Kristina Arnebrant

Growth rate and response of bacterial communities to pH in limed and ash treated forest soils

Abstract Culturable and total bacterial counts, bacterial growth rate and tolerance to pH, as well as microbial biomass, were studied in two coniferous forest soils. The pH had been changed by addition of lime and wood-ash from 4.3–4.4 to 7.0 in one soil and from 3.9–4.4 to 6.1 in the other. Higher total microbial activities and higher bacterial growth rates, measured as soil respiration rate and thymidine incorporation rate, respectively, were found in the treatments with increased pH. Similar effects of soil pH on the thymidine incorporation rate were found using two different methods, by measuring rates in either a soil slurry with all bacteria present or using a subsample of bacteria extracted from soil after homogenization-centrifugation. The number of culturable bacteria was up to 5.1 times higher in the high pH soils, while the acridine orange direct counts were unaffected by the treatments. Thus, the proportions of culturable bacteria increased in the limed and ash-treated soils compared to the untreated controls. An altered bacterial community composition due to liming was indicated by an altered response to pH, where the pH of the soil was correlated to the optimum pH for growth of the bacterial community. The ATP content of the soil was unaffected or increased in treatments with high pH compared to the control, while microbial biomass estimated by the substrate induced respiration technique (SIR) was always higher in limed and ash-treated plots.

Nitrogen amendments reduce the growth of extramatrical ectomycorrhizal mycelium

The effect of three different nitrogen sources on the growth of external ectomycorrhizal mycelium was studied in Perspex micorocosms. Nonsterile peat was used as substrate. Five different fungal isolates growing in symbiosis with pine seedlings were investigated: two isolates of Paxillus involutus, one of Suillus bovinus and two unidentified ectomycorrhizal fungi isolated from ectomycorrhizal root tips. Three different nitrogen sources were used: ammonium as (NH4)2SO4, nitrate as NaNO3 and a complete nutrient solution (Ingestad 1979), and three different nitrogen concentrations, 1, 2 or 4 mg N/g dry wt. of peat. The mycelial growth of all fungi was found to be negatively affected by the nitrogen amendments, although the sensitivity to nitrogen varied between the isolates. One of the unidentified isolates was extremely sensitive and growth was completely inhibited by all nitrogen treatments. In contrast, the growth of one of the P. involutus isolates was only slightly reduced by the nitrogen amendments. The different nitrogen sources all reduced growth, and since no significant difference was found between the nitrogen sources or between the different nitrogen concentrations the results were pooled to give one value that summarized the effect of nitrogen on mycelial growth. Thus, the mycelial growth of one of the two P. involutus isolates was reduced to approximately 80% of the growth in the control, the other P. involutus and one of the unidentified fungi, vgk 2 89.10, were reduced to 40–50% of the control growth, S. bovinus to 30% of the control and the most sensitive fungus, the unidentified isolate vg 1 87.10, was reduced to 3% of the growth in the control treatment. In all experiments, the shoot to root ratio generally increased, mainly as a result of increased shoot growth.

Changes in microfungal community structure after fertilization of scots pine forest soil with ammonium-nitrate or urea

The microfungal population structure of two Scots pine forest soils in Southern Sweden was studied 8–13 yr after nitrogen fertilization. The different fertilizers used. ammonium nitrate and urea. influenced the species composition in different ways. The main effect of the NH4NO3 treatment was an increased isolation frequency of Penicillium spinulosum and 4Oidiodendron echinulatum, while Penicillium cf. brevi-compactum. Mortierella subgenus Mortierella. Chaunopycnis alba and Oidiodendron griseum were isolated less frequently in ammonium nitrate-amended treatments compared to the non-fertilized control. In the urea-fertilized plots the isolation frequency of P. cf. brevi-compactum increased, while that of C. alba. O. echinulatum, O. griseum and P. spinulosum decreased in comparison with control plots. The changes in species composition were most evident at the highest rate of application, 600 kg N ha−1, especially when NH4NO3 was used as a fertilizer.

