plants of castor bean (Ricinus communis L). J. Exp. Bot. 48, 75–91. doi: 10.1093/Jxb/48.1.75
Johnson, N. C., and Graham, J. H. (2013). The continuum concept remains a useful framework for studying mycorrhizal functioning. Plant Soil 363, 411–419. doi: 10.1007/s11104-012-1406-1
Johnson, N. C., Graham, J. H., and Smith, F. A. (1997). Functioning of mycorrhizal associations along the mutualism-parasitism continuum. New Phytol. 135, 575–586. doi: 10.1046/j.1469-8137.1997.00729.x
Johnson, N. C., Wilson, G. W. T., Wilson, J. A., Miller, R. M., and Bowker, M. A. (2015). Mycorrhizal phenotypes and the law of the minimum. New Phytol. 205, 1473–1484. doi: 10.1111/nph.13172
Kaschuk, G., Kuyper, T. W., Leffelaar, P. A., Hungria, M., and Giller, K. E. (2009). Are the rates of photosynthesis stimulated by the carbon sink strength of rhizobial and arbuscular mycorrhizal symbioses? Soil Biol. Biochem. 41, 1233–1244. doi: 10.1016/j.soilbio.2009.03.005
Kaschuk, G., Leffelaar, P. A., Giller, K. E., Alberton, O., Hungria, M., and Kuyper, T. W. (2010). Responses of legumes to rhizobia and arbuscular mycorrhizal fungi: a meta-analysis of potential photosynthate limitation of symbioses. Soil Biol. Biochem. 42, 125–127. doi: 10.1016/j.soilbio.2009.10.017
Kiers, E. T., Duhamel, M., Beesetty, Y., Mensah, J. A., Franken, O., Verbruggen, E.,et al. (2011). Reciprocal rewards stabilize cooperation in the mycorrhizal symbiosis. Science 333, 880–882. doi: 10.1126/science.1208473
Knegt, B., Jansa, J., Franken, O., Engelmoer, D. J. P., Werner, G. D. A., Bücking, H.,et al. (2015). Host plant quality mediates competition between arbuscular mycorrhizal fungi. Fungal Ecol. doi: 10.1016/j.funeco.2014.09.011
Korhonen, J., Kytoviita, M. M., and Siikamaki, P. (2004). Are resources allocated differently to symbiosis and reproduction in Geranium sylvaticum under different light conditions? Can. J. Bot. 82, 89–95. doi: 10.1139/B03-142
Lekberg, Y., Hammer, E. C., and Olsson, P. A. (2010). Plants as resource islands and storage units - adopting the mycocentric view of arbuscular mycorrhizal networks. FEMS Microbiol. Ecol. 74, 336–345. doi: 10.1111/j.1574-6941.2010.00956.x
Lendenmann, M., Thonar, C., Barnard, R. L., Salmon, Y., Werner, R. A., Frossard, E.,et al. (2011). Symbiont identity matters: carbon and phosphorus fluxes between Medicago truncatula and different arbuscular mycorrhizal fungi. Mycorrhiza 21, 689–702. doi: 10.1007/s00572-011-0371-5
Martens, S. N., Breshears, D. D., and Meyer, C. W. (2000). Spatial distributions of understory light along the grassland/forest continuum: effects of cover, height, and spatial pattern of tree canopies. Ecol. Modell. 126, 79–93. doi: 10.1016/S0304-3800(99)00188-X
McCormick, M. P., Thomason, L. W., and Trepte, C. R. (1995). Atmospheric effects of the Mt-Pinatubo eruption. Nature 373, 399–404. doi: 10.1038/373399a0
McGonigle, T. P., Miller, M. H., Evans, D. G., Fairchild, G. L., and Swan, J. A. (1990). A new method which gives an objective measure of colonization of roots by vesicular-arbuscular mycorrhizal fungi. New Phytol. 115, 495–501. doi: 10.1111/j.1469-8137.1990.tb00476.x
Millar, J. A., and Ballhorn, D. J. (2013). Effect of mycorrhizal colonization and light limitation on growth and reproduction of lima bean (Phaseolus lunatus L.). J. Appl. Bot. Food Q. 86, 172–179. doi: 10.5073/Jabfq.2013.086.023
Mortimer, P. E., Perez-Fernandez, M. A., and Valentine, A. J. (2009). Arbuscular mycorrhizae affect the N and C economy of nodulated Phaseolus vulgaris (L.) during NH4+ nutrition. Soil Biol. Biochem. 41, 2115–2121. doi: 10.1016/j.soilbio.2009.07.021
Ohno, T., and Zibilske, L. M. (1991). Determination of low concentrations of phosphorus in soil extracts using malachite green. Soil Sci. Soc. Am. J. 55, 892–895. doi: 10.2136/sssaj1991.03615995005500030046x
Olsson, P. A., Rahm, J., and Aliasgharzad, N. (2010). Carbon dynamics in mycorrhizal symbioses is linked to carbon costs and phosphorus benefits. FEMS Microbiol. Ecol. 72, 123–131. doi: 10.1111/j.1574-6941.2009.00833.x
Paul, E. A., and Kucey, R. M. N. (1981). Carbon flow in plant microbial associations. Science 213, 473–474. doi: 10.1126/science.213.4506.473
Pearson, J. N., and Jakobsen, I. (1993a). The relative contribution of hyphae and roots to phosphorus uptake by arbuscular mycorrhizal plants, measured by dual labeling with 32P and 33P. New Phytol. 124, 489–494. doi: 10.1111/j.1469-8137.1993.tb03840.x
