Roots Of Pinus Sylvestris Research Articles

Iron and reactive oxygen responses in Pinus sylvestris root cortical cells infected with different species of Heterobasidion annosum sensu lato

Defence mechanisms in trees are not well understood. We assessed whether distribution of iron ions and their co-localisation with reactive oxygen species in Pinus sylvestris root cells reflect differential preferences of the pathogens Heterobasidion annosum sensu stricto, H. parviporum and H. abietinum to the host. Strains of H. annosum s.s. characterised by a greater preference for P. sylvestris induced accumulation of superoxide (O(2)(-)) in host cells 6 h after inoculation, whereas two peaks in accumulation of O(2)(-) (after 4 and 48 h) were observed after infection with strains of the pathogens H. parviporum and H. abietinum, which have a lower preference for P. sylvestris. Moreover, strains of H. annosum s.s. caused increased production of hydrogen peroxide (H(2)O(2)) in P. sylvestris cells, in contrast with strains of the other two species (H. parviporum and H. abietinum). Following inoculation with H. annosum s.s. strains, H(2)O(2) was correlated negatively with O(2)(-) and correlated positively with ferrous iron (Fe(2+)). Co-localisation of Fe(3+) with H(2)O(2) may suggest that they are involved in inducing hypersensitive responses and eventually cell death in roots inoculated with H. annosum s.s. strains, in contrast with H. parviporum, in which other mechanisms operate when the host is parasitised.

High resolution, quantitative reconstruction of erosion rates based on anatomical changes in exposed roots at Draix, Alpes de Haute-Provence — critical review of existing approaches and independent quality control of results

Extensive areas of the French Alps are underlain by Jurassic black marls. Wherever these “terres noires” crop out they become subject to intense erosion, causing major sedimentation in regional reservoirs and river systems. In the badlands near Draix (Alpes de Haute-Provence, France), measured sediment rates were obtained at the plot scale by surface elevation change-based methods and at the catchment scale by monitoring sedimentation in dams. In this study, we use a dendrogeomorphic approach based on anatomical changes in exposed roots of Pinus sylvestris L. to accurately quantify continuous denudation rates. A total of 123 cross sections (75 from buried and 48 from exposed roots of 23 trees) were sampled in the Moulin basin. The size and position of roots at the time of exposure was determined via anatomical variations in the annual growth rings of roots. In cross sections of buried roots, a sharp reduction of earlywood tracheid lumen area — a growth signature which has traditionally been used to determine the moment of root denudation — was observed as soon as erosion reduced soil cover to ≤ 3 cm. As a consequence, estimates of eroded soil thickness had to be adjusted to take account of this bias. Bias-adjusted, averaged, medium-term erosion rates derived from exposed roots vary between 6 and 7 mm y –1 at Moulin basin depending on the importance accorded to the uplift of roots after exposure. Values are significantly correlated to slope angle and match with erosion rates derived from monitored iron stakes (5.7 mm y –1) or measurements of sediment yield in retention dams (4.7 mm y –1) at the outlet of the Moulin basin. Besides demonstrating that the interpretation of anatomical signatures in tree roots to erosion have to be revised, this paper also shows that dendrogeomorphic analyses of roots are indeed a powerful tool for the quantification of minimal rates of soil erosion in environments where measurements of past activity are not available.

Effect of tree species and mycorrhizal colonization on the archaeal population of boreal forest rhizospheres.

Group 1.1c Crenarchaeota are the predominating archaeal group in acidic boreal forest soils. In this study, we show that the detection frequency of 1.1c crenarchaeotal 16S rRNA genes in the rhizospheres of the boreal forest trees increased following colonization by the ectomycorrhizal fungus Paxillus involutus. This effect was very clear in the fine roots of Pinus sylvestris, Picea abies, and Betula pendula, the most common forest trees in Finland. The nonmycorrhizal fine roots had a clearly different composition of archaeal 16S rRNA genes in comparison to the mycorrhizal fine roots. In the phylogenetic analysis, the 1.1c crenarchaeotal 16S rRNA gene sequences obtained from the fine roots formed a well-defined cluster separate from the mycorrhizal ones. Alnus glutinosa differed from the other trees by having high diversity and detection levels of Crenarchaeota both on fine roots and on mycorrhizas as well as by harboring a distinct archaeal flora. The similarity of the archaeal populations in rhizospheres of the different tree species was increased upon colonization by the ectomycorrhizal fungus. A minority of the sequences obtained from the mycorrhizas belonged to Euryarchaeota (order Halobacteriales).

