白刺花根瘤菌与非根瘤菌在不同培养条件下互作模式的转变

Transformation of the interaction pattern between rhizobia and non-rhizobia of Sophora davidii under different culture conditions

【目的】除根瘤菌外,豆科植物根瘤中还存在着大量的非根瘤菌,它们存在的意义及其潜在的生态学功能还不清楚,尤其是它们与根瘤菌间的互作机制更需揭示。【方法】以从陕北旱区野生白刺花根瘤中分离得到的根瘤菌和非根瘤菌为研究对象,采用共培养、二分隔平板实验和单独培养的方法,于正常培养条件下从白刺花根瘤中筛选出有互作效应的菌株,测定其对p H和Na Cl的耐受性以及对各种氮源的利用情况,并通过根瘤菌的菌落直径、生长曲线和多糖产量来表征其互作效应,进一步探明互作菌株在盐碱和营养胁迫条件下互作效应转变的机制。【结果】在盐碱胁迫下,非根瘤菌Pseudomonas oryzihabitans BT-147和Priestia aryabhattai BT-59对Rhizobium azibense BT-170互作效果由正常培养条件下的抑制转变为促生,与对照组相比试验组R.azibense BT-170菌落直径分别增加了0.803 mm和1.034 mm。Bacillus siamensis BT-9-1在正常培养条件下抑制Mesorhizobium metallidurans YC-39的生长,而在盐碱胁迫下表现为促生作用,与对照组相比试验组M.metallidurans YC-39菌落直径增加了1.019 mm,多糖产量由1.088μg/m L增加到2.555μg/m L。在以谷氨酸作为唯一氮源时,Pseudomonas oryzihabitans BT-147和Priestia aryabhattai BT-59对R.azibense BT-170的互作效果转变为促生作用,R.azibense BT-170试验组与对照组菌落直径差分别达到1.348 mm和2.196 mm,其多糖产量从对照组的0.559μg/m L分别增加到0.821μg/m L和3.341μg/m L。【结论】在盐碱和氮源的胁迫下,非根瘤菌Pseudomonas oryzihabitans BT-147和Priestia aryabhattai BT-59对R.azibense BT-170互作效果由正常培养条件下的抑制转变为促生,并显著提高了R.azibense BT-170多糖的产量(P<0.05),不同培养条件下根瘤菌与非根瘤菌互作模式的转变,提高了根瘤菌的抗逆性,扩大了根瘤菌可利用氮源的范围,揭示了非根瘤菌在根瘤微生态中所发挥的作用,并从体外简化了根瘤微生态中复杂的相互作用。

[Objective] In addition to rhizobia, non-rhizobia have been simultaneously isolated from the same root nodules, and the existence of non-rhizobia in root nodules of Fabaceae plants was a universal phenomenon. Their colonization meaning and potential ecological functions have been unknown so far. Moreover, the real relationships between rhizobia and non-rhizobia remain unclear.Therefore, the interactions and mechanism of their co-evolution should be investigated intensively and further studied. This paper was designed to reveal the interactions between rhizobia and non-rhizobia in root nodule microbiome of Sophora davidii and to further explore the shift in the interaction pattern between them under different culture conditions. [Methods] The rhizobia and non-rhizobia isolated from the root nodules of wild Sophora davidii in arid areas of Northern Shaanxi Province was taken as the subjects. The interacting strains were screened based on co-culture, two-compartment culture and pure culture, and were investigated for the tolerance to pH and NaCl and the utilization of various nitrogen sources. The colony diameter, growth curve and polysaccharide production of the rhizobia were used to evaluate the interaction effects, so as to further explore the transformation mechanism underlying the interaction effects of strains under saline-alkali and nutritional stresses. [Results] Under saline-alkali stress, the interaction effects of the non-rhizobia Pseudomonas oryzihabitans BT-147 and Priestia aryabhattai BT-59 on Rhizobium azibense BT-170 changed from inhibition under normal culture conditions to promotion. The colony diameter of R. azibense BT-170 increased by 0.803 mm and 1.034 mm, respectively in test groups as compared with the condition in control group. When Bacillus siamensis BT-9-1 was co-cultured with Mesorhizobium metallidurans YC-39 under saline-alkali stress, the interaction pattern changed to promotion from inhibition under normal culture conditions. Compared with control group, the test group had increased colony diameter of M.metallidurans YC-39 by 1.019 mm, and the exopolysaccharide production rose from 1.088 μg/mL to 2.555 μg/mL. When Glutamic acid was used as the only nitrogen source, the non-rhizobia Pseudomonas oryzihabitans BT-147 and Priestia aryabhattai BT-59 had promotion effects on R. azibense BT-170,and the difference in colony diameter of R. azibense BT-170 between test group and control group reached 1.348 mm and 2.196 mm, respectively. The exopolysaccharides yield of R. azibense BT-170increased from 0.559 μg/mL in control group to 0.821 μg/mL and 3.341 μg/mL, respectively.[Conclusion] Under saline-alkali and nitrogen source stresses, the interaction effects of the non-rhizobia Pseudomonas oryzihabitans BT-147 and Priestia aryabhattai BT-59 on R. azibense BT-170 changed to promotion from inhibition under normal culture conditions and enhanced the exopolysaccharides production of R. azibense BT-170(P<0.05). The shift in the interaction pattern between rhizobia and non-rhizobia under different culture conditions improved the stress resistance of rhizobia and expanded the range of nitrogen sources available to rhizobia, revealing the role of non-rhizobia in root nodule microbiome and simplifying the complex interactions in root nodule microbiome in vitro.