Cadmium and copper change root growth and morphology of Pinus pinea and Pinus pinaster seedlings

Heavy metal loads in forest soils have been increasing over time due to atmospheric inputs. Accumulation in the upper soil layers could affect establishment of seedlings and forest regeneration. Mediterranean species show a high initial root development, allowing seedlings to reach the moisture of deeper soil layers. In the present work seedlings of stone pine ( Pinus pinea L.) and maritime pine ( Pinus pinaster Ait.), were grown in culture solution supplied with 0.0, 0.1, 1 or 5 μ M CdSO₄ or with 1 μ M CdSO₄ and 1 μ M CuSO₄ combined. In both species tap-root elongation was drastically reduced in the 5 μ M Cd²⁺ and in the (Cd²⁺+ Cu²⁺) treatments. A supply of 0.1 or 1 μ M Cd²⁺, however, enhanced root elongation in Pinus pinea without significantly influencing root elongation in Pinus pinaster . In both species the root density (weight per unit length) and the width of the cortex increased in response to Cd²⁺ exposure. In Pinus pinaster the mitotic index decreased at the higher Cd²⁺ concentrations and when Cd²⁺ and Cu²⁺ were combined. The data suggest that cell elongation is more sensitive to Cd²⁺ than cell division. The number and length of the lateral roots were also affected by Cd²⁺ treatment to a higher degree in Pinus pinaster than in Pinus pinea, reflecting the different Cd- tolerance of the two species.     

基于指数施肥法的一年生油松菌根化容器苗的供N研究

为了探讨一年生菌根化油松容器苗的最适苗木供N量和供N速率,采用4种不同指数施肥量进行了试验.通过测定苗木生物量和N含量,显示总供N量为80 mg·株-1的处理组在生物量积累和苗木N含量上显著优于其它各组（P〈0.05）;在100 d时,该处理可获得0.47 mg·株-1·d-1的N最适添加速率;在该速率下,苗木生物量、N含量分别可以达到845.60 mg·株-1和6.51 mg·株-1.根据生物量和N含量随着供N量增加的回归拟合结果,在107 d的生长过程中,67.96 mg·株-1至84.15 mg·株-1的供N总量可以使得一年生油松幼苗获得较高的生物量积累水平和N含量.     

Priority effects determine the outcome of ectomycorrhizal competition between two Rhizopogon species colonizing Pinus muricata seedlings

Competition is often considered a fundamental process influencing assemblage structure, yet little is known about competition among ectomycorrhizal (EM) fungi. Here, we examine competitive interactions between Rhizopogon occidentalis and Rhizopogon salebrosus in a 6-month microcosm study. Pinus muricata seedlings were grown in three EM treatments: R. occidentalis, R. salebrosus, and R. occidentalis and R. salebrosus. At 2, 4, and 6 months, five seedlings per treatment were harvested and the EM root tip biomass of each species was determined. Root tips in the two-species treatment were identified using molecular techniques. R. occidentalis had similar EM root tip biomass when grown alone or in the presence of R. salebrosus. By contrast, R. salebrosus had significantly lower EM root tip biomass when grown with R. occidentalis than when grown alone, indicating it was a competitive inferior under the conditions tested. Competition was driven by differences in timing of colonization resulting in a strong priority effect for R. occidentalis. Our results, together with two earlier studies, indicate competition may play a more important role in EM interactions than previously recognized.     

Role of auxin and gibberellin in regenerative differentiation of tracheids in Pinus pinea seedlings

A new experimental system was developed for studying the hormonal mechanisms which control tracheid differentiation. In this system the tracheids redifferentiated from parenchyma cells in the hypocotyl of young Pinus pinea L. seedlings. The experimentally induced tracheids have unique shapes and patterns, and are therefore easily distinguished from the primary and secondary tracheids formed before the experiments. Auxin (0·1–1% NAA) alone sufficed to cause the redifferentiation of short tracheids, usually in discontinuous patterns across the hypocotyl. Gibberellin by itself did not induce redifferentiated tracheids. Combinations of auxin with gibberellin (0·1% NAA+0·1–1% GA₃) promoted the differentiation of long tracheids (up to threefold greater than those induced by auxin) in continuous patterns along the stem axis. Gibberellin in the presence of auxin promoted tracheid elongation by stimulating intrusive growth of both the upper and lower ends of the differentiating tracheids. The role of auxin and gibberellin in controlling the evolution of tracheary elements, from tracheids to vessels and fibres, is discussed.     

