Responses of spruce seedlings (Picea abies) to exhaust gas under laboratory conditions--II. Ultrastructural changes and stomatal behaviour

This study examines the effects of exhaust gas exposure on the epistomatal wax structure and mesophyll ultrastructure in needles of Norway spruce (Picea abies (L.) Karst.) seedlings. Stomatal diffusive resistance was also measured. Two independent exhaust gas fumigations were performed: 100 and 200 ppb measured as NO(x), for 10 days and 50, 100 and 200 ppb NO(x) for 19 days. The obstructive effect of exhaust gas exposure on epistomatal wax tubules was apparent. The stomata became covered by flat and solid wax resulting from the structural degradation of the wax crystalloids. Increasing the exhaust gas concentration in the chamber atmosphere exacerbated the degradation of the wax structure. Exhaust gas exposure induced aggregation and electron translucence of plastoglobuli, swelling of thylakoids, increase of cytoplasmic lipids and slight increase of vesiculation of cytoplasm in mesophyll cells of current and previous year needles. These changes were exemplified in current year needles. Damage to the epicuticular waxes and mesophyll ultrastructure of spruce needles most likely reflects the NO(x) and volatile hydrocarbon fraction. The alterations in epicuticular waxes and mesophyll ultrastructure can be related to accelerated senescence of the youngest, photosynthetically active, needle generation. The exhaust gas also resulted in decreased diffusive stomatal resistance at night which indicates that the exhaust gas exposure disturbed the gas exchange of spruce seedlings. The results show that even relatively short-term exposure to realistic concentrations of exhaust gas in the atmosphere can induce rather severe injuries to the needle surface structure as well as ultrastructure at the cellular level.     

Growth variation of Scots pine across a pollution gradient on the Kola Peninsula, Russia

Decadal exposure to emissions from a non-ferrous smelter has damaged the forest ecosystems surrounding the city of Monchegorsk located on the Kola Peninsula in northwestern Russia. We use the methods of tree-ring analysis to study the areal extent and timing of recent growth reductions of Scots pine (Pinus sylvestris L.) in the region surrounding the smelter in Monchegorsk. Reduced growth of Scots pine was observed up to 30 km southwest of the smelter. This directional gradient of forest damage is related to the dispersal of pollutants which is influenced by the prevailing northern winds and local topographic features. Old Scots pines (age 200 years +) appeared to be more sensitive than younger ones: growth reductions of old trees had started earlier and reductions were observed farther from the smelter than for younger trees. The findings are compared to a classification which describes the state of forest ecosystems based on the occurrence of certain plant species; the classification matched well with the observed growth trends. Pollution-induced changes in the climatic signal in tree-rings are also studied. The strong dependence of growth on mid-summer temperatures, typical for Scots pine on high latitudes, proved to be insensitive to effects of pollution. Changes in the climate-growth relationship took place decades after growth trends had started to decline.     

Relations between Scots pine needle element concentrations and decreased needle longevity along pollution gradients

Scots pine (Pinus sylvestris L.) shoots were sampled along transects near one urban pollution source and two smelters. Needle Mg, P and K concentrations decreased from the second to the fourth age class linearly with needle survival along the urban pollution gradient. Still, over 80% of the average concentration of these nutrients remained in the fourth needle age class. Decreased needle longevity was closely related to the increased heavy metal concentrations near the smelters. Near the urban pollution source, it was related to the increased annual needle mass and the increased needle nutrient concentrations. Decreased Mn accumulation along with needle age was detected near all pollution sources. Leaching of Mn from needles and especially from soil as a cause of decreased needle concentrations is discussed.     

The effect of simulated acid rain on the biochemical composition of Scots pine (Pinus sylvestris L.) needles

The effects of prolonged simulated acid rain on the biochemistry of Scots pine needles were studied in Finnish Lapland. Pine trees were exposed by spraying the foliage and soil with either clean water or simulated acid rain (SAR; both sulphuric and nitric acids) over the period 1985-1991. The concentrations of carbohydrates (starch, glucose, fructose, sucrose) in one-year-old pine needles were not affected by SAR-treatments. The SAR-treatments did not have significant effects on protein bound amino acids, which was true also for most of the free amino acids. However, the citrulline concentration was over three-fold greater in the foliage of pines exposed to SAR of pH 3 compared to irrigated controls. The concentrations of total phenolics, individual low molecular weight phenolics and soluble proanthocyanidins were not affected by the treatments, but insoluble proanthocyanidins had increased in acid-treated trees. Some of the studied biochemical compounds showed significant differences between two sub-areas (similar treatments) only 120 m apart.     

