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.     

Effects of simulated rain acidity on ectomycorrhizae of red spruce seedlings potted in natural soil

Formation of ectomycorrhizae of red spruce (Picea rubens) grown in natural soil was measured after seedlings were exposed to 25 or 50 applications of simulated rain of pH 5.5, 3.5 or 2.5. Ectomycorrhizae were quantified as the total number of ectomycorrhizal tips per centimeter, and as the number of ectomycorrhizal tips for each morphotype and for Cenococcum geophilum. Rain solutions were applied to the soil alone, to foliage and stem alone, or to entire potted seedlings. Final soil pH was linearly related to rain solution acidity. Lower base saturation, calcium and zinc content, and higher exchangeable acidity were observed after pH 2.5 treatments if the soil was exposed. Rain solutions and the subsequent changes in soil characteristics did not affect the total numbers of ectomycorrhizal tips. Four morphotypes of ectomycorrhizae observed for these seedlings were unaffected by simulated rain. However, the numbers of ectomycorrhizal tips formed by C. geophilum tended to increase with rain solution acidity after 50 applications. Method of rain deposition did not affect ectomycorrhizae, suggesting both plant and soil mediated responses may favor certain mycobionts. The results of this study indicate that short-term acidic deposition does not induce significant changes in the frequency of ectomycorrhizae, but higher numbers of C. geophilum tips suggest there may be changes in the relative occurrence of specific morphotypes of fungus species.     

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.     

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.     

Urban polluted forest soils induce elevated root peroxidase activity in Scots pine (Pinus sylvestris L.) seedlings.

Plant biomass. mycorrhizal status and root peroxidase activity were measured in ectomycorrhizal Scots pine (Pinus sylvestris L.) seedlings grown in urban polluted and native, non-polluted forest soils with added ammonium or potassium sulphates simulating N and S deposition of urban areas. Peroxidase activity in the fine roots of seedlings planted in polluted forest soils was higher than in those planted in non-polluted soils and correlated positively with the activities measured in an earlier study in the roots of mature Scots pines growing at the sites from where the soils were collected. Growth of seedlings and mycorrhizal status were not affected by the origin of soil. Exposing the seedlings to winter acclimation conditions for 6 weeks elevated peroxidase activity in the roots. The addition of ammonium or potassium sulphate to non-polluted soils did not induce elevated root peroxidase activity, although at the levels of 0.5 and 1.0 g of ammonium sulphate a slight increasing trend was observed. We suggest, that indirect biotic factors, i.e. changes in the community structure of soil fungi, early stages of recognition, and defence reactions of pine roots against saprophytic and pathogenic fungi may be participating in the elicitation of peroxidase (POD) activity, although the possible role of heavy metals cannot be excluded.     

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.     

Assessment by laboratory simulation of approaches to amelioration of peat acidification

This study was conducted to determine whether acidic cloudwater and ozone (O3) influence the growth of red spruce (Picea rubens L.) seedlings growing at a high elevation site in the southern Appalachian Mountains. A field exclusion chamber study was established at Whitetop Mountain, VA (elevation 1689 m) which included the following treatments: (1) clouds and O3 excluded (COE); (2) exposure to ambient O3 with clouds excluded (CE); (3) exposure to clouds and O3 (CC); and (4) ambient air plots (AA) that served as a control to evaluate possible chamber effects. After 2 years, seedlings exposed to ambient levels of O3 and cloudwater (AA and CC) did not differ in biomass accumulation, diameter growth, or epicuticular wax amounts from seedlings grown in chambers where pollution levels were reduced (CE and COE). Treatments receiving cloudwater (AA and CC) had statistically lower current-year needle concentrations of Ca and Mg, indicating that the cloudwater exposure dynamics occurring at this site elicited reductions in needle Ca and Mg. Ozone had negligible impact on all of the seedling parameters measured.     

Growth and physiology of Northern Red Oak: preliminary comparisons of mature tree and seedling responses to ozone

Considerable progress has been made during the past decade in the development of mechanistic models that allow complex chemical, physical, and biological processes to be evaluated in the global change context. However, quantitative predictions of the response of individual trees, stands, and forest ecosystems to pollutants and climatic variables require extrapolation of existing data sets, derived largely from seedling studies, to increasing levels of complexity with little or no understanding of the uncertainties associated with these extrapolations. Consequently, a project designed to address concerns associated with scaling from seedling to mature tree responses was initiated. During the 1990 and 1991 growing seasons, mature northern red oak (Quercus rubra L.) trees and seedlings were exposed to subambient, ambient, and twice ambient ozone (O(3)) concentrations. The initial focus of the study was to identify possible trends and obvious differences between mature trees and seedlings, both in terms of growth and physiology and in response to O(3). Generally, mature trees exhibited a greater decrease in photosynthesis rates over the growing season than did the seedlings. Ozone treatments had no consistent effect on gas exchange rates of seedlings, but the twice ambient O(3) treatment resulted in reduced photosynthesis rates in the mature tree. Despite no effect of O(3) on seedling gas exchange rates, total seedling biomass was significantly less at the end of the 1991 growing season for those seedlings exposed to twice ambient O(3) levels. Disproportionate reductions in root biomass also resulted in reduced root to shoot ratios at elevated O(3) concentrations.     