Spatiotemporal variability of stone pine (Pinus pinea L.) growth response to climate across the Iberian Peninsula

Climate warming and increasing aridity have impacted diverse ecosystems in the Mediterranean region since at least the 1970s. Pinus pinea L. has significant environmental and socio-economic importance for the Iberian Peninsula, so a detailed understanding of its response to climate change is necessary to predict its status under future climatic conditions. However, variability of climate and uncertainties in dendroclimatological approach complicate the understanding of forest growth dynamics. We use an ensemble approach to analyze growth-climate responses of P. pinea trees from five sites along a latitudinal gradient in Spain over time. The growth responses to April-June precipitation totals were stronger in the north than in the south. Since the 1950s, the sensitivity of growth to April-June precipitation increased in the north and decreased in the south. Meteorological drought usually started in May in the southern sites, but in June-July in the northern sites. The water deficit in the southern sites is thus greater and more limiting for tree growth, and this likely accounts for the lower growth sensitivity during these months. Our results indicate that P. pinea has a high degree of plasticity, suggesting the species will withstand changing climatic conditions. However, growth response to drought regimes varies among P. pinea populations, suggesting that different populations have different capacities for acclimation to warmer and drier climate, and this may influence future vegetation composition.     

Contrasting response of seedlings of two tropical species Clusia minor and Clusia multiflora to mycorrhizal inoculation in two soils with different pH

Differences in mycotrophic growth and response to phosphorus (P) fertilization were studied in seedlings of two woody native species: Clusia minor L. and Clusia multiflora H.B.K. from a cloud montane forest of tropical America. Greenhouse investigation was undertaken to determine the relationships between mycorrhizal dependency of host species associated with P utilization and growth in two different soils contrasting in pH (acidic and neutral) and nutrient content. Four treatments were performed: sterilized soil; sterilized soil plus 375 mg/kg of triple superphosphate (TSP); sterilized soil inoculated with Scutellospora fulgida (20 g/pot); and sterilized soil plus S. fulgida and TSP, with 10 replications per treatment for the two species. Results showed that both Clusia species presented high growth response to increasing P availability, which indicates that the root morphology (magnolioid roots) of these species is not a limiting factor for the incorporation of P from soils. Plants inoculated with arbuscular mycorrhizal fungi (AMF) in acidic soil had significantly increased shoot and root biomass, leaf area and height, in comparison to the biomass of P-fertilized plants and nonmycorrhizal plants. In neutral soil, seedlings of C. minor and C. multiflora were negatively affected by inoculation with AMF. In contrast, a significant decrease in growth was observed when inoculated plants were compared with noninoculated plants on neutral soil. Results indicate that an increase in the availability of a limiting nutrient (P) can turn a balanced mutualistic relationship into a less balanced nonmutualistic one.     

马尾松菌根化苗木对干旱的生理响应及抗旱性评价

采用温室盆栽方法,研究了持续干旱及复水处理后,接种褐环乳牛肝菌7、牛肝菌1、鸡油菌、彩色豆马勃和土生空团菌的马尾松苗木生理变化,并对菌根化苗木进行抗旱性评价.结果表明:在持续干旱条件下,马尾松苗木的丙二醛和相对质膜透性随之增加,但菌根化苗木的丙二醛和相对质膜透性均显著低于未接种苗木(对照);复水后,菌根化苗木中丙二醛和质膜透性较对照迅速降低.在持续干旱胁迫前21 d,马尾松苗木超氧阴离子自由基产生速率增加,同时也诱导了菌根化苗木中过氧化物歧化酶、过氧化物酶和硝酸还原酶活性显著增加.随着胁迫时间的延长,苗木复水后的恢复能力各异.在胁迫14 d复水后,苗木过氧化物歧化酶、过氧化物酶和硝酸还原酶的活性均得以恢复.菌根化苗木抗旱性的强弱为牛肝菌7＞牛肝菌1＞鸡油菌＞土生空团菌＞彩色豆马勃.过氧化物歧化酶和丙二醛与马尾松菌根化苗木抗旱性关联度较大,可以作为抗旱鉴定的主要指标.     

Role of ethylene and auxin in regenerative differentiation and orientation of tracheids in Pinus pinea seedlings

A low auxin concentration (0.1% naphthalene acetic acid) induced tracheids with longitudinal polarity parallel to the hypocotyl axis in young Pinus pinea seedlings. Application of 0.1% ethrel laterally and 0.1% naphthalene acetic acid apically disturbed axial tracheid polarity and promoted the differentiation of tracheids with a lateral orientation. Ethrel by itself, with no auxin background, did not affect tracheid differentiation. Apical application of 1% gibberellic acid with the low auxin, reversed the polarity disorder induced by ethrel. Disturbance of axial tracheid polarity was observed under a high auxin concentration (0.5% naphthalene acetic acid) which was similar to the combined effect of ethrel and auxin. The high auxin concentration increased tracheid number significantly. This effect was curtailed following treatment with inhibitors of ethylene formation (Co²⁺; 1-aminoethoxy- vinylglycine) and action (Ag²⁺). The role of ethylene in controlling the differentiation of radial tracheids, which characterize the vascular rays of pines, is discussed.     

