Research On Characteristics And Application Of Urea By Black Soldier Fly Based On ¹⁵N Isotope Tracer Technique

Nitrogen is an essential nutrient for any animal or plant to complete its life activities,and it is also essential for the development of the black soldier fly larvae（BSFL）.Nitrogen deficiency will seriously affect the growth and development of BSFL and its ability to treat waste.Due to high nitrogen content,low cost and simple application of urea,it has been widely used worldwide.But there are few studies on whether it can be used as a nitrogen supplement for BSFL conversion waste.In order to explore the utilization ability of BSFL to urea,the ¹⁵N isotope tracing technology was used for the first time to verify the utilization ability of BSFL on urea,quantified the distribution of urea nitrogen in larvae,substrates and losses,explored the physicochemical characteristics and microbial changes of substrates during the conversion of BSFL to urea,and finally studied the effect of urea addition on cow manure with high carbon to nitrogen ratio of BSFL.The main results are as follows:1.The ability of the black soldier fly to use ureaDifferent amounts of uera were added to rice husk and glucose base matrix,the results showed that the maximum fresh weight of single larvae in the urea groups were higher than that in contril group,which was 2.2 to 2.4 times higher than that in the control group.At the optimal amount of urea added for larval growth,the isotope urea was instead fed with BSFL,and the total protein harvested was 4.9 times higher the control group and the content of various amino acids in BSFL was significantly increased.The ¹⁵N content of the larvae and all parts（gut,fat,Integument）increased significantly,which directly proved that the BSFL can use urea,and the utilization efficiency could reach 0.59±0.05.The distribution of ¹⁵N-urea showed that 59.4%±4.9%transfer to the larval body,19.9%±1.3%residual in the substrate and 20.7%±5.5%losses.2.Dynamic analysis of matrix physicochemical characteristics during urea conversion by black soldier flyCompared with the BSFL（CK-L）group in the rice shell system,adding urea（U+L）could promote the utilization of carbon in the substrate,and significantly change the physicochemical properties of the substrate,especially pH and EC.The added urea was rapidly decomposed into NH₄⁺under the action of microorganisms,and the decomposition rate of urea in the U+L group was slower than that in the CK-U（with the BSFL and urea in the rice shell system）group.In the U+L group,15.3%of urea nitrogen was lost in the form of NH₃.Correlation analysis showed that NH₃emissions are mainly affected by substrate pH,while BSFL would rapidly raise the pH in the matrix above neutral,resulting in earlier emission of NH₃ in the U+L group than CK-U,and more cumulative emissions,but the cumulative emissions of CH₄,CO₂ and N₂O decreased by 47.4%±0.2%,33.6%±0.1%and 30.5%±0.1%,respectively.3.Dynamic analysis of microbial community structure and functional characteristics during urea conversion by black soldier flyThis chapter compared the alpha diversity of microbial communities and the relative abundance changes of microbial community structure composition at the phylum and genera levels during the transformation of CK-L,CK-U and U+L.During the whole transformation process,BSFL reduced the diversity and richness of the substrate microorganisms.In the microbial community composition,BSFL significantly changed the matrix microbial community structure.According to the analysis of several nitrogen metabolism related enzymes obtained by PICRUSt1 prediction,the possible metabolic route of urea is rapidly decomposed into NH₄⁺under the action of urease,and then formed glutamine with glutamate catalyzed by glutamine synthetase,and utilized by BSFL;Meanwhile,NH₄⁺is also converted to NH₃,and correlation analysis shows that NH₃emission is closely correlated with Acinetobacter and Klebsiella,their R values were 0.91and 0.86,respectively.4.Effect of urea addition on the transformation of cow manure by black soldier flyCompared with the natural placement of cow manure（CK1）,added BSFL（CK2）accelerated the degradation and conversion of cow manure,and the degradation efficiency increased by 11.4%±1.7%（WW）and 108.7%±8.0%（DW）.The addition of urea had no significant difference in the survival rate of BSFL compared with the CK2 group,and all could reach more than 95.6%.The addition of 3 g（U3）and 5 g（U5）urea groups both promoted body growth,and the total fresh weight of the harvested larvae in the U3 group was 12.7%±2.8%higher than that in the CK2 group.The addition of urea promoted the utilization of carbon in cow manure by BSFL,and the carbon reduction rate of in U3 and U5 groups increased by 63.9%±17.3%and 27.8%±5.8%compared with CK2 group,respectively.The addition of urea was also promoted the degradation of cow manure by BSFL,and the degradation efficiency of all group were 10.0%-66.0%（WW）higher than CK2.U1（addition of 1 g urea）and U3 groups showed higher degradation efficiency than CK2,with 13.4%±1.4%and 11.5%±1.6%higher,respectively.The results of scanning electron microscope pictures further showed that the introduction of BSFL to cow manure could accelerate the decomposition of lignocellulose,and added urea could further promote the decomposition of lignocellulose in cow manure by BSFL within the uera addition range of this experiment.In summary,BSFL can efficiently use urea and convert urea-N into its own biomass.The metabolic pathway may be that urea is rapidly decomposed to produce NH₄⁺,then synthesized glutamine and used by BSFL,while some NH₄⁺is mainly affected by pH and converted into NH₃ form loss.Urea can be used as a nitrogen source supplement to increase the yield of larvae,promote the conversion of waste,and achieve clean recovery of carbon resources.This provides direction for BSFL to treat more organic wastes with a high carbon-to-nitrogen ratio,and provides theoretical guidance for reducing nitrogenous gas emissions during conversion.