Szymon Zubek

Response of endangered plant species to inoculation with arbuscular mycorrhizal fungi and soil bacteria

Three endangered plant species, Plantago atrata and Pulsatilla slavica, which are on the IUCN red list of plants, and Senecio umbrosus, which is extinct in the wild in Poland, were inoculated with soil microorganisms to evaluate their responsiveness to inoculation and to select the most effective microbial consortium for application in conservation projects. Individuals of these taxa were cultivated with (1) native arbuscular mycorrhizal fungi (AMF) isolated from natural habitats of the investigated species, (2) a mixture of AMF strains available in the laboratory, and (3) a combination of AMF lab strains with rhizobacteria. The plants were found to be dependent on AMF for their growth; the mycorrhizal dependency for P. atrata was 91%, S. umbrosus-95%, and P. slavica-65%. The applied inocula did not significantly differ in the stimulation of the growth of P. atrata and S. umbrosus, while in P. slavica, native AMF proved to be the less efficient. We therefore conclude that AMF application can improve the ex situ propagation of these three threatened taxa and may contribute to the success of S. umbrosus reintroduction. A multilevel analysis of chlorophyll a fluorescence transients by the JIP test permitted an in vivo evaluation of plant vitality in terms of biophysical parameters quantifying photosynthetic energy conservation, which was found to be in good agreement with the results concerning physiological parameters. Therefore, the JIP test can be used to evaluate the influence of AMF on endangered plants, with the additional advantage of being applicable in monitoring in a noninvasive way the acclimatization of reintroduced species in nature.

Hypericin and pseudohypericin concentrations of a valuable medicinal plant Hypericum perforatum L. are enhanced by arbuscular mycorrhizal fungi

Hypericum perforatum L. (St. John’s-wort, Hypericaceae) is a valuable medicinal plant species cultivated for pharmaceutical purposes. Although the chemical composition and pharmacological activities of H. perforatum have been well studied, no data are available concerning the influence of arbuscular mycorrhizal fungi (AMF) on this important herb. A laboratory experiment was therefore conducted in order to test three AMF inocula on H. perforatum with a view to show whether AMF could influence plant vitality (biomass and photosynthetic activity) and the production of the most valuable secondary metabolites, namely anthraquinone derivatives (hypericin and pseudohypericin) as well as the prenylated phloroglucinol—hyperforin. The following treatments were prepared: (1) control—sterile soil without AMF inoculation, (2) Rhizophagus intraradices (syn. Glomus intraradices), (3) Funneliformis mosseae (syn. Glomus mosseae), and (4) an AMF Mix which contained: Funneliformis constrictum (syn. Glomus constrictum), Funneliformis geosporum (syn. Glomus geosporum), F. mosseae, and R. intraradices. The application of R. intraradices inoculum resulted in the highest mycorrhizal colonization, whereas the lowest values of mycorrhizal parameters were detected in the AMF Mix. There were no statistically significant differences in H. perforatum shoot mass in any of the treatments. However, we found AMF species specificity in the stimulation of H. perforatum photosynthetic activity and the production of secondary metabolites. Inoculation with the AMF Mix resulted in higher photosynthetic performance index (PItotal) values in comparison to all the other treatments. The plants inoculated with R. intraradices and the AMF Mix were characterized by a higher concentration of hypericin and pseudohypericin in the shoots. However, no differences in the content of these metabolites were detected after the application of F. mosseae. In the case of hyperforin, no significant differences were found between the control plants and those inoculated with any of the AMF applied. The enhanced content of anthraquinone derivatives and, at the same time, better plant vitality suggest that the improved production of these metabolites was a result of the positive effect of the applied AMF strains on H. perforatum. This could be due to improved mineral nutrition or to AMF-induced changes in the phytohormonal balance. Our results are promising from the biotechnological point of view, i.e. the future inoculation of H. perforatum with AMF in order to improve the quality of medicinal plant raw material obtained from cultivation.

Medicinal plants as hosts of arbuscular mycorrhizal fungi and dark septate endophytes

Arbuscular mycorrhizal and dark septate endophyte associations of 31 medicinal plant species collected from the Garden of Medicinal Plants of the Faculty of Pharmacy, Jagiellonian University, Collegium Medicum in Kraków were investigated. Arbuscular mycorrhiza (AM) was found in 30 species; 23 were of the Arum-type, 5—Paris and 2 taxa revealed intermediate morphology. Many plants were strongly colonized by arbuscular mycorrhizal fungi (AMF). The mycelium of dark septate endophytes (DSE) was observed in 21 taxa. However, the percentage of root colonization by these fungi was low. Spores of 15 species of AMF (Glomeromycota) were found in the rhizosphere of the investigated plants. Our results are the first detailed report of both AMF and DSE associations of these plant species. The use of AMF and DSE during the process of medicinal plant cultivation for pharmaceutical purposes is discussed.

Arbuscular mycorrhizal fungi alter thymol derivative contents of Inula ensifolia L.

