Effects of soil-applied lead on seedling growth and ectomycorrhizal colonization of loblolly pine

Six-month-old loblolly pine (Pinus taeda) were grown for 15 weeks in two native soils amended with 0, 30, 60, 120, 240 or 480 mg kg(-1) Pb as PbCl2. Ectomycorrhizae were quantified, by morphotype, as the total number of tips per centimeter, and as the number of tips for each morphotype and for Cenococcum geophilum. Total numbers of non-ectomycorrhizal short roots and necrotic tips were recorded. Total height and biomass exhibited a non-linear response to soil-applied lead. Growth generally was greatest in the controls and higher treatments, and least in the intermediate treatments. In both soils, Pb concentrations in roots increased linearly with increasing levels of soil-applied Pb. Neither foliage nor stems exhibited significant increases in Pb concentrations with increasing levels of Pb. Significant linear decreases in total numbers of ectomycorrhizal tips and significant linear increases in non-ectomycorrhizal short roots and necrotic tips occurred with increasing levels of Pb in the soil. The majority of individual morphotypes decreased with increasing Pb. However, the number of ectomycorrhizal tips formed by C. geophilium increased with increasing soil Pb levels after 15 weeks of treatment. These results indicate that short-term loblolly pine seedling growth is not inhibited by increasing Pb levels. Ectomycorrhizal formation decreased, and alterations in species composition occurred as a result of increasing concentrations of soil-applied Pb. These effects on ectomycorrhizae may cause long-term changes in nutrient and water balances, which could reduce tree vigor.     

Growth, ectomycorrhizae and nonstructural carbohydrates of loblolly pine seedlings exposed to ozone and soil water deficit

Loblolly pine (Pinus taeda) seedlings from three full-sib families were exposed to 0, 50, 100 or 150 ppb ozone (O(3)) (5 h/d, 5 d/week for 6 or 12 weeks). Soil water potential was maintained near pot capacity (-0.03 MPa) or soil was allowed to dry to approximately -1.0 MPa and resaturated. Chlorotic mottling and flecking of needles due to O(3) injury were observed for seedlings from all pine families. Soil water deficit lessened the intensity of O(3) symptoms, possibly due to stomatal closure. Exposure to O(3) and soil water deficit each resulted in less seedling volume growth and dry weight, and changed the nonstructural carbohydrate content of seedlings compared with controls. Increasing O(3) concentrations resulted in a linear reduction in foliar starch content but did and affect hexose or sucrose content. Soil water deficit resulted in less starch and soluble sugar contents in above- and below-ground plant parts compared with controls. Soil water deficit did not affect numbers or percentages of roots that formed ectomycorrhizal tips. A linear dose-response relationship between O(3) and ectomycorrhizae was observed. The number of ectomycorrhizal tips/cm long root and the percentage of feeder roots that formed ectomycorrhizae were lower as O(3) concentration increased. Overall, each stress alone caused less seedling growth and carbohydrate content compared with controls, but only O(3) was responsible for suppression of ectomycorrhizae.     

Ectomycorrhizal colonization of loblolly pine seedlings during three growing seasons in response to ozone, acidic precipitation, and soil Mg status

A three-year study was initiated in 1987 to evaluate the impact of O3, acidic precipitation, and soil Mg on ectomycorrhizal colonization of loblolly pine (Pinus taeda L.) seedlings. Thirty-six open-top chambers equipped with a rainfall exclusion/addition system were utilized to administer three levels of O3 (subambient, ambient, or twice ambient) and two precipitation acidity levels (pH 3.8 or 5.2) to seedlings growing in 24-liter plastic pots containing soil having either 35 or 15 mg kg(-1) of exchangeable Mg. Seedlings exposed to the twice ambient O3 treatment exhibited smaller percentages of total ectomycorrhizal short roots at the end of each year of the study, but trends were statistically significant in 1989 only. Changes in number of specific ectomycorrhizal morphotypes in response to O3 were not consistent from year to year. Acidic precipitation treatments had no effect on number or percent of mycorrhizal short roots, and responses of two morphotypes to soil Mg treatments were probably due to differences in the soil environment rather than a result of changes in aboveground processes. Temporal shifts in morphotype frequencies were observed for seedlings in all treatments and indicate that mycorrhizal succession occurred during the study period.     