Pearson, J. N., and Jakobsen, I. (1993b). Symbiotic exchange of carbon and phosphorus between cucumber and 3 arbuscular mycorrhizal fungi. New Phytol. 124, 481–488. doi: 10.1111/j.1469-8137.1993.tb03839.x
Rausch, C., Daram, P., Brunner, S., Jansa, J., Laloi, M., Leggewie, G.,et al. (2001). A phosphate transporter expressed in arbuscule-containing cells in potato. Nature 414, 462–466. doi: 10.1038/35106601
Reinhard, S., Weber, E., Martin, P., and Marschner, H. (1994). Influence of phosphorus supply and light-intensity on mycorrhizal response in Pisum-Rhizobium-Glomus symbiosis. Experientia 50, 890–896. doi: 10.1007/Bf01923475
Smith, S. E., Smith, F. A., and Jakobsen, I. (2004). Functional diversity in arbuscular mycorrhizal (AM) symbioses: the contribution of the mycorrhizal P uptake pathway is not correlated with mycorrhizal responses in growth or total P uptake. New Phytol. 162, 511–524. doi: 10.1111/j.1469-8137.2004.01039.x
Somasegaran, P., and Hoben, H. J. (1994). Handbook for Rhizobia: Methods in Legume-Rhizobium Technology. New York: Springer. doi: 10.1007/978-1-4613-8375-8
Son, C. L., Smith, F. A., and Smith, S. E. (1988). Effect of light-intensity on root-growth, mycorrhizal infection and phosphate-uptake in onion (Allium cepa L). Plant Soil 111, 183–186. doi: 10.1007/Bf02139935
Son, C. L., and Smith, S. E. (1988). Mycorrhizal growth-responses - interactions between photon irradiance and phosphorus-nutrition. New Phytol. 108, 305–314. doi: 10.1111/j.1469-8137.1988.tb04167.x
Thonar, C., Schnepf, A., Frossard, E., Roose, T., and Jansa, J. (2011). Traits related to differences in function among three arbuscular mycorrhizal fungi. Plant Soil 339, 231–245. doi 10.1007/s11104-010-0571-3
Tuomi, J., Kytoviita, M. M., and Hardling, R. (2001). Cost efficiency of nutrient acquisition and the advantage of mycorrhizal symbiosis for the host plant. Oikos 92, 62–70. doi: 10.1034/j.1600-0706.2001.920108.x
Viereck, N., Hansen, P. E., and Jakobsen, I. (2004). Phosphate pool dynamics in the arbuscular mycorrhizal fungus Glomus intraradices studied by in vivo 31P NMR spectroscopy. New Phytol. 162, 783–794. doi: 10.1111/j.1469-8137.2004.01048.x
Voisin, A. S., Cazenave, A. B., Duc, G., and Salon, C. (2013). Pea nodule gradients explain C nutrition and depressed growth phenotype of hypernodulating mutants. Agron. Sustain. Dev. 33, 829–838. doi: 10.1007/s13593-013-0146-9
Walder, F., Niemann, H., Natarajan, M., Lehmann, M. F., Boller, T., and Wiemken, A. (2012). Mycorrhizal networks: common goods of plants shared under unequal terms of trade. Plant Physiol. 159, 789–797. doi: 10.1104/pp.112.195727
Wang, X. R., Pan, Q. A., Chen, F. X., Yan, X. L., and Liao, H. (2011). Effects of co-inoculation with arbuscular mycorrhizal fungi and rhizobia on soybean growth as related to root architecture and availability of N and P. Mycorrhiza 21, 173–181. doi: 10.1007/s00572-010-0319-1
Wright, D. P., Scholes, J. D., and Read, D. J. (1998). Effects of VA mycorrhizal colonization on photosynthesis and biomass production of Trifolium repens L. Plant Cell Environ. 21, 209–216. doi: 10.1046/j.1365-3040.1998.00280.x
Keywords: arbuscular mycorrhizal symbiosis, light intensity, shading duration, mycorrhizal benefits, phosphorus acquisition, biomass production, root colonization, nitrogen fixation
Citation: Konvalinková T, Püschel D, Janoušková M, Gryndler M and Jansa J (2015) Duration and intensity of shade differentially affects mycorrhizal growth- and phosphorus uptake responses of Medicago truncatula. Front. Plant Sci. 6:65. doi: 10.3389/fpls.2015.00065
Received: 22 August 2014; Accepted: 25 January 2015;
Published online: 13 February 2015.
Jan Kofod Schjoerring, University of Copenhagen, Denmark
Iver Jakobsen, Technical University of Denmark, Denmark
Daniel J. Ballhorn, Portland State University, USA
Copyright © 2015 Konvalinková, Püschel, Janoušková, Gryndler and Jansa. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
*Correspondence: Tereza Konvalinková, Laboratory of Fungal Biology, Institute of Microbiology, Academy of Sciences of the Czech Republic, Vídeňská 1083, Prague 142 20, Czech Republic e-mail: konvalinkova@biomed.cas.cz
People also looked at
There is no supplementary material currently available for this article
Loading supplemental data...
