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Isolation, genetic variation and expression of TIR-NBS-LRR resistance gene analogs from western white pine ( Pinus monticola Dougl. ex. D. Don.).

Western white pine ( Pinus monticola Dougl. ex. D. Don., WWP) shows genetic variation in disease resistance to white pine blister rust ( Cronartium ribicola). Most plant disease resistance (R) genes encode proteins that belong to a superfamily with nucleotide-binding site domains (NBS) and C-terminal leucine-rich repeats (LRR). In this work a PCR strategy was used to clone R gene analogs (RGAs) from WWP using oligonucleotide primers based on the conserved sequence motifs in the NBS domain of angiosperm NBS-LRR genes. Sixty-seven NBS sequences were cloned from disease-resistant trees. BLAST searches in GenBank revealed that they shared significant identity to well-characterized R genes from angiosperms, including L and M genes from flax, the tobacco N gene and the soybean gene LM6. Sequence alignments revealed that the RGAs from WWP contained the conserved motifs identified in angiosperm NBS domains, especially those motifs specific for TIR-NBS-LRR proteins. Phylogenic analysis of plant R genes and RGAs indicated that all cloned WWP RGAs can be grouped into one major branch together with well-known R proteins carrying a TIR domain, suggesting they belong to the subfamily of TIR-NBS-LRR genes. In one phylogenic tree, WWP RGAs were further subdivided into fourteen clusters with an amino acid sequence identity threshold of 75%. cDNA cloning and RT-PCR analysis with gene-specific primers demonstrated that members of 10 of the 14 RGA classes were expressed in foliage tissues, suggesting that a large and diverse NBS-LRR gene family may be functional in conifers. These results provide evidence for the hypothesis that conifer RGAs share a common origin with R genes from angiosperms, and some of them may play important roles in defense mechanisms that confer disease resistance in western white pine. Ratios of non-synonymous to synonymous nucleotide substitutions (Ka/Ks) in the WWP NBS domains were greater than 1 or close to 1, indicating that diversifying selection and/or neutral selection operate on the NBS domains of the WWP RGA family.

Amino Acid Sequence↗

Water source utilization by Pinus jeffreyi and Arctostaphylos patula on thin soils over bedrock.

Stable isotopes were used to evaluate water sources for co-occurring Jeffrey pine (Pinus jeffreyi Grev & Balf.) and greenleaf manzanita (Arctostaphylos patula Greene) in the southern Sierra Nevada, California, where soils averaged only 75 cm thick but were underlain by up to 5 m of weathered granitic bedrock. Soils and underlying weathered bedrock were sampled three times during both the 1997 and 1998 growing seasons, in 25 cm increments, from 0 to 400 cm or until hard bedrock was reached, and plant stem tissue was sampled simultaneously. Extracted water from the soil/bedrock substrate and plant tissue was analyzed for delta(18)O and/or deltaD, and depth of water source was determined by inference in conjunction with moisture status of the substrate. Water source utilization over the growing seasons for both plants generally followed a pattern similar to that observed for water depletion. Predominant water use was initially from the surface soils. Progressively deeper water sources, including weathered bedrock to a depth of several meters, were exploited as the season progressed and the overlying substrate was depleted of moisture. Early in the growing season, stable isotope values were slightly lower for pine than for manzanita (e.g., average deltaD in June 1997 was -81 per thousand for pine and -77 per thousand for manzanita), and suggest that the functional rooting depth for pine may have been slightly greater than for manzanita. In September 1997, manzanita deltaD values averaged -57 per thousand while pine values averaged -85 per thousand, indicating that manzanita opportunistically utilized summer precipitation while pine used more dependable bedrock water. In 1998, soils remained moist through July due to a late snowfall. Unlike the previous year, pine and manzanita deltaD values were not significantly different in mid- and late-growing season, and both plants exploited bedrock-derived water as soil water was depleted. Water held within bedrock was essential for meeting plant transpirational requirements over the summer drought.

Adaptation, Physiological↗

Nutrient conservation increases with latitude of origin in European Pinus sylvestris populations.

