Paxillus Involutus Strains Research Articles

Ectomycorrhizal Fungal Strains Facilitate Cd2+ Enrichment in a Woody Hyperaccumulator under Co-Existing Stress of Cadmium and Salt.

Cadmium (Cd2+) pollution occurring in salt-affected soils has become an increasing environmental concern in the world. Fast-growing poplars have been widely utilized for phytoremediation of soil contaminating heavy metals (HMs). However, the woody Cd2+-hyperaccumulator, Populus × canescens, is relatively salt-sensitive and therefore cannot be directly used to remediate HMs from salt-affected soils. The aim of the present study was to testify whether colonization of P. × canescens with ectomycorrhizal (EM) fungi, a strategy known to enhance salt tolerance, provides an opportunity for affordable remediation of Cd2+-polluted saline soils. Ectomycorrhization with Paxillus involutus strains facilitated Cd2+ enrichment in P. × canescens upon CdCl2 exposures (50 μM, 30 min to 24 h). The fungus-stimulated Cd2+ in roots was significantly restricted by inhibitors of plasmalemma H+-ATPases and Ca2+-permeable channels (CaPCs), but stimulated by an activator of plasmalemma H+-ATPases. NaCl (100 mM) lowered the transient and steady-state Cd2+ influx in roots and fungal mycelia. Noteworthy, P. involutus colonization partly reverted the salt suppression of Cd2+ uptake in poplar roots. EM fungus colonization upregulated transcription of plasmalemma H+-ATPases (PcHA4, 8, 11) and annexins (PcANN1, 2, 4), which might mediate Cd2+ conductance through CaPCs. EM roots retained relatively highly expressed PcHAs and PcANNs, thus facilitating Cd2+ enrichment under co-occurring stress of cadmium and salinity. We conclude that ectomycorrhization of woody hyperaccumulator species such as poplar could improve phytoremediation of Cd2+ in salt-affected areas.

Amelioration of nitrate uptake under salt stress by ectomycorrhiza with and without a Hartig net.

Summary Salt stress is an important environmental cue impeding poplar nitrogen nutrition. Here, we characterized the impact of salinity on proton‐driven nitrate fluxes in ectomycorrhizal roots and the importance of a Hartig net for nitrate uptake.We employed two Paxillus involutus strains for root colonization: MAJ, which forms typical ectomycorrhizal structures (mantle and Hartig net), and NAU, colonizing roots with a thin, loose hyphal sheath. Fungus‐colonized and noncolonized Populus × canescens were exposed to sodium chloride and used to measure root surface pH, nitrate (NO 3 −) flux and transcription of NO3 − transporters (NRTs; PcNRT1.1, ‐1.2, ‐2.1), and plasmalemma proton ATPases (HAs; PcHA4, ‐8, ‐11). Paxillus colonization enhanced root NO 3 − uptake, decreased surface pH, and stimulated NRTs and HA4 of the host regardless the presence or absence of a Hartig net. Under salt stress, noncolonized roots exhibited strong net NO 3 − efflux, whereas beneficial effects of fungal colonization on surface pH and HAs prevented NO 3 − loss. Inhibition of HAs abolished NO 3 − influx under all conditions.We found that stimulation of HAs was crucial for the beneficial influence of ectomycorrhiza on NO 3 − uptake, whereas the presence of a Hartig net was not required for improved NO 3 − translocation. Mycorrhizas may contribute to host adaptation to salt‐affected environments by keeping up NO 3 − nutrition.

Paxillus involutus-Facilitated Cd2+ Influx through Plasma Membrane Ca2+-Permeable Channels Is Stimulated by H2O2 and H+-ATPase in Ectomycorrhizal Populus × canescens under Cadmium Stress.