Transcriptional analysis of Pinus sylvestris roots challenged with the ectomycorrhizal fungus Laccaria bicolor.

BackgroundSymbiotic ectomycorrhizal associations of fungi with forest trees play important and economically significant roles in the nutrition, growth and health of boreal forest trees, as well as in nutrient cycling. The ecology and physiology of ectomycorrhizal associations with Pinus sp are very well documented but very little is known about the molecular mechanisms behind these mutualistic interactions with gymnosperms as compared to angiosperms.ResultsUsing a micro-array approach, the relative abundance of 2109 EST transcripts during interaction of Pinus sylvestris roots with the ectomycorrhizal fungus was profiled. The results reveal significant differential expression of a total of 236 ESTs, 96 transcripts differentially abundant after 1 day of physical contact with the fungus, 134 transcripts after 5 days and only 6 after 15 days at early stages of mantle formation on emerging lateral roots. A subset of cell wall modification and stress related genes was further assessed by quantitative reverse transcription PCR at late stages of mycorrhizal development coinciding with Hartig net formation. The results reveal down regulation of gene transcripts involved in general defence mechanism (e.g. antimicrobial peptide) as well as those involved in cell wall modification (e.g. glycine rich protein, xyloglucan endo transglycosylase).ConclusionThis study constitutes the first attempt to characterize the transcriptome of the plant partner in the Pinus sylvestris – Laccaria bicolor model system. We identified 236 ESTs which are potentially important for molecular regulation of a functional symbiotic association in conifer host. The results highlight similarities with other studies based on angiosperm model systems, nevertheless some differences were found in the timing and spatial scale of gene regulation during ectomycorrhiza development in gymnosperms. The present study has identified a number of potentially important molecular events responsible for the initiation and regulation of biochemical, physiological and morphological changes during development of a fully functional symbiosis that are relevant for gymnosperm hosts.

Cloning of Pinus sylvestris SCARECROW gene and its expression pattern in the pine root system, mycorrhiza and NPA‐treated short roots

The SCARECROW (SCR) gene is central to root radial patterning. Its expression has not been investigated in conifers with morphologically different root types. Additional interest in SCR functions in the Pinus sylvestris root system comes from the effect of ectomycorrhiza formation on the short root apical structure. Here, the P. sylvestris SCR gene (PsySCR) was cloned and its expression investigated by northern blot and in situ hybridization of primary, lateral and short roots and mycorrhiza. Short root dichotomization was induced by auxin transport inhibitor (N-1-naphthylphthalamic acid (NPA)). PsySCR has conserved GRAS family protein motifs at the C-terminus and a variable N-terminus. PsySCR expression occurred in young root tissue and mycorrhiza. In root sections the PsySCR signal runs through the tip in initials for stele and root cap column and becomes upwards-restricted to endodermis in all root types. The PsySCR expression pattern suggests for the first time a regulatory role for SCR in maintaining the endodermal characteristics and radial patterning of roots with open meristem organization. The specific PsySCR localization is also an excellent marker for investigation of the dichotomization process in short roots.

Fungi in decayed roots of conifer seedlings in forest nurseries, afforested clear‐cuts and abandoned farmland

