两株解磷细菌的解磷活性及作用机制研究

解磷细菌在增加土壤可溶性磷含量、提高磷肥利用效率方面具有重要作用。为选筛高效解磷菌、探讨其解磷机制,本文利用平板溶磷圈法筛选解磷细菌,采用钼锑抗比色法研究其解磷活性,苯磷酸二钠法研究其磷酸酶活性,利用薄层层析分析其产生的有机酸,根据生理生化特征和16S r RNA基因序列系统发育分析,确定其分类学地位。结果表明,菌株JXJ-11和JXJ-15对植酸钙的降解活性很强,3 d后培养液中可溶性磷浓度分别增加219 mg·L~(-1)和216 mg·L~(-1);对磷酸钙降解活性较弱,最高可溶性磷浓度仅为植酸钙的21.79%~30.37%;解磷细菌可分泌酸性、中性和碱性磷酸酶,降解不溶性磷,可能产生丙酸和琥珀酸等有机酸,降低培养液p H,增加可溶性磷浓度。两株细菌均为革兰氏阴性杆菌,无芽孢,产生硫化氢,其中菌株JXJ-11的16S rRNA基因序列与Sphingomonas melonis DAPP-PG 224T和S.aquatilis JSS7T相似性最高(99.79%),菌株JXJ-15的16S rRNA基因序列与Klebsiella pneumoniae subsp.pneumoniae DSM 30104T相似性最高(99.73%),根据以上信息,确定菌株JXJ-11和JXJ-15分别是鞘氨醇单胞菌属和克雷白氏杆菌属的成员。菌株JXJ-11和JXJ-15的解磷机制包括分泌有机酸和磷酸酶,其中JXJ-11在微生物磷肥研制方面具有潜在应用价值。

Phosphate solubilization activities and action mechanisms of two phosphate-solubilizing bacteria

Phosphate-solubilizing bacteria play an important role in increasing soluble phosphorus content in soils and utilization efficiency of phosphatic fertilizers. Halo zone on agar plate was used to screen solubilizing-phosphate bacteria and the molybdenum antimony resistance colorimetric and disodium phenyl phosphate methods were used to analyze solubilizingphosphate activities and phosphatase activities of the bacteria. The organic acids produced by solubilizing-phosphate bacteria were analyzed using thin layer chromatography (TLC). The taxonomic status of the solubilizing-phosphate bacteria was determined on the basis of physiological and biochemical characteristics and phylogenetic analysis of 16S rRNA gene sequences. The aim of the study was to select bacteria strains solubilizing both inorganic and organic phosphate,and to explore their phosphate solubilization activities and mechanisms. The results showed that two bacteria (designated as JXJ-11 and JXJ-15 strains) exhibited higher activities in solubilizing phytin and the soluble phosphorus contents in the culture broths increased respectively by 219 mg·L^-1 and 216 mg·L^-1 after three days of incubation. The two strains showed lower activities of solubilizing calcium phosphate and the highest soluble phosphorus contents in the culture broths were only 21.79%-30.37% of phytin in media. Both JXJ-11 and JXJ-15 strains secreted acid,neutral and alkaline phosphatases to degrade insoluble phosphorus and probably produced organic acids (e.g.,propionic and succinic acids) which reduced pH values of culture broths while increasing soluble phosphorus contents. Both JXJ-11 and JXJ-15 strains were gram-stain negative,non-endospore forming and positive for the production of hydrogen sulfide. While 16S rRNA gene sequence of JXJ-11 strain was closely related to Sphingomonas melonis DAPP-PG 224^T and S. aquatilis JSS7^T (99.79%),JXJ-15 strain was closely related to Klebsiella pneumoniae subsp. pneumoniae DSM 30104^T (99.73%). Based on the results above,JXJ-11 and JXJ-15 strains were identified as members of the genus Sphingomonas and Klebsiella,respectively. The action mechanisms of phosphate solubilization of JXJ-11 and JXJ-15 strains included secretion of organic acids and phosphatases. The study showed that JXJ-11 strain had a potential application value in developing microbial phosphatic fertilizers.