Using a Non-invasive Micro-test Technique, flux profiles of Cd2+, Ca2+, and H+ were investigated in axenically grown cultures of two strains of Paxillus involutus (MAJ and NAU), ectomycorrhizae formed by these fungi with the woody Cd2+-hyperaccumulator, Populus × canescens, and non-mycorrhizal (NM) roots. The influx of Cd2+ increased in fungal mycelia, NM and ectomycorrhizal (EM) roots upon a 40-min shock, after short-term (ST, 24 h), or long-term (LT, 7 days) exposure to a hydroponic environment of 50 μM CdCl2. Cd2+ treatments (shock, ST, and LT) decreased Ca2+ influx in NM and EM roots but led to an enhanced influx of Ca2+ in axenically grown EM cultures of the two P. involutus isolates. The susceptibility of Cd2+ flux to typical Ca2+ channel blockers (LaCl3, GdCl3, verapamil, and TEA) in fungal mycelia and poplar roots indicated that the Cd2+ entry occurred mainly through Ca2+-permeable channels in the plasma membrane (PM). Cd2+ treatment resulted in H2O2 production. H2O2 exposure accelerated the entry of Cd2+ and Ca2+ in NM and EM roots. Cd2+ further stimulated H+ pumping activity benefiting NM and EM roots to maintain an acidic environment, which favored the entry of Cd2+ across the PM. A scavenger of reactive oxygen species, DMTU, and an inhibitor of PM H+-ATPase, orthovanadate, decreased Ca2+ and Cd2+ influx in NM and EM roots, suggesting that the entry of Cd2+ through Ca2+-permeable channels is stimulated by H2O2 and H+ pumps. Compared to NM roots, EM roots exhibited higher Cd2+-fluxes under shock, ST, and LT Cd2+ treatments. We conclude that ectomycorrhizal P. × canescens roots retained a pronounced H2O2 production and a high H+-pumping activity, which activated PM Ca2+ channels and thus facilitated a high influx of Cd2+ under Cd2+ stress.

Inoculation with a Pb-tolerant strain of Paxillus involutus improves growth and Pb tolerance of Populus\u2009\xd7\u2009canescens under in vitro conditions

AimsEctomycorrhizal fungi can improve poplar growth and tolerance to heavy metal stress, and may be useful during the afforestation and phytoremediation of polluted regions with poplar trees. In this study, we determined the effects of the symbiotic interaction between Populus × canescens trees and Paxillus involutus strains different in their tolerance to lead.MethodsIn vitro inoculated and non-inoculated plants were treated with 0.75 mM Pb(NO3)2. The root colonization rate of the two fungal strains, as well as their impacts on poplar health and lead accumulation were examined.ResultsBased on the colonization level, the roots were classified into one of three categories: non-mycorrhized, changed (ie, fungal cells were present on the root surface, but the Hartig net did not fully develop), and fully mycorrhized. The lead-tolerant P. involutus strain colonized roots better than the non-tolerant strain (ie, changed and fully mycorrhized roots). Moreover, plants inoculated with the tolerant fungal strain grew better than the control plants (217 % increase in dry weight over the controls), and accumulated lead in the roots and stems.ConclusionsInoculation of P. × canescens trees with a Pb-tolerant strain of P. involutus improves host plant growth and may increase Pb phytostabilization potential.

Performance of ectomycorrhizal alders exposed to specific Canadian oil sands tailing stressors under in vivo bipartite symbiotic conditions

Canadian oil sands tailings are predominately sodic residues contaminated by hydrocarbons such as naphthenic acids. These conditions are harsh for plant development. In this study, we evaluated the effect of inoculating roots of Alnus viridis ssp. crispa and Alnus incana ssp. rugosa with ectomycorrhizal fungi in the presence of tailings compounds. Seedlings were inoculated with 7 different strains of Paxillus involutus and Alpova diplophloeus and were grown under different treatments of NaCl, Na2SO4, and naphthenic acids in a growth chamber. Afterwards, seedling survival, height, dry biomass, leaf necrosis, and root mycorrhization rate were measured. Paxillus involutus Mai was the most successful strain in enhancing alder survival, health, and growth. Seedlings inoculated with this strain displayed a 25% increase in survival rate, 2-fold greater biomass, and 2-fold less leaf necrosis compared with controls. Contrary to our expectations, A. diplophloeus was not as effective as P. involutus in improving seedling fitness, likely because it did not form ectomycorrhizae on roots of either alder species. High intraspecific variation characterized strains of P. involutus in their ability to stimulate alder height and growth and to minimize leaf necrosis. We conclude that in vivo selection under bipartite symbiotic conditions is essential to select effective strains that will be of use for the revegetation and reclamation of derelict lands.