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.     

Soil contamination by crude oil: impact on the mycorrhizosphere and on the revegetation potential of forest trees

In vitro and greenhouse biotests were carried out to study the effects of various concentrations of crude oil on the mycorrhizosphere and the ability of ectomycorrhizal fungi to colonise Norway spruce and poplar seedlings grown on contaminated soil. Ectomycorrhizal fungi grown in pure cultures showed a variety of reactions to crude oil, ranging from growth stimulation to total inhibition of growth, depending on the species of fungi. Germination of poplar and spruce seeds was not significantly affected. The growth of spruce seedlings was not affected by crude oil, whereas that of poplar seedlings was significantly reduced at high concentrations. None of the concentrations had any effect on the degree of ectomycorrhizal and endomycorrhizal colonisation of poplar. With spruce, however, the ectomycorrhizal fungi showed species-specific reactions to increasing concentrations, in accordance with the results of the pure culture test. The length of time between soil contamination and seeding affects both seedling growth and the mycorrhizal infection potential of the soil. The results confirm the importance of mycorrhizal fungi in the bioremediation of soils contaminated by crude oil.     

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 two growing seasons of experimental ozone fumigation on photosynthetic characteristics of white oak seedlings

White oak (Quercus alba L.) seedlings were exposed to charcoal-filtered air or to above-ambient ozone concentrations for 19-20 weeks during each of two growing seasons in continuously stirred tank reactors in greenhouses. Ozone treatments were 0.15 ppm (300 microg m(-3)) for 8 h day(-1), 3 days week(-1) in 1988, and continuous 15% above ambient in 1989. The seedlings were grown in forest soil watered twice weekly with simulated rain of pH 5.2. Responses of net photosynthesis to photosynthetically active radiation and intercellular CO(2) concentration were measured three times each year. There were no significant differences in light-saturated net photosynthesis or stomatal conductance, dark respiration, quantum or carboxylation efficiencies, and light or CO(2) compensation points on any date between control and ozone-exposed seedlings.     

Interactive effects of ozone and elevated carbon dioxide on the growth and physiology of black cherry, green ash, and yellow-poplar seedlings

Potted seedlings of black cherry (Prunus serotina Ehrh.) (BC), green ash (Fraxinus pennsylvanica Marsh.) (GA), and yellow-poplar (Liriodendron tulipifera L.) (YP) were exposed to one of the four treatments: (1) charcoal-filtered air (CF) at ambient CO(2) (control); (2) twice ambient O(3) (2 x O(3)); (3) twice ambient CO(2) (650 microl l(-1)) plus CF air (2 x CO(2)); or (4) twice ambient CO(2) (650 microl l(-1)) plus twice ambient O(3) (2 x CO(2) + 2 x O(3)). The treatments were duplicated in eight continuously stirred tank reactors for 10 weeks. Gas exchange was measured during the last 3 weeks of treatment and all seedlings were destructively harvested after 10 weeks. Significant interactive effects of O(3) and CO(2) on the gas exchange of all three species were limited. The effects of elevated CO(2) and O(3), singly and combined, on light-saturated net photosynthesis (A(max)) and stomatal conductance (g(s)) were inconsistent across species. In all three species, elevated O(3) had no effect on g(s). Elevated CO(2) significantly increased A(max) in GA and YP foliage, and decreased g(s) in YP foliage. Maximum carbon exchange rates and quantum efficiencies derived from light-response curves increased, while compensation irradiance and dark respiration decreased in all three species when exposed to 2 x CO(2). Elevated O(3) affected few of these parameters but any change that was observed was opposite to that from exposure to 2 x CO(2)-air. Interactive effects of CO(2) and O(3) on light-response parameters were limited. Carboxylation efficiencies, derived from CO(2)-response curves (A/C(i) curves) decreased only in YP foliage exposed to 2 x CO(2)-air. In general, growth was significantly stimulated by 2 x CO(2) in all three species; though there were few significant growth responses following exposure to 2 x O(3) or the combination of 2 x CO(2) plus 2 x O(3). Results indicate that responses to interacting stressors such as O(3) and CO(2) are species specific.     