Long-term evaluation of the needle surface wax condition of Pinus sylvestris around different industries in Lithuania

The aim of our study was to evaluate the annual dynamics of needle surface wax erosion and wettability in Scots pines exposed to a gradient of industrial pollutants emitted from the main factories of Lithuania: a nitrogen fertilizer factory, an oil refinery and a cement factory. Decreased emissions (in the case of the oil refinery and the cement factory) were reflected in the increased structural surface area (SSA, i.e. area covered by tubular waxes) on the needles. The nearly constant amount of emissions from the nitrogen fertilizer factory within the 1994-2000 period corresponded to negligible annual differences in SSA. Annual changes in the hydrophobicity of needles on the investigated transects were small. Despite the decreased pollution within the 7-year period, industrial emissions are still causing significantly accelerated wax erosion and increased wettability in needles sampled from the stands most heavily affected by pollutants.     

Secondary air pollutants

The study has been performed in a supposedly clean-air region of a Northern Finnish forest with a homogeneous stand of Scots pine. Stomatal epicuticular wax layer erosion is described using a classification system of five erosion stages. The percentage of stomatal wax within each erosion stage is calculated and the results are treated statistically, which makes the morphological study of the needle surface semi-quantitative. Severe wax degradation has already been found in the current year, increasing with the age of the needles. In this area, the wax layer erosion is correlated with secondary air pollutants, as analyses have shown high trichloroacetate (TCA) levels in needles from the same trees. The wax layer analyses are in accordance with earlier findings which have demonstrated differences in tolerance to TCA.     

Ecological condition of forests around the eastern part of the Gulf of Finland

The effects of air pollutants on forests around the eastern part of the Gulf of Finland were studied by measurement of the sulphur and calcium content of pine needles and evaluation of the ecological conditions of pine forests. Several parameters for pine trees and their needles were chosen as well as the species composition and condition of epiphytic lichens. Very high pine needle S- and Ca-contents were measured in the vicinity of the Narva and Slantsy plants. In this region both the acid and basic pollutant load is massive, partly neutralizing each other. It is suggested that the total load will, sooner or later, cause unexpected environmental damage. Wide 'lichen desert' areas were detected around Narva and Slantsy. Near the margins of these areas extraordinary epiphytes on pines were observed namely Xanthoria parietina (L.) Th.Fr. and red-coloured green alga Trentepohlia umbrina. They are regarded as indicators of alkaline pollution. The lowest pine needle S- and Ca-contents of the study area were measured in south-eastern Finland. The condition of pine forests and their needles was, however, better on the neighbouring Karelian Isthmus although the species number of epiphytic lichens was very low and the condition of the lichens was poor. It is suggested that these most sensitive indicators of air pollutants are damaged by pollutants from St Petersburg and Narva. Vast virgin forests of the Karelian Isthmus act as pollutant sinks reducing the effect of pollutants on trees. On the Finnish side intensive forest management has been carried on for many decades making forests and trees more sensitive to pollutants.     

Total vs. internal element concentrations in Scots pine needles along a sulphur and metal pollution gradient

Analysis of foliar elements is a commonly used method for studying tree nutrition and for monitoring the impacts of air pollutants on forest ecosystems. Interpretations based on the results of foliar element analysis may, however, be different in nutrition vs. monitoring studies. We studied the impacts of severe sulphur and metal (mainly Cu and Ni) pollution on the element concentrations (Al, Ca, Cu, Fe, K, Mg, Mn, Ni, P, Pb, S and Zn) in Scots pine (Pinus sylvestris L.) foliage along an airborne sulphur and metal pollution gradient. Emphasis was put on determining the contribution of air-borne particles that have accumulated on needle surfaces to the total foliage concentrations. A comparison of two soil extraction methods was carried out in order to obtain a reliable estimate of plant-available element concentrations in the soil. Element concentrations in the soil showed only a weak relationship with internal foliar concentrations. There were no clear differences between the total and internal needle S concentrations along the gradient, whereas at the plot closest to the metal smelter complex the total Cu concentrations in the youngest needles were 1.3-fold and Ni concentrations over 1.6-fold higher than the internal needle concentrations. Chloroform-extracted surface wax was found to have Ni and Cu concentrations of as high as 3000 and 600 microg/g of wax, respectively. Our results suggest that bioindicator studies (e.g. monitoring studies) may require different foliar analysis techniques from those used in studies on the nutritional status of trees.     

