Philipp Franken

Bradysia difformis Frey (Diptera: Sciaridae) is a fungus gnat that poses a significant threat to greenhouse cultures, and is attracted to soils devoid of peat. Fungal strains from the German Collection of Microorganisms and Cell Culture (DSMZ), such as Beauveria bassiana, Metarhizium flavoviride, Mucor hiemalis, and Niesslia tinuis, as well as Serendipita indica, were screened for entomopathogenic activity against B. difformis and their capacity to colonize Petunia hybrida cv. “Mitchell” and Ocimum basilicum plants. The survival rates of Bradysia difformis (three instar larvae) treated with Metarhizium flavoviride were 45.33% at 14 days following inoculation with 1 × 10⁶ spores/mL of each fungal strain, when compared to others. We concluded that the fungal strain M. flavoviride could serve as an entomopathogenic fungus with the highest virulence against B. difformis larvae. Although M. flavoviride did not show a beneficial effect as an endophyte, interestingly, the strain Niesslia tinuis exhibited plant growth benefits in Petunia hybrida cv. “Mitchell” by enhancing its shoot length up to 13.18 ± 0.72 cm, whereas the control treatment had a shoot length up to 10.68 ± 0.39. Enzymatic assays confirmed the ability of M. flavoviride to produce cuticle-degrading enzymes such as chitinase and protease. Together, these findings highlight the potential of EMPF—particularly M. flavoviride—as a sustainable biocontrol tool well-suited for peat-free horticultural systems, offering an eco-friendly alternative to chemical insecticides where fungus gnat pressure is typically high.

Fungal endophytes can improve plant tolerance to abiotic stress conditions. Dark septate endophytes (DSEs) belong to phylogenetically non-related groups of root colonizing fungi among the Ascomycota with high melanin-producing activities. They can be isolated from roots of more than 600 plant species in diverse ecosystems. Still the knowledge about their interaction with host plants and their contribution to stress alleviation is limited. The current work aimed to test the abilities of three DSEs (Periconia macrospinosa, Cadophora sp., Leptodontidium sp.) to alleviate moderate and high salt stress in tomato plants. By including an albino mutant, the role of melanin for the interaction with plants and salt stress alleviation could also be tested. P. macrospinosa and Cadophora sp. improved shoot and root growth 6 weeks after inoculation under moderate and high salt stress conditions. No matter how much salt stress was applied, macroelement (P, N, and C) contents were unaffected by DSE inoculation. The four tested DSE strains successfully colonized the roots of tomato, but the colonization level was clearly reduced in the albino mutant of Leptodontidium sp. Any difference in the effects on plant growth between the Leptodontidium sp. wild type strain and the albino mutant could, however, not be observed. These results show that particular DSEs are able to increase salt tolerance as they promote plant growth specifically under stress condition. Increased plant biomasses combined with stable nutrient contents resulted in higher P uptake in shoots of inoculated plants at moderate and high salt conditions and higher N uptake in the absence of salt stress in all inoculated plants, in P. macrospinosa-inoculated plants at moderate salt condition and in all inoculated plants except the albino mutants at high salt condition. In summary, melanin in DSEs seems to be important for the colonization process, but does not influence growth, nutrient uptake or salt tolerance of plants.

Endophytic fungi that colonize the plant root live in an environment with relative high concentrations of different sugars. Analyses of genome sequences indicate that such endophytes can secrete carbohydrate-related enzymes to compete for these sugars with the surrounding plant cells. We hypothesized that typical plant sugars can be used as carbon source by root endophytes and that these sugars also serve as signals to induce the expression and secretion of glycolytic enzymes. The plant-growth-promoting endophytes Serendipita indica and Serendipita herbamans were selected to first determine which sugars promote their growth and biomass formation. Secondly, particular sugars were added to liquid cultures of the fungi to induce intracellular and extracellular enzymatic activities which were measured in mycelia and culture supernatants. The results showed that both fungi cannot feed on melibiose and lactose, but instead use glucose, fructose, sucrose, mannose, arabinose, galactose and xylose as carbohydrate sources. These sugars regulated the cytoplasmic activity of glycolytic enzymes and also their secretion. The levels of induction or repression depended on the type of sugars added to the cultures and differed between the two fungi. Since no conventional signal peptide could be detected in most of the genome sequences encoding the glycolytic enzymes, a non-conventional protein secretory pathway is assumed. The results of the study suggest that root endophytic fungi translocate glycolytic activities into the root, and this process is regulated by the availability of particular plant sugars.

Fungus-bacterium interactions are widespread, encompass multiple interaction types from mutualism to parasitism, and have been frequent targets for microbial inoculant development. In this study, using in vitro systems combined with confocal laser scanning microscopy and real-time quantitative PCR, we test whether the nitrogen-fixing bacterium Kosakonia radicincitans can provide protection to the plant-beneficial fungus Serendipita indica, which inhabits the rhizosphere and colonizes plants as an endophyte, from the fungus-feeding bacterium Collimonas fungivorans. We show that K. radicincitans can protect fungal hyphae from bacterial feeding on solid agar medium, with probable mechanisms being quick hyphal colonization and biofilm formation. We furthermore find evidence for different feeding modes of K. radicincitans and C. fungivorans, namely “metabolite” and “hyphal feeding”, respectively. Overall, we demonstrate, to our knowledge, the first evidence for a bacterial, biofilm-based protection of fungal hyphae against attack by a fungus-feeding, bacterial predator on solid agar medium. Besides highlighting the importance of tripartite microbial interactions, we discuss implications of our results for the development and application of microbial consortium-based bioprotectants and biostimulants.