Notify me on publication
If you already have an account, please login.
You don't have a Frontiers account ? You can register here.
Select one of your emails
You have multiple emails registered with Frontiers:
Comment deleted on


Who Watched the Watchmen? Me!
On Friday night I was as predictable as most of the people in my feedreader and was camped down at 5PM for the evening showing of Watchmen - and I enjoyed it.
It's been a good few years since I read the original graphic novel so I'm not as likely to pick out little errors and omissions (like the Silk Spectre looking for a lighter in the book and just being nosy in the film) but I thought the story was a very good, and close, adaptation. The fight scenes were well done and extremely short, sharp and brutal and the film utilised its 18 rating well.
I'm interested in how easy to follow the film will be to someone who's never been exposed to the source material. There were lots of little bits and pieces that might not fit together that easily. I thought the pacing was decent, I was never bored and didn't get too fidgety.
I thought the idea of using Dr Manhattans power as the basis for the destruction was an acceptable alternative to the novels ending. Trying to stick to the novel would have required a lot more leg work and slowed the pace. I was a little surprised at how most of the heros seemed to have some degree of super-strength but that's artistic license. As for the killing of Rorschach I can see why they did it that way and increased the drama but I didn't like it. The original handled it much better and without all the over the top responses.
Although I missed the giant squid I'll still give it 8⁄10 for fans of the novel, 6⁄10 for others.


Decorate your homeschool room with these sturdy and colorful cardstock reminders of the formation of the cursive alphabet. This set contains upper and lowercase letters on 9 cards and numbers 1–10 on 2 cards, specifically designed for 3rd graders. Now with an updated "white-board" look, this set of cards is perfect for display for student reference. Help your child write neatly and beautifully with this classroom brightener!


Mérieux NutriSciences works through easy and direct relationships with customers as the utmost transparency at every stage is an important element when offering practical and innovative solutions to specific needs.
To achieve this goal, listening and understanding customers' needs are of utmost importance: Mérieux NutriSciences thus created its Customer Care, a team with qualified skills and dedicated to each customer. You can contact our Customer Care to receive technical support and updated and timely information.
Each customer is followed by a Customer Care operator who has been selected on the basis of the skills connected to customers' productive sector and specificity of products.
Mérieux NutriSciences operators have cutting-edge skills in a particular sector or product category and provide customers with detailed and specific support for every need.
Thanks to this organization specializing in various sectors, customers from the food industry, the primary sector, cereal, fruit and vegetable and other sectors, as well as large retailers and caterers will find a partner able to tackle their specific problems and a valuable guidance in interpreting tests' results.


text:
Five medals for Britain on day two at European Juniors
Posted by Paul Halford | Jul 17, 2015 | 0
Ojie Edoburun and Laviai Nielsen live up to their billing as pre-event favourites in their events as they inspire a British medal rush in Sweden
Ojie Edoburun in the 100m and Laviai Nielsen in the 400m each led a Great Britain 1-2 on the second day of the European Junior Championships in Eskilstuna, Sweden, on Friday.
Edoburun led fellow Briton Joseph Dewar by 0.1 as he clocked 10.36. Reuben Arthur just missed out on making it a GB monopoly of the podium as he finished fourth with 10.67. It was Emil von Barth who took bronze for the home nation as Sweden also had three in the final.
Edoburun was two tenths outside his PB, but for the Shaftesbury Barnet Harrier it was all about becoming Britain's 12th men's 100m champion in the 23 editions of these championships.
He said: "It feels really good that I've finally got my first title. I got the silver in Donetsk two years ago (World Youth Championships) and I feel like I've grown a lot since then. I've used the experiences from the World Youths and [European] Juniors to put together a really good champs here in Eskilstuna."
Edoborun, who was third at the Sainsbury's British Championships – the national event for seniors held earlier this month – believes he has a good chance of getting into Britain's relay squad for the IAAF World Championships next month.
Dewar, who previously concentrated on long jump, at which he has a 7.36m PB, was delighted with his silver medal on his major championships debut.
In the women's 400m, Nielsen clocked 52.58 as she headed silver medallist Cheriece Hylton (53.16). Lina Nielsen, the identical twin sister of the winner, was eighth in the final, running from lane eight and having been laid low with a cold, with 54.72.
Laviai said: "It's what I've wanted all season. I've had so many doubts throughout the season,but on the startline I said to myself, 'You have to believe in yourself.'"
The 19-year-old revealed she was concerned when she had to withdraw injured from the England Under-20 Championships last month. She said: "I was thinking, 'Am I going to recover in time?' But my coach and everyone around me had more belief in me than I've had in myself so I have to thank everyone."
Hylton, whose identical twin Shannon later won her 200m heat in 23.24, was delighted to take 0.38 off her PB.
Britain's Jacob Fincham-Dukes set a PB to win silver in the long jump with 7.75m, pushing Russia's Anatoliy Ryapolov to equal his own best of 7.96m to take gold.