Responses of microbial populations in the rhizosphere to deposition of simulated acidic rain onto foliage and/or soil

Air pollutants or some chemicals applied to plant foliage can alter the ecology of the rhizosphere. Experiments were conducted to distinguish among possible foliage-mediated versus soil- or root-mediated effects of acid deposition on microorganism in the rhizosphere. Seedlings of a sorghum x sudangrass hybrid in pots of non-sterile soil-sand mix in a greenhouse were exposed to simulated rain solution adjusted with H2SO4 + HNO3 to pH 4.9, 4.2, 3.5 or 2.8. Solutions were applied as simulated rain to foliage and soil, foliage only (soil covered by plastic, and deionized water applied directly to the soil), or soil only (solution applied directly to the soil). Solutions were applied on 16 days during a 6-week period (1.5 cm deposition in 1 h per application). Plant shoot and root dry weights and population densities of selected types of bacteria, filamentous actinomycetes and fungi in the rhizosphere were quantified after exposures were completed. Deposition of simulated acidic rain onto foliage alone had no effect on plant biomass or microbial population densities in the rhizosphere (colony-forming units per gram of rhizosphere soil). However, plant growth was stimulated and all microbial populations in the rhizosphere increased 3- to 8-fold with increased solution acidity (relative to pH 4.9 solution) when solution penetrated the soil. Statistical analyses indicated that the acid dose-population response relationships for soil-only and foliage-and-soil applications were not different. Thus, no foliage-mediated effect of simulated acidic rain on rhizosphere ecology was detected.     

Effects of ozone and simulated acid rain on birch seedling growth and formation of ectomycorrhizae

Four-week-old paper birch (Betula papyrifera Marsh.) seedlings, inoculated or non-inoculated with the ectomycorrhizal fungus Pisolithus tinctorius (Pers.) Coker & Couch and grown in steamed or non-steamed soil, were exposed to ozone (O(3)) and/or simulated acid rain (SAR). Plants were exposed to O(3) for 7 h per day on 5 days per week for 12 weeks. O(3) concentrations were maintained between 0.06 and 0.08 ppm. SAR was applied 10 min per day on 2 days per week. O(3), SAR, soil regime and mycorrhizal treatment did not significantly affect any of the measured variables. Interactions between O(3) and SAR, SAR and mycorrhizal treatment, soil regime and mycorrhizal treatment and ozone and soil regime had significant effects. Treatment of seedlings with pH 3.5 SAR caused increases in growth which were more apparent in birch exposed to O(3). Mucorrhizal treatment caused increased growth in non-steamed soil, while growth appeared to decrease in steamed soil. Birch seedlings grew much better in steamed soil. The implications of increased growth in steamed soil may demonstrate the importance of looking at the secondary effects of pollutants on soil-borne organisms.     

Effects of rhizobium, arbuscular mycorrhizal fungi and anion content of simulated rain on subterranean clover

An experiment was conducted to determine the extent to which rhizobia, mycorrhizal fungi, and anions in simulated rain affect plant growth response to acid deposition. Germinating subterranean clover seeds were planted in steam-pasteurized soil in pots and inoculated with Rhizobium leguminosarum, Glomus intraradices, Glomus etunicatum, R. leguminosarum + G. intraradices, R. leguminosarum + G. etunicatum, or no microbial symbionts. Beginning 3 weeks later, plants and the soil surface were exposed to simulated rain in a greenhouse on 3 days week(-1) for 12 weeks. Rain solutions were deionized water amended with background ions only (pH 5.0) or also adjusted to pH 3.0 with HNO3 only, H2SO4 only, or a 50/50 mixture of the two acids. Glomus intraradices colonized plant roots poorly, and G. intraradices-inoculated plants responded like nonmycorrhizal plants to rhizobia and rain treatments. Variation in plant biomass attributable to different rain formulations was strongest for G. etunicatum-inoculated plants, and the effect of rain formulation differed with respect to nodulation by rhizobia. The smallest plants at the end of the experiment were noninoculated plants exposed to rains (0.38 g mean dry weight total for 3 plants pot(-1)). Among nonnodulated plants infected by G. etunicatum, those exposed to HNO3 rain were largest, followed by plants exposed to HNO3 + H2SO4, pH 5.0, and H2SO4 rain, in that order. Among plants inoculated with both R. leguminosarum + G. etunicatum, however, the greatest biomass occurred with pH 5.0 rains, resulting in the largest plants in the study (1.00 g/3 plants). Treatment-related variation among root and shoot biomass data reflected those for whole-plant biomass. Based on quantification of biomass and N concentrations in shoot and root tissues, total N content of plants inoculated with G. etunicatum alone and exposed to the HNO3 + H2SO4 rains was approximately the same as plants inoculated with R. leguminosarum + G. entunicatum and exposed to pH 5 rains. Thus, the acid-mixture rains and rhizobia under no acid deposition provided approximately equal amounts of N in biomass. The significant interactions among rain formulation and the symbiotic status of the plants suggest that conclusions concerning the impact of acid deposition on plants in the environment cannot be considered reliable because most experiments on which such assessments are based have not tested confounding influences of microorganisms and precipitation characteristics.     

