Supported by the National Natural Science Foundation of China (No. U1703232)
为了认识和理解丛枝菌根真菌（AM真菌）根外菌丝表面定殖的细菌是否同时具有固氮和解磷能力，从田间生长的玉米菌根根外菌丝表面分离鉴定了固氮菌，评价了其固氮和解磷能力。从AM真菌根外菌丝表面分离出23株可在无氮培养基中生长的细菌，分属于变形菌门（Proteobacteria）、厚壁菌门（Firmicutes）和放线菌门（Actinobacteria）下的9个不同的属（Brevundimonas、Microbacterium、Sphingomonas、Paenibacillus、Variovorax、Xenophilus、Hydrocarboniphaga、Arthrobacter和Bacillus）；11株细菌具有固氮酶活性，固氮酶活性最高值为（2.97 ±1.32）nmol·mg-1·h-1；12株细菌具有活化有机磷的能力，11株细菌具有溶解难溶性无机磷的能力，其中8株细菌同时具有解有机磷和难溶性无机磷能力，并且解有机磷能力高于解难溶性无机磷能力；16株细菌具有分泌IAA（吲哚乙酸）能力，IAA分泌量最高值为（688.00 ±19.17）μg·mL-1；6株细菌同时具备固氮酶活性、解有机磷、解难溶性无机磷和分泌IAA能力。上述结果表明，AM真菌根外菌丝表面定殖有多种具备固氮、解磷和促生能力的细菌，这些具有多重功能的细菌可能极大地扩展菌根途径吸收土壤养分的能力。
[Objective] The objectives of this study were to investigate whether the bacteria colonized on the surface of extraradical hyphae of arbuscular mycorrhizal (AM) fungi do have the dual functions of N2 fixation and phosphates solubilization. AM fungi release through extraradical hyphae exudates that are rich in carbohydrates and hence provide a habitat for bacteria to colonize in. The interaction between AM fungi and bacteria in the hyphosphere has been found to promote mineralization of organic phosphorus and organic nitrogen, enhance nutrient acquisition of AM fungi, and further improve nutritional status of the host plant. However, it remains unclear whether the bacteria colonized on the surface of extraradical hyphae of AM fungi do have the dual functions of N2 fixation and phosphate solubilization. [Method] AM fungal hyphae were collected with PVC tubes and nylon mesh in a maize field, and then N2-fixing bacteria colonized on the surface of extraradical hyphae of AM fungi were isolated with a nitrogen-free selective medium, purified for DNA extraction and identified through 16S rRNA gene sequencing. NifH gene of the bacterial DNA was amplified with specific primers. Nitrogenase activity of the N2-fixing bacteria was determined via acetylene reduction assay, phosphorus solubilizing ability was by measuring the diameter of the bacteria colony and phosphorus solubilizing halo in NBRIP solid medium, and IAA (Indoleacetic acid) secretion ability was with the Salkowski colorimetric method. [Result] Twenty-three strains of N2-fixing bacteria, capable of surviving in N-free media, were isolated from the surface of AM fungal hyphae, and sorted into nine different genera (Paenibacillus and Bacillus in Firmicutes; Microbacterium and Arthrobacter in Actinobacteria; Sphingomonas, Brevendimonas, Variovorax, Xenophilus and Hydrocarboniphaga in Proteobacteria), among which, bacteria in Brevendimonas (six strains, accounting for 26% of the total) and Microbacterium (six strains, for 26%)were the highest in relative abundance; eleven strains showed nitrogenase activity, peaking up to (2.97 ±1.32) nmol·mg-1·h-1; twelve strains did the ability to solubilize organic phosphorus (phytin); eleven strains did the ability to solubilize sparingly soluble inorganic phosphates (tricalcium phosphate); eight strains did the ability to solubilize both phytin and tricalcium phosphate, with the former being higher than the latter; sixteen strains did the ability to secrete IAA, peaking up to (688.00 ±19.17) μg·mL-1; and six strains did all the functions, nitrogenase activity, phytin solubilization, sparingly soluble inorganic phosphates solubilization, and IAA secretion, and they belong to five different genera (Brevundimonas, Microbacterium, Sphingomonas, Paenibacillus, and Arthrobacter) and among them, Paenibacillus was the highest in ability of N2 fixation, sparingly soluble phosphates solubilization, and IAA secretion. [Conclusion] A variety of strains of bacteria with nitrogen-fixing, sparingly soluble phosphates-solubilizing and growth-promoting ability are found colonized on the surface of extraradical hyphae of AM fungi in the maize field. These bacteria with multiple functions may have the ability to greatly expand the pathways for mycorrhizae to absorb nutrients in the soil.
石晶晶,张林,江飞焰,王晓,冯固. AM真菌菌丝际细菌具有固氮解磷双重功能[J].土壤学报,2021,58(5):1289-1298. DOI:10.11766/trxb20200316004 SHI Jingjing, ZHANG Lin, JIANG Feiyan, WANG Xiao, FENG Gu. Dual Functions of Bacteria Colonized on AM Fungal Hyphae-Fixing N2 and Solubilizing Phosphate[J]. Acta Pedologica Sinica,2021,58(5):1289-1298.复制