The effect of zeatin and iso-pentenyladenine on IAA transport from the shoot to the root of Pinus pinea seedlings

The tips of the tap roots of Pinus pinea seedlings were dipped in zeatin or iso-pentenyladenine solutions. Immediately after cytokinin application to the root tip or after a 24 h lag phase, [2-¹⁴C]IAA was applied to the shoot apex. Treating with zeatin resulted in an increase in [2-¹⁴C]IAA transport from the shoot to the root. Iso-pentenyladenine also caused a slight increase in transport of radioactivity to the root but this was less pronounced compared to the results obtained with zeatin. With zeatin treatment increasing amounts of radioactivity accumulated in the lateral root emerging zone of the tap root (Section III). This was in sharp contrast to the treatment with iso-pentenyladenine where little radioactivity accumulated in this section of the root. Recovery of radioactivity 48 h after applying [2-¹⁴C]IAA showed that 33% of the recovered radioactivity co-chromatographed with authentic IAA. The implications of the effect of different cytokinins on the distribution of radioactivity along the tap root of Pinus pinea following [2-¹⁴C]IAA application to the shoot are discussed.Abbreviations Z zeatin - iP iso-pentenyladenine - TCL thin-layer chromatography     

Effects of elevated nickel and cadmium concentrations on growth and nutrient uptake of mycorrhizal and non-mycorrhizal Pinus sylvestris seedlings

The effects of Ni and Cd on growth and nutrient uptake of mycorrhizal and non-mycorrhizal Pinus sylvestris L. seedlings were investigated in a pot experiment. Seedlings were either inoculated with Laccaria bicolor (Maire) Orton or left uninoculated before being planted in pots containing a mixture of sandy soil from the B-horizon of a coniferous forest, small stones and pure quartz sand. The pots were supplied with small amounts of a balanced nutrient solution every 24 h using peristaltic pumps. Nickel or Cd were added as chlorides to the nutrient solution at levels of 85 M Ni (Ni 1), 170 M Ni (Ni 2), or 8.9 M Cd. Mycorrhizal colonisation of the roots was nearly 100% in the mycorrhizal treatments. The mycorrhizal seedlings grew significantly better than the non-mycorrhizal ones. The weight of mycorrhizal seedlings in the Ni 2 treatment was 29% lower than that of the mycorrhizal controls, but still 34% greater than that of the non-mycorrhizal seedlings not exposed to metals. There was an overall, statistically significant, negative effect of metals on plant yield. Mycorrhizal plants had lower root:shoot (R:S) ratios than non-mycorrhizal plants and the R:S ratio was increased by metal exposure, particularly in the non-mycorrhizal seedlings. Plant concentrations of Cd or Ni were not affected by mycorrhizal colonisation, but total uptake of Cd and Ni was higher in bigger mycorrhizal seedlings. Nickel decreased P concentration in all seedlings and Cd decreased P concentration in the non-mycorrhizal seedlings. Generally, the mycorrhizal seedlings grew better than non-mycorrhizal ones and had better P, K, Mg and S status. Root growth was not significantly affected by the metal treatments. The reduction in mean shoot growth of non-mycorrhizal plants, relative to the metal-free control, appeared higher than in mycorrhizal plants but was not statistically significant due to high variation in the non-mycorrhizal plants not exposed to metals. The main mycorrhizal effect was thus increased nutrient uptake and growth of the seedlings.     

构树幼苗对接种丛枝菌根真菌的生长响应

利用石灰岩适生植物构树进行菌根真菌摩西球囊霉(GM)、地表球囊霉(GV)和透光球囊霉(GD)的单独接种(SI)、混合接种(CI)和不接种(N)处理试验,测定了构树幼苗生长3个月后的形态及生长指标.结果表明:接种VA真菌促进了宿主构树的生长,单株地上部分、地下部分生物量和全株生物量等生长指标均较对照组显著提高,4种处理下构树幼苗生物量较对照提高了2.49～8.19倍,其中CI处理生物量最大,而单株叶片数目与对照组没有显著差异.不同接种处理的构树幼苗生长响应不同.在SI处理中,GD对构树幼苗生长效应最大,而CI处理较SI处理对构树幼苗的促进效应更加明显.CI处理中地径、苗高和总叶面积分别是对照组的1.5、2.2和6.0倍.各处理中根冠比最大的为CI(0.446).说明宿主植物与菌种存在一定的选择性.     