Nutrient availability varies across climatic gradients, yet intraspecific adaptation across such gradients in plant traits related to internal cycling and nutrient resorption remains poorly understood. We examined nutrient resorption among six Scots pine (Pinus sylvestris L.) populations of wide-ranging origin grown under common-garden conditions in Poland. These results were compared with mass-based needle N and P for 195 Scots pine stands throughout the species' European range. At the common site, green needle N (r(2)=0.81, P=0.01) and P (r(2)=0.58, P=0.08) concentration increased with increasing latitude of population origin. Resorption efficiency (the proportion of the leaf nutrient pool resorbed during senescence) of N and P of Scots pine populations increased with the latitude of seed origin (r(2) > or = 0.67, P < or = 0.05). The greater resorption efficiency of more northerly populations led to lower concentrations of N and P in senescent leaves (higher resorption proficiency) than populations originating from low latitudes. The direction of change in these traits indicates potential adaptation of populations from northern, colder habitats to more efficient internal nutrient cycling. For native Scots pine stands, results showed greater nutrient conservation in situ in cold-adapted northern populations, via extended needle longevity (from 2 to 3 years at 50 degrees N to 7 years at 70 degrees N), and greater resorption efficiency and proficiency, with their greater resorption efficiency and proficiency having genotypic roots demonstrated in the common-garden experiment. However, for native Scots pine stands, green needle N decreased with increasing latitude (r(2)=0.83, P=0.0002), and P was stable other than decreasing above 62 degrees N. Hence, the genotypic tendency towards maintenance of higher nutrient concentrations in green foliage and effective nutrient resorption, demonstrated by northern populations in the common garden, did not entirely compensate for presumed nutrient availability limitations along the in situ latitudinal temperature gradient.

Adaptation, Physiological↗

Delta(13)C and tree-ring width reflect different drought responses in Quercus ilex and Pinus halepensis.

Holm oak (Quercus ilexL.) and Aleppo pine (Pinus halepensisMill) are representative of two different functional types of trees extensively found in the Mediterranean: evergreen sclerophyllous and drought-adapted conifers. The former is considered a partially drought-tolerant species, whereas the latter is a typically drought-avoiding, water-saving species. We postulated that contrasting strategies in response to water deficits in Q. ilex and P. halepensis would lead to a differential sensitivity to changes in water availability. To test this hypothesis, we compared the response of both species in growth rate (measured as radial increments) and intrinsic water use efficiency WUE(i), as inferred from carbon isotope discrimination (Delta(13)C) in wood samples] among sites from different provenance regions in NE Spain. We found significant differences in Delta(13)C and growth among provenance regions, partly explained by contrasting water availability. Wood Delta(13)C was positively related with precipitation and the ratio between precipitation and potential evapotranspiration (P/ E). However, these relationships were stronger in P. halepensis (for P / E, r(2)=0.67, P <0.001) than in Q. ilex ( r(2)=0.42, P <0.01). In addition, radial growth was positively related with precipitation and Delta(13)C in P. halepensis ( r(2)=0.32 and r(2)=0.35, respectively, P <0.01), but not in Q. ilex. We concluded that P. halepensis was more sensitive than Q. ilex to water availability, showing faster increase in WUE(i) in response to water stress. We also found that the effect of north/south aspect on Delta(13)C and growth was site-specific, and unrelated to climatic variables.

Adaptation, Physiological↗

Effectiveness of six species of rodents as dispersers of singleleaf piñon pine (Pinus monophylla).

The effectiveness of six species of rodents as dispersers of singleleaf piñon pine (Pinus monophylla) was investigated in field enclosures. The rodent species were Panamint kangaroo rat (Dipodomys panamintinus), Great Basin pocket mouse (Perognathus parvus), pinyon mouse (Peromyscus truei), deer mouse (Peromyscus maniculatus), least chipmunk (Tamias minimus), and white-tailed antelope ground squirrel (Ammospermophilus leucurus). Five measures of seed harvesting and caching behavior were assessed and used to calculate the effect of species on the probability of effective dispersal (i.e., movement of seeds to potential establishment sites) of piñon pine seeds. These measures were proportion of seeds harvested, proportion of harvested seeds that were scatter hoarded, relative dispersion of scatter-hoarded seeds, seedling emergence and seedling establishment. To evaluate the effects of the last two variables, a 2-year study of seed germination and seedling establishment was conducted in the field. Seedling survival was 72-90% for seedlings emerging from 1-4 cm deep but was markedly lower from surface caches and for depths >4 cm. All seedlings in the open microsite perished before the end of the first year, whereas 38-97% of seedlings under shrubs survived. Cache size did not affect seedling survival over the 2 years, but large caches reduced the total number of potential establishment sites. All six species harvested over 90% of the piñon seeds presented and scatter hoarded from 31% (pocket mouse) to 66% (chipmunk) of the seeds they harvested. Piñon mice and deer mice distributed seeds to more cache sites (i.e., put fewer seeds in caches) than the other species and, consequently, created more potential establishment sites. All species scatter hoarded seeds at depths suitable for seedling emergence, but pocket mice and piñon mice cached under shrubs more than the other species, the microhabitat where seedlings had the greatest probability of establishing. Factors that had the greatest effect on the probability of effective dispersal were microhabitat selection and cache size.