Damage to Eastern White Pine by Sulfur Dioxide,Semimature-Tissue Needle Blight,and Ozone

Eastern white pine (Pinus strobus L.) forests were severely damaged by atmospheric sulfur dioxide up to distances of 25 miles northeast of large smelters located in the Sudbury mining district of Ontario. Damage to white pine was measured in terms of foliage, bark, and biological injuries, radial and volume growth decrement, and tree mortality. The foliar symptoms of sulfur dioxide injury on white pine trees sometimes resembles those caused by a physiogenic disease, semimature-tissue needle blight (abbreviated to SNB). Studies on the nature and etiology of SNB were conducted in white pine forests in the Upper Ottawa Valley, which are remote from smelter operations which might pollute the atmosphere. These studies included the determination of the role that naturallyoccurring atmospheric ozone plays in the occurrence of SNB. Differences between the symptoms of sulfur dioxide injury, SNB, and ozone damage are outlined.     

UV-absorbing compounds and waxes of Scots pine needles during a third growing season of supplemental UV-B

Methanol-extractable UV-absorbing compounds, wax tube distribution and the chloroform-soluble waxes of the needles of mature Scots pines were studied in a UV-B field experiment in Oulu (65 degrees N). Throughout the experiment, UV-B lamp banks were placed over the same selected branch and each year needle samples were taken from the same branch. In the third exposure year, needle samples were taken twice a month from 3-day-old needles (18 June) to fully developed needles (13 August). On 28 September, the previous year's needles (c + 1, c + 2) were also collected. There was a significant negative correlation between the amount of waxes and UV-absorbing compounds. A high amount of UV-absorbing compounds was observed early and late in the season when the amount of waxes was low and epicuticular waxes were undeveloped (youngest needles) or already eroded (c + 2 needles). The amount of UV-absorbing compounds (A310/cm2 and A320/cm2) was significantly (30-day-old needles) or slightly (all the other needle ages) higher in the ambient needles compared to the needles under supplemental UV-B. This possibly indicated the already inhibited pigment synthesis in the UV-B-treated needles during the third year of supplemental UV-B. This observation could mean that the protective mechanisms may not be effective under accumulated UV-B dose.     

Haloacetic acids,phytotoxic secondary air pollutants

Haloacetic acids are atmospheric oxidation products of airborne C₂-halocarbons which are important solvents and propellants. Levels of trichloroacetate (TCA) in conifer needles from mountain ranges in Germany (Black Forest, Erzgebirge) and from two sites in Finland are compared; TCA is present in conifer needles at concentrations up to 0.7 μmol/kg, MCA up to 0.2 μmol/kg. At the Finnish sites, TCA-concentrations and branch degeneration symptoms of Scots pine are correlated. Monochloroacetate (MCA) has been determined in needle samples from Southern Germany in concentrations exceeding its phytotoxicity threshold towards photoautotrophic organisms. Data on atmospheric chloroacetate levels in Germany are also given; ambient air levels of chloroacetic acids range from about 2 pmol/m³ (TCA) to 390 pmol/m³ (MCA). TCA and dichloroacetic acid (DCA) arise from atmospheric oxidation of airborne C₂-chlorocarbons, while the source of MCA is not yet known; several tentative pathways are suggested.     

Growth and photosynthetic responses of two pine species (Pinus koraiensis and Pinus rigida) in a polluted industrial region in Korea

We investigated the effects of pollutants on two pine species (Pinus koraiensis and Pinus rigida) in an industrial region in Korea, using a physiological approach. The concentrations of fluorine (F) and chlorine (Cl) in the atmosphere, in precipitation and soil water at the damaged site were all significantly higher than at a control site. Moreover, the concentrations of F, Cl and Mn in pine needles were significantly higher, and essential elements and chlorophyll in needles were significantly lower at the damaged site than at the control site. The photosynthetic capacities, shoot length and survival statistics of needles of the two pines were all significantly reduced at the damaged site compared to the control site, especially P. rigida. Based on our comparison of photosynthetic responses and the concentrations of F, Cl and Mn in needles of the two pine species, P. koraiensis is more resistant to excess Mn in its needles than P. rigida.     