Extramatrical mycelial growth, biomass allocation and nitrogen uptake in ectomycorrhizal systems in response to collembolan grazing

The effects of Collembola grazing activities on the ectomycorrhizal symbiosis were studied in laboratory microcosms. Pinus contorta seedlings in association with Paxillus involutus were grown in a sandy soil and the Collembola Onychiurus armatus was added in different densities. To study effects on nutrient uptake by the extramatrical mycorrhizal mycelium, cups containing 15NH4+ and phytin amended soil were evenly distributed in the microcosms. These cups were covered with a net that allowed the mycelium to penetrate but neither Collembola nor plant roots. Extramatrical hyphal growth was impeded at a high density of O. Armatus. Low densities of O. armatus increased the extramatrical hyphal growth, the colonization rate of side plants and the biomass of P. involutus. However, the amount of P. involutus on/in the plant roots was not affected. Thus, low densities of collembolans induced a shift towards a larger proportion of P. involutus growing extramatrically. The presence of O. armatus in low numbers enhanced the uptake and transfer of 15N by P. involvus to the plants by up to 76%. The Collembola population growth was not higher in mycorrhizal compared with non-mycorrhizal microcosms while nematode population size was reduced in mycorrhizal compared with non-mycorrhizal microcosms. (Less)

Effects of different fertilizer treatments on ectomycorrhizal colonization potential in two Scots pine forests in Sweden

Ectomycorrhizal fungal colonization potential was studied in two low-productivity Scots pine (Pinus sylvestris L.) forests in Sweden treated with different fertilizers. Young Pinus sylvestris seedlings were used as baits for ectomycorrhizal fungi for one growing season and the total ectomycorrhizal colonization level and frequencies of different types were recorded. One of the experimental sites had been fertilized once with 600 kg nitrogen (N) ha-1, either as ammonium nitrate or urea, 13 years previously. At this site no difference in the total number of ectomycorrhizal root tips was found between the different treatments and approximately 70% of all root tips were ectomycorrhizal. The other experimental site had been continuously fertilized for 15 years with a balanced nutrient mixture, applied either daily as a solution (IF treatment), or as solid fertilizers once a year (F treatment). The IF treatment had received 1700 kg N ha-1 and the F treatment 950 kg N ha-1 until our study was conducted. At this site a significant decrease in total ectomycorrhizal infection level was found as an effect of the fertilizer treatment. In the non-fertilized plots, 70% of all root tips were ectomycorrhizal and the corresponding figure for fertilized plots was 55%. At both sites the frequency of one of the eight classified ectomycorrhizal types decreased significantly in all fertilized plots except in the urea treatment. (Less)

Copper Tolerance of Microfungi Isolated from Polluted and Unpolluted Forest Soil

Microfungi were isolated from unpolluted and copper polluted forest soils and their tolerance was tested on agar plates. Species from polluted sites were usually tolerant to Cu. The species were Paecilomyces farinosus, Beauveria bassiana, Verticillium spp. and Penicillium pactum. Sensitive species were found among Mortierella, Oidiodendron and Penicillium. usually rare in polluted sites. Little evidence for adaptation to Cu was found among isolates with short or long history of pollution.

Soil microbial activity in eleven Swedish coniferous forests in relation to site fertility and nitrogen fertilization

The effect of forest fertilization on soil microbial activity and biomass was studied in 11 coniferous forests with different site fertility. Soil microbial activity was measured as respiration rate and the microbial biomass was estimated using ATP content and substrate‐induced respiration rate (SIR). At all sites, except those with the highest site fertility, the microbial activity and biomass were reduced in the fertilized treatments compared with the control. The general trend indicated that the lower the site fertility, the more pronounced was the reduction of microbial biomass and activity. Some possible explanations for this effect are discussed.