Animals↗

Host physiological condition regulates parasitic plant performance: Arceuthobium vaginatum subsp. cryptopodum on Pinus ponderosa.

Much research has focused on effects of plant parasites on host-plant physiology and growth, but little is known about effects of host physiological condition on parasite growth. Using the parasitic dwarf mistletoe Arceuthobium vaginatum subsp. cryptopodum (Viscaceae) and its host Pinus ponderosa, we investigated whether changes in host physiological condition influenced mistletoe shoot development in northern Arizona forests. We conducted two studies in two consecutive years and used forest thinning (i.e., competitive release) to manipulate host physiological condition. We removed dwarf mistletoe shoots in April, before the onset of the growing season, and measured the amount of regrowth in the first season after forest thinning (Study I: n=38 trees; Study II: n=35 trees). Thinning increased tree uptake of water and carbon in both studies, but had no effect on leaf N concentration or delta13C. Mistletoe shoot growth was greater on trees with high uptake of water and carbon in thinned stands than trees with low uptake in unthinned stands. These findings show that increased resource uptake by host trees increases resources to these heterotrophic dwarf mistletoes, and links mistletoe performance to changes in host physiological condition.

Arizona↗

The role of below-ground competition during early stages of secondary succession: the case of 3-year-old Scots pine (Pinus sylvestris L.) seedlings in an abandoned grassland.

In abandoned or extensively managed grasslands, the mechanisms involved in pioneer tree species success are not fully explained. Resource competition among plants and microclimate modifications have been emphasised as possible mechanisms to explain variation of survivorship and growth. In this study, we evaluated a number of mechanisms that may lead to successful survival and growth of seedlings of a pioneer tree species (Pinus sylvestris) in a grass-dominated grassland. Three-year-old Scots pines were planted in an extensively managed grassland of the French Massif Central and for 2 years were either maintained in bare soil or subjected to aerial and below-ground interactions induced by grass vegetation. Soil temperatures were slightly higher in bare soil than under the grass vegetation, but not to an extent explaining pine growth differences. The tall grass canopy reduced light transmission by 77% at ground level and by 20% in the upper part of Scots pine seedlings. Grass vegetation presence also significantly decreased soil volumetric water content (Hv) and soil nitrate in spring and in summer. In these conditions, the average tree height was reduced by 5% compared to trees grown in bare soil, and plant biomass was reduced by 85%. Scots pine intrinsic water-use efficiency (A/g), measured by leaf gas-exchange, increased when Hv decreased owing to a rapid decline of stomatal conductance (g). This result was also confirmed by delta 13C analyses of needles. A summer 15N labelling of seedlings and grass vegetation confirmed the higher NO3 capture capacity of grass vegetation in comparison with Scots pine seedlings. Our results provide evidence that the seedlings' success was linked to tolerance of below-ground resource depletion (particularly water) induced by grass vegetation based on morphological and physiological plasticity as well as to resource conservation.

Carbon Dioxide↗

The role of genetic and chemical variation of Pinus sylvestris seedlings in influencing slug herbivory.

This study investigated the genetic and chemical basis of resistance of Pinus sylvestris seedlings to herbivory by a generalist mollusc, Arion ater. Using feeding trials with captive animals, we examined selective herbivory by A. ater of young P. sylvestris seedlings of different genotypes and correlated preferences with seedling monoterpene levels. We also investigated the feeding responses of A. ater to artificial diets laced with two monoterpenes, Delta(3)-carene and alpha-pinene. Logistic regression indicated that two factors were the best predictors of whether seedlings in the trial would be consumed. Individual slug variation (replicates) was the most significant factor in the model; however, alpha-pinene concentration (also representing beta-pinene, Delta(3)-carene and total monoterpenes due to multicollinearity) of needles was also a significant factor. While A. ater did not select seedlings on the basis of family, seedlings not eaten were significantly higher in levels of alpha-pinene compared to seedlings that were consumed. We also demonstrated significant genetic variation in alpha-pinene concentration of seedlings between different families of P. sylvestris. Nitrogen and three morphological seedling characteristics (stem length, needle length and stem diameter) also showed significant genetic variation between P. sylvestris families. Artificial diets laced with high (5 mg g(-1) dry matter) quantities of either Delta(3)-carene or alpha-pinene, were eaten significantly less than control diets with no added monoterpenes, supporting the results of the seedling feeding trial. This study demonstrates that A. ater selectively feed on P. sylvestris seedlings and that this selection is based, in part, on the monoterpene concentration of seedlings. These results, coupled with significant genetic variation in alpha-pinene concentration of seedlings and evidence that slug herbivory is detrimental to P. sylvestris fitness, are discussed as possible evidence for A. ater as a selective force on the evolution of defensive chemistry in P. sylvestris.

