Effects Of Exogenous Chitosan On Symbiotic Nitrogen Fixation Of Vegetable Soybean Under Low Nitrogen Stress

The increasingly deteriorating environment has forced people to abandon the production concept of high fertilizer investment and shift towards the production goal of low fertilizer and green environmental protection.In this study,ammonium nitrogen and nitrate nitrogen were used as nitrogen sources,and the symbiotic nitrogen fixation system of’Nipponbare’-HH103 was used as test material to study the effect of chitosan on symbiotic nitrogen fixation of vegetable soybean under low nitrogen stress.The aim is to clarify the promoting effect of chitosan on symbiotic nitrogen fixation of vegetable soybeans under low nitrogen stress,and to elucidate the dynamic essence and causal relationship of chitosan improving the nitrogen fixation ability of vegetable soybeans under low nitrogen stress,enrich the theory and methods of chitosan to improve nitrogen fixation capacity of leguminous plants,and explore new ways to reduce fertilizer nitrogen application in production.The main results are as follows:1.Under ammonium nitrogen and nitrate nitrogen,with the nitrogen concentration increases,the aboveground and underground biomass,nodule number,nodule fresh weight,and root activity of vegetable soybeans were first increased and then decreased.The nitrogen content in the aboveground and underground parts of the plant,as well as the ammonia content in the cytoplasm of the root nodule cells,gradually increased;The activity of nitrogenase in nodules,the hemoglobin content in nodules,and the ammonia content in nodules gradually decreased.The results indicated that suitable nitrogen concentration was beneficial to nodulation and biomass accumulation of vegetable soybean,but nitrogen application was not conducive to nodule nitrogen fixation.Through comprehensive evaluation,it was concluded that 50 mg·L^-1ammonium nitrogen and nitrate nitrogen were the optimal nitrogen concentration for symbiotic nitrogen fixation of vegetable soybean,while 25 mg·L^-1ammonium nitrogen and nitrate nitrogen could be used as the low nitrogen stress concentration for subsequent studies.2.Under the low nitrogen stress of ammonium nitrogen and nitrate nitrogen,the nodulation,nitrogen fixation and plant growth of vegetable soybean were significantly inhibited,while different concentrations of exogenous chitosan treatment significantly promoted the aboveground biomass,underground biomass,plant nitrogen content,nodule number,nodule fresh weight and root activity of vegetable soybean under low nitrogen stress.in addition,the suitable induction concentration of chitosan under low nitrogen stress of ammonium nitrogen and nitrate nitrogen was different.The suitable concentration of ammonium nitrogen low nitrogen stress was 250 mg·L^-1,and the suitable concentration of nitrate nitrogen low nitrogen stress was 200 mg·L^-1.3.Low nitrogen stress of ammonium nitrogen and nitrate nitrogen significantly inhibited the symbiotic nitrogen fixation of vegetable soybeans,while exogenous chitosan treatment significantly increased the nitrogen fixation-related enzyme activity and substance content of vegetable soybeans under ammonium nitrogen and nitrate nitrogen low nitrogen stress.The average increase of nitrogen content in the aboveground and underground parts during the stress period was 49.55%,43.68%and 38.02%,39.25%,respectively,and the average increase of nodule bean hemoglobin content was 32.41%and 24.63%,respectively.The average increase of nitrogenase activity in nodules was 11.20%and 16.87%respectively,and the average increase of ammonia content in nodules was 18.82%and 21.88%respectively.The hemoglobin content,nitrogenase activity and ammonium content in nodules treated with chitosan under low nitrogen stress（T3）were significantly higher than those under suitable nitrogen concentration（T1）,indicating that the induction effect of chitosan under low nitrogen stress was significantly higher than that under suitable nitrogen concentration.4.The photosynthesis of vegetable soybean was significantly inhibited by low nitrogen stress of ammonium nitrogen and nitrate nitrogen.After chitosan treatment,chlorophyll content,net photosynthetic rate（P_n）,stomatal conductance（G_s）,transpiration rate（T_r）and ribulose1,5-bisphosphate carboxylase（Rubisco）activity were significantly increased under low nitrogen stress,and intercellular CO₂content（C_i）was significantly decreased.The average increase of P_nwas 14.68%and 14.19%,and the average increase of Rubisco activity was 25.68%and 24.89%.The results showed that exogenous chitosan significantly promoted the photosynthesis of vegetable soybean under low nitrogen stress and improved the efficiency of carbon assimilation.5.Under low nitrogen stress,the nitrogen metabolism of roots and leaves of vegetable soybean was significantly inhibited.After chitosan treatment,the soluble protein content,nitrate reductase（NR）,glutamine synthetase（GS）,glutamate synthase（GOGAT）,glutamate dehydrogenase（GDH）,glutamate oxaloacetate transaminase（GOT）and glutamate pyruvate transaminase（GPT）activities of roots and leaves under low nitrogen stress of ammonium nitrogen and nitrate nitrogen were significantly increased,showing that chitosan significantly promoted the nitrogen metabolism of vegetable soybean under low nitrogen stress.In summary,exogenous chitosan provides sufficient carbon and energy for symbiotic nitrogen fixation by promoting photosynthesis.By increasing the activity of nitrogenase and the content of hemoglobin in root nodules,the assimilation and fixation of N₂are accelerated,promoting nitrogen metabolism in roots and leaves,thereby accelerating nitrogen conversion and utilization.This may be one of the important reasons why chitosan can promote symbiotic nitrogen fixation in vegetable soybeans under low nitrogen stress.