Fungi colonizing decayed roots of Pinus sylvestris and Picea abies seedlings were assessed by pure‐culture isolation and direct sequencing of DNA extracted from roots collected from three environments: bare‐root forest nurseries; afforested clear‐cuts; and abandoned farmland. Pure‐culture isolation from 1500 roots collected from 480 seedlings (240 of each tree species) yielded 1110 isolates which, based on mycelial morphology and ITS rDNA sequencing, were found to represent 87 distinct taxa. Direct ITS rDNA sequencing from decayed sections of 140 roots (70 of each tree species) yielded 160 sequences representing 58 taxa. Direct sequencing revealed a significantly higher fungal diversity per root segment than pure‐culture isolation. Overall, a total of 131 taxa were found, 92 of which (70·2%) were identified at least to genus level. Only 14 taxa (10·7%) were detected by both methods, while 73 (55·7%) were detected exclusively by isolation and 44 (33·6%), exclusively by sequencing. The pathogens Fusarium oxysporum (25·6%) and Nectria radicicola (14·9%) were the most common isolates. In contrast, direct sequencing most frequently detected endophyte Phialocephala fortinii (33·1%) and Chalara sp. NS234A2 (10·0%). There were no significant differences in species richness between roots from the different environments, but there was a marked effect on fungal community structure. The results demonstrate that different fungi inhabit decayed roots of conifer seedlings in different environments, and that pure‐culture isolation and direct sequencing are complementary methods that are both necessary for a complete description of the fungal communities colonizing diseased conifer roots.

Chemical Characteristics and Biological Activity of Organic Substances Extracted from Soils by Root Exudates

Plants have evolved with roots in close contact with the solid phase of the soil. Therefore, root exudates may be a better medium for extracting low molecular size (LMS) organic fractions than currently used alkaline solutions. Our objective was to compare the chemical and biological activity of LMS extracts using maize (Zea mays L.), Picea abies, and Pinus sylvestris root exudates to humic substances (HS) extracted with alkaline solution. Gas chromatographic/mass spectrometric (GC/MS) spectra revealed that the LMS fractions had a greater variety of fatty acids than the HS. Fourier transform infrared (FT‐IR) spectra of LMS fractions also indicated different amounts of functional groups by comparison with HS. The possible biological role of LMS fractions with respect to HS was assessed by measuring hormone‐like activity and nitrate uptake in P. sylvestris seedlings. The LMS fractions from agricultural soil stimulated nitrate uptake and nitrate reductase (NR) and glutamine synthetase (GS) activities, whereas those from a forest soil increased ammonium uptake, NR, and glutamate dehydrogenase (GDH) activities. The stimulation of nitrate and ammonium uptake via a NR‐GS or NR‐GDH metabolic pattern was consistent with the different chemical composition of the LMS fractions. This indicates LMS fractions in soil have consequential affects on the plant's capacity to adapt to different environmental conditions.

Sheet erosion rates determined by using dendrogeomorphological analysis of exposed tree roots: Two examples from Central Spain

This paper describes the determination of sheet erosion rates by using dendrogeomorphological methods on exposed tree roots. Two sites on the northern slope of the Guadarrama Mountains, Central Spain, were studied: a popular trail in a Scots pine forest (Senda Schmidt, Valsaín) growing on granites and gneisses, and an open holm-oak forest on granitic slopes (Monterrubio). These sites were selected because they showed high denudation morphologies due to accelerated soil-erosion processes caused by human influence (trampling by continuous trekking and overgrazing), resulting in exposed roots. The method applied is based on the morphological pattern of roots, defined by the growth-ring series of the sampled roots. In order to confirm the validity of the criteria used and to make the estimations of erosion more accurate, several anatomical indicators of exposed and non-exposed Pinus sylvestris roots were characterized. The study entailed a statistical analysis of exposure time and erosion depth. The influence of environmental factors affecting the variation in velocity of the erosion processes was also examined. With a significance level of 95%, the mean erosion rates were in the range of 1.7–2.6 mm/year (29–44 t/ha/year) on Senda Schmidt over the last 101 years, and 1.1–1.8 mm/year (19–31 t/ha/year) in Monterrubio over the last 42 years. Using a multifactor analysis of variance, we observed a change in the erosion rates as a function of position on the path along Senda Schmidt. In Monterrubio, however, we reached no significant conclusions, apart from an inverse relationship between erosion and slope gradient that was difficult to interpret. Climate conditions in Senda Schmidt and the accuracy of dating Scots pine indicate that the evaluation on P. sylvestris roots is fairly reliable, which is not the case for oak roots. Although this paper is based on the application of an existing method, its novelty lies in being the first attempt in Spain to estimate ‘accelerated’ sheet erosion rates (due to recreational activities and overgrazing) using dendrogeomorphological techniques, supplemented by anatomical indicators for P. sylvestris.