Exposure of two upland plant species to acidic fogs

A system is described for exposing large numbers of plants to acidic fogs. The system allows low volumes of treatment solutions to be provided at particle sizes chiefly in the 5-30 microm range (equivalent to fog/cloud droplets). Plants of Poa alpina L. and Epilobium brunnescens were propagated from material collected in Snowdonia, North Wales and exposed to fog treatments at pH values of 2.5, 3.5, 4.5 and 5.6. There were 3 x 4 h exposures per week which provided a total of 6 mm deposition. Supplementary watering was with pH 4.5 simulated acid rain (24 mm per week). After 21 weeks, there was increased lowering and a greater dry weight for plants of E. brunnescens exposed to the pH 2.5 fog in comparison with other treatments. Also, the plants used assimilated material to form shoots rather than roots. A similar increase in dry weight accumulation in the pH 2.5 treatment was found in P. alpina after 63 weeks but this was not associated with changes in assimilate partitioning.     

The individual and combined effects of ozone and simulated acid rain on growth, gas exchange rate and water-use efficiency of Pinus armandi Franch

The seedlings of Pinus armandi Franch. were exposed to ozone (O(3)) at 300 ppb for 8 h a day, 6 days a week, and simulated acid rain of pH 3.0 or 2.3, 6 times a week, alone or in combination, for 14 weeks from 15 June to 20 September 1993. The control seedlings were exposed to charcoal-filtered air and simulated rain of pH 6.8 during the same period. Significant interactive effects of O(3) and simulated acid rain on whole plant net photosynthetic rate were observed, but not on other determined parameters. The exposure of the seedlings to O(3) caused the reductions in the dry weight growth, root dry weight relative to the whole plant dry weight, net photosynthetic rate, transpiration rate in light, water-use efficiency and root respiration activity, and increases in shoot/root ratio, and leaf dry weight relative to the whole plant dry weight without an appearance of acute visible foliar injury, but did not affect the dark respiration rate and transpiration rate in the darkness. The decreased net photosynthetic rate was considered to be the major cause for the growth reduction of the seedlings exposed to O(3). On the other hand, the exposure of the seedlings to simulated acid rain reduced the net photosynthetic rate per unit chlorophyll a + b content, but did not induce the significant change in other determined parameters.     

Mycorrhizal populations and fine root development on Norway spruce exposed to controlled doses of gaseous pollutants and simulated acidic rain treatments

Effects after long-term fumigation and simulated acidic rain (pH 4.0) treatment in open-top chambers (OTC) during 5 vegetation periods on ectomycorrhizal (EM) frequency, fine root structure and distribution of short roots were recorded for Norway spruce. Simultaneous application of ozone and SO(2) resulted in a significant decrease (P< or =0.01) in the proportion of EM root tips and an increase in the numbers of necrotic short roots. The observed differences were partly due to more than additive effects of ozone and SO(2). On the basis of EM and non-mycorrhizal (NM) short roots in the irrigated controls (OTC-V, pH 5.6, Ah-horizon: EM + NM/cm=0.05) the acid treatments resulted in drastic effects (P< or =0.05) on the branching density and the proportion of EM root tips and NM short roots. Relative numbers of short roots infected by indigenous EM fungi with trees in the fumigation treatments at pH 4.0 were similar: 1.5-1.9 EM tips/cm and differed noticeably from the control: 5.2 EM tips/cm. Again, increasing numbers of necrotic and NM short roots were noted. Parallel measurements by the ingrowth-core technique confirm the argument, that EM formation seems to be a reliable indicator of pollution-induced stress.     