He said: "I wasn't in medal contention going into it, but once a few of the people dropped out in the qualifying it was anybody's medal. In qualifying and the war- up I felt really good so I knew anything could happen today."
Switzerland's Caroline Agnou's 6123 to win the heptathlon took her into the world all-time top 20 at junior level. Boosted by a 13.86m shot and 49.34 javelin, the Swiss junior record was just 25 points less than the score which gave Morgan Lake world junior gold last year.
Hungary's Bence Halasz set a championship best to win the hammer with 79.60m, but UK junior hammer record-holder Taylor Campbell was disappointed to miss out on a medal. His 75.63m was more than three metres down on the best that ranked him third of the finalists coming into the event and was only good enough for sixth. European No.1 Germany's Alexej Mikhailov also performed below-par in 11th.
In an eventful women's discus competition, the big favourite Claudine Vita of Germany suffered a scare before taking gold with 57.47m. Slovenia's Veronika Domjan led until round five when Vita – who was six metres ahead at the top of the world rankings with 62.31m – edged ahead with 56.64m. She improved to 57.47m in the final round and, despite Domjan improving her own national senior record to 56.63m, it was enough.
Moments after the GB 1-2 in the men's 100m, their female counterparts failed to follow them on to the podium.
Daryll Neita was devastated to place fourth in 11.69 – just a hundredth behind the bronze medallist as Polish pre-race favourite Ewa Swoboda took gold in 11.52. Britain's Imani Lansiquot was fifth with 11.74.
Russia's Valentina Kosolapova took a 2cm win over fellow Russian Kristina Malaya to win the triple jump with 13.27m.
World junior champion Morgan Lake survived a nervy start to her campaign in the high jump as she needed three attempts to go clear at her opening height of 1.79m.
The athlete who also scooped world junior heptathlon gold last year, said: "We'd been sitting around for quite a while so I wasn't that fresh going into the first attempt and just brushed it over. The second one was a bit rushed and the run-up wasn't great, so I had to leave it up to the final attempt.
"I'm glad I did start at 1.79m. I wasn't thinking about 1.74m. I usually start at 1.80m so I don't think I pushed myself too much at the starting height, but think I might go a bit lower in the final if there's a long wait in between."
The UK junior record-holder at 1.94m should start as a big favourite in the final as her closest challenger has jumped 1.92m.
All three British pole vaulters went through to the final. UK junior record-holder Adam Hague and Charlie Myers were joined by Harry Coppell, who survived a nervous moment at 5.05m – 37cm below his best – as he needed three attempts.
It was a similar story in the 800m where all of the British trio looked comfortable. UK senior champion Kyle Langford will be joined in the semi-finals by compatriots Spencer Thomas and Daniel Rowden.
As with the men's 100m, the half-lap final will also feature three British runners as Tommy Ramdhan (21.18) and Elliot Powell (21.06) won their heats and Frankie Owsley (21.37) joined them as a fastest loser.
Also progressing from the GB team were Jack Lawrie, whose winning 51.91 in the 400m hurdles heats is just 0.14 outside his PB, and Charlotte McLennaghan, who coasted to a win in her 200m heat in 23.64.
However, there was rare disappointment for the British team as Jack Hutton, who had been a medal contender in the 110m hurdles, pulled out just before the heats with a hamstring injury.
British steeplechaser Haran Dunderdale was just ninth in his heat but just went through to the final as the last fastest loser, but compatriot Jacob Jibb was last in his despite looking comfortable in the lead for much of the race.
» Full results can be found at www.european-athletics.org
Tags: Eskilstuna, European Junior Championships, Photography credit: Mark Shearman
PreviousRunners flock to Olympic Park for Morrisons Great Newham London Run
NextWorld 1500m record: Genzebe Dibaba magnificent in Monaco
Paul Halford
Charlie Grice on his magical metric mile in Monaco...
Grab a drink and make yourself comfortable then enjoy this #LongRead as @CharlieGrice1 tells @Jason_AW about his 3:30 1500m and how beating Steve Ovett's time was a performance a decade in the making ➡ https://t.co/LL9WZDwY30
Mark Shearman MBE@AthleticsImages·
4 years ago today Josh Kerr won the 1500m. at the Eur. junior champs. in Sweden @scotathletics @AthleticsWeekly @joshk97 @brooksrunning

redpajama_set_name: RedPajamaCommonCrawl


text:
In Nature All Things Must Follow an Order, Even Viruses. Why Was This Not Observed With Treating SARS-CoV-2?
Home|In Nature All Things Must Follow an Order, Even Viruses. Why Was This Not Observed With Treating SARS-CoV-2?
"After all, if we can live through the massive fear campaign for contamination, we can sure handle receiving the truth."
We try to complicate matters but nature is not that complicated when you understand the science of natural law or the patterns of function. We have survived for many millennia with viruses: it is estimated that humanity has 380 trillion viruses within and outside the human body. If this is the case, why did we change everything with the discovery of SARS-CoV-2 that had worked in the past? In the midst of a pandemic don't you think it is irresponsible to try to introduce an experimental treatment while the virus is still active and changing its DNA? It would have been much better to allow the virus to run its course, while protecting the most vulnerable, than to place the entire global population into a petri dish for observation and discovery.