A GC-MS method for determination of amino acid uptake by plants.

In this study, we present a rapid, robust and sensitive method for quantification of plant amino acid uptake using universally (U) (13C, 15N)-labelled amino acids and gas chromatography-mass spectrometry (GC-MS). Amino acids were analysed as their tert-butyldimethylsilyl (tBDMS) derivatives and displayed detection limits in the range 10-100 fmol on column, depending on the amino acid. The technique allows for simultaneous detection and quantification of both unlabelled and isotopically labelled species of amino acids. This makes simple quantification of plant amino acid uptake from an isotopically labelled source possible. The analytical variation was low, concerning total amino acid concentrations (relative standard deviation, rsd, less than 5.3%) as well as enrichment of U-13C, 15N-labelled glycine (Gly), arginine (Arg) and glutamic acid (Glu) (rsd<2.1%). An application of the GC-MS method was conducted on non-mycorrhizal Pinus sylvestris roots supplied with U-13C, 15N-labelled amino acids. Intact, labelled amino acids were traced in root extracts. This provided conclusive evidence of plant root uptake of intact amino acids. Uptake rates of the three amino acids Gly, Glu and Arg in the range 0.5-37.9 &mgr;mol g-1 dry weight h-1 were recorded. These rates are comparable with those recorded in earlier studies of amino acid uptake, using other methods, as well as uptake rates measured for nitrate and ammonium.

Subcellular compartmentation of elements in non‐mycorrhizal and mycorrhizal roots of Pinus sylvestris: an X‐ray microanalytical study. I. The distribution of phosphate

The inter‐ and intracellular distribution of phosphate in non‐mycorrhizal and ectomycorrhizal roots of Pinus sylvestris was analysed by energy‐dispersive X‐ray spectroscopy after cryofixation, freeze‐drying, pressure infiltration with resin and dry cutting of the material. The results show (1) that in non‐mycorrhizal roots the distribution of phosphate across the root cross section is relatively homogenous, (2) that the intracellular distribution of phosphate depends on the external supply, and (3) that the phosphate absorption can be increased by a supply of NH4. Under phosphate starvation, the phosphate is mainly localized in the cell cytoplasm, whereas under high external phosphate this element accumulates in the vacuoles of cortical cells. It can be inferred that at low external conditions phosphate is mainly located in the cytoplasm of cells to protect metabolism from deficiency, that phosphate is translocated to the shoot via symplastic and apoplastic pathways, and that especially the vacuolar pool size is regulated by the external phosphate supply. The nutrition of the host plant under phosphate starvation was improved by a mycorrhizal infection with Suillus bovinus. However, this effect might not be the same for all ectomycorrhizal infections and supply conditions. With high external phosphate an ectomycorrhizal infection with S. bovinus had no effect on intracellular phosphate within the roots, the shoot contents of mycorrhizal plants were less than those of non‐mycorrhizal seedlings and translocation rate across the mycorrhizal interface appeared to be independent of the availability of phosphate. Intracellular phosphate in fungal cells of mycorrhizal roots was independent of the external supply and seemed to be regulated by the formation of polyphosphates. A supply of (NH4)2HPO4, which led to an increase in cytoplasmic phosphate levels in the hyphae of the Hartig net of the Paxillus involutus mycorrhiza also led to higher phosphate in the plant cell compartments. Therefore, it can be inferred that the translocation of phosphate from fungus to host plant across the mycorrhizal interface is regulated by the phosphate concentration in the cytoplasm of the Hartig net and by the efflux rate into the interfacial apoplast.

