Hydroxylation of bisphenol A by hyper lignin-degrading fungus Phanerochaete sordida YK-624 under non-ligninolytic condition

Bisphenol A (BPA) is one of the representative compounds of the endocrine disrupting compounds group and the highest volume chemicals produced worldwide. As a result, BPA is often detected in many soil and water environments. In this study, we demonstrated the transformation of BPA from liquid cultures inoculated with hyper lignin-degrading fungus Phanerochaete sordida YK-624. Under non-ligninolytic condition, approximately 80% of BPA was eliminated after 7 d of incubation. High-resolution electrospray ionization mass spectra and nuclear magnetic resonance analyses of a metabolite isolated from the culture supernatant suggested that BPA was metabolized to hydroxy-BPA, 4-(2-(4-hydroxyphenyl)propan-2-yl)benzene-1,2-diol, which has a much lower estrogenic activity than BPA. In addition, we investigated the effect of the cytochrome P450 inhibitor piperonyl butoxide (PB) on the hydroxylation of BPA, markedly lower transformation activity of BPA was observed in cultures containing PB. These results suggest that cytochrome P450 plays an important role in the hydroxylation of BPA by P. sordida YK-624 under non-ligninolytic condition.     

Molecular and physiological approaches to understand the ecology of methanol degradation during the biofiltration of air streams

A 13.4 L biofilter treating an off-gas stream supplemented with methanol under two different situations was studied in terms of MeOH removal efficiency, microbial ecology and odor removal. During Period 1 (P1) the reactor was packed with wood bark chips with no pH control, treating an off-gas resulting from the aerobic chamber of a membrane biological reactor treating sewage and located outdoor, whereas during Period 2 (P2) a compressed air stream fed with MeOH was treated using PVC rings and maintaining pH at neutral values. Both systems operated at 96 g MeOH m⁻³ h⁻¹ achieving removal efficiencies of around 90% during P1 and 99.9% during P2. The relative activity of biomass developed in both systems was assessed using respirometric analysis with samples obtained from both biofilms. Higher biomass activity was obtained during P2 (0.25-0.35 kg MeOH kg⁻¹ VSS d⁻¹) whereas 1.1 kg MeOH kg⁻¹ VSS d⁻¹ was obtained in the case of P1. The application of molecular and microscopic techniques showed that the eukaryotes were predominant during P1, being the yeast Candida boidinii the most abundant microorganism. A specific Fluorescence in situ hybridization probe was designed for C. boidinii and tested successfully. As a result of the neutral pH, a clear predominance of prokaryotes was detected during P2. Interestingly, some anaerobic bacteria were detected such as Desulfovibrio, Desulfobacteraceae species and also some archaea such as Methanosarcina.     

Immunotoxicity in ascidians: antifouling compounds alternative to organotins: III--the case of copper(I) and Irgarol 1051

After the widespread ban of TBT, due to its severe impact on coastal biocoenoses, mainly related to its immunosuppressive effects on both invertebrates and vertebrates, alternative biocides such as Cu(I) salts and the triazine Irgarol 1051, the latter previously used in agriculture as a herbicide, have been massively introduced in combined formulations for antifouling paints against a wide spectrum of fouling organisms. Using short-term (60 min) haemocyte cultures of the colonial ascidian Botryllus schlosseri exposed to various sublethal concentrations of copper(I) chloride (LC₅₀ = 281 μM, i.e., 17.8 mg Cu L⁻¹) and Irgarol 1051 (LC₅₀ > 500 μM, i.e., >127 mg L⁻¹), we evaluated their immunotoxic effects through a series of cytochemical assays previously used for organotin compounds. Both compounds can induce dose-dependent immunosuppression, acting on different cellular targets and altering many activities of immunocytes but, unlike TBT, did not have significant effects on cell morphology. Generally, Cu(I) appeared to be more toxic than Irgarol 1051: it significantly (p < 0.05) inhibited yeast phagocytosis at 0.1 μM (∼10 μg L⁻¹), and affected calcium homeostasis and mitochondrial cytochrome-c oxidase activity at 0.01 μM (∼1 μg L⁻¹). Both substances were able to change membrane permeability, induce apoptosis from concentrations of 0.1 μM (∼10 μg L⁻¹) and 200 μM (∼50 mg L⁻¹) for Cu(I) and Irgarol 1051, respectively, and alter the activity of hydrolases. Both Cu(I) and Irgarol 1051 inhibited the activity of phenoloxidase, but did not show any interactive effect when co-present in the exposure medium, suggesting different mechanisms of action.     