Ion fluxes in Paxillus involutus-inoculated roots of Populus × canescens under saline stress

The Paxillus involutus strains MAJ and NAU improve ion homeostasis under salt stress although only strain MAJ forms a functional ectomycorrhiza with poplar roots, whereas strain NAU induces defense reactions. The goal of the present study was to determine whether beneficial effects of MAJ and NAU on root nutrient element fluxes are induced during early stages of fungal-root interaction or require long-term co-culture. Salt-induced flux profiles of H+, Na+, K+, and Ca2+ were examined in the salt-sensitive poplar Populus×canescens after inoculation for 10d and 20d with the P. involutus strains, MAJ and NAU and after short-term (24h) and long-term (7d) salt stress. Inoculation with P. involutus for 10d and 20d increased the capacity of P.×canescens roots to retain K+ after short and long-term exposure to NaCl stress (100mM). P. involutus-inoculated plants reduced the influx of Na+, especially in the NAU-inoculated roots. The salt-elicited Na+ efflux corresponded with an apparent decline of H+ extrusion in NAU-inoculated roots, which was presumably the result of increased Na+/H+ exchange activity across the plasma membrane. After 10 days of fungal inoculation, P.×canescens roots exhibited an enhanced Ca2+ uptake ability upon salt treatments, whereas a prolonged inoculation time of 20d caused a marked Ca2+ efflux from P.×canescens roots. The P. involutus-elicited Ca2+ enrichment was probably replaced by Na+ at the later stage of fungal colonization. Ca2+ enrichment is known to mediate K+/Na+ homeostasis in poplar roots under salt stress and therefore both NAU- and MAJ-impeded Na+ accumulation compared with non-inoculated roots. NAU provided greater benefit to the inoculated roots to the maintenance of the K+/Na+ homeostasis because of the pronounced Na+ extrusion during the early stage of fungal colonization when the Ca2+ enrichment was greater than in MAJ-inoculated roots. In accordance with flux data, the whole-plant assessment revealed that inoculation with P. involutus attenuated NaCl-induced leaf damage in P.×canescens. Overall, our results support that the formation of a mature ectomycorrhiza is not required for the amelioration of the protection from salinity stress.

Effect of NaCl on growth and ion relations in two salt-tolerant strains of Paxillus involutus

The effect of NaCl on growth, biomass and ion relations of two salt-tolerant isolates of Paxillus involutus, MAJ and NAU were investigated. The two Paxillus strains were exposed to the following concentrations of NaCl: 0, 100, 200 and 500 mmol·L−1. Growth of MAJ and NAU was enhanced by 100 mmol·L−1 NaCl but severely inhibited at the concentration of 500 mmol·L−1. NAU exhibited a greater capacity to exclude Na+ and Cl− under all salinity levels, whereas the salt-includer MAJ had a higher capacity in nutrient uptake under salt stress. The ratios Na+/K+, Na+/Ca2+ and Na+/Mg2+ in NaCl-treated MAJ and NAU did not increase at levels of 100 and 200 mmol·L−1 NaCl but markedly increased at 500 mmol·L−1. This suggests that the two strains, especially MAJ, enhanced nutrient uptake corresponding to the increased Na+ influx at moderate salinity. We conclude that both MAJ and NAU are able to tolerate 500 mmol·L−1 NaCl but there are species-specific differences in retaining ionic homeostasis in the two Paxillus strains. NAU is a salt-excluder, MAJ is a salt-includer but retains a high capacity in nutrient selectivity under saline conditions. Their definite mechanisms to enhance salt tolerance of mycorrhizal hosts need further study.

Effect of heavy metals on the glutathione status in different ectomycorrhizal Paxillus involutus strains

The glutathione (GSH) status and heavy metal tolerance were investigated in four Paxillus involutus strains isolated from different heavy-metal-polluted and non-polluted regions of Europe. The heavy metal burden in the habitats did not affect significantly either the heavy metal (Cr2O 7 2− , Cd2+, Hg2+, Pb2+, Zn2+, Cu2+) tolerance and accumulation or the GSH production of the strains tested. Exposures to heavy metals increased the intracellular GSH concentrations in 12 from 24 experimental arrangements (four strains exposed to six heavy metals) independently of the habitats of the strains. The importance of GSH in heavy metal tolerance (high MIC values, ability to accumulate heavy metals and to grow in the presence of heavy metals) was thus demonstrated in this ectomycorrhizal fungus.

Ectomycorrhizal fungal species and strains differ in their ability to produce free and conjugated polyamines.