Shift in ectomycorrhizal community composition in Scots pine (Pinus sylvestris L.) seedling roots as a response to nickel deposition and removal of lichen cover

Scots pine seedlings were exposed to wet-deposited nickel (Ni) and removal of lichen cover in a dry heath Scots pine forest. Ni deposition affected the colonization of roots by indigenous ectomycorrhizal fungi in contrasting ways in intact and skimmed quadrats. Highest frequencies of tubercle morphotypes of ectomycorrhiza were found in quadrats exposed to 100 mg m(-2) year(-1) Ni in lichen covered treatment, while in skimmed quadrats these peaked after the treatment with 10 mg Ni m(-2) year(-1). Removal of the lichen layer increased the value of diversity index (H') of ectomycorrhizal fungal community, probably due to the increase in the evenness of the morphotype distribution. Lichen removal seemed also to improve the condition of the short roots, as the frequencies of poor and senescent short roots were decreased by the removal.     

Yields of field-grown soybeans exposed to simulated acidic deposition

Rainfall acidity response functions for crop yield and growth are necessary to predict the overall impacts on crop yields of ambient and/or anticipated levels of acidic rain. The experiment described herein was performed at Brookhaven National Laboratory during 1984 and 1985 to determine the effects of simulated rainfalls of pH 5.6, 4.4, 4.1 and 3.3 on seed yields of four cultivars of field-grown soybeans (Glycine max Merrill). Soybeans were chosen because previous results suggested that certain varieties of this crop are sensitive to rainfall acidity and because they are an economically important crop in the USA. Sixteen plots per treatment were used. Plants were grown using standard agronomic practices under automatically movable exclusion shelters which minimize changes in the plant's microclimate. Analysis of variance of seed yields of the four cultivars showed significant treatments for all four cultivars. For all cultivars, seed yields were lower when exposed to simulated rainfalls of pH 4.4, 4.1 and 3.3 compared with rainfalls of pH 5.6. Compared with pH 5.6 rainfall (controls), yields of Amsoy exposed to pH 4.4, 4.1 and 3.3 were lower by 13, 11 and 12%, respectively. The percentage yield reductions for Asgrow, Corsoy and Hobbit were not as great as those of Amsoy. Averaged over all cultivars tested, there was a 9% seed reduction for plants exposed to simulated rain at pH 4.1 (comparable to ambient) compared with pH 5.6.     

Response of sugar maple to multiple year exposures to ozone and simulated acidic precipitation

Potted sugar maple seedlings were exposed to ozone and acidic precipitation in open-top chambers for three consecutive growing seasons. Periodic measurements of photosynthesis, dark respiration, through-fall and soil solution chemistry, and annual measurements of the weight of plant parts were made. Experimental treatments caused few and minor effects on above- or below-ground growth of the seedlings, even after three growing seasons. There were trends for reduced photosynthesis in trees exposed to elevated concentrations of ozone and increased photosynthesis in those exposed to the lowest pH simulated rain treatment. The chemistries of soil-solutions and through-fall were not altered significantly by treatment. Although major effects were not observed, sugar maple may respond to exposures that take place over a significant part of its life cycle.     

Effects of acid mist on mature grafts of Sitka spruce. Part I. Frost hardiness and foliar nutrient concentrations

