| Soil Salinization had became a major problem in utilization of soils, improving plant salt tolerance is simple, economical and ecological cycle method to utilization of salt-affected soil. Some researches showed that inoculation with rhizobia of leguminous grass could improve its salt tolerance, but the nodule was more sensitive to salt stress than plant, therefore, the salt tolerance in rhizobia has become the key point of the salt tolerance research of whole symbiosis system. This study firstly confirmed salt tolerance of rhizobia strain RJS9-2of Stylosanthes guianensias, then investigated the characteristics and salt tolerance mechanism of this strain. finally this research studied the effect of inoculation with rhizobia of Stylosanthes guianensias cv. Reyan V on the nutrition characterstic and physiological indexes, combining with the tolerance mechanism of rhizobia strain RJS9-2to investigate the mechanism of higher salt tolerance of rhizobia-inoculated Stylo. The main results were as follow:1) Stylo rhizobia strain RJS9-2had superior salt tolerance, could survived in the YAM medium with0.4mol/L NaCl, and the logarithmic phase was decreased along with increasing NaCl concentration. Moreover, other rhizobia strains could not survived in YAM medium with NaCl less more than0.1mol/L.2) Stylo rhizobia strain RJS9-2belongs to Bradyrhizobium yuanmingense, departed from strains PN13-3、LZ3-2、BS1-1. Compared with other strain, RJS29owned its benefit characteristic to resist salt stress:more stereoscopic thallus after gram stain, and deep stained poly (β-hydroxybutyric acid) particles among cells; it could produced acid or alkaline in pure culture under different time and salt concentration; had a superior abilities to secrete IAA and solubilize Ca-P, furthermore, the amount of IAA increased along with enhanced salt concentration in some content.3)Stylo rhizobia strain RJS9-2could improve the tolerance to salt stress through accumulating compatible solutes proline, ectoin, lycine and mycose. At the same time, RJS9-2could utilize the iron and molybdenum from Stylo to sustain its growth and N2-fixation under salt stress conditions, improving the salt tolerance of rhizobia. After analyzing2D images with software, we found14protein spots showed significant expression levels change,4proteins were up-regulated,10proteins were down-regulated, among them, we could speculate the possible mechanism for salt tolerance of RJS9-2according to the analysis of protein expression:RJS9-2could produce bi-component reaction to response to the outside stress under salt stress conditions; cell increased the expression of ABC transporter protein to transport amount of compatible solutes to improving the intercellular osmotic pressure; meanwhile elevated expression of CO dehydrogenase and Superoxide dismutase were contributed to transform the toxic CO to CO2and scavenged the ROS from salt stress, thus improved the salt tolerance of rhizobia.4) It is found that the salt damage degree, plant fresh weight, stem and leaf fresh weight and mortality of plant with salt treatment after inoculation with rhizobia for two weeks, both higher than the non-inoculation plant. But there were no difference of each indexes for inoculation and salt stress apply at same time, so inoculation with rhizobia before salt treatment was more benefit for improving the salt tolerance of Stylo. Under0.10mol/L salt stress treatment, the available nodule number and the ratios of it to total nodule number of Stylosanthes guianensias cv. Reyan||and Reyan V inoculated with RJS9-2were significantly higher than other strains. And Under0.20mol/L salt stress treatment, the fresh weight, N content and P content of plant with RJS9-2were higher than other treatment. Finally The salt tolerance coefficient and the main factor analysis confirmed that inoculation with rhizobia strains RJS9-2markerly improved the salt tolerance of Stylosanthes guianensias cv Reyan V) The mechanism underlying the salt tolerance of Stylosanthes guianensias cv Reyan V inoculated with RJS9-2under salt stress were as follow:①Inoculation with RJS9-2increased the content of chlorophyll in stylo leaves, guarantee the photosynthesis and relieve the salt-induced nutrition stress by N2-fixation and phosphate absorption.②Inoculation with RJS9-2deduced the uptake of Na+、Cl-, and increased the absorptionof K+, promoting the balance of ion and nutrient uptake in plant, and significantly increased the ratios of P/Cl-、K+/Na+、P/Na+、Ca2+/Na+.③Inoculation with RJS9-2adjusted pH of culture medium, sustaining the pH value at the similar level of non-salt treatment conditions.④Inoculation with RJS9-2significantly increased the root activity of stylo, and decreased the browning rate of root suffered from salt stress; under salt stress, the available nodule number was significantly positive with salt tolerance coefficient, a large amount of available nodule under salt stress was the key reason for the improved salt tolerance of rhizobia-inoculated stylo.⑤Inoculated with strain RJS9-2, Stylosanthes guianensias cv Reyan V under salt stress can significantly increase the accumulation of proline and betaine in the plant, ensuring the penetration balance of cells. In pure culture conditions, the proline and betaine content in the plant and strain are respectively positive correlated, indicating that salt-tolerant rhizobium inoculation and the accumulation of compatible solutes may have a relation of reciprocal induction.⑥Inoculation with RJS9-2improved the SOD and POD activities, and the increase of POD was higher than SOD, suggested inoculated stylo mainly eliminate ROS through POD; in addition, the MDA of plant significantly reduced to protect lipid for salt tolerance improvement.⑦Inoculation with RJS9-2secreted a massive of IAA, the salt damage degree and fresh weight of stylo treated with same amount IAA as secreted were markedly higher than which out of IAA treatment, indicated the secreted IAA of inoculated stylo could improve the salt tolerance of stylo.In conclusion, Stylo rhizobia strain RJS9-2belongs to Bradyrhizobium yuanmingense, have a superior salt tolerance. This strain contained a large amount of poly-hydroxybutyric acid (PHB). it secreted acid or alkaline, IAA, and presented the phosphate and organic phosphorus solubilizing abilities; the strain RJS9-2also could improve the tolerance to salt stress through accumulating compatible solutes proline, ectoin, lycine and mycose; at protein level, RJS9-2produced bi-component reaction, with increased the expression of ABC transporter protein, CO dehydrogenase and Superoxide dismutase to improved the salt tolerance of rhizobia. Inoculation with rhizobia before salt treatment was more benefit for improving the salt tolerance of stylo, the mechanism underlying salt tolerance were increased available nodule number; deduced the uptake of Na\Cl-, and increased the absorption of K+; increased the content of chlorophyll and N,P content, sustaining the balance of ion and nutrient uptake in plant; sustaining the pH value of root environment; secreted IAA; improved the SOD and POD activities with reduced MDA content. |