Animals↗

Connections between climatic variables and the growth and needle dynamics of Scots pine (Pinus sylvestris L.) in Estonia and Lapland.

The relationships between climatic variables and Scots pine (Pinus sylvestris L.) growth and needle dynamics were studied in three stands in Estonia and in four stands located near the northern timberline in Lapland. The trees sampled in Estonia had low correlations with the analysed climatic variables (air temperature, precipitation and indices of atmospheric circulation). Moreover, the weak cross-correlation of the time-series of the Estonian sample trees indicated that Scots pine is affected mainly by local factors in that region. In Lapland, however, height increment and needle production correlated strongly among trees within a stand (mean r=0.45 and 0.46, respectively) and between stands (r=0.32 and 0.37). Radial increment also showed a high inter-correlation among the trees within a stand in Lapland (r=0.45). Both height increment and needle production were strongly influenced by the temperature regime of the previous summer in Lapland (mean r=0.64 and 0.64, respectively). Radial increment was correlated with the mean July temperature of the current year (mean r=0.29). The correlations between the indices of atmospheric circulation and tree attributes were weak, while the strongest correlation was between the Ponta Delgada NAO index (PD-NAO) and height increment and needle production in Lapland. Height increment, needle production and radial increment have increased since the 1990s in the trees growing in Lapland. This may indicate a positive effect of climate warming on tree growth in Lapland. In Estonia, where climatic conditions do not limit tree growth, the climate warming seems not to directly influence the growth and needle dynamics of Scots pine.

Climate↗

Spatiotemporal transfer of carbon-14-labelled photosynthate from ectomycorrhizal Pinus densiflora seedlings to extraradical mycelia.

Seedlings of Pinus densiflora colonized by an unidentified ectomycorrhizal fungus (T01) were labelled photosynthetically with 14C. Movement of 14C-labelled photosynthates within the underground part of the seedlings was investigated by temporal autoradiography using an imaging plate. Within 1 day, 14C was transferred from the shoot to the underground part that included roots, mycorrhizae, and the extraradical mycelium; within 3 days, the 14C in the underground part reached its maximum density. Mycorrhizae and actively growing root tips were large C sinks. Three days after 14C labelling, counts of 14C radioactivity in the underground part of the mycorrhizal seedlings were 2.6 times those of nonmycorrhizal seedlings. The mycorrhizae of mycorrhizal plants accumulated 5.2 times the 14C counts in the short-root tips of nonmycorrhizal plants. 14C counts in various areas of the extraradical mycelium demonstrated that all 14C-photosynthate transfer from the host root to the extraradical mycelium occurred within 3 days after 14C labelling, and that there was only a short lag of < 1 day between 14C accumulation in the basal and distal parts of the mycelium. Although more 14C accumulated in the distal than in the basal parts, 14C counts per unit hyphal biomass were similar between the two. These results suggest that 14C spread rapidly throughout the entire mycelium. Thirteen days after 14C labelling, we estimated 14C allocation to extraradical mycelia by taking autoradiographs after removing host roots. About 24% of 14C counts in the underground part of the mycorrhizal seedlings had been allocated to extraradical mycelia in this system, indicating that the fugal mycelium is an important sink for photosynthates.

Autoradiography↗

The ectomycorrhizal symbiosis between Lactarius deliciosus and Pinus sylvestris in forest soil samples: symbiotic efficiency and development on roots of a rDNA internal transcribed spacer-selected isolate of L. deliciosus.