Individuals of Inula ensifolia L. (Asteraceae), a valuable xerothermic plant species with potential therapeutic value, were inoculated under laboratory conditions with different strains of arbuscular mycorrhizal fungi (AMF): (1) Glomus intraradices UNIJAG PL-Bot, (2) G. intraradices UNIJAG PL-Kap, (3) Glomus clarum UNIJAG PL13-2, and (4) AMF crude inoculum from natural stands of I. ensifolia. We found AMF species specificity in the stimulation of thymol derivative production in the roots of I. ensifolia. There was an increase in thymol derivative contents in roots after G. clarum inoculation and at the same time the decreased production of these metabolites in the G. intraradices treatments. Moreover, no correlation between the extent of AMF colonization and the effects of the fungal symbionts on the plant was observed. A multilevel analysis of chlorophyll a fluorescence transients (JIP test) permitted an evaluation of plant vitality, expressed in photosynthetic performance index, influenced by the applied AMF strains, which was found to be in good agreement with the results concerning thymol derivative production. The mechanisms by which AMF trigger changes in phytochemical concentration in plant tissues and their consequences for practice are discussed.

ROLE OF MYCORRHIZAL FUNGI IN PHYTOREMEDIATION AND TOXICITY MONITORING OF HEAVY METAL RICH INDUSTRIAL WASTES IN SOUTHERN POLAND

The chapter summarizes research carried out within the last 15 years on the role of mycorrhizal fungi in phytoremediation of zinc wastes located in southern Poland. The impact of various technologies on tailing material toxicity was described and physico-chemical features of the wastes were characterized. Symbiotic fungi are an important component of soil microbiota. As shown in a range of studies, properly developed mutual symbiosis enhances the survival of plants in polluted areas by for instance improving nutrient acquisition and water relations, thus supporting the success of bioremediation. In addition, mycorrhizal fungi were found to play an important role in heavy metal detoxification and the establishment of vegetation in strongly polluted areas. Fungal strains isolated from old zinc wastes also decrease heavy metal uptake by plants growing on metal rich substrata, limiting the risk of increasing the levels of these elements in the food chain. The effectiveness of the bioremediation techniques depends on the appropriate selection of both the plant and the fungal partners. The experiments pointed out that plants conventionally introduced in such places disappear relatively soon, while those appearing during natural succession are better adapted to harsh conditions. Symbiotic partners selected on the basis of such research are often the best choice for future phytoremediation technologies. Moreover, mycorrhizas of different types are also helpful in tailing material toxicity monitoring.

Enhanced concentrations of elements and secondary metabolites in Viola tricolor L. induced by arbuscular mycorrhizal fungi

Background and AimsViola tricolor L. (heartsease, wild pansy) is a valuable medicinal plant obtained for pharmaceutical purposes by cultivation. Given that the species is usually strongly colonized by arbuscular mycorrhizal fungi (AMF), we tested in a pot experiment whether these microorganisms were able to influence V. tricolor mass, vitality, and the concentrations of selected elements, phenolic acids and flavonoids.MethodsThe following treatments were prepared: (1) control: sterile soil without AMF inoculation; (2) Rhizophagus irregularis BEG144; (3) Funneliformis mosseae BEG12; and (4) both isolates. Using a combination of physiological, phytochemical and biophysical methods, we evaluated the effects of these AMF on the performance of heartsease.ResultsThe intensity of mycorrhizal colonization and arbuscule formation was higher when F. mosseae and R. irregularis were introduced separately than when both isolates were present. None of the AMF treatments had an impact either on V. tricolor vitality as expressed by photosynthetic performance index (PI) or on its shoot and root mass. However, in general, a negative correlation was found between the extent of mycorrhizal colonization and shoot mass. We found AMF species specificities in their influence on element, phenolic acid and flavonoid concentrations. Viola tricolor showed no response to F. mosseae. The plants inoculated with R. irregularis had higher concentrations of P, Zn, Mg, and Ca, as well as p-hydroxybenzoic acid and rutin, in comparison to control. Dual AMF species inoculation increased concentrations of Cu, Mg and rutin.ConclusionsThe enhanced production of secondary metabolites in V. tricolor shoots may be due to improved mineral nutrition by AMF and/or a result of general plant defense reaction to fungal colonization. The tendency towards biomass decrease in AMF treatments could be explained by the allocation of plant carbon both to the maintenance of symbionts and enhanced production of secondary compounds.

Arbuscular Mycorrhizal and Dark Septate Endophyte Colonization along Altitudinal Gradients in the Tatra Mountains

Abstract The evaluation of fungal root endophytes of two multizonal mountain plant species (Soldanella carpatica and Homogyne alpine) in relation to altitude was conducted. The comparison of root colonization by coarse arbuscular mycorrhizal fungi (AMF) and the fine AMF endophyte (Glomus tenue), as well as the presence of dark septate endophytes (DSE) were assessed along altitudinal gradients (1000–2050 m a.s.l.) on calcareous and non-calcareous substrata in the Tatra Mts. (Western Carpathians). Additionally, AMF species composition in the rhizosphere of the investigated plants was determined. Coarse AMF dominated over the fine endophyte in roots of S. carpatica and H. alpina. In the case of S. carpatica, there was a tendency for coarse AMF colonization decline with increasing altitude, while the reverse trend was observed for the fine endophyte. In contrast, the altitudinal patterns of the two types were opposite in H. alpina. Fifteen AMF species associated with the rhizosphere of S. carpatica were identified at the sites located in the Western Tatra Mountains, whereas spores of only four species were isolated from the rhizosphere of H. alpina in the High Tatra Mountains. None of the identified AMF species was observed to occur both in the High and Western Tatra Mts. DSE accompanied AMF in the roots of S. carpatica and H. alpina at each site; however, the root colonization by this group of fungi was low. The DSE colonization did not have a consistent relationship with altitude in both plant species. The results suggest that at the investigated altitudes factors such as the type of substrata, host plants, and local plant species composition may play a more important role in determining root colonization as well as the establishment of a local AMF community than the climatic changes with increasing elevation above sea level.