We must understand the origins of viruses
Viruses are quite diverse. Unlike all other biological entities, some of them (like poliovirus) have RNA genomes and some (like herpesvirus) have DNA genomes. Further, some viruses (like influenza virus) have single-stranded genomes, while others (like smallpox) have double-stranded genomes. Viruses, do, however, share a few features, e.g. that they can replicate only within a host cell. We also know that no known virus contains ribosomes, the necessary component of a cell's protein-making translational machinery.[1]
If viruses don't contain ribosomes, then why would you use an experimental mRNA-ribonucleic acid as possible treatment? According to Lynne Maquat, scientist from the University of Rochester, who studies the RNA of viruses "understanding RNA structure and function helps us understand how to throw a therapeutic wrench into what the COVID-19 RNA does—make new virus that can infect more of our cells and also the cells of other human beings."[2] My question is this: what if you get the genetic sequences wrong? "Once the virus is in our cells, the entire process of infection and re-infection depends on the viral RNA," Maquat says.[3] Scary stuff to be experimenting with on a global scale wouldn't you say?
Protection and the inconsistencies
But we must protect ourselves somehow, you might say. What about the masks then—aren't they useful? Let's take a look at some math. Viruses are quite small, with a diameter of less than 125 nanometers (nm) or (0.125 microns) but their range is between 0.06 and 0.14 microns. Now here are the numbers for the N95 mask filters: they are down to 0.3 microns which means that they block few, if any, virions (virus particles). With this in mind why would the media push the wearing of masks and let the people think they were protected when they knew perfectly well that they were ineffective in blocking the viral transmission of SARS-CoV-2?
COVID-19 rarely spreads through surfaces. They knew this at the beginning of the virus since this was not the first time the coronavirus was introduced into society—MERS and SARS were both from the same family. So why were we shown images of workers in hazmat suits spraying every possible surface, and people changing their work clothes before entering their own homes? They stated afterwards that the virus was airborne with very low viral transmission on surfaces and yet most stores and restaurants continue requesting hand sanitizers to be used while staff uses disinfectants on touch pads after every payment, wearing face masks, in an attempt to make healthy people believe they are all contaminated.
So who takes responsibility for all these claims? It seems that nobody does. Nature magazine, for example, tried to ask the CDC regarding the inconsistencies in its statements about the risks posed by fomites (clothes, surfaces) but the agency has not responded.
Have we also lost our minds in an attempt to blocking the existence of natural herd immunity and existing COVID-19 antibodies, why is that? Does it interfere with the business side of "health"?
What about natural immunity?
We know that natural herd immunity and COVID-19 (coronavirus) can be reached when enough people in the population have recovered from a disease and have developed protective antibodies against future infection. We can assume after living through SARS and MERS that a good percentage of people have natural antibodies for the Covid virus, although the strain of SARS-CoV-2 will be different in nature. Our adaptive immunity would respond and the recovery from infection would be much shorter for those with the natural antibodies than for those without.
An Israeli study revealed that natural immunity based on prior infection is considerably better than 2 doses of the Covid mRNA vaccine. According to Science Magazine, the study found that "never-infected people who were vaccinated in January and February were, in June, July, and the first half of August, six to 13 times more likely to get infected than unvaccinated people who were previously infected with the coronavirus."[4]
Tell me this: does it make any sense only to use PCR testing which has proven to be often false positive in many situations than to test the general public for natural immune antibodies for COVID-19? If they test for natural immunity, why on earth would they be shunned and overlooked, what possible reason would it do?
It would be nice to see those in public office to come clean. After all, if we can live through the massive fear campaign for contamination, we can sure handle receiving the truth. I always say follow the money, and in this case money and control go hand in hand. What do you think?
At the end it all comes down to one truth: if we don't work proactively to become and stay healthy, if we don't change our lifestyle in a way that is full of healthy foods, good amount of sleep and movement, quality supplements and as low of a stress level as possible, no amount of hand sanitizers, face masks and PCR tests is going to save us. The emphasis should be on healing, it should be about taking care of the root of a problem, and not about covering up its results.
Prevention Protocol for COVID-19: https://lifechoice.net/prevention-protocol-for-covid-19/
Wessner, David R. 2010. The Origins of Viruses. Nature Education3(9):37.
University of Rochester. COVID-19 vaccine: What's RNA research got to do with it? https://www.rochester.edu/newscenter/covid-19-rna-coronavirus-research-428952/
Arutz Sheva Staff. 2021. Israeli study: Natural immunity gives better protection than COVID shot. https://www.israelnationalnews.com/news/312637
[1] Wessner, David R. 2010.
[2] University of Rochester. 2020.
[4] Arutz Sheva Staff. 2021.
By Eldon Dahl|2022-02-14T15:05:54-07:00February 2nd, 2022|Uncategorized|0 Comments


Democrats are breathing a sigh of relief this week after a reported hack of the DNC's valuable voter database turned out to be a false alarm.
WASHINGTON – Democrats are breathing a sigh of relief this week after a reported hack of the Democratic National Committee's valuable voter database turned out to be a false alarm triggered accidentally by the Michigan Democratic Party.