The effect on plant growth of pre-inoculation of Pinus sylvestris with the ectomycorrhizal (ECM) edible basidiomycete Lactarius deliciosus (isolate D45) under controlled conditions, and the development on roots of this basidiomycete, were investigated in gamma-irradiated and unsterilized containers containing different forest soil cores or a perlite-vermiculite mixture. Five months after planting, L. deliciosus mycorrhizal plants exhibited greater growth than the non-mycorrhizal ones in all soil types, i.e. up to a 325% increase in shoot height in the sterilized soils. The experiment demonstrated the dependency of P. sylvestris seedlings upon ECM symbiosis for their survival in gamma-irradiated, microbiologically disturbed soil samples. Furthermore, in two soils, the growth of L. deliciosus-inoculated seedlings was greater in the sterilized soil samples than in the non-sterilized ones, i.e. 46% and 132% increase in shoot height under sterilized soil conditions. In containers randomly sampled from each soil type, the degree of root colonization by the inoculated isolate, calculated as the number of mycorrhizal root tips divided by the total number of root tips x100, ranged from 80% to 35%. Within the short term, the inoculated isolate developed rapidly on roots, dominated, and hampered ectomycorrhiza formation by various unidentified (but not Lactarius) resident ECM fungi in unsterilized soil types. Results indicate that the ECM species L. deliciosus is worth investigating to ascertain if other isolates benefit pine growth like the isolate D45, and are therefore also attractive candidates for forestry applications in the Mediterranean area.

Basidiomycota↗

In vitro evidence of mycoparasitism of the ectomycorrhizal fungus Laccaria laccata against Mucor hiemalis in the rhizosphere of Pinus sylvestris.

Interactions between the ectomycorrhizal fungus Laccaria laccata and the soil fungus Mucor hiemalis f. hiemalis in co-culture, and in the rhizosphere of in vitro-grown Pinus sylvestris seedlings were investigated by light- and scanning electron-microscopy. In co-culture, mycelial growth away from the L. laccata colony reduced the number of aerial hyphae at the contact zone and increased the density and compactness of the mycelium-characterized gross morphology of the saprobic fungus. Although the growth of M. hiemalis was suppressed, no penetration of M. hiemalis hyphae after the colony was entered by L. laccata was observed. Instead, dense coiling of L. laccata hyphae around sporangiophores, overpowering them and causing them to disappear, was quite common. On nonmycorrhizal roots, sporangiospores germinated heavily and formed long hyphae for 2 days post inoculation, whereas their germination was totally inhibited on mycorrhizal roots. At 3 days after inoculation, only sporangia were seen with mycelial mats firmly attached to the roots by the mantle hyphae, whereas some remnants of sporangiophores, ruptured sporangial walls and degraded hyphae of M. hiemalis were overgrown by the mantle hyphae. During the next 3 days, the mantle-hyphae-invading sporangia formed short, thin branches that grew directly towards individual spores, tapering off upon contact.

Agaricales↗

Evaluation of mycelial inocula of edible Lactarius species for the production of Pinus pinaster and P. sylvestris mycorrhizal seedlings under greenhouse conditions.

Different methods to inoculate seedlings of Pinus pinaster and P. sylvestris with edible Lactarius species under standard greenhouse conditions were evaluated. Fungal inoculations were performed both under pure culture synthesis in vitro, followed by transplantation of acclimatized seedlings, and directly in the greenhouse using different techniques for inocula production (mycelial slurries, vegetative inoculum grown in peat-vermiculite and alginate-entrapped mycelium). In vitro inoculations with L. deliciosus produced thoroughly colonized seedlings. However, a sharp decrease in mycorrhizal colonization was detected on transplanted seedlings after 4 month's growth in the greenhouse. On the other hand, all the inocula applied directly in the greenhouse, except the alginate-entrapped mycelium, produced a variable number of mycorrhizal seedlings and colonization rates after the first growing season, depending on the plant-fungal combination and the inoculation method. Inoculations with vegetative inocula of the strain 178 of L. deliciosus were the most effective in producing mycorrhizal seedlings. All the seedlings inoculated with this strain were colonized although the colonization rates were relatively low. The commercial feasibility of the different inoculation methods for the production of seedlings colonized with edible Lactarius species is discussed.

Agaricales↗

Differential responses of three fungal species to environmental factors and their role in the mycorrhization of Pinus radiata D. Don.