Invasive plants affect arbuscular mycorrhizal fungi abundance and species richness as well as the performance of native plants grown in invaded soils

We studied the effects of invasions by three plant species: Reynoutria japonica, Rudbeckia laciniata, and Solidago gigantea, on arbuscular mycorrhizal fungi (AMF) communities in habitats located within and outside river valleys. Arbuscular mycorrhizal colonization, AMF abundance and species richness in soils were assessed in adjacent plots with invaders and native vegetation. We also quantified the performance (expressed as shoot mass, chlorophyll fluorescence, and the concentration of elements in shoots) of two common, mycorrhizal native plants, Plantago lanceolata and Trifolium repens, grown in these soils. The invasions of R. japonica, R. laciniata, and S. gigantea influenced AMF communities compared to native vegetation, but the changes depended on the mycorrhizal status of invaders. The effects of non-mycorrhizal R. japonica were the most pronounced. Its invasion reduced AMF abundance and species richness. In the plots of both mycorrhizal plants, R. laciniata and S. gigantea, we observed decreased AMF species richness in comparison to native vegetation. The AMF community alterations could be due to (i) depletion of organic C inputs to AMF in the case of R. japonica, (ii) plant secondary metabolites that directly inhibit or selectively stimulate AMF species, or (iii) changes in soil physicochemical properties induced by invasions. The effect of invasion on AMF abundance and species richness did not generally differ between valley and outside-valley habitats. The invasions affected photosynthetic performance and the concentrations of elements in the shoots of P. lanceolata or T. repens. However, the directions and magnitude of their response depended on both species identity and the mycorrhizal status of invaders.

Root-inhabiting fungi in alien plant species in relation to invasion status and soil chemical properties

In order to recognize interactions between alien vascular plants and soil microorganisms and thus better understand the mechanisms of plant invasions, we examined the mycorrhizal status, arbuscular mycorrhizal fungi (AMF) colonization rate, arbuscular mycorrhiza (AM) morphology and presence of fungal root endophytes in 37 non-native species in Central Europe. We also studied the AMF diversity and chemical properties of soils from under these species. The plant and soil materials were collected in southern Poland. We found that 35 of the species formed AM and their mycorrhizal status depended on species identity. Thirty-three taxa had AM of Arum-type alone. Lycopersicon esculentum showed intermediate AM morphology and Eragrostis albensis developed both Arum and Paris. The mycelia of dark septate endophytes (DSE) were observed in 32 of the species, while sporangia of Olpidium spp. were found in the roots of 10. Thirteen common and worldwide occurring AMF species as well as three unidentified spore morphotypes were isolated from trap cultures established with the soils from under the plant species. Claroideoglomus claroideum, Funneliformis mosseae and Septoglomus constrictum were found the most frequently. The presence of root-inhabiting fungi and the intensity of their colonization were not correlated with soil chemical properties, plant invasion status, their local abundance and habitat type. No relationships were also found between the presence of AMF, DSE and Olpidium spp. These suggest that other edaphic conditions, plant and fungal species identity or the abundance of these fungi in soils might have an impact on the occurrence and intensity of fungal root colonization in the plants under study.

Fungal Root Endophyte Colonization of Fern and Lycophyte Species from the Celaque National Park in Honduras

Abstract Root endophyte colonization was investigated in 32 fern and lycophyte species of 11 families collected from the Celaque National Park in Honduras. Arbuscular mycorrhizae (AM) were found in 11 plant species (34%) of Anemiaceae, Gleicheniaceae, Ophioglossaceae, Pteridaceae, Selaginellaceae, Thelypteridaceae, and Woodsiaceae. The abundance of arbuscular mycorrhizal fungi (AMF) in roots varied with particular species, ranging from 4% (Sticherus underwoodianus) to 78% (Thelypteris patens). The morphological AM colonization pattern of all investigated species was of the Paris-type. The mycelium of dark septate endophytes (DSE) was found in 19 species (58%), and was observed both in the roots of plants that were colonized by AMF and were devoid of AM association. However, in both cases the percentage of root colonization by these fungi was low. Exceptions were Asplenium serra, Elaphoglossum erinaceum, Lellingeria prionodes, and Lycopodium thyoides, where abundant DSE hyphae were observed. Our results are the first detailed report of both AMF and DSE associations of these plant species. Moreover, the mycorrhizal status of 27 plant species is reported for the first time.