Utilization Of Acidolysis Amino Acids From Dead Pigs To Produce Bio-organic Fertilizers And Their Effects On Soil Microbial Community

Highly intensive animal production systems in China have resulted in large amounts of dead pigs,which cause serious environmental issues.However,few studies have focused on the utilization of dead pigs.On the other hand,continuous cropping obstacle of tomato and cucumber have caused a serious threat to crop production.Thus,it is necessary to explore strategies to solve the problems.The main job of this study was to utilize amino acids hydrolyzed from dead pigs to produce bio-organic fertilizers(BIOs)and to evaluate their effects on soil microbial community.In the first part,a novel method to hydrolyze the dead pigs into amino acids by H2SO4 was investigated;the acidolysis amino acids were utilized to produce BIOs;and,the microbial community during BIOs production were studied by Illumine-MiSeq sequencing.In the second part,effects of novel Bacillus amyloliquefaciens SQR9 bio-organic fertilizer(SQR9 BIO)produced in the first part on the biomass,yield and disease incidence of tomato,as well as the microbial communities of tomato bulk and rhizosphere soil were evaluated.In the third part,the effect of novel Trