Research And Application Of SP Characteristic Value Of Fungal Heterotrophic Nitrification Pathway

Nitrous oxide?N₂O?is a profound greenhouse gas.Soil is one of the most important sources of N₂O emissions.There have been lots of unknown uncertainties at the estimate of soil N₂O emissions for numerous ambiguous N₂O production processes.Previous studies have suggested that fungal heterotrophic nitrification may be an important process to produce N₂O in humid subtropical and tropical regions.Due to lacking available methods,fungal heterotrophic nitrification has not been well defined.This project takes subtropical acid soils in China as the object,uses N₂O isotopomer method,and determines the SP?Site Preference?of N₂O from fungal heterotrophic nitrification in pure culture as well as soil culture.Then the contribution of N₂O emissions from two soils with different land use in subtropical acid soil of China were investigated by the way of two-endmember isotope mixing model based on?¹⁵N-SP of N₂O,and the SP of fungal heterotrophic nitrification.In this paper,Aspergillus flavus,Trichoderma harzianum,and Fusarium oxysporum from diverse genera were incubated in pure culture.Among these strains,Aspergillus flavus?ATCC 26214?and Aspergillus flavus belonging to the same genera were selected.It has been revealed that four fungi can obviously produce N₂O via heterotrophic nitrification after applying organic nitrogen?peptone,urea,?-alanine?or inorganic nitrogen??NH₄?₂SO₄?.Besides,the species have an obvious impact on their N₂O production rate.The highest N₂O production rate occurred in the two strains of Aspergillus flavus,especially when urea or peptone was applied.The influence of nitrogen substrates on N₂O production rate was inconsistent with the effect of strains since the given one seems to produce N₂O in specific favorite nitrogen substrate.In addition,substrate type and reaction progress can apparently influence?¹⁵N and?¹⁸O values of N₂O.Even the same strain applying the same substrate in different batches produced N₂O with differed?¹⁵N and?¹⁸O values.Unlike?¹⁵N and?¹⁸O values,a high and relatively constant SP values were observed in all treatments,ranging from23.5?30.1‰.Species,substrate type and reaction process have few impacts on SP.To verify the robust of SP values in fungal heterotrophic nitrification,Aspergillus flavus?ATCC 26214?,Trichoderma harzianum and Fusarium oxysporum were inoculated into?-ray sterilized acid forest soil with peptone as organic nitrogen substrate.The results have illustrated that these fungi were capable of generating N₂O,and the yield tends to be changeable with time.Aspergillus flavus?ATCC 26214?and Trichoderma harzianum produced highest N₂O in the middle period of culture,while Fusarium oxysporum produced highest N₂O in the early period of culture.The isotopic signatures(?¹⁵N,?¹⁸O,and SP)are significantly different at the beginning of culture,and the SP values gradually reached within a similar range?23.79?28.06‰?in the late stage of soil culture,closely to that obtained from fungi pure culture?23.5?30.1‰?.The contribution of N₂O emissions from two subtropical acid soils?upland angriculture soil and forest soil?was investigated by the two-endmember isotope mixing model based on?¹⁵N-SP.It has been demonstrated that autotrophic nitrification and denitrification are dominated N₂O emission sources in upland angriculture soil,while the main N₂O production pathway in forest soil is heterotrophic nitrification.After integrating the SP values of fungal heterotrophic nitrification into the two-endmember isotope mixing model,the relative contribution to N₂O emission from two subtropical acid soils is generally consistent with what we obtained from the ¹⁵N labeling with numerical model.It's the first time to accurately trace the dominate N₂O production pathway of subtropical acid forest soil in Jiangxi province and furtherly verify the reliability of SP value of heterotrophic nitrification after integrating that with other N₂O production processes.In this research,a definite SP value of fungal heterotrophic nitrification was determined by both pure culture and soil culture.Then,this value was integrated into the two-endmember isotope mixing model based on?¹⁵N-SP values and helped to investigate the N₂O emissions from subtrophical acid forest and agiculture soils.Our results indicate that N₂O produced by fugal heterotrophic nitrification showed obvious different SP value with that other N₂O production processes.This is of vital importance to understand soil N₂O emissions,accurately predict the soil N₂O flux and effectively establish mitigation strategies for N₂O emissions.