Fine Root Production and Nitrogen Content in Roots of Pinus sylvestris L. after Clear-felling

Effects of clear-felling on fine root production and N content in roots were studied in a Scots pine (Pinus sylvestris L.) stand. A plot was clear-felled and root samples were compared with a control plot. Even if reduced by 50%, a significant fine root (diameter<2 mm) growth was noticed up to 1 yr after the clear-felling. For live roots (diameters 2-4 and 4-6 mm) nitrogen content in root-wood and root-bark samples from the clear-felling, compared with the reference plot, were 30-50% already higher 2 months after the clear-felling. The difference did not increase in later comparisons. Nitrogen content in buttresses did not differ during the period studied, except for a higher percentage in bark at the clear-felling after two summers. A second study confirmed the results. It is suggested that there exists a continued nitrogen uptake in live roots after a clear-felling.

Reactions of Pinus sylvestris (Scots Pine) Root Tissues to the Presence of Mutualistic, Saprotrophic and Necrotrophic Micro-organisms

Using Scots pine seedlings as experimental hosts, the general reactions and recognition patterns of living conifer tissues challenged in vitro with either parasitic, saprotrophic, epiphytic or mutualistic microbial species were investigated. Three distinct reaction and recognition patterns were observed. In one, a necrosis type of browning reaction occurred that restricted vascular penetration of the seedlings when inoculated with a saprotroph (Marasmius androsaceus) or a mycorrhizal-associated bacterium (Pseudomonas fluorescens). The second pattern, a kind of necrosis in which the host defence system was ultimately ineffective, was typically observed in seedlings challenged with the S or P types of the root rot fungus (Heterobasidion annosum), the fine root parasite (Fusarium avenaceum) or the saprotrophic wood decay fungus (Phlebia gigantea). The third type, described as non-recognition, was observed on seedlings showing no necrosis and was particularly common with seedlings inoculated with either mycorrhizal fungi (Suillus granulatus, Piloderma croceum) or saprotrophs (Trichoderma aureoviride, Coriolus versicolor), or an unidentified rod-shaped bacterial isolate. Since necrosis is often linked to rapid accumulation of H2O2, increases in peroxidase activity were determined. Only seedlings challenged with the S-type of H. annosum had a systemic increase in peroxidase levels of needles, shoot and roots. Similarly, only H. annosum, F. avenaceum, P. gigantea and the Pseudomonas bacterium caused a two- to four-fold increase in the peroxidase level of inoculated roots, results suggesting that a relationship exists between the degree of necrosis and the ratio of peroxidase induction. But the higher levels of peroxidase activity was not correlated with increased resistance to disease development. Zusammenfassung Reaktionen von Wurzelgewebe der Gemeinen Kiefer (Pinus sylvestris) auf die Anwesenheit mutualistischer, saprophytischer und nekrotropher Mikroorganismen An Samlingen der Gemeinen Kiefer als experimentelle Wirtspflanzen untersuchten wir allgemeine Reaktionen und Erkennungsmuster lebender Nadelbaumgewebe, die in vitro mit parasitischen, saprophytischen, epiphytischen oder mutualistischen Mikroorganismen herausgefordert wurden. Dabei zeigten sich drei verschiedene Reaktions-und Erkennungsmuster. Das erste war ein Nekrosetyp, bei dem die Verbraunungsreaktion die Penetration in die Gefase der Samlinge begrenzte. Dieser Typ trat auf nach Inokulation mit einem Saprophyten (Marasmius androsaceus) oder einem mit Mykorrhizen assoziierten Bakterium (Pseudomonas fluorescens). Das zweite Muster war ein Typ von Nekrose, bei dem das Abwehrsystem des Wirtes inaktiviert wurde. Es kam typischerweise bei Samlingen vor, die mit dem S- oder dem P-Typ des Wurzelschwammes (Heterobasidion annosum), dem auf Feinwurzeln parasitierenden Pilz Fusarium avenaceum oder dem saprophytischen, holzzerstorenden Pilz Phlebia gigantea herausgefordert worden waren. Der dritte Typ entsprach einer Nicht-Erkennung, hier zeigten die Samlinge keine Nekrose. Diese Reaktion kam vor allem bei Samlingen vor, die mit Mykorrhizapilzen (Suillus granulatus, Piloderma croceum), Saprophyten (Trichoderma aureoviride, Coriolus versicolor) oder einem Isolat eines nicht identifizierten Stabchenbakteriums inokuliert worden waren. Da Nektrosen oft mit einer raschen Akkumulation von H2O2 gekoppelt sind, wurde auch der Anstieg der Peroxidaseaktivitat bestimmt. Lediglich die mit dem S-Typ von H. annosum herausgeforderten Samlinge zeigten einen systemischen Anstieg der Peroxidasewerte in Nadeln, oberirdischen Trieben und Wurzeln. Nur H. annosum, F. avenaceum, P. gigantea und das Pseudomonas-Bakterium riefen einen 2- bis 4-fachen Anstieg der Peroxidasewerte bei inokulierten Wurzeln hervor. Diese Ergebnisse zeigen, das ein Zusammenhang zwischen dem Ausamas der Nekrose und dem Ausmas der Peroxidaseinduktion besteht. Die hohere Peroxidaseaktivat war jedoch nicht mit erhohter Krankheitsresistenz gekoppelt.

