In vitro mycorrhization of micropropagated plants: studies on Castanea sativa Mill.

In vitro mycorrhization can be made by several axenic and nonaxenic techniques but criticism exists about their artificiality and inability to reproduce under natural conditions. However, artificial mycorrhization under controlled conditions can provide important information about the physiology of symbiosis. Micropropagated Castanea sativa plants were inoculated with the mycorrhizal fungus Pisolithus tinctorius after in vitro rooting. The mycorrhizal process was monitored at regular intervals in order to evaluate the mantle and hartig net formation, and the growth rates of mycorrhizal and nonmycorrhizal plants. Plant roots show fungal hyphae adhesion at the surface after 24 hours of mycorrhizal induction. After 20 days a mantle can be observed and a hartig net is forming although the morphology of the epidermal cells remains unaltered. At 30 days of root–fungus contact the hartig net is well developed and the epidermal cells are already enlarged. After 50 days of mycorrhizal induction, growth was higher for mycorrhizal plants than for nonmycorrhizal ones. The length of the major roots was lower in mycorrhizal plants after 40 days. Fresh and dry weights were higher in mycorrhizal plants after 30 days. The growth rates of chestnut mycorrhizal plants are in agreement with the morphological development of the mycorrhizal structures observed at each mycorrhizal time. The assessment of symbiotic establishment takes into account the formation of a mantle and a hartig net that were already developed at 30 days, when differences between fresh and dry weights of mycorrhizal and nonmycorrhizal plants can be quantified. In vitro conditions, mycorrhization influences plant physiology after 20 days of root–fungus contact, namely in terms of growth rates. Fresh and dry weights, heights, stem diameter and growth rates increased while major root growth rate decreased in mycorrhizal plants.Springe

The influence of the ectomycorrhizal fungus Rhizopogon subareolatus on growth and nutrient element localisation in two varieties of Douglas fir (Pseudotsuga menziesii var. menziesii and var. glauca) in response to manganese stress

Acidification of forest ecosystems leads to increased plant availability of the micronutrient manganese (Mn), which is toxic when taken up in excess. To investigate whether ectomycorrhizas protect against excessive Mn by improving plant growth and nutrition or by retention of excess Mn in the hyphal mantle, seedlings of two populations of Douglas fir (Pseudotsuga menziesii), two varieties, one being menziesii (DFM) and the other being glauca (DFG), were inoculated with the ectomycorrhizal fungus Rhizopogon subareolatus in sand cultures. Five months after inoculation, half of the inoculated and non-inoculated seedlings were exposed to excess Mn in the nutrient solution for further 5 months. At the end of this period, plant productivity, nutrient concentrations, Mn uptake and subcellular compartmentalisation were evaluated. Non-inoculated, non-stressed DFM plants produced about 2.5 times more biomass than similarly treated DFG. Excess Mn in the nutrient solution led to high accumulation of Mn in needles and roots but only to marginal loss in biomass. Colonisation with R. subareolatus slightly suppressed DFM growth but strongly reduced that of DFG (−50%) despite positive effects of mycorrhizas on plant phosphorus nutrition. Growth reductions of inoculated Douglas fir seedlings were unexpected since the degree of mycorrhization was not high, i.e. ca. 30% in DFM and 8% in DFG. Accumulation of high Mn was not prevented in inoculated seedlings. The hyphal mantle of mycorrhizal root tips accumulated divalent cations such as Ca, but not Mn, thus not providing a barrier against excessive Mn uptake into the plants associated with R. subareolatus

Comparative Metaproteomic Analysis on Consecutively Rehmannia glutinosa-Monocultured Rhizosphere Soil

National Natural Science Foundation of China [30772729, 30671220, 31070403]; Natural Science Foundation of Fujian province, China [2008J0051]Background: The consecutive monoculture for most of medicinal plants, such as Rehmannia glutinosa, results in a significant reduction in the yield and quality. There is an urgent need to study for the sustainable development of Chinese herbaceous medicine. Methodology/Principal Findings: Comparative metaproteomics of rhizosphere soil was developed and used to analyze the underlying mechanism of the consecutive monoculture problems of R. glutinosa. The 2D-gel patterns of protein spots for the soil samples showed a strong matrix dependency. Among the spots, 103 spots with high resolution and repeatability were randomly selected and successfully identified by MALDI TOF-TOF MS for a rhizosphere soil metaproteomic profile analysis. These proteins originating from plants and microorganisms play important roles in nutrient cycles and energy flow in rhizospheric soil ecosystem. They function in protein, nucleotide and secondary metabolisms, signal transduction and resistance. Comparative metaproteomics analysis revealed 33 differentially expressed protein spots in rhizosphere soil in response to increasing years of monoculture. Among them, plant proteins related to carbon and nitrogen metabolism and stress response, were mostly up-regulated except a down-regulated protein (glutathione S-transferase) involving detoxification. The phenylalanine ammonia-lyase was believed to participate in the phenylpropanoid metabolism as shown with a considerable increase in total phenolic acid content with increasing years of monoculture. Microbial proteins related to protein metabolism and cell wall biosynthesis, were up-regulated except a down-regulated protein (geranylgeranyl pyrophosphate synthase) functioning in diterpenoid synthesis. The results suggest that the consecutive monoculture of R. glutinosa changes the soil microbial ecology due to the exudates accumulation, as a result, the nutrient cycles are affected, leading to the retardation of plant growth and development. Conclusions/Significance: Our results demonstrated the interactions among plant, soil and microflora in the proteomic level are crucial for the productivity and quality of R. glutinosa in consecutive monoculture system