Mature grafts of five clones of Sitka spruce (Picea sitchensis Bong. Sarg.) were exposed to simulated acid mist composed of an equimolar mixture of sulphuric acid and ammonium nitrate at pH 2.5 and pH 5.0 in open-top chambers from May to November 1991. Treatments were applied on consecutive days, four times a week. The pH 2.5 treatment provided an overall dose three times higher than that received by forests in upland areas of Britain. Frost hardiness was assessed in November by freezing detached current year shoots at a range of temperatures and assessing the rate of electrolyte leakage Foliar nutrient concentrations were determined on the same shoots. Acid mist at pH 2.5 significantly reduced frost hardiness in four of the five clones; the temperature causing 50% shoot death (LT50) was increased by 0 to 7 degrees C. The clones varied in their level of hardiness, one clone being exceptionally frost sensitive. The frost hardiness of the frost sensitive clone was found to be less perturbed by acid mist than the hardiness of the more frost resistant clones. Mature grafts showed a smaller reduction in hardiness at an equivalent dose than that found previously with Sitka spruce seedlings. Compared with seedlings, grafts had lower absolute concentrations of foliar sulphur. Exposure to acid mist at pH 2.5 increased %S in current year foliage by <0.05% compared with absolute increases of more than 0.10% in current year foliage of seedlings. We conclude that the effect of acid mist on frost hardiness is likely to be less on mature trees than on seedlings and that the increased frost risk to mature trees of Sitka spruce from occult deposition alone is small.     

Shift in ectomycorrhizal community composition in Scots pine (Pinus sylvestris L.) seedling roots as a response to nickel deposition and removal of lichen cover

Scots pine seedlings were exposed to wet-deposited nickel (Ni) and removal of lichen cover in a dry heath Scots pine forest. Ni deposition affected the colonization of roots by indigenous ectomycorrhizal fungi in contrasting ways in intact and skimmed quadrats. Highest frequencies of tubercle morphotypes of ectomycorrhiza were found in quadrats exposed to 100 mg m⁻² year⁻¹ Ni in lichen covered treatment, while in skimmed quadrats these peaked after the treatment with 10 mg Ni m⁻² year⁻¹. Removal of the lichen layer increased the value of diversity index (H′) of ectomycorrhizal fungal community, probably due to the increase in the evenness of the morphotype distribution. Lichen removal seemed also to improve the condition of the short roots, as the frequencies of poor and senescent short roots were decreased by the removal.     

Impact of ozone, acid mist and soil characteristics on growth and development of fine roots and ectomycorrhiza of young clonal Norway spruce

As part of the joint 14-month exposure experiment on Norway spruce (Picea abies (L.) Karst.) sensitivity to pollution (two levels of ozone plus acid mist) of growth and development of the fine-root system and of mycorrhizae, have been investigated in two forest soils from areas showing forest decline. This study shows that differences in fine-root biomass and the occurrence of species of ectomycorrhizae were mainly due to prevailing conditions within the acid or calcareous substrate. The pollution treatment resulted in higher numbers of short root tips in only one soil, whereas the percentage of ectomycorrhizal roots with a well-developed mantle was low (10-23%) in both soils, irrespective of exposure of trees to ozone and acid mist. There was no consistent response, with the two clones examined, in terms of mycorrhizal frequency, beaded short roots and renewed growth. Regarding the effects on root growth, data cannot be used unrestrictedly for extrapolation to a more complex field situation.     

Influence of acidity level in simulated rain on disease progress in four plant pathosystems

Field investigations were performed in 1984-1985 on the effects of acidity level in simulated rain on disease dynamics in four pathosystems: alfalfa leaf spot (ALS), peanut leaf spot (PLS), potato late blight (PLB), and soybean brown spot (SBS). Studies were performed in an ambient rain exclusion apparatus with simulated rain acidity levels of pH 2.8, 3.6, 4.2, 4.8, or 5.6 and four plots per pH level. In 1984 for ALS and PLB, rain was simulated three times a week at 6-8 mm per event. For PLS and SBS in 1984 and for PLB, PLS, and SBS in 1985, rain was simulated twice a week at 13-16 mm per event with a 10 min pause halfway through each event. Disease was assessed three times a week. In 1984, no significant effect of acidity level in simulated rain on disease severity was detected in any pathosystem. Severity of PLB differed among treatments in 1985 with significant quadratic and cubic components for the dose-response relationship. PLS severity decreased with increasing level of acidity in simulated rain in 1985 and the dose-response relationship was linear. No differences in severity of ALS or SBS due to acidity of simulated rain were found. Thus, disease response to acidity of simulated rain is system dependent.     

