台湾相思根瘤菌的系统发育、氮代谢基因及表型特征  

Phylogeny,Nitrogen Metabolism Genes,and Phenotypic Characteristics of Rhizobia Associated with Acacia confusa

作　　者：李彤欣 钟冰俞 兰光飞 马红亮[1,2] 尹云锋[1,2] 高人[1,2] LI Tongxin;ZHONG Bingyu;LAN Guangfei;MA Hongliang;YIN Yunfeng;GAO Ren(School of Geographic Sciences,School of Carbon Neutrality Future Technology,Fujian Normal University,Fuzhou 350117,China;State Key Laboratory for Subtropical Mountain Ecology of the Ministry of Science and Technology and Fujian Province,Fujian Normal University,Fuzhou 350117,China)

机构地区：[1]福建师范大学地理科学学院、碳中和未来技术学院,福州350117 [2]福建师范大学湿润亚热带山地生态国家重点实验室培育基地,福州350117

出　　处：《亚热带资源与环境学报》2025年第1期145-155,共11页Journal of Subtropical Resources and Environment

基　　金：国家自然科学基金项目(32171758,31570607)。

摘　　要：台湾相思是中国重要的造林树种之一,其共生根瘤菌具有多样性,氮代谢途径也具有多样性。对从一年生台湾相思(Acacia confusa)苗木根瘤中分离出的5株内生细菌(ZPR1-ZPR5)进行系统发育分析,并对其中2株(ZPR3和ZPR4)进行种鉴定及氮代谢功能基因和表型特征描述,为该树种共生根瘤菌的基础研究和应用提供科学参考。利用根瘤切片法分离内生菌,采用16S rRNA测序分析确定属水平的分类地位;在全基因组测序基础上,利用平均核苷酸一致性(Average Nucleotide Identity,ANI)分析进一步确定菌株ZPR3和ZPR4种水平的分类地位;利用京都基因和基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)注释了解其氮代谢相关基因分布特征;利用常规方法对其表型特征进行描述。结果表明,各分离菌株与根瘤菌属(Rhizobium)的菌种R.tropici、R.rhizogenes、R.freirei、R.lusitanum之间的16S rRNA序列相似性都大于99.5%,因此可以确定这5株内生菌都属于根瘤菌属。菌株ZPR3和ZPR4的ANI值为100%,与34个相似根瘤菌(R.spp.)的ANI值(83.65%~1.50%)都没有达到95%的种鉴定标准,为同一根瘤菌潜在新种。菌株ZPR3和ZPR4没有固氮基因簇(nif),但具有结瘤基因(nodDET)和固氮调节基因(fixABLJ);也没有反硝化功能基因,但具有硝酸盐、亚硝酸盐同化基因(nasCDEST)和铵同化、氨基酸代谢基因(GDH2、glnA、gltBD)。该菌为快生菌,好氧生长,无固氮酶乙炔还原活性,以酵母粉/苹果酸为单一氮/碳源时生长最好,耐酸碱范围广。该菌氮代谢功能基因特殊,揭示其未来在无机氮同化还原研究中具备潜在价值,同时,应继续开展回接实验验证菌株的实际结瘤和固氮能力,以更明确地了解菌株的独特性和应用价值。Acacia confusa is one of the important afforestation tree species in China,harbors diverse symbiotic rhizobia with varied nitrogen metabolism pathways.Phylogenetic analysis was conducted on five endophytic bacteria(ZPR1-ZPR5)isolated from root nodules of one-year-old Acacia confusa seedlings.Species identification and description of nitrogen metabolism functional genes and phenotypic characteristics were performed for two of strains(ZPR3 and ZPR4),providing scientific references for basic research and application of symbiotic rhizobia of this tree species.Endophytes were isolated by root nodule sectioning method,and the taxonomic status at the genus level was determined by 16S rRNA sequencing analysis.Based on whole-genome sequencing,average nucleotide identity(ANI)analysis was used to further determine the taxonomic status at the species level of strains ZPR3 and ZPR4.The distribution characteristics of nitrogen metabolism-related genes were understood by kyoto encyclopedia of genes and genomes(KEGG)annotation.Phenotypic characteristics were described by conventional methods.The results showed that the 16S rRNA sequence similarity between each isolated strain and Rhizobium tropici,R.rhizogenes,R.freirei,and R.lusitanum was greater than 99.5%.Therefore,it can be determined that these five endophytes all belong to the genus Rhizobium.The ANI value of strains ZPR3 and ZPR4 is 100%.Their ANI values with 34 similar rhizobia(R.spp.)(83.65%~91.50%)do not reach the species identification standard of 95%,indicating that they are potential new species of the same Rhizobium.Strains ZPR3 and ZPR4 do not have a nitrogen fixation gene cluster(nif),but have nodulation genes(nodDET)and nitrogen fixation regulatory genes(fixABLJ).They also lacked denitrification functional genes but have nitrate and nitrite assimilation genes(nasCDEST)and ammonium assimilation and amino acid metabolism genes(GDH2,glnA,gltBD).Phenotypic analysis revealed that these strains are fast-growing,aerobic bacteria with no detectable nitrogenase activity,

关 键 词：台湾相思(Acacia confusa) 根瘤菌 16S rRNA 全基因组测序 氮代谢 

分 类 号：S718[农业科学—林学]