Disappearance of polycyclic aromatic hydrocarbons sorbed on surfaces of pine [Pinua thunbergii] needles under irradiation of sunlight: Volatilization and photolysis

The solar photodegradation of 16 polycyclic aromatic hydrocarbons (PAHs), sorbed on surfaces of pine [Pinua thunbergii] needles was investigated. The PAHs were produced by combustion of polystyrene and exposed onto the surfaces of pine needles. The disappearance of PAHs sorbed on the pine needle surfaces is mainly caused by volatilization and photolysis, with photolysis playing a major role. The volatilization rates correlate with PAH molecular weight significantly. The photolysis of the 16 PAHs follows first-order kinetics and their photolysis half-lives (t_1/2,P) range from 12.9 h for naphthalene to 65.4 h for fluorene. The PAHs have similar half-lives whether they are sorbed on spruce or pine needles. Compared with water, the cuticular waxes of pine needles can stabilize photolysis of PAHs and facilitate accumulation of PAHs. t_1/2,P for selected PAHs correlate with semi-empirically calculated energy of the highest occupied orbital (E_HOMO). Photochemical behaviors of PAHs are dependent not only on their molecular structures but also the physical–chemical properties of the substrate on which they are adsorbed.     

Indication of airborne pollution by birch and spruce in the vicinity of copper smelter

The aim of the study was determination of air pollution impact of the copper smelter in Bor and its surroundings (Serbia) by assessing the suitability of birch (Betula pendula Roth.) and spruce (Picea abies L.) for the purposes of biomonitoring and comparing it with previously published data from the same study area. The concentrations of Cu, Zn, Pb and Mn in leaves/needles, branches, roots and soil were determined. Sampling was performed during 2009 in two zones with high load of air pollution due to copper mining and smelting activities, and one background zone. Metal accumulation and translocation was evaluated in terms of biological factors. In addition, plant enrichment factor was calculated. According to the results, plant foliage was not enriched through soil, which indicates absorption from the air, with both species acting as excluders of Cu, Pb, Zn and Mn. Leaves were more enriched with all the metals than needles, indicating a better response of birch to airborne pollution than spruce. Cluster analysis showed different level of pollution at the sites, while correlations between Cu and Pb obtained by Principal Component Analysis indicated their anthropogenic origin. Regarding previously published results, beside birch leaves, pine needles (which showed higher level of response to pollution compared to linden leaves) could be applied in air biomonitoring surveys near copper smelters.     

Variation in CO2 assimilation rate induced by simulated dew waters with different sources of hydroxyl radical (*OH) on the needle surfaces of Japanese red pine (Pinus densifora Sieb. et Zucc.)

The hydroxyl radical (*OH) is generated in polluted dew on the needle surfaces of Japanese red pine (Pinus densiflora Sieb. et Zucc.). This free radical, which is a potent oxidant, is assumed to be a cause of ecophysiological disorders of declining trees on the urban-facing side of Mt. Gokurakuji, western Japan. Mists of *OH-generating N(III) (HNO2 and NO2-) and HOOH + Fe + oxalate solutions (50 and 100 microM, pH 5.1-5.4) simulating the dew water were applied to the foliage of pine seedlings grown in open-top chambers in the early morning. Needles treated with 100 microM N(III) tended to have a greater maximum CO2 assimilation rate (Amax), a greater stomatal conductance (g(s)) and a greater needle nitrogen content (Nneedle), suggesting that N(III) mist acts as a fertilizer rather than as a phytotoxin. On the other hand, needles treated with 100 microM HOOH + Fe + oxalate solution showed the smallest Amax, g(s), and Nneedle, suggesting that the combination of HOOH + Fe + oxalate caused a decrease in needle productivity. The effects of HOOH + Fe + oxalate mist on pine needles were very similar to the symptoms of declining trees at Mt. Gokurakuji.     

Chemical and morphological characteristics of key tree species of the Carpathian Mountains