The DNC has been on the lookout for cyber attacks this election year after being hacked in 2015 and 2016 by the Russian government. U.S. intelligence agencies said the Kremlin interfered in the election to try to help Donald Trump and hurt Hillary Clinton.
This week, the DNC was notified by an independent security firm that there was a spearphishing attempt to lure people into entering their passwords for the voter database onto a fake website.
The DNC notified the FBI and the state parties about the phishing attempt on Wednesday. Later that day, the Michigan Democratic Party told the DNC that they had been running a cybersecurity test on their site – which is linked to the DNC – and failed to tell the national committee.
Brandon Dillon, chairman of the Michigan Democratic Party, said Thursday that the state party has taken steps to fortify its cybersecurity, "especially as the Trump Administration refuses to crack down on foreign interference in our elections."
"Despite our misstep and the alarms that were set off, it's most important that all of the security systems in place worked," Dillon said in a statement.
Bob Lord, the DNC's chief security officer, said sensitive data that is critical to candidates and state parties was never compromised. He said the DNC took steps to protect its site as soon as it was informed of the suspected phishing attempt.
"There are constant attempts to hack the DNC and our Democratic infrastructure, and while we are extremely relieved that this wasn't an attempted intrusion by a foreign adversary, this incident is further proof that we need to continue to be vigilant in light of potential attacks," Lord said in a statement.
The DNC was criticized heavily in 2016 for failing to take strong cybersecurity measures or heed warnings from the FBI and other intelligence agencies to protect its systems. Tom Perez, who was elected as DNC chairman in early 2017, promised to increase efforts to shield the party from cyber attacks.
Leaders of the FBI, CIA and National Security Agency have all warned that they expect Russia and other foreign adversaries to try to interfere in this year's midterm elections.
Just this week, Microsoft announced that it had uncovered new Russian hacking attempts targeting U.S. political groups. The unsuccessful attempts were aimed at two conservative think tanks: the Hudson Institute and the International Republican Institute.


So...I have been swamped with this...finally my thank you cards for the wedding arrived and lucky they have because I think it is honestly going to take me two weeks to thank all of the amazing guests we had attend our wedding! My goal is to have these done and mailed out before December 01...and I am pretty optimistic I will meet that goal, however I don't think I have done this much writing since my university days! It has been such a rewarding process though, and a wonderful trip down memory lane considering I still don't recollect about 95% of the day itself. It is still a blur, and something that at times really does make me sad. It is so nice to read all of the cards and everyones well wishes and sweet words of wisdom and praise.
On another note...last weekend I threw myself into creative mode and finished my December Daily album, as seen on Ali Edwards blog. I am so happy to have it ready and am eagerly awaiting December 01/2008 so I can start recording all of the big and little moments this December has to offer. I know I will be so thankful that I did this project and will be one of the greatest gifts I can give myself. Thank you Ali for some amazing inspiration once again!
Love your album...and can't wait to see more as you fill it up!


Dr. Mark Anthony Neal analyzes the ways in which black masculinity has been read and misread through popular culture. The most 'legible' black men are often rendered as criminal. He has examined figures such as Jay-Z, R&B Svengali R. Kelly, Luther Vandross, and characters from the The Wire, among others. In conjunction with the museum's current exhibition Question Bridge: Black Males.


Accreditation: The School is a member of and accredited by the National Association of Independent Schools (NAIS) and the New York State Association of Independent Schools (NYSAIS).
Diversity: More than 25 local communities are represented at East Woods School.
Faculty and Staff: 49 Full & Part-Time.
Languages Offered to Our Students: Spanish and French. Spanish is introduced to the curriculum in our Nursery program. French is introduced in the Third grade, when the students are taught Spanish for half the year and French for half the year.
Sports: Our campus is home to four playing fields and two gymnasiums. Fourth through Eighth graders participate on interscholastic sports teams in the fall, winter, and spring. Teams include: soccer, ice hockey, cross country, basketball, baseball, softball, and lacrosse.
Distinctive Programs: Multi-media technology; music and drama productions; fine arts curriculum; student council; overnight field trips for ages 10 and above. We offer after school extended care for Nursery and Pre-K, and after school enrichment classes for grades Kindergarten through Fourth, after-care programs, and an extensive summer program.
Facilities and Tools: 2 science labs, 1 Mac lab, 1 PC lab, 1 LEGO Robotics Room, 1 Maker Space, 1 dance studio, 1 art studio, 1 wood shop, 1 band room, 4 playing fields, 2 gymnasiums, 2 pools, 1 pond, 1:1 Chromebooks grades 5-8, 2 3-D printers, 1 drone, 1 GoPro, and Smart Boards, ipads, and laptops in the classrooms.


Welcome to the Bennett family. Meeting them gives the sense of being an "adopted" Bennett. Such a down to earth family with so many siblings. Awesome. This is the story of the oldest son, Sebastian.
Sebastian is a self made billionaire in the jewellery business. He is the CEO of the company with some of his siblings in supportive roles. He is not like many others in this position. He is very family orientated and he doesn't forget where he came from and how he got to where he is now.