Propiconazole inhibits the sterol 14α-demethylase in Glomus irregulare like in phytopathogenic fungi

The increasing concentrations impact (0.02, 0.2 and 2 mg L⁻¹) of a Sterol Biosynthesis Inhibitor (SBI) fungicide, propiconazole, was evaluated on development and sterol metabolism of two non-target organisms: mycorrhizal or non-mycorrhizal transformed chicory roots and the arbuscular mycorrhizal fungus (AMF) Glomus irregulare using monoxenic cultures. In this work, we provide the first evidence of a direct impact of propiconazole on the AMF by disturbing its sterol metabolism. A significant decrease in end-products sterols contents (24-methylcholesterol and in 24-ethylcholesterol) was observed concomitantly to a 24-methylenedihydrolanosterol accumulation indicating the inhibition of a key enzyme in sterol biosynthesis pathway, the sterol 14α-demethylase like in phytopathogenic fungi. A decrease in end-product sterol contents in propiconazole-treated roots was also observed suggesting a slowing down of the sterol metabolism in plant. Taken together, our findings suggest that the inhibition of the both AM symbiotic partners development by propiconazole results from their sterol metabolism alterations.     

Photocatalytic removal of organic pollutants in aqueous solution by Bi(4)Nb(x)Ta((1-x))O(8)I

In this work, Bi₄Nb_xTa_(1−x)O₈I photocatalysts have been synthesized by solid state reaction method and characterized by powder X-ray diffraction, scanning electron microscope and UV-Vis near infrared diffuse reflectance spectroscopy. The photocatalytic activity of these photocatalysts was evaluated by the degradation of methyl orange (MO) in aqueous solutions under visible light, UV light and solar irradiation. The effects of catalyst dosage, initial pH and MO concentration on the removal efficiency were studied, and the photocatalytic reaction kinetics of MO degradation as well. The results indicated that Bi₄Nb_xTa_(1−x)O₈I exhibited high photocatalytic activity for the removal of MO in aqueous solutions. For example, the removal efficiency of MO by Bi₄Nb_0.1Ta_0.9O₈I was as high as 92% within 12 h visible light irradiation under the optimal conditions: initial MO concentration of 5-10 mg L⁻¹, catalyst dosage of 6 g L⁻¹ and natural pH (6-8), the MO molecules could be completely degradated by Bi₄Nb_0.1Ta_0.9O₈I within 40 min under UV light irradiation, and the photodegradation efficiency reaches to 60% after 7 h solar irradiation. Furthermore, the photocatalytic degradation of Bisphenol A (BPA) was also investigated under visible light irradiation. It is found that 99% BPA could be mineralized by Bi₄Nb_0.1Ta_0.9O₈I after 16 h visible light irradiation. Through HPLC/MS, BOD, TOC, UV-Vis measurements, we determined possible degradation products of MO and BPA. The results indicated that MO was degradated into products which are easier to be biodegradable and innocuous treated, and BPA could be mineralized completely. Furthermore, the possibility for the photosensitization effect in the degradation process of MO under visible light irradiation has been excluded.     

Uptake of 17β-estradiol and biomarker responses in brown trout (Salmo trutta) exposed to pulses

In streams, chemicals such as 17β-estradiol (E2) are likely to occur in pulses. We investigated uptake and biomarker responses in juvenile brown trout (Salmo trutta) of 3- or 6-h pulses of concentrations up to 370 ng E2 L⁻¹. Uptake by the fish was estimated from disappearance of E2 from tank water. A single 6-h pulse of 370 ng E2 L⁻¹ increased the plasma vitellogenin concentration, liver Erα- and vitellogenin-mRNA. Exposure to 150-160 ng E2 L⁻¹ for 6 h increased vitellogenin in one experiment but not in another. Two 6-h pulses had a larger effect one pulse. Brown trout in the size range 24-74 g took up E2 linearly with time and exposure concentration with a concentration ratio rate of 20.2 h⁻¹. In conclusion, the threshold for induction of estrogenic effects in juvenile brown trout at short term pulse exposure appears to be in the range 150-200 ng E2 L⁻¹.     