Dark septate endophytes (DSEs) represent a diverse group of root-endophytic fungi that have been isolated from plant roots in many different natural and anthropogenic ecosystems. Melanin is widespread in eukaryotic organisms and possesses various functions such as protecting human skin from UV radiation, affecting the virulence of pathogens, and playing a role in development and physiology of insects. Melanin is a distinctive feature of the cell walls of DSEs and has been thought to protect these fungi from abiotic stress. Melanin in DSEs is assumed to be synthesized via the 1,8-dihydroxynaphthalene (DHN) pathway. Its function in alleviation of salt stress is not yet known. The aims of this study were: (i) investigating the growth responses of three DSEs (Periconia macrospinosa, Cadophora sp., and Leptodontidium sp.) to salt stress, (ii) analyzing melanin production under salt stress and, (iii) testing the role of melanin in salt stress tolerance of DSEs. The study shows that the three DSE species can tolerate high salt concentrations. Melanin content increased in the hyphae of all DSEs at 100 mM salt, but decreased at 500 mM. This was not reflected in the RNA accumulation of the gene encoding scytalone dehydratase which is involved in melanin biosynthesis. The application of tricyclazole, a DHN-melanin biosynthesis inhibitor, did not affect either salt stress tolerance or the accumulation of sodium in the hyphae. In addition, melanin biosynthesis mutants of Leptodontidium sp. did not show decreased growth performance compared to the wild-type, especially not at high salt concentrations. This indicates that DSEs can live under salt stress and withstand these conditions regardless of melanin accumulation.

Dark septate endophytes (DSEs) are often trace element (TE)-tolerant fungi and are abundant in TE-polluted environments. The production of melanin, a black polymer found in cell walls, was hypothesized by several authors to play a role in the TE tolerance of DSEs. To test this hypothesis, we established a series of experiments using albino strains and melanin inhibitors and examined the responses to Cd and Zn. Six DSEs belonging to genera Cadophora sp., Leptodontidium sp. and Phialophora mustea, were evaluated. The strains mainly produced 1,8-dihydroxynaphthalene (DHN) melanin whereas 3,4-dihydroxyphenylalanin melanin was also synthetized. Cd and Zn decreased melanin synthesis in most of the strains. A reduction in melanin concentration in hyphae through the use of tricyclazole, an inhibitor of DHN-melanin synthesis, did not reduce the tolerance of the strains to Cd and Zn. Similarly, albino mutants of Leptodontidium sp. were not more sensitive to Cd and Zn than the WT strain. Moreover, tricyclazole-treated colonies accumulated less Cd but more Zn compared to untreated colonies. The Cd and Zn contents of Leptodontidium albino strains were variable and similar to that of the WT. The results suggest that melanin production is not an important functional trait that contributes to Cd and Zn tolerance, but might contribute to Cd accumulation.

Silencing of SlSUT2 expression in tomato plants leads to a dwarfed phenotype, reduced pollen vitality and reduces pollen germination rate. Male sterility of flowers, together with a dwarfed growth behavior is reminiscent to brassinosteroid defective mutant plants. Therefore we aimed to rescue the SlSUT2 silencing phenotype by local brassinosteroid application. The phenotypical effects of SlSUT2 down-regulation could partially be rescued by epi-brassinolide treatment suggesting that SlSUT2 interconnects sucrose partitioning with brassinosteroid signaling. We previously showed that SlSUT2 silenced plants show increased mycorrhization and, this effect was explained by a putative sucrose retrieval function of SlSUT2 at the periarbuscular membrane. More recently, we reported that the symbiotic interaction between Solanaceous hosts and AM fungi is directly affected by watering the roots with epi-brassinolide. Here we show that the SlSUT2 effects on mycorrhiza are not only based on the putative sucrose retrieval function of SlSUT2 at the periarbuscular membrane. Our analyses argue that brassinosteroids as well as SlSUT2 per se can impact the arbuscular morphology/architecture and thereby affect the efficiency of nutrient exchange between both symbionts and the mycorrhizal growth benefit for the plant.

Dark septate endophytes (DSEs) present a group of widespread root-colonizing fungi. The role of these endophytes in ecosystems and their interactions with plant pathogens are not well understood. In the current study, we assessed the antagonistic potential of the model DSE Cadophora sp. against the tomato soilborne pathogens Rhizoctonia solani, Pythium aphanidermatum and Verticillium dahliae. To investigate their interactions, we conducted in vitro assays followed by a greenhouse experiments in which tomato plants were inoculated with different combinations of the DSE and pathogens. RNA accumulation of selected tomato pathogenesis-related genes and of Cadophora sp. genes with putative antifungal function was analyzed. Cadophora sp. inhibited the growth of the fungal pathogens in vitro and vice versa; a negative impact of the pathogens on the growth of the DSE was also detected. In roots, however, this mutual negative interaction could not be observed. Expression analyses of plant genes could not explain this differential effect, but among the Cadophora sp. genes analyzed, a gene coding for a chalcone synthase was downregulated in planta. The data indicate that plants can change the interaction between fungi and, therefore, in vitro detected antagonism does not necessarily reflect the situation inside the plant.