Ava is a consultant for a reputable company and she is assigned to the Bennett for the next collection launch. When she walks in the office for the first time, she is pleasantly surprised. Sebastian and his brother Logan were in one of their banters. Both well built and incredibly good looking but it's Sebastian who makes her heart flutter. Sebastian has long given up on having relationships as most women look at the size of his wallet instead of him as a person. Ava stirs something in him and he sets out to find the reason why. He is persistent!
The more they interact, the stronger the attraction. Nothing can really come out of getting involved as Ava works under a strict no "fraternisation with clients rule". Doing so will cost her the job that she has worked so hard for. Besides, she is very independent and has her own plans on how she wants to settle down. Her past relationships have her left with little or no faith in men. Will they take a chance and throw cautions to the wind and see where a relationship might take them?
I fell in love with Sebastian and Ava as well as the rest of the Bennetts. This story is full of fun and laughter, uncertainties and insecurities. More importantly, love and loyalty. This is not the first book which I have read by this author and she manages once again to pull at my heart strings. I would fully recommend this book to other readers. Believe me the Bennett family is worth meeting. I await with bated breath for the next story, which I gather will be about Logan. Quite the lovable character!
Being rich and famous is not always as rosy as it looks. What the public is allowed to see and what really goes on behind closed doors can be totally different stories. Rock Wedding is a good example of such a case. It's also about how different individuals deal with grief.
Abe is the keyboard player of the Schoolboy Choir, a successful and famous rock band. He has suffered some losses which have left him angry as well as the fear of losing loved ones. He used drugs and alcohol to numb the pain, until he had a very close call.
Sarah married Abe at a young age and worshipped him but even then she felt lonely. When things got out of hand, she walked out and is now divorced. After what happened between them, she believes that he has never really been in love with her. Now that she is older and wiser, she has vowed to never put herself in a vulnerable position again.
In the previous book, Abe came across Sarah at a concert and witnessed something that made him really angry and is trying to look out for her. Having been bitten once, Sarah is not prepared to repeat the experience but Abe is a changed man. Convincing her is not going be so easy! Who can blame her?
When they were together, neither of them confided in the other about their pasts. Abe, because it was hard for him and Sarah, because she feared being misjudged for her unpleasant background. As they spend time together this time round, they start to open up to each other. Abe openly admits that he was the reason for their break up. If anything, he feels guilty for making her life so miserable, something that he intends to put right if only she would give him another chance.
Rock wedding is a very emotive and romantic story. Finding out about Sarah's past before she met Abe and what she went through and how she felt during and after their time together is devastating. The reasons why Abe behaved the way he did is just as heartbreaking. This journey to their second chance is a hard one. The myriad of emotions is very palpable. The connection between Sarah and Abe is electric. In some ways, reading this book is like going down memory lane. All the characters in the previous books are involved and seeing how far they have come since the beginning makes this series even more realistic.
The series is wrapped up beautifully but..... Ms Singh has indicated that there is more to come for Gabriel and Charlotte. Gabe's hunky brother will be at the forefront. Yay! Can't wait to read more from this author.
The author of the Sweet Evil trilogy from HarperTeen brings you a Young Adult modern folktale full of Irish magic and romance.
While most seventeen-year-old American girls would refuse to let their parents marry them off to a stranger, Robyn Mason dreams of the mysterious McKale in Ireland, wondering how he'll look and imagining his Irish accent. Prearranged bindings are common for magical families like her own, however when she travels to the whimsical Emerald Isle she discovers there's more to her betrothal and McKale's clan than she'd been led to believe.
Irish Folklore is not very often portrayed in Young Adult Novels and this is what makes See Me so interesting. It shows how being different leads to discrimination and brings lack of self esteem as a result.
Robyn is the oldest of the two Mason sisters. Instead of going off to college like all her other friends, she is leaving the US to go to Ireland. Not on an adventure but by obligation. Failing to do this could bring danger to her family.
McKale is a Leprechaun. Being taller than the rest of his clan makes him feel left out. He has grown up as a loner for lack of confidence and the feeling of not being able to fit in with the rest.
Robyn is from magical blood since her parents work for the fairies. When she was an infant, the consort to the king of the fairies took a shine to her and gave her parents no choice in a proposed binding. Coming up to her eighteenth birthday, Robyn is ready to honour this obligation. Although her parents have prepared her from a very young age, she only finds out about McKale's nature on their way to Ireland. Dread sets in, as she is very tall and being betrothed to a Leprechaun is not what she expected.
When McKale first meets Robyn, his shyness stops him from showing his reaction. He has always believed that his betrothal is a duty and his duty lies with his clan. However, as he gets to know Robyn, he realises that she is different from what he had imagined. Romance starts to blossom but the Shoe Mistress who is a fairy has other ideas, she wants McKale for herself and she always gets what she wants. She turns vicious and her threats can't be ignored. What she has in mind is completely ludicrous and selfish. Instantly McKale and Robyn are at a disadvantage and have to find a way to twart her plan.
The fairies treat the humans very badly and brainwash them in becoming slaves. They use their powers as a means of control.