Fine root studies in situ and in the laboratory

The fine roots and myocorrhizae of beech, spruce and fir trees exposed to ozone, sulphur dioxide and simulated acid precipitation in open-top chambers (OTC) were examined both in situ by rhizoscopy and in the laboratory using root samples from soil cores. Prior to measurements the trees were treated for about one year. During the second year of treatment the fine root production of all three tree species was determined rhizoscopically. The OTC experiments were concluded after an additional three years at which time fine root and small root dry matter as well as the absolute and relative frequencies of mycorrhizae of spruce and fir were determined from soil cores. The vitality of spruce mycorrhizae was examined by fluorescein diacetate staining. In addition total contents of essential cations of spruce mycorrhizae were measured. Long-term exposure to SO(2), SO(2) + O(3), and simulated acid precipitation led to an increased mycorrhizal production by fir. On spruce, a decreased number of mycorrhizae was found in the chambers polluted with SO(2), but a high proportion of dead fine roots indicated an increased root production with an intensified turnover or a delayed decomposition of spruce mycorrhizae. The cation analyses showed an accumulation of Ca(2+) and Zn(2+) in the mycorrhizae of spruce exposed to ozone.     

Foliar injury, leaf gas exchange and biomass responses of black cherry (Prunus serotina Ehrh.) half-sibling families to ozone exposure

Open pollinated families of black cherry seedlings were studied to determine genotypic differences in foliar ozone injury and leaf gas exchange in 1994 and growth response following three growing seasons. An O(3)-sensitive half-sibling family (R-12) and an O(3)-tolerant half-sibling family (MO-7) planted in natural soil were studied along with generic nursery stock (NS) seedlings. Ozone exposure treatments were provided through open top chambers and consisted of 50, 75, and 97% of ambient ozone, and open plots from May 9 to August 26, 1994. Ambient ozone concentrations reached an hourly peak of 88 ppb with 7-hour averages ranging from 39 to 46 ppb. Seedlings in the 50 and 75% of ambient chambers were never exposed to greater than 80 ppb O(3). Visible foliar ozone injury (stipple) was significantly higher for R-12 seedlings than MO-7 seedlings and increased with increasing ozone exposures. For the chamber treatments averaged over all families, there was no significant difference in stomatal conductance and net photosynthetic rates, but there was a significant decrease in root biomass, and a significant decrease in root/shoot ratio between the 50 and 97% of ambient chambers. Stomatal conductance and net photosynthetic rates were significantly different between families with R-12 seedlings generally greater than MO-7 seedlings. The R-12 seedlings had a 7.5 mmol m(-2) increase in ozone uptake compared to MO-7, and at the same cumulative O(3) exposure R-12 exhibited 40.9% stippled leaf area, whereas MO-7 had 9.2% stippled leaf area. Significant differences were observed in stem volume growth and total final biomass between the open-top chambers and open plots. Although R-12 had the most severe foliar ozone injury, this family had significantly greater stem volume growth and total final biomass than MO-7 and NS seedlings. Root:shoot ratio was not significantly different between MO-7 and R-12 seedlings.     

Scots pine responses to CO2 enrichment--I. Ectomycorrhizal fungi and soil fauna

Ectomycorrhizal Scots pine seedlings were grown in unfertilized forest soil at ambient and double (ca 700 ppm) atmospheric concentrations of CO2. The biomass of seedlings and fungal biomass both in the roots and in the soil and the numbers of certain groups of soil animals were measured under summer conditions and after an artificial winter acclimation period. No biomass parameter showed any significant change due to CO2 elevation. Increases were found during the winter acclimation period in total and fine root biomasses, fungal biomass in the soil and total fungal biomass both in the roots and in the soil, while the ratio of needle biomass: fungal biomass and the shoot: root ratio decreased. The N concentration in previous-year needles was lower in the double CO2 environment than with ambient CO2. Enchytraeids almost disappeared in the double CO2 environment during winter acclimation, while the numbers of nematodes increased at the same time in both treatments.     

Response of mycorrhizal Norway spruce seedlings to various nitrogen loads and sources

Norway spruce seedlings were grown under greenhouse conditions in Rootrainers with a vermiculite-peat moss mixture under various N-regimes for 6 months. Either ammonium or nitrate was applied in loads of 100 or 800 kg N ha(-1) year(-1) to seedlings which were either non-mycorrhizal or inoculated with the mycorrhizal fungi Hebeloma crustuliniforme or Laccaria bicolor. The use of increasing N loads enhanced shoot and total biomass, whereas root/shoot ratio, number of short roots and mycorrhization decreased. A significant enhancement of the concentration and content was obvious for the element N, whereas a significant decrease was obvious for P and Zn concentrations. The use of ammonium, as opposed to nitrate, significantly enhanced the biomass and the numbers of short roots, and reduced the root/shoot ratios, but did not influence the mycorrhization. It further significantly enhanced the N concentrations in roots and shoots. Fungal inoculation with H. crustuliniforme or L. bicolor compared to non-inoculated controls significantly enhanced shoot and total biomass, but reduced root/shoot ratios. The mycorrhization further significantly enhanced N and P concentrations and contents, but reduced Mn. Overall, the mycorrhization improved the P nutrition of the seedlings independently on the applied N loads or N sources. Dose response curves using ammonium nitrate as N source with a maximum load of 1600 kg N ha(-1) year(-1) applied on seedlings associated with H. crustuliniforme revealed that the maximum growth was reached at a load of 800 kg N ha(-1) year(-1) with a simultaneous decrease of the mycorrhization. In both shoots and roots, N concentrations increased constantly with increasing N loads, while P, Ca, and Zn concentrations decreased constantly.     