Acid rain and pollen germination in corn

The properties of an acid rain episode that could influence the germination of pollen in corn, Zea mays L., were evaluated by treating silks with a simulated acid rain and measuring the subsequent germination of pollen on the silks. The data indicated that acid rain creates an inhospitable environment for pollen germination on the silk surface. Reduced germination appeared directly related to the acidity of the rain, but not the sulphate concentration. Rinsing silks with a pH 5.6 rain after treatment with a pH 2.6 rain did not increase pollen germination above that on silks treated only with a pH 2.6 rain, suggesting the reduced germination was due to physical and/or chemical modifications of the silk surface and not to residual acid on the tissue. Pollen germination on silks was inhibited even when silk tissue was exposed to a simulated rain of pH 2.6 for <1.5min.     

Effects of simulated acid rain and ozone on foliar chemistry of field-grown Pinus ponderosa seedlings and mature trees

We investigated the additive and interactive effects of simulated acid rain and elevated ozone on C and N contents, and the C:N ratio of one-year-old and current-year foliage of field-grown mature trees and their half-sib seedlings of a stress tolerant genotype of ponderosa pine. Acid rain levels (pH 5.1 and 3.0) were applied weekly to foliage only (no soil acidification or N addition), from January to April, 1992. Plants were exposed to two ozone levels (ambient and twice-ambient) during the day from September 1991 to November 1992. The sequential application of acid rain and elevated ozone mimicked the natural conditions. Twice-ambient ozone significantly decreased foliar N content (by 12-14%) and increased the C:N ratio of both one-year-old and current-year foliage of seedlings. Although similar ozone effects were also observed on one-year-old foliage of mature trees, the only statistically significant effect was an increased C:N ratio when twice-ambient ozone combined with pH 3.0 rain (acid rain by ozone interaction). Enhancing the effect of twice-ambient ozone in increasing the C:N ratio of one-year-old foliage of mature trees in June was the only significant effect of acid rain.     

A Collaborative Effort to Model Plant Response to Acidic Rain

Radish plants were exposed three times per week to simulated acidic rain at pH values of 2.6 to 5.4 over the course of four weeks in trials performed at Argonne, Illinois; Ithaca and Upton, New York; Corvallis, Oregon; Oak Ridge, Tennessee; and Toronto, Canada. Uniform genotype, soil media and planting techniques, treatment procedures, biological measurements, and experimental design were employed. Growth of plants differed among trials as a result of variation in greenhouse environmental conditions according to location and facilities. Larger plants underwent greater absolute but lower relative reductions in biomass after exposure to the higher levels of acidity. A generalized Mitscherlich function was used to model the effects of acidity of simulated rain or dry mass of hypocotyls using data from three laboratories that performed duplicate trials. The remaining data, from three other laboratories that performed only one trial each, were used to test the model. When the laboratory by trial effect was removed (influence of different growth. conditions), lack of fit to the Mitscherlich function was insignificant. Thus, a single mathematical model satisfactorily characterized the relationship between acidity and mean plant response. The pH value associated with a 10 percent reduction in mass was 3.3 ± 0.3 for hypocotyls. No value was estimated for shoots because effects oh shoots were not significant. The results of this study demonstrate that a generalized exposure-response model can be developed in the presence of large variations in environmental conditions when plant culture and exposure to simulated rain are standardized among laboratories.     

Antioxidant responses to simulated acid rain and heavy metal deposition in birch seedlings

This study measured the responses of different anti-oxidants in 2-year-old birch (Betula pendula Roth) seedlings subjected to simulated acid rain (pH 4.0) and heavy metals (Cu/Ni), applied alone or in combination for 2 months. The applied concentrations of pollutants did not significantly affect seedling biomass or total glutathione levels. Acid rain alone increased superoxide dismutase (SOD) activity both in leaves and roots, while heavy metals alone inhibited SOD activity in roots. Both acid rain and heavy metals applied singly increased ascorbate peroxidase (APX) and guaiacol peroxidase (GPX) activities in leaves but decreased activities in roots. In contrast, acid rain and heavy metal treatments increased glutathione reductase (GR) activity in roots but not in leaves. Spraying birch seedlings with a mixture of acid rain and heavy metals increased SOD, APX and GPX activities in leaves and GR activity in roots. However, the effects of mixed pollutants on enzyme activities usually were less than the summed effects of individual pollutants. Enzyme responses also depended on where pollutants were applied: spraying pollutants onto the shoots initiated higher responses in SOD, APX and GPX than did application to the soil surface, while the opposite was true for GR.