Selection experiments for the optimum combination of AMF-plant-substrate for the restoration of coal mines

A complex substrate consisting of fly ash, coal gangue and excess sludge was used as an experimental soil in pot culture experiments. Different soil compositions were tested by observing the growth of arbuscular mycorrhizal （AM） fungi inoculated white clover, rye grass or com. The biomass of the host plants, the mycorrhizal colonization （MC） rate and the mycorrhizal depen- dency （MD） were measured. The research addresses the preferable AMF-plant-substrate combination appropriate for restoration of coal mines. We used two inoculation methods： single-inoculation with Glomus versiforme or Glomus mosseae and a dual inocula- tion with both Ctv and Gm. The results show that G^m is the preferable fungi and that dual inoculation does not show advantages for the restoration of coal mines. White clover inoculated with AM fungi is the most suitable condition for restoration of coal mines. The best weight ratio of fly ash, coal gangue and excess sludge was found to be 20：60：20. The optimum treatment conditions of AMF-plant-activated-substrate are described.

Ectomycorrhizal fungi and dark septate endophyte inoculation improve growth and tolerance of Pinus tabulaeformis under cadmium stress

Forest trees can establish symbiotic associations with dark septate endophytes(DSEs)and ectomycorrhizal fungi(ECMF)simultaneously.However,the combined effects of these two fungi on the growth and cadmium(Cd)tolerance of host plants remain largely unexplored.To address this knowledge gap,a pot experiment was conducted to examine the effects of the interaction between an ECMF strain(Suillus granulatus)and a DSE strain(Pseudopyrenochaeta sp.)on Pinus tabulaeformis under Cd stress,by assessing plant growth and physiological parameters,nutrient uptake,and soil properties.Notably,the colonization rates of both fungal strains were found to increase in response to Cd stress,with the extent of this increase being influenced by the specific fungal species and the Cd level in the soil.Compared to the non-inoculation treatment,single inoculation with fungal strain resulted in enhanced biomass,root development,and nutrient contents in P.tabulaeformis seedlings under Cd stress.Furthermore,a synergistic effect was observed when these seedlings were co-inoculated with S.granulatus and Pseudopyrenochaeta sp.,as indicated by significantly greater measurements in various indicators compared to both the single and non-inoculation treatments.Fungal inoculation effectively regulated the antioxidant defense responses and photosynthesis of P.tabulaeformis seedlings subjected to Cd stress,particularly in the co-inoculation treatment.In addition,fungal inoculation facilitated the Cd accumulation in P.tabulaeformis,suggesting a promising potential for the implementation of bioremediation strategies in the areas contaminated with heavy metals.The findings from this study indicate that the utilization of root symbiotic fungi obtained from stress environments could potentially enhance the growth performance and tolerance of P.tabulaeformis towards heavy metals,and co-inoculation of both fungal groups may result in even more pronounced synergistic effects on the overall fitness of the plant.

Occurrence of dark septate endophytes in Phragmites australis in the Baiyang Lake and their resistance to Cd stress

Heavy metal pollution poses a serious hazard to human health,and microbial remediation of heavy metals in soil has been widely studied.A group of ascomycetes classified as dark septate endophytes(DSEs)colonize plant roots and benefit host plants under abiotic stress conditions.In this study,Phragmites australis,a common remediation plant in the Baiyang Lake in North China,was investigated.Soils and roots of P.australis were collected in typical heavy metal-contaminated sites,and the species diversity and community structure of DSEs in P.australis roots were studied.In addition,DSE strains were isolated,cultured,and tested for their tolerance to Cd stress.The results showed that DSEs occurred extensively in P.australis roots,forming typical dark septate hyphae,with a total colonization rate of 19.7%–83.1%.Morphological and internal transcribed spacer sequencing analyses were used to identify 10 species within 9 genera of DSE fungi.Among these fungi,6 strains with considerable resistance to Cd stress were identified.The biomasses of Poaceascoma helicoides,Alternaria doliconidium,and Acrocalymma vagum strains increased as the Cd levels increased.These results can not only help to understand plant-DSE interactions in wetland environments,but also provide a theoretical basis for making full use of DSE fungi to alleviate heavy metal contamination in soil.