Three ectomycorrhizal (ECM) isolates of Rhizopogon luteolus, R. roseolus and Scleroderma citrinum were found to differ markedly in their in vitro tolerance to adverse conditions limiting fungal growth, i.e. water availability, pH and heavy metal pollution. S. citrinum was the most sensitive, R. luteolus intermediate and R. roseolus the most tolerant species. Pinus radiata D. Don seedlings were inoculated in the laboratory and in a containerised seedling nursery with spore suspensions of the three ECM species. Colonisation percentage was considerably lower under nursery conditions, probably due to competition by native fungi. The effects of nursery ECM inoculation on seedling growth depended on the fungal species. Only R. roseolus-colonised plants showed a significantly higher shoot growth than non-mycorrhizal plants. All three fungi induced significantly higher root dry weights relative to control plants. Despite the low mycorrhizal colonisation, mycorrhization with all three species improved the physiological status of nursery-grown seedlings, e.g. enhanced root enzyme activity, shoot nutrient and pigment content, net photosynthesis rate and water use efficiency. Of the three fungal species, R. roseolus was the most effective; this species was also the most adaptable and showed the greatest range of tolerance to adverse environmental conditions in pure culture. It is, therefore, proposed as a promising fungal species for ECM inoculation of P. radiata in the nursery.

Basidiomycota↗

Species richness and seasonal abundance of ectomycorrhizal fungi in plantations of Eucalyptus dunnii and Pinus taeda in southern Brazil.

The abundance and diversity of ectomycorrhizal fungi (EMF) was assessed based on the collection of basidiocarps during 12 months comprising the spring of 1995, and the summer, autumn, and winter of 1996, in three stands of young, middle-aged, and rotation age plantations of Pinus taeda and Eucalyptus dunnii, in the state of Santa Catarina, southern Brazil. A total of 3,085 collections yielded 34 presumed EMF taxa in ten genera, including mushroom-like and sequestrate species. Fruiting patterns of EMF differed with host and season, and host specificity was apparent in some. The overall relative importance (RI) and the Shannon diversity index (H) suggested that stands of E. dunnii had a more diverse aboveground EMF community than those of P. taeda. Overall, species of Scleroderma and Laccaria were not only the most abundant but also had the highest biomass values. The results show that a small number of species of abundant biomass and a larger number of species of less-abundant biomass characterize each forest class.

Biodiversity↗

Ectomycorrhizal fungal biomass in roots and uptake of P from apatite by Pinus sylvestris seedlings growing in forest soil with and without wood ash amendment.

Forest soil from an experimental Norway spruce forest with four levels of wood ash addition (0, 1, 3 and 6 tonnes ha(-1)) was used to inoculate pine (Pinus sylvestris) seedlings with indigenous ectomycorrhizal (EM) fungi. Uptake of 32P and 86Rb in a root bioassay was used to estimate the demand for P and K by seedlings grown in the different soils. Utilisation of P from apatite was tested in a laboratory system where uptake by the ectomycorrhizal mycelium was separated from uptake by roots. The demand for P and K in the seedlings was similar regardless of the ash treatment. Variation in EM levels, estimated as fungal biomass (ergosterol) in roots, was large in the different soils, but not related to ash addition. Uptake of P from apatite was, on average, 23% of total seedling P and was not related to EM levels. It was concluded that the improved P uptake from apatite by EM fungi found in earlier studies is probably not a general phenomenon among EM fungi. The small effect of ash addition on EM levels and P uptake suggests that addition of granulated wood ash is a forest management treatment that will have only minor influence on ectomycorrhizal symbiosis.

Apatites↗

The mycorrhizal fungus Tricholoma matsutake stimulates Pinus densiflora seedling growth in vitro.

While it has been suggested that Matsutake mycorrhizae might not be functional and that Matsutake may behave as a saprobic fungus in soil or even have some pathogenic activity on seedlings, we investigated the consequences of Matsutake inoculation on Pinus densiflora growth. Seventy-five days after inoculation, hyphae were anchored on short roots and well-developed Hartig net palmettis were observed. Compared to both control treatments--seedlings treated with distilled water and seedlings treated with autoclaved mycelium--inoculation significantly stimulated seedling total dry weight by 70.9% and 98.0%, respectively. These findings attest that some type of symbiotic relationship must be functional and favour host growth, ruling out claims of pathogenicity under the sterile conditions used here.

Agaricales↗

Ectomycorrhizal symbiosis in vitro between Tricholoma matsutake and Pinus densiflora seedlings that resembles naturally occurring 'shiro'.

We established an in vitro ectomycorrhizal symbiosis between Tricholoma matsutake and Pinus densiflora. Mycorrhiza formed in a substrate of Modified Norkrans' C medium and granite-based soil had features similar to those observed previously only in naturally occurring mycorrhizal system called 'shiro,' and promoted the growth of plants with smaller root/shoot ratios. The in vitro formation of 'shiro' is essential for the development of T. matsutake system to produce mushrooms and is useful for the propagation and plantation of the mycorrhizal seedlings.

Culture Media↗