Concentrations of Al, B, Ca, Cu, Fe, K, Mg, Mn, N, Na, P, S and Zn in the foliage of white fir (Abies alba), Norway spruce (Picea abies) and common beech (Fagus sylvatica) from 25 sites of the Carpathian Mts. forests (Czech Republic, Poland, Romania, Slovakia and Ukraine) are discussed in a context of their limit values. S/N ratio was different from optimum in 90% of localities when compared with the European limit values. Likewise we found increase of Fe and Cu concentrations compared with their background levels in 100% of locations. Mn concentrations were increased in 76% of localities. Mn mobilization values indicate the disturbance of physiological balance leading to the change of the ratio with Fe. SEM-investigation of foliage waxes from 25 sites in the Carpathian Mts. showed, that there is a statistically significant difference in mean wax quality. Epistomatal waxes were damaged as indicated by increased development of net and amorphous waxes. The most damaged stomata in spruce needles were from Yablunitsa, Synevir and Brenna; in fir needles from Stoliky, and in beech leaves from Malá Fatra, Morské Oko and Beregomet. Spruce needles in the Carpathian Mts. had more damaged stomata than fir needles and beech leaves. Spruce seems to be the most sensitive tree species to environmental stresses including air pollution in forests of the Carpathian Mountains. Foliage surfaces of three forest tree species contained Al, Si, Ca, Fe, Mg, K, Cl, Mn, Na, Ni and Ti in all studied localities. Presence of nutrition elements (Ca, Fe, Mg, K and Mn) on foliage surface hinders opening and closing stomata and it is not physiologically usable for tree species.     

Influence of solar UV radiation on the nitrogen metabolism in needles of Scots pine (Pinus sylvestris L.)

Needles of 20-year-old Scots pine (Pinus sylvestris L.) saplings were studied in an ultraviolet (UV) exclusion field experiment (from 2000 to 2002) in northern Finland (67 °N). The chambers held filters that excluded both UV-B and UV-A, excluded UV-B only, transmitted all UV (control), or lacked filters (ambient). UV-B/UV-A exclusion decreased nitrate reductase (NR) activity of 1-year-old needles of Scots pines compared to the controls. The proportion of free amino acids varied in the range 1.08-1.94% of total proteins, and was significantly higher in needles of saplings grown under UV-B/UV-A exclusion compared to the controls or UV-B exclusion. NR activity correlated with air temperature, indicating a “chamber effect”. The study showed that both UV irradiance and increasing temperature are significant modulators of nitrogen (N) metabolism in Scots pine needles.     

Assessing the critical level of SO2 for Scots pine in situ

A field survey was performed in eastern Finland, where measured ambient SO2 concentrations were 1.4-3.8 microg m(-3) a(-1) and bulk S deposition 0.17-0.32 g m(-2) a(-1) in 1991-1993. The accumulation of sulphur (S) in needles of Scots pine (Pinus sylvestris L.) was studied with XRF, IC and FESEM analyses and the needle damage examined under a light microscope and by SEM. Foliar N concentrations were also measured. Foliar total S concentrations were observed to be above the normal S level (500-700 microg g(-1)) over almost the whole area. Slight chlorosis and/or necrosis of the needle tips and stomatal areas, changes in the needle surface waxes and localization of S into needle tips and mesophyll cells around the stomata suggested the impact of S deposition, as did the calculations of St/Nt, and 'predicted' and 'excess' S. A concentration of about 900 microg g(-1) may be considered a critical level for foliar St in areas with low N supply.     

Chlorohydrocarbons, PCB congeners, polychlorodioxins, furans and dibenzothiophenes in pine needles in the vicinity of a metal reclamation plant

Chlorohydrocarbons, PCB congeners, polychlorodioxins, furans and dibenzothiophenes in pine needles in the vicinity of a metal reclamation plant were analyzed by GC/ECD and GC/MS. Wax and the rest of the needles were analyzed separately. As a rule the concentrations of -HCH (0.5–13.6 ng/g), γ-HCH (0.4–7.3 ng/g), HCB (0.2–3.4 ng/g), PCB congeners (0.2–67 ng/g), PCDDs and PCDFs were higher in the older needles. The ratio of the concentration in the wax to the concentration in the rest of the needles was higher in the younger needles. The concentrations of PCB congeners, TeCDDs and TeCDFs were higher in the needles collected in the area nearest to the plant. The samples, obviously, contained tri- and tetrachlorodibenzothiophenes but these were not determined quantitatively due to interfering peaks in GC/MS.     

Effect of smelter emissions on the hemogram of the deer mouse (Peromyscus maniculatus)

Blood samples were obtained from 284 deer mice (Peromyscus maniculatus) live-trapped near two base metal smelters in northern Manitoba, Canada. Hemoglobin and hematocrit values were significantly higher in mice trapped within 40-70 km of the smelters compared to those in control areas 185 to 190 km from the smelter. These increases appear to be normal physiological responses to a mild respiratory stress, as in a true secondary polycythemia of anoxic tupe. The mice are sensitive bioindicators of sulphur dioxide emissions that apparently induce the higher blood values.