Chlorophyll accumulation in cotyledons, hypocotyls and primary needles of Pinus pinea seedlings in light and dark

Seeds of Pinus pinea L. were germinated and grown in darkness or under natural photoperiodic (day/night) conditions for 8 weeks. Samples were taken for analysis at frequent intervals and chlorophyll (Chl) and protochlorophyllide (Pchlide) contents of cotyledons, hypocotyls and primary needles determined spectrophotometrically alter separating the esterified from unesterified pigments by partitioning between ammoniacal acetone and petroleum ether. Chlorophyll accumulated in cotyledons, hypocotyls and primary needles of dark as Well as light grown seedlings. The maximum Chl concentrations achieved in darkness in these organs were approximately 60, 20 and 10% respectively, of concentrations achieved in light. The tinsel of rapid Chl accumulation by cotyledons in darkness coincided with the emergence of the radicle from the seed. Light had no effect on the timing of this process or on the duration of the lag period. However, it extended the exponential growth phase of cotyledons by at least a week and. once the seedlings were about 2 weeks old, enabled Chl to accumulate at a faster rate. Time courses of Chi and Pchlide accumulation by cotyledons in darkness were similar. Pchlide did not accumulate prior to the onset of Chi accumulation and Chl slopped accumulating once the Pchlide pool reached maximum size. The implications of these findings are discussed and a hypothesis to account for the inability of primary needles of smaller seeded pines to accumulate Chl in darkness is proposed.     

Effects of mycorrhizal fungi inoculation and soil amendment with hydrogel on leaf anatomy,growth and physiology performance of olive plantlets under two contrasting water regimes

Abiotic stresses present a real environmental problem in agriculture field. In our paper, we examine the significance of arbuscular mycorrhizal fungi (AMF) and soil amendment with water retaining superpolymers (hydrogel) on growth and physiology performance of olive plantlets. Our experiment was carried out in nursery conditions, to test the impact of hydrogel (TH) and mycorrhizal fungi (TM), used individually or combined (THM), and compare them with non inoculated plants (TC), to understand and reduce the water stress damage in olive plantlets (cv. Chemlali). We also evaluate interactions between hydrogel, mycorrhizal treatments and water regimes. Results of mycorrhization (M%) show that roots colonized by Rhizophagus irregularis of well-watered plants were about 40.87%. In combined treatment (THM), M% was about 32.14%. Compared to TC treatment, TM treatment enhances significantly the dry weights of the whole plant under the two water regimes. The TM treatment had the highest relative water content (66.50%) and Chl (a?+?b) (0.83 mg g^??1) in stressed conditions. We found also that under water stress, the maximal quantum efficiency of the photosystem II measurements in leaves were significantly improved by 50.70% in TH treatment compared to control. For phenolic contents, TH treatment decreased significantly total phenols by 50.10% compared to TC. Our study gives evidence that the use of AMF and the hydrogel separately or in combination may enhance the capacity to avoid drought damages of olive plantlets and improve olive performances.     

Regulation of phosphoenolpyruvate carboxylase in Pinus halepensis needles submitted to ozone and water stress

Effects of ozone and/or drought stresses on phosphoenolpyruvate carboxylase (PEPc, EC 4.1.1.31) regulation in Pinus halepensis Mill. needles were assessed over 3 months in controlled conditions. Whereas moderated water stress applied to Aleppo pine had no effect on PEPc activity compared to the control, which was probably related to the high tolerance of this species to drought, ozone stress induced a dramatic increase of PEPc activity in pine needles. This stimulation of the anaplerotic pathway could provide substrates to repair processes, well known for being enhanced upon ozone exposure. The ozone-increased PEPc activity was related, to a certain extent, to an increase in protein and mRNA levels. The possible role of the stimulation of the phosphorylation status of the enzyme in the increased PEPc activity under ozone was also investigated. Following the demonstration of the existence of the phosphorylation site at the N terminal part of Aleppo pine PEPc, it was shown that, under ozone treatment, the light/dark PEPc activity ratio and the Ki (malate) for PEPc were increased. This strengthens the hypothesis of an ozone-related post-translational process, which could be part of an adaptation of the plants to prolonged stress. When ozone and water stress were applied in combination, the enhancement in PEPc activity was only related to changes in gene expression. This difference in PEPc regulation, compared to the effect of single stress, could be the consequence of the specific action of each stress on the enzyme. This study brings new insights into the regulation of PEPc in a C₃ plant, Aleppo pine under these stresses. A different regulatory mechanism of PEPc is occurring according to the stress. The physiological implications of the increase in PEPc activity in response to ozone and/or water stress are discussed.     