Production of free and conjugated polyamines by one strain of Laccaria proxima (Boud.) Maire, three strains (H, O, K) of Paxillus involutus (Batsch) Fr., and one strain of Pisolithus tinctorius was studied in vitro. Spermidine (Spd) was the main polyamine in the 4-week-old mycelium of all the fungi. It was mainly present in the free form, but it also occurred in conjugated forms. Paxillus involutus strain H released large amounts of free putrescine (Put), and the Pisolithus tinctorius released a compound probably related to cadaverine (Cad). On the other hand, these two fungi contained less conjugated polyamines than the other fungi. In addition to the amounts, the forms (perchloric acid soluble and insoluble) of conjugated polyamines in the mycelium varied between species and strains. L. proxima contained nearly as much insoluble conjugated Spd as free Spd, whereas Paxillus involutus strains O and K contained relatively large amounts of soluble conjugated Spd. The results suggest that ectomycorrhizal fungal species and strains differ in their ability and need to produce conjugated polyamines. The small amounts of soluble conjugated polyamines found in the culture filtrates indicate that some specific conjugated polyamines may be involved in polyamine translocation across the plasma membrane.

Application of ectomycorrhizal fungi in rooting of Scots pine fascicular shoots

The ectomycorrhizal fungi, Pisolithus tinctorius (Pers.) Coker & Couch and three Paxillus involutus (Batsch) Fr. strains, were used to enhance in vivo rooting in fascicular shoots of 49 Scots pine (Pinus sylvestris L.) genotypes representing seed families from southern and northern Finland. Inoculation with specific fungi either increased the rooting percentage or accelerated root formation expressed as an increased number of adventitious roots per cutting. Only one of the six seed families gave no positive response. The relationship between the in vitro production of free and conjugated forms of indole-3-acetic acid (IAA) by the fungi and rooting was also investigated. Pisolithus tinctorius was the only significant producer of IAA in the absence of exogenous tryptophan. All the Paxillus involutus strains also synthesized both free and conjugated IAA but did that actively only from exogenous tryptophan. In vitro production of IAA did not correlate with root induction in vivo, but the rooting responses depended on the fungus strains and the Scots pine seed families and (or) genotypes within families. This indicates that in vitro IAA production capacity cannot be the only criterion when selecting ectomycorrhizal fungi for rooting in vivo and that specific genotype-genotype interactions play a key role in root initiation.

Application of ectomycorrhizal fungi in rooting of Scots pine fascicular shoots

The ectomycorrhizal fungi, Pisolithus tinctorius (Pers.) Coker & Couch and three Paxillus involutus (Batsch) Fr. strains, were used to enhance in vivo rooting in fascicular shoots of 49 Scots pine (Pinus sylvestris L.) genotypes representing seed families from southern and northern Finland. Inoculation with specific fungi either increased the rooting percentage or accelerated root formation expressed as an increased number of adventitious roots per cutting. Only one of the six seed families gave no positive response. The relationship between the in vitro production of free and conjugated forms of indole-3-acetic acid (IAA) by the fungi and rooting was also investigated. Pisolithus tinctorius was the only significant producer of IAA in the absence of exogenous tryptophan. All the Paxillus involutus strains also synthesized both free and conjugated IAA but did that actively only from exogenous tryptophan. In vitro production of IAA did not correlate with root induction in vivo, but the rooting responses dep...

Mycorrhizal formation by Paxillus involutus strains in relation to their IAA-synthesizing activity.

IAA production by the ectomycorrhizal fungus Paxillus invotutus grown in pure culture was investigated by testing IAA accumulation or IAA-synthesizing activity. All the Paxillus involutus strains studied released indole compounds either when incubated with 0.5 mM tryptophan or in its absence. HPLC analyses of indole compounds all yielded two main peaks identified by reference to controls as being IAA and indole-3-carboxylic acid and a small peak of indole-3-aldehyde. Mass spectrometry unequivocally confirmed IAA production in the culture filtrates and in the incubation solution used to study in vitro IAA-synthesizing activity. The IAA-synthesizing activity was found to vary widely, from 7.6 to 137.4 nmol IAA h-1 g-1 f. wt, between strains. The mycorrhizal activity of these strains with seedlings of Pinus sylvestris cultured under aseptic conditions was tested. Fungal strains that were characterized in pure culture conditions by high LAA-synthesizing activity induced more fine roots and significantly higher numbers of mycorrhizas than strains which showed a low activity of IAA synthesizing enzymes. The increased ectomycarrhiza formation was generally accompanied by an increased growth of seedlings.