The exposition of a calcareous Mediterranean soil to toxic concentrations of Cr, Cd and Pb produces changes in the microbiota mainly related to differential metal bioavailability

The involvement of the bacterial community of an agricultural Mediterranean calcareous soil in relation to several heavy metals has been studied in microcosms under controlled laboratory conditions. Soil samples were artificially polluted with Cr(VI), Cd(II) and Pb(II) at concentrations ranging from 0.1 to 5000 mg kg⁻¹ and incubated along 28 d. The lowest concentrations with significant effects in soil respirometry were 10 mg kg⁻¹ Cr and 1000 mg kg⁻¹ Cd and Pb. However, only treatments showing more than 40% inhibition of respirometric activity led to significant changes in bacterial composition, as indicated by PCR-DGGE analyses. Presumable Cr- and Cd-resistant bacteria were detected in polluted microcosms, but development of the microbiota was severely impaired at the highest amendments of both metals. Results also showed that bioavailability is an important factor determining the impact of the heavy metals assayed, and even an inverted potential toxicity ranking could be achieved if their soluble fraction is considered instead of the total concentration. Moreover, multiresistant bacteria were isolated from Cr-polluted soil microcosms, some of them showing the capacity to reduce Cr(VI) concentrations between 26% and 84% of the initial value. Potentially useful strains for bioremediation were related to Arthrobacter crystallopoietes, Stenotrophomonas maltophilia and several species of Bacillus.     

Comparative bioremediation potential of four rhizospheric microbial species against lindane

Four microbial species (Kocuria rhizophila, Microbacterium resistens, Staphylococcus equorum and Staphylococcus cohnii subspecies urealyticus) were isolated from the rhizospheric zone of selected plants growing in a lindane contaminated environment and acclimatized in lindane spiked media (5-100 μg mL⁻¹). The isolated species were inoculated with soil containing 5, 50 and 100 mg kg⁻¹ of lindane and incubated at room temperature. Soil samples were collected periodically to evaluate the microbial dissipation kinetics, dissipation rate, residual lindane concentration and microbial biomass carbon (MBC). There was a marked difference (p < 0.05) in the MBC content and lindane dissipation rate of microbial isolates cultured in three different lindane concentrations. Further, the dissipation rate tended to decrease with increasing lindane concentrations. After 45 d, the residual lindane concentrations in three different spiked soils were reduced to 0%, 41% and 33%, respectively. Among the four species, S. cohnii subspecies urealyticus exhibited maximum dissipation (41.65 mg kg⁻¹) and can be exploited for the in situ remediation of low to medium level lindane contaminated soils.     

Distribution of Se and its species in Myriophyllum spicatum and Ceratophyllum demersum growing in water containing Se (VI)

The uptake of Se (VI) by two aquatic plants, Myriophyllum spicatum L. and Ceratophyllum demersum L., and its effects on their physiological characteristics have been studied. Plants were cultivated outdoors under semi-controlled conditions and in two concentrations of Na selenate solution (20 μg Se L⁻¹ and 10 mg Se L⁻¹). The higher dose of Se reduced the photochemical efficiency of PSII in both species, while the lower dose had no effect on PSII. Addition of Se had no effect on the amounts of chlorophyll a and b. The concentration of Se in plants grown in 10 mg Se L⁻¹, averaged 212 ± 12 μg Se g⁻¹ DM in M. spicatum (grown from 8-13 d), and 492 ± 85 μg Se g⁻¹ DM in C. demersum (grown for 31 d). Both species could take up a large amount of Se. The amount of soluble Se compounds in enzyme extracts ranged from 16% to 26% in control, and in high Se solution from 48% to 36% in M. spicatum and C. demersum, respectively. Se-species were determined using HPLC-ICP-MS. The main soluble species in both plants was selenate (∼37%), while SeMet and SeMeSeCys were detected at trace levels.     

Toxic effects of chemical pesticides (trichlorfon and dimehypo) on Dunaliella salina

Dunaliella salina, a unicellular green alga of environmental tolerance, was employed as test organism to investigate the toxicity effects of trichlorfon and dimehypo widely used in agriculture and veterinary as pesticides. The influences of trichlorfon and dimehypo on cell growth, β-carotene level, cell morphology changes, and activities of superoxide dismutase (Sod) and catalase (Cat) were investigated. At the concentrations less than 0.050 g L⁻¹ trichlorfon or 0.0005 g L⁻¹ dimehypo, cell responses were similar to control. When treated with 0.075-0.100 g L⁻¹ trichlorfon or 0.001-0.004 g L⁻¹ dimehypo, cell growth and β-carotene levels declined at first and then revived. When concentrations were higher than 0.125 g L⁻¹ trichlorfon or 0.005 g L⁻¹ dimehypo, both cell growth and β-carotene levels decreased until they were undetectable. The 10-d IC50 of trichlorfon and dimehypo on D. salina were 0.179 g L⁻¹ and 0.032 g L⁻¹. Both pollutants could stimulate the increase of Cat activity at a low concentration. Tolerance of D. salina to trichlorfon was obviously higher than that of dimehypo.     