Reactions of Pinus sylvestris (Scots Pine) Root Tissues to the Presence of Mutualistic, Saprotrophic and Necrotrophic Micro-organisms

Reactions of Pinus sylvestris (Scots Pine) Root Tissues to the Presence of Mutualistic, Saprotrophic and Necrotrophic Micro-organisms

Developmentally regulated proteins during differentiation of root system and ectomycorrhiza in Scots pine (Pinus sylvestris) with Suillus bovinus

Total and radiolabelled proteins from Pinus sylvestris roots, ectomycorrhiza and the ectomycorrhizal fungus Suillus bovinus, were analyzed after separation by 2‐D gel electrophoresis. The purpose was to address the contribution of individual proteins to the development of ectomycorrhiza. In Pinus sylvestris, ectomycorrhiza forms only in the short roots and therefore special attention was paid to the proteins of short roots before formation of ectomycorrhiza. A main result, from comparisons of protein amounts in main and lateral root tips and short roots, was that the specialized growth pattern of short roots is associated with the production of five short root‐specific proteins, and that several proteins of the main and lateral roots are repressed in short roots. At the morphologically different stages of the ectomycorrhiza, only few changes appeared in the amounts of the host and the symbiont proteins. Only one ectomycorrhiza‐specific protein could be distinguished.

Endogone maritima, a new species in the Endogonales from Poland

Endogone maritima sp. nov. is described and illustrated. This fungus was recovered from around the roots of endomycorrhizal Ammophila arenaria, Corynephorus canescens, Juncus balticus, J. articulatus , and ectomycorrhizal Pinus sylvestris colonizing a deflation hollow of the Slowinski National Park in Poland. Endogone maritima produces sunflower to brownish yellow zygosporangia enveloped by a separable hyphal mantle consisting of interwoven hyphae appearing in section as a dense net. The zygosporangia occur both singly in the soil and in small, non-peridial sporocatps typically containing 2-7 zygosporangia.

Bacterial colonization patterns of intactPinus sylvestrismycorrhizospheres in dry pine forest soil: an electron microscopy study

The bacterial populations associated with different plant and fungal habitats of intact Pinus sylvestris – Suillus bovinus or Pinus sylvestris – Paxillus involutus ectomycorrhizospheres grown in natural forest soil were examined by scanning and transmission electron microscopy. Surfaces of nonmycorrhizal Pinus sylvestris roots hosted large numbers of morphologically distinct bacteria. Bacteria were detected on the mantle surfaces and at inter- and intra-cellular locations in the mantle and Hartig net of Suillus bovinus mycorrhizas. The fungal strands were colonized by only a few bacteria unlike the outermost external fine hyphae on which extensive monolayers of bacteria were attached. The mycorrhizas of Paxillus involutus were mostly devoid of bacteria, but the intact external mycelium supported both bacterial colonies and solitary bacteria. Intracellular bacteria were not present in Paxillus involutus hyphae. In both mycorrhizal systems, bacterial aggregation and attachment to hyphae were mediated with electron-dense or -translucent material. Our study shows that the Pinus sylvestris mycorrhizospheres formed by two different ectomycorrhizal fungi are clearly dissimilar habitats for mycorrhizosphere-associated bacteria. Additionally, the spatially and physiologically defined mycorrhizosphere habitats were shown to host distinct populations of bacteria.Key words: ectomycorrhiza, intracellular bacteria, Paxillus involutus, soil bacteria, Suillus bovinus.