Uptake of N-15-labelled alanine, ammonium and nitrate in Pinus sylvestris L. Ectomycorrhiza growing in forest soil treated with nitrogen, sulphur or lime

The uptake of 15N-labelled alanine, ammonium and nitrate was studied in ectomycorrhizal morphotypes of intact Pinus sylvestris seedlings. PCR-RFLP analysis of the ITS-region of fungal rDNA was used to identify the morphotypes. Seedlings were grown in forest soil collected at an experimental site in southern Sweden. The treatments compared were a control, N fertilisation (600 kg N ha-1 as urea), sulfur application (1200 kg S ha-1) and lime application (6000 kg CaCO3 ha-1). The forest, which had been dominated by Picea abies, was clear-cut two years before the forest soil was sampled. Soil was also collected from an adjacent standing forest. The aim of the present study was to detect changes in the ectomycorrhizal communities in forest soils and relate these changes to the functional parameter of uptake of nitrogen from organic (alanine and protein) and inorganic (ammonium and nitrate) sources.Liming resulted in the detection of a morphotype not found in other samples, and one morphotype was only found in samples from the standing forest (the fungi in these two morphotypes could not be identified). All mycorrhizal root tips showed a higher 15N concentration after exposure to different nitrogen forms than non-mycorrhizal long roots. Uptake of15 N from a labelled solution of alanine or ammonium was higher (about tenfold) than uptake from a 15N-labelled solution of nitrate. Uptake of ammonium and alanine varied between 0.2 and 0.5 mg N g-1 h-1 and between 0.1 and 0.33 mg N g-1 h-1, respectively, among the different morphotypes.In seedlings grown in the control soil and in soil from standing forest, alanine and ammonium were taken up to a similar degree from a supply solution by all morphotypes, whereas ammonium uptake was higher than alanine uptake in seedlings grown in lime-treated soil (about twofold) and, to a lesser extent, in the nitrogen- and sulfur-treated soils. The higher ammonium uptake by morphotypes from the limed soil was confirmed in pure culture studies. In cases where ammonium was used as the N source, an isolate of the S. variegatus morphotype collected in the limed soil produced more biomass compared with isolates of S. variegatus collected in nitrogen- or sulphur-treated soil. One isolate of a silvery white morphotype produced about equal amounts of biomass on alanine and ammonium, whereas all S. variegatus isolated performed better with ammonium as their N source. Based on the results it is hypothesised that liming can induce a shift in the ectomycorrhizal community, favouring individuals that mainly utilise inorganic nitrogen over those that primarily utilise organic nitrogen.

Measurements of ATP in forest humus

The measurement of ATP as an estimate of microbial biomass C in humus was studied. Three different acid extractants, H2SO4, TCA and H3PO4 were compared, as well as two different buffers, Tris and arsenate. No difference in extraction efficiency was found when the light output was converted to ATP content using internal ATP standards, but the different extraction agents and buffers influenced the light output. Several pretreatments and storage conditions were investigated. Incubation at 25°C for 1–7 days gave ca 50% higher ATP values than incubation at 4°C. Sieving did not affect the result. For long-term storage of humus, 4°C is recommended instead of freezing, since freezing reduced the ATP content to ca 80% of the amount found in humus stored at 4°C after 2 months. Freezing of the humus extract resulted in even larger losses of ATP. Humus from eight coniferous forests in Sweden was investigated with respect to ATP content and microbial biomass, estimated with the substrate induced respiration rate method (SIR). A close linear relationship (r = 0.95) was found between the two types of measurement. Calculated from this relationship the amount of ATP (at 25°C) was 3.2μmg−1 biomass C.

Microbial biomass and ATP in smelter-polluted forest humus

Many aspects of microbial activity in soil have been studied in connection with heavy metal pollution, but few investigations have included microbial biomass. To study how biomass-C and ATP were affected over a wide range of metal concentrations, these variables have been measured around the Gusum brass mill in south Sweden. Near the smelter more than 20,000 ppm Cu + Zn g{sup {minus}1} dry soil have been found. This area has been extensively studied form microbiological, zoological and botanical points of view.