Levels of organochlorine pesticides and polychlorinated biphenyls in the critically endangered Iberian lynx and other sympatric carnivores in Spain

Accumulation of organochlorine compounds is well studied in aquatic food chains whereas little information is available from terrestrial food chains. This study presents data of organochlorine levels in tissue and plasma samples of 15 critically endangered Iberian lynx (Lynx pardinus) and other 55 wild carnivores belonging to five species from three natural areas of Spain (Doñana National Park, Sierra Morena and Lozoya River) and explores their relationship with species diet. The Iberian lynx, with a diet based on the consumption of rabbit, had lower PCB levels (geometric means, plasma: <0.01 ng mL⁻¹, liver: 0.4 ng g⁻¹ wet weight, fat: 87 ng g⁻¹ lipid weight) than other carnivores with more anthropic and opportunistic foraging behavior, such as the red fox (Vulpes vulpes; plasma: 1.11 ng mL⁻¹, liver: 459 ng g⁻¹, fat: 1984 ng g⁻¹), or with diets including reptiles at higher proportion, such as the Egyptian mongoose (Herpestes ichneumon; plasma: 7.15 ng mL⁻¹, liver: 216 ng g⁻¹, fat: 540 ng g⁻¹), or the common genet (Genetta genetta; liver: 466 ng g⁻¹, fat: 3854 ng g⁻¹). Chlorinated pesticides showed interspecific variations similar to PCBs. Organochlorine levels have declined since the 80s in carnivores from Doñana National Park, but PCB levels are still of concern in Eurasian otters (Lutra lutra; liver: 3873-5426 ng g⁻¹) from the industrialized region of Madrid.     

Canine toys and training devices as sources of exposure to phthalates and bisphenol A: Quantitation of chemicals in leachate and in vitro screening for endocrine activity

Chewing and mouthing behaviors exhibited by pet dogs are likely to lead to oral exposures to a variety of environmental chemicals. Products intended for chewing and mouthing uses include toys and training devices that are often made of plastics. The goal of the current study was to determine if a subset of phthalates and bisphenol A (BPA), endocrine disrupting chemicals commonly found in plastics, leach out of dog toys and training devices (bumpers) into synthetic canine saliva. In vitro assays were used to screen leachates for endocrine activity. Bumper leachates were dominated by di-2-ethylhexyl phthalate (DEHP) and BPA, with concentrations reaching low μg mL⁻¹ following short immersions in synthetic saliva. Simulated chewing of bumpers during immersion in synthetic saliva increased concentrations of phthalates and BPA as compared to new bumpers, while outdoor storage had variable effects on concentrations (increased DEHP; decreased BPA). Toys leached substantially lower concentrations of phthalates and BPA, with the exception of one toy which leached considerable amounts of diethyl phthalate. In vitro assays indicated anti-androgenic activity of bumper leachates, and estrogenic activity of both bumper and toy leachates. These results confirm that toys and training devices are potential sources of exposure to endocrine disrupting chemicals in pet dogs.     

Effect of BPA on the germination,root development,seedling growth and leaf differentiation under different light conditions in Arabidopsis thaliana

Bisphenol A (BPA) is a well-known environmental toxic substance, which exerts unfavorable effects through endocrine disruptor (ER)-dependent and ER-independent mechanisms to threaten ecological systems seriously. BPA may also interact with other environmental factors, such as light and heavy metals, to have a synergetic effect in plants. However, there is little data concerning the toxic effect of BPA on the primary producers-plants and its possible interaction with light-dependent response. Here, the effects of BPA on germination, fresh weight, tap root length, and leaf differentiation were studied in Arabidopsis thaliana under different parts of light spectrum (dark, red, yellow, green, blue, and white light). Our results showed that low-dose BPA (1.0, 5.0 μM) caused an increase in the fresh weight, the tap root length and the lateral root formation of A. thaliana seedlings, while high-dose BPA (10.0, 25.0 μM) show an inhibition effect in a dose-dependent manner. Unlike karrikins, the effects of BPA on germination fresh weight and tap roots length under various light conditions are similar, which imply that BPA has no notable role in priming light response in germination and early seedling growth in A. thaliana. Meanwhile, BPA exposure influences the differentiation of A. thaliana leaf blade significantly in a light-dependent manner with little to no effect in dark and clear effect under red illumination.     