Effects of interactions between arbuscular mycorrhizal fungi and bacteria on the growth of Lotus corniculatus L.:From the perspective of regulating rhizosphere fungal community

Arbuscular mycorrhizal fungi(AMF)provide essential nutrients to crops and are affected by fertilizers.Phosphate-solubilizing bacteria(PSB),nitrogen-fixing bacteria(NFB),and AMF have mutually beneficial relationships with plants,but the effects of their interactions on plant growth by regulating rhizosphere fungal community have not been sufficiently studied.In this study,a greenhouse pot experiment was conducted to investigate the interactions between AMF and bacteria(PSB and NFB)on the growth of Lotus corniculatus L.Specifically,the role of rhizosphere fungal community in the growth of Lotus corniculatus L.was explored using Illumina MiSeq high-throughput sequencing.The results showed that combined inoculation of AMF with PSB and NFB increased plant biomass,plant height,and fungal colonization rate.The richness,complexity,and stability of rhizosphere fungal community also increased after combined inoculation of AMF with PSB and/or NFB,particularly with PSB.In addition,combined inoculation of AMF with PSB and NFB enriched the abundance of beneficial microorganisms,with Chaetomium and Humicola showing the greatest alterations.The structural equation model showed that the interactions of AMF with PSB and NFB promoted plant growth by affecting fungal network structure and soil enzyme activities involved in carbon,nitrogen,and phosphorus cycling.These findings provide evidence for the effects of interactions of AMF with PSB and NFB on rhizosphere fungal community and plant growth.

Influence of Arbuscular Mycorrhizal Fungi on Growth, Mineral Nutrition and Chlorogenic Acid Content of Lonicera confusa Seedlings Under Field Conditions

Lonicera confusa, a traditional Chinese medicine herb for treating cold, flu, acute fever, and so forth, is often grown artificially in acidic soils and suffers from phosphorus (P) deficiency. A five-year field experiment was carried out to study the colonization rate, growth, nutrition, and chlorogenic acid content of Lonicera confusa seedlings inoculated with arbuscular mycorrhizal (AM) fungi, Glomus etunicatum and Glomus intraradices. Before transplanting into a field, both AM-inoculated and uninoculated control plants were cultured in nursery beds. In the plants inoculated with the AM fungi, the colonization rate decreased linearly with time and a greater decrease was observed in the plants inoculated with G. intraradices than with G. etunicatum, while the AM colonization increased from 0% to 12.1% in the uninoculated control plants 5 years after transplanting. Plant height, crown diameter, number of new branches, and flower yield increased significantly by AM inoculation as compared to the uninoculated control. Phosphorus concentrations in leaves and flowers increased, and plant uptake of nutrients, e.g., nitrogen (N), P, and potassium (K), was also enhanced significantly by AM inoculation. The Lonicera confusa seedlings had a better response to inoculation of G. intraradices than G. etunicatum in both growth and chlorogenic acid content in flowers. In contrast, both plant P uptake and P concentrations in leaves and flowers were similar between two fungal inoculations. The positive responses of Lonicera confusa to AM inoculation in growth, nutrient uptake, flowering, and chlorogenic acid content in flowers suggested that AM inoculation in nursery beds could promote the plant growth and increase chlorogenic acid content in flowers of Lonicera confusa when grown on acidic and P-deficient soils.

Seasonal and Tissue Age Influences on Endophytic Fungi of Pinus tabulaeformis (Pinaceae) in the Dongling Mountains, Beijing

Endophytic fungi associated with Pinus tabulaeformis Carr. were investigated in the Dongling Mountains of Beijing. A total of 16200 tissue segments of P. tabulaeformis from four sample collections were processed, and 10659 fungal isolates were recovered. The overall colonization and isolation rates of endophytic fungi from high to low were spring〉winter〉autumn〉summer and in different tissues were bark〉needle〉xylem, irrespective of sampling seasons. The colonization rates of endophytic fungi of needles increased with age in the four sample collections. There were no significant differences of the colonization rates of endophytic fungi among 1-, 2-, and 3-year-old bark and xylem, except for significantly lower colonization rates in 1-year-old bark than in 2- and 3-year-old bark in summer. A similar trend of the isolation rates of endophytic fungi occurred. A total of 24 fungal taxa were recorded, of these five taxa Alternaria alternata, Leptostroma sp., Pestalotiopsis besseyi, Phoma lingam, and Phomopsis archeri, were consistently isolated as the common fungi in each sample collection. Our results suggest that some fungi show a certain degree of tissue recurrence or specificity, and the composition of endophytic assemblages is not conspicuously influenced by the seasonal factor.