不同指数施肥方法下长白落叶松播种苗的需肥规律

为了探讨长白落叶松苗木的最适N含量和供N速率,应用3种不同指数施肥方法在大田条件下进行试验。结果表明:经过111d的生长,供N总量为30mg.株-1的指数施肥处理下的苗木可以获得相对较高的生物量积累和N含量水平,该处理还可以使得苗木在103d的生长中获得0.29 m.g株-.1d-1的N添加速率。在该速率下,苗木生物量、N含量和N吸收效率分别可以达到637.80 m.g株-1、8.10 m.g株-1和41.77%。根据生物量和N含量随着供N量增加的二项式回归拟合结果,在111d的生长过程中24.3—33.7m.g株-1的供N总量可以使得苗木获得较高的生物量积累水平和N利用效率。试验设置的总供N量为30 m.g株-1的处理优于60 m.g株-1或70 m.g株-1。     

土壤水分变化对长白山主要树种蒙古栎幼树生长的影响

选择长白山红松阔叶林主要优势树种蒙古栎为研究对象,人工控制3种施水量研究蒙古栎幼树形态、生物量效应和光合生理特征对土壤含水量变化的响应．结果表明,不同土壤含水量变化显著影响蒙古栎叶片、枝、根的生物量及其分配格局和叶片光合气体交换特征．水分胁迫改变幼树树冠结构,抑制幼树树高、地径、叶片大小、地上和地下生物量;同时,蒙古栎幼树根冠生物量比随着土壤水分含量的减少显著提高;供水量减少对幼树净光合速率、CO2利用率和碳利用率等特征有显著的负向影响;而叶片气孔导度、蒸腾速率和水分利用率对不同土壤含水量反应较复杂,只在土壤含水量较低时,幼树气孔导度、蒸腾速率明显降低,叶片水分利用率升高,表现出蒙古栎树种是干旱可变植物,长期水分胁迫可提高树种的耐旱能力．     

红松幼苗对CO_2浓度升高的生理生态反应

研究了用开顶箱控制CO2 浓度在 5 0 0和 70 0 μmol·mol-1左右时红松幼苗的生理生态反应 .结果表明 ,高浓度CO2 (5 0 0、70 0 μmol·mol-1CO2 )和对照 (对照开顶箱、裸地 )条件下 ,红松幼苗的净光合速率与气孔导度之间的变化不同 .红松幼苗在 5 0 0 μmol·mol-1CO2 条件下 ,RuBPcase活性最高 ,呈现光合上调反应 ,日平均净光合速率最大 ,叶绿素及可溶性糖含量最高 ;而生长在 70 0 μmol·mol-1CO2 的红松幼苗呈现光合下调反应 ,光合作用明显低于对照植株 ,其酶活性及物质含量均最低     

Growth response of drought-stressed Pinus sylvestris seedlings to single- and multi-species inoculation with ectomycorrhizal fungi

Many trees species form symbiotic associations with ectomycorrhizal (ECM) fungi, which improve nutrient and water acquisition of their host. Until now it is unclear whether the species richness of ECM fungi is beneficial for tree seedling performance, be it during moist conditions or drought. We performed a pot experiment using Pinus sylvestris seedlings inoculated with four selected ECM fungi (Cenococcum geophilum, Paxillus involutus, Rhizopogon roseolus and Suillus granulatus) to investigate (i) whether these four ECM fungi, in monoculture or in species mixtures, affect growth of P. sylvestris seedlings, and (ii) whether this effect can be attributed to species number per se or to species identity. Two different watering regimes (moist vs. dry) were applied to examine the context-dependency of the results. Additionally, we assessed the activity of eight extracellular enzymes in the root tips. Shoot growth was enhanced in the presence of S. granulatus, but not by any other ECM fungal species. The positive effect of S. granulatus on shoot growth was more pronounced under moist (threefold increase) than under dry conditions (twofold increase), indicating that the investigated ECM fungi did not provide additional support during drought stress. The activity of secreted extracellular enzymes was higher in S. granulatus than in any other species. In conclusion, our findings suggest that ECM fungal species composition may affect seedling performance in terms of aboveground biomass.