The use of inositol hexaphosphate as a phosphorus source by mycorrhizal and non‐mycorrhizal Scots Pine (Pinus sylvestris)

1. The external mycelia of the ectomycorrhizal fungi Thelephora terrestris and Suillus luteus, associated with Pinus sylvestris roots, exhibited a substantial extracellular acid phosphatase activity. The activity was positively correlated with the ergosterol concentration in the growth substratum and decreased with an increasing P nutrition. 2. The pioneer species T. terrestris grew best at a high Pi nutrition level whereas S. luteus, a ‘late‐stage’ mycobiont, produced more active biomass at a low Pi nutrition level. 3. The phytase activity of the external mycelia could not be detected; at the root surface a phytase activity was observed. Mycorrhizas had significantly higher activities than uninfected roots. 4. The addition of a relatively high concentration of a soluble phytate to the growth substratum resulted in an increased relative growth rate (RGR) in both mycorrhizal and non‐mycorrhizal plants. The influence of the mycorrhizal fungi on the use of the phytate‐P was small, despite the phytase activity of the mycorrhizal feeder roots. 5. The addition of phytate fixed on a HPLC resin did not result in an increase of the RGR and P uptake neither in the non‐mycorrhizal nor in the mycorrhizal Pines. The experiment did not support the hypothesis that phytate, which has a low solubility in soils, is a useful P source for ectomycorrhizal plants.

Phosphate uptake and polyphosphate metabolism of mycorrhizal and nonmycorrhizal roots of pine and of Suillus bovinus at varying external pH measured by in vivo 31 P-NMR

Comparative in vivo 31P-NMR analyses of mycorrhizal and nonmycorrhizal roots of Pinus sylvestris and the fungus of Suillus bovinus in pure culture were used to investigate alterations in phosphate metabolism due to changes in external pH in the range 3.5–8.5. All control samples maintained a constant pH in both cytoplasm and vacuole. Mycorrhizal roots and pure fungus, but not nonmycorrhizal roots, transformed accumulated inorganic phosphate into mobile polyphosphate with a medium chain length. Phosphate uptake rates and polyphosphate accumulation responded differently to external pH. In all cases, maximal phosphate uptake occurred at an external pH close to 5.5. At an external pH of 8.5, both roots and fungus showed a distinct lag in phosphate uptake, which was abolished when the external pH was lowered to 7.5. An irreversible effect on phosphate uptake as a consequence of variation in external pH was also observed. The central role of the fungus in regulating mycorrhizal phosphate metabolism is discussed.

Tubulin and actin protein patterns in Scots pine (Pinus sylvestris) roots and developing ectomycorrhiza with Suillus bovinus

Tubulin and actin protein patterns in Scots pine (Pinus sylvestris) roots and developing ectomycorrhiza with Suillus bovinus

Determination of chitin in fungi and mycorrhizal roots by an improved HPLC analysis of glucosamine

A method to measure chitin content in fungi and ectomycorrhizal roots with high-performance liquid chromatography (HPLC) was developed. Measurements of fluorescence of 9-fluorenylmethylchloroformate (FMOC-CI) derivatives of glucosamine were made on acid hydrolysates of pure chitin, chitin-root mixtures and fungal-root mixtures. The method was applied on 5 isolates of ectomycorrhizal fungi, and ectomycorrhizal and non-mycorrhizal Pinus sylvestris roots. Interference from amino acids was removed by pre-treatment of samples with 0.2 N NaOH. This pre-treatment did not reduce the recovery of chitin, nor did plant material affect the recovery of chitin. The HPLC method was compared with a colorimetric chitin-method by measurements on root-fungal mixtures, with known fungal content. The HPLC method gave estimates of fungal biomass which were equal to the expected while the colorimetric method showed values significantly (p<0.001) lower than the expected. The present chitin method offers a sensitive and specific tool for the quantification of chitin in fungi and in ectomycorrhizal roots.