This is an easy to read and beautiful story. It has love, laughter, loyalty, heartache and fun. The difficulties which Robyn and McKale have to face make their emotions feel real. I like the loving relationship in Robyn's family and her interactions with her sister, Cass. The Leprechaun's way of life gives a different insight to the modern world. I really enjoyed reading this novel and hope that we get to read the story of Cass and Rock!
Cass is a happily married school teacher. After a tough time looking after her mother, she is now beginning to enjoy life with her doting husband and the support of her long time friend, Rachel.
Matthew is happily married to Cass and works for the oil rigs. He appears to be the kind and considerate husband.
After a night out with her colleagues at the end of term, Cass is caught in a storm. Visibility is very poor through the heavy downpour. Although, she has been warned to stick to the main roads, she is worried about having an accident and takes the treacherous country road. She notices a car parked in a lay by and stops to see if the driver is in need of assistance. Only, the driver doesn't get out of her car or shows any sign of distress. Cass makes an informed decision to drive on and send for help when she gets home.
These experiences become more and more frequent and she becomes more and more distressed, worrying about her sanity. At least she has Matthew and Rachel for support! As luck would have it, she stumbles upon some totally baffling and upsetting information which might shed light on the murder and her own dilemma.
This novel is very well paced. The characters are very well portrayed. It's full of suspense, mystery and angst. Although a little predictable in places, the revelation at the end is just Wow! I didn't see that one coming. Totally worth reading and would appeal to lovers of Psychological Thrillers.
I was kindly issued with a complimentary eARC from the publisher via NetGalley and the views expressed are my personal opinion.
B A Paris is the internationally bestselling author of Behind Closed Doors, her debut novel. She was brought up in England and moved to France where she spent some years working in Finance before re-training as a teacher and setting up a language school with her husband. They still live in France and have five daughters. Her second novel, The Breakdown is out February 9th, 2017.
London, November 1965. Millions of Londoners are living and working on the fringes of the fashionable West End. Tales of the Moors murders fill the papers. Beyond the glamour of the all-white Carnaby Street scene lie other worlds of Caribbean music, Turkish coffee houses, Soho prostitution and stolen identities.
One Saturday evening, American star Iolanthe Green finishes a performance, walks out onto the Charing Cross Road and disappears. At first everybody cares where she went and then nobody does. A diverse group of emigre Londoners an Irish policeman, a Turkish coffee house owner and a Jamaican accountant are drawn together to search for a woman who's quickly being forgotten. They are led by Iolanthe's dresser, Anna Treadway a woman who knows quite a bit about reinvention and together they will travel into a world of underground music clubs, back street abortionists, seaside ghost towns and police brutality.
This story centres around life in London as it was in the 60s. It's about an actress who goes missing and what follows her disappearance.
Anna works at the Galaxy Theatre as a Dresser to the actress Iolanthe who is also known as Lanny. She lives in a flat above a Turkish Cafe, owned by Ottmar. She is an avid reader and not known for socialising.
When Lanny fails to turn up for work, alarms are raised as she is known for being punctual. The police starts an investigation into the disappearance but soon loses interest.
Brennan who is Irish, is the sergeant in charge of the case and has changed his first name to Barnaby to avoid discrimination. He first meets Anna when gathering information about Lanny and Anna realises that some of the information is not quite accurate. She is worried about her and decides to start her own investigations.
She goes to one of the nightclubs, last visited by Lanny where she meets the black accountant, Aloysius. It just so happens that he has some information which might lead to the whereabouts of Lanny. Anna enrols him for assistance and they embark on the most frustrating and difficult journey. They go from one place to another in the freezing cold whilst they try to piece things together. The reaction they receive is abismal. Aloysius is subjected to cruelty, violence and humiliation because of the colour of his skin, nothing else. The cruelty and ignorance are sickening and inhumane.
As the story unfolds, it becomes very apparent that Barnaby, Ottmar and Aloysius have one thing in common. Fighting to fit into society, amongst other issues! Lanny is not who she appears to be and as Anna learns more about her, she realises that they too have something in common. Something, well hidden from Anna's past. This only reinforces her determination to find Lanny. Aloysius is only too happy to assist despite the cruel treatment imposed upon him. He is also being blackmail even when he is wrongly accused of a crime that he didn't commit. It would appear that this is quite acceptable because of the colour of his skin!
It's a very poignant and palpable story. There is corruption, racism, illegal abortions, illicit transactions, poverty and class division. Family life is difficult. Relationships break down but true friendships prevail. It's very well portrayed and the characters are believable. I was engrossed and touched by this storyline and would recommend to other readers.
I am an author and dramatist. I grew up in London before studying English at Oxford and Playwriting Studies at Birmingham University. I now live in the beautiful Vale of Glamorgan with my husband and our two daughters.
My travel and food memoir Fragrant Heart was published by Summersdale in 2014. My first novel Miss Treadway and the Field of Stars will be published by 4th Estate (UK), Harper Books (US) and HarperCollins (Canada) in early 2017.
I'm a regular contributor to BBC Radio 4 (sometimes as Miranda Davies) and I'm currently undertaking a PhD in the history of BBC radio adaptation at Cardiff University where I'm attempting to catalogue everything that BBC Radio has ever adapted.
I'm represented for fiction and non fiction by Caroline Hardman at Hardman & Swainson.