Effects of acid rain on growth and nutrient concentrations in Scots pine and Norway spruce seedlings grown in a nutrient-rich soil

The effects of artificially applied acid precipitation on growth and nutrient concentrations of Scots pine (Pinus sylvestris L.) and Norway spruce (Picea abies [L.] Karst.) seedlings were investigated in a long-term acid irrigation experiment in field conditions. Seedlings of northern and southern origin were planted in boxes containing peat and composted soil rich in nutrients, and sprinkler irrigated with water acidified with nitric and sulphuric acids to pH 3 or pH 4 for periods varying from two to three and a half growing seasons during 1986-1989. Water irrigated (pH 5.4-7.6) and non-irrigated groups of seedlings were also included in the experiment. At the end of the experiment needles, main and lateral shoots and roots were collected from the seedlings for the determination of height growth and biomass partitioning, and for the analysis of S, N, Mg, P, K, Ca, Mn and Fe concentrations. The treatment effects compared to the irrigated control were studied using multivariate analyses of variance and covariance. In the pine seedlings the total dry matter production increased by 25-70% compared with the irrigated controls when the total wet deposition to the seedlings exceeded 67 kg S ha(-1) and 36 kg N ha(-1) (e.g. after two growing seasons' exposure of the pH 3 treatment). The increase was mainly due to an increase in needle dry weight (54-72% greater at pH 3) and root weight (20-65% greater at pH 3), whereas the height growth or shoot weight growth were less affected. The northern provenance pine seedlings responded more clearly to the pH 3 irrigation than the southern ones. The treatments had no consistent effects on any of the growth variables studied in the spruce seedlings, however. The pines had higher root and foliage Ca concentrations as a result of the acid irrigation, whereas in spruce, acid rain decreased the Ca concentration in needles and shoots. Root Mn and Fe concentrations were higher in both species as a result of the pH 3 treatment. A higher soil conductivity and Ca concentration resulted from the prolonged pH 3 treatment. The results strongly support the hypothesis that the long-term growth and nutrient allocation response of conifers to acid precipitation is dependent both on the tree species and on the nutritional status of the soil.     

Interactive effects of CO2 and O3 on a ponderosa pine plant/litter/soil mesocosm.

To study individual and combined impacts of two important atmospheric trace gases, CO2 and O3, on C and N cycling in forest ecosystems; a multi-year experiment using a small-scale ponderosa pine (Pinus ponderosa Laws.) seedling/soil/litter system was initiated in April 1998. The experiment was conducted in outdoor, sun-lit chambers where aboveground and belowground ecological processes could be studied in detail. This paper describes the approach and methodology used, and presents preliminary data for the first two growing seasons. CO2 treatments were ambient and elevated (ambient + 280 ppm). O3 treatments were elevated (hourly averages to 159 ppb, cumulative exposure > 60 ppb O3, SUM 06 approximately 10.37 ppm h), and a low control level (nearly all hourly averages <40 ppb. SUM 06 approximately 0.07 ppm h). Significant (P < 0.05) individual and interactive effects occurred with elevated CO2 and elevated O3. Elevated CO2 increased needle-level net photosynthetic rates over both seasons. Following the first season, the highest photosynthetic rates were for trees which had previously received elevated O3 in addition to elevated CO2. Elevated CO2 increased seedling stem diameters, with the greatest increase at low O3. Elevated CO2 decreased current year needle % N in the summer. For 1-year-old needles measured in the fall there was a decrease in % N with elevated CO2 at low O3, but an increase in % N with elevated CO2 at elevated O3. Nitrogen fixation (measured by acetylene reduction) was low in ponderosa pine litter and there were no significant CO2 or O3 effects. Neither elevated CO2 nor elevated O3 affected standing root biomass or root length density. Elevated O3 decreased the % N in coarse-fine (1-2 mm diameter) but not in fine (< 1 mm diameter) roots. Both elevated CO2 and elevated O3 tended to increase the number of fungal colony forming units (CFUs) in the AC soil horizon, and elevated O3 tended to decrease bacterial CFUs in the C soil horizon. Thus, after two growing seasons we showed interactive effects of O3 and CO2 in combination, in addition to responses to CO2 or O3 alone for a ponderosa pine plant/litter/soil system.