Genetic Diversity Analysis And ACC Deaminase Activity Research Of The Rhizobia Associated With Acacia Melanoxylon

Acacia melanoxylon belongs to the Acacia genus of the plant subfamily Mimosaceae. It is one of the high-valued timber tree introduced to southern China at the end of last century. However, inoculating rhizobia stains, in particular, needs further invesitgation, as it is effective way improving the survival and growth of black acacia. Hence, genetic diversity of the rhizobia associated with A. melanoxylon grown in South China is urgently needed.In our study, 174 bacterial isolates were obtained from A. melanoxylon root nodules colleced from 15 sampling sites in Guangdong, Fujian and Jiangxi provinces of China.Isolated rhizobia were characterized by genotype and phenotype analysis. The results indicated that the microsymbionts associated with A. melanoxylon showed abundant genetic diversity. All of the isolates were identified as alpha-Proteobacteria belonging to Bradyrhizobium, Rhizobium, Mesorhizobium, and as beta-Proteobacteria belonging to Burkholderia and Herbaspirillum. 13 microsymbionts were able to nodulate with A. melanoxylon, including Bradyrhizobium cytisi, Br. betae, Br. huanghuaihaiense, Mesorhizobium plurifarium, Rhizobium multihospitium, Burkholderia stabilis, Herbaspirillum seropedicae and 3 novel species in Bradyrhizobium, 4 novel species in Mesorhizobium. We also find it especially interesitng that the rhizobia species were in evidence with geographical regions.According to the Polyphasic Taxonomy, 7 novel species were identified from the strains associated with A. melanoxylon. 3 novel species in Bradyrhizobium and 4 novel species in Mesorhizobium were found. Therefore, it is suggested that these groups of strains represents several novel species for which the name Bradyrhizobium ganzhouense sp. nov.(strains RITF211, RITF806, RITF807), Bradyrhizobium sp. nov. II(strain RITF1558), Bradyrhizobium sp. nov. III(strain RITF1622), Mesorhizobium huizhouense sp.nov.(strain RITF711, RITF712), Mesorhizobium sp. nov. II(strain RITF907), Mesorhizobium sp. nov. III(strain RITF1212) and Mesorhizobium sp. nov. IV(strain RITF1564) are proposed.The inoculation and cross-nodulation tests showed that A. melanoxylon could easily symbiosed with various rhizobia species, and rhizobia isolated from A. melanoxylon may nodulate a variety of Acacia trees, such as Acacia aneura, Acacia victoriae and Acacia implexa. The results suggested that A. melanoxylon was a promiscuous host for rhizobia.In order to promote plant growth, rhizobia may cleave the plant ACC through the ACC deaminase, and thereby lowering the level of ethylene in the plant. A colorimetric ninhydrin assay was used to quantify ACC deaminase activity for all isolates and the phylogeny of acdS genes were assessed. As a result, 151 strains among 174 isolates showed ACC deaminase activity, but there was rather different in enzyme activity. However, the acdS gene of 121 strains was detected which were conserved in Mesorhizobium genus. The phylogeny of acdS genes was significantly correlated with geographical origin. A further research on correlation between the level of ACC deaminase activity and the prevalence of acdS genes is urgently needed.