Effects of nitrogen deposition and soil fertility on cover and physiology of Cladonia foliacea (Huds.) Willd., a lichen of biological soil crusts from Mediterranean Spain

We are fertilizing a thicket with 0, 10, 20 and 50 kg nitrogen (N) ha⁻¹ yr⁻¹ in central Spain. Here we report changes in cover, pigments, pigment ratios and FvFm of the N-tolerant, terricolous, lichen Cladonia foliacea after 1-2 y adding N in order to study its potential as biomarker of atmospheric pollution. Cover tended to increase. Pigments increased with fertilization independently of the dose supplied but only significantly with soil nitrate as covariate. β-carotene/chlorophylls increased with 20-50 kg N ha⁻¹ yr⁻¹ (over the background) and neoxanthin/chlorophylls also increased with N. (Neoxanthin+lutein)/carotene decreased with N when nitrate and pH seasonalities were used as covariates. FvFm showed a critical load above 40 kg N ha⁻¹ yr⁻¹. Water-stress, iron and copper also explained variables of lichen physiology. We conclude that this tolerant lichen could be used as biomarker and that responses to N are complex in heterogeneous Mediterranean-type landscapes.     

Partitioning behaviour of perfluorinated alkyl contaminants between water, sediment and fish in the Orge River (nearby Paris, France)

This paper reports on the partitioning behaviour of 15 perfluorinated compounds (PFCs), including C₄-C₁₀ sulfonates and C₅-C₁₄ carboxylic acids, between water, sediment and fish (European chub, Leuciscus cephalus) in the Orge River (nearby Paris). Total PFC levels were 73.0 ± 3.0 ng L⁻¹ in water and 8.4 ± 0.5 ng g⁻¹ in sediment. They were in the range 43.1-4997.2 ng g⁻¹ in fish, in which PFC tissue distribution followed the order plasma > liver > gills > gonads > muscle. Sediment-water distribution coefficients (log K_d) and bioaccumulation factors (log BAF) were in the range 0.8-4.3 and 0.9-6.7, respectively. Both distribution coefficients positively correlated with perfluoroalkyl chain length. Field-based biota-sediment accumulation factors (BSAFs) are also reported, for the first time for PFCs other than perfluorooctane sulfonate. log BSAF ranged between −1.3 and 1.5 and was negatively correlated with the perfluoroalkyl chain length in the case of carboxylic acids.     

Detection of transfluthrin and metofluthrin genotoxicity in the ST cross of the Drosophila Wing Spot Test

In this study, different concentrations of transfluthrin and metofluthrin have been assayed for genotoxicity by using the Wing Spot Test on Drosophila melanogaster. Standard cross was used in the experiment. Third-instar larvae that were trans-heterozygous for the two genetic markers mwh and flr³ were treated at different concentrations (0.0103 mg mL⁻¹, 0.103 mg mL⁻¹ for transfluthrin and 6 μg mL⁻¹, 60 μg mL⁻¹ for metofluthrin) of the test compounds. Feeding ended with pupation of the surviving larvae and the genetic changes induced in somatic cells of the wing’s imaginal discs lead to the formation of mutant clones on the wing blade. Results indicated that two experimental concentrations of transfluthrin and 60 μg mL⁻¹ metofluthrin showed mutagenic and recombinogenic effects in both the marker-heterozygous (MH) flies and the balancer-heterozygous (BH) flies.     

Growth and photosynthetic responses to copper in wild grapevine

The present study evaluates the tolerance and accumulation potential of Vitis vinifera ssp. sylvestris under moderate and high external Cu levels. A greenhouse experiment was conducted in order to investigate the effects of a range of external Cu concentrations (0–23 mmol L⁻¹) on growth and photosynthetic performance by measuring gas exchange, chlorophyll fluorescence parameters and photosynthetic pigments. We also measured the total copper, nitrogen, phosphorus, sulphur, calcium, magnesium, iron, potassium and sodium concentrations in the plant tissues. All the experimental plants survived even with external Cu concentrations as high as 23 mmol L⁻¹ (1500 mg Cu L⁻¹), although the excess of metal resulted in a biomass reduction of 35%. The effects of Cu on growth were linked to a reduction in net photosynthesis, which may be related to the effect of the high concentration of the metal on photosynthetic electron transport. V. vinifera ssp. sylvestris survived with leaf Cu concentrations as high as 80 mg kg⁻¹ DW and growth parameters were unaffected by leaf tissue concentrations of 35 mg Cu kg⁻¹ DW. The results of our study indicate that plants of V. vinifera ssp. sylvestris from the studied population are more tolerant to Cu than the commercial varieties of grapevine that have been studied in the literature, and could constitute a basis for the genetic improvement of Cu tolerance in grapevine.     

Effects of dioxin exposure in Eisenia andrei: integration of biomarker data by an Expert System to rank the development of pollutant-induced stress syndrome in earthworms

A battery of biomarkers has recently been developed in the earthworm Eisenia andrei. In this study, different biomarkers (i.e. Ca²⁺-ATPase activity, lysosomal membrane stability-LMS, lysosomal lipofuscin and neutral lipid content) were utilized to evaluate the alterations in the physiological status of animals, induced by exposure for 3 d to different sublethal concentrations of TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) (1.5 × 10⁻³, 1.5 × 10⁻², 1.5 × 10⁻¹ ng mL⁻¹) utilizing the paper contact toxicity test. Lysosome/cytoplasm volume ratio and DNA damage were also evaluated as a biomarker at the tissue level and as a biomarker of genotoxicity, respectively. Moreover, the NR retention time assay conditions were optimized for the determination of in vivo LMS in earthworm coelomocytes. The results demonstrate that LMS and Ca²⁺-ATPase activity were early warning biomarkers able to detect the effects of minimal amounts of TCDD and that biomarkers evaluated at the tissue level are important for following the evolution of the stress syndrome in earthworms. To evaluate the health status of the animals, an Earthworm Expert System (EES) for biomarker data integration and interpretation was developed. The EES proved to be a suitable tool able to rank, objectively, the different levels of the stress syndrome in E. andrei induced by the different concentrations of TCDD.     

Influence of environmental factors on the response of a natural population of Daphnia magna (Crustacea: Cladocera) to spinosad and Bacillus thuringiensis israelensis in Mediterranean coastal wetlands

The present study was undertaken to assess the impact of a candidate mosquito larvicide, spinosad (8, 17 and 33 μg L⁻¹) on a field population of Daphnia magna under natural variations of water temperature and salinity, using Bti (0.16 and 0.50 μL L⁻¹) as the reference larvicide. Microcosms (125 L) were placed in a shallow temporary marsh where D. magna was naturally present. The peak of salinity observed during the 21-day observation period may have been partly responsible for the decrease of daphnid population density in all the microcosms. It is also probably responsible for the absence of recovery in the microcosms treated with spinosad which caused a sharp decrease of D. magna abundance within the first two days following treatment whereas Bti had no effect. These results suggest that it may be difficult for a field population of daphnids to cope simultaneously with natural (water salinity and temperature) and anthropogenic (larvicides) stressors.     

Does nitrogen deposition increase forest production? The role of phosphorus

Effects of elevated N deposition on forest aboveground biomass were evaluated using long-term data from N addition experiments and from forest observation plots in Switzerland. N addition experiments with saplings were established both on calcareous and on acidic soils, in 3 plots with Fagus sylvatica and in 4 plots with Picea abies. The treatments were conducted during 15 years and consisted of additions of dry NH₄NO₃ at rates of 0, 10, 20, 40, 80, and 160 kg N ha⁻¹ yr⁻¹. The same tree species were observed in permanent forest observation plots covering the time span between 1984 and 2007, at modeled N deposition rates of 12-46 kg N ha⁻¹ yr⁻¹. Experimental N addition resulted in either no change or in a decreased shoot growth and in a reduced phosphorus concentration in the foliage in all experimental plots. In the forest, a decrease of foliar P concentration was observed between 1984 and 2007, resulting in insufficient concentrations in 71% and 67% of the Fagus and Picea plots, respectively, and in an increasing N:P ratio in Fagus. Stem increment decreased during the observation period even if corrected for age. Forest observations suggest an increasing P limitation in Swiss forests especially in Fagus which is accompanied by a growth decrease whereas the N addition experiments support the hypothesis that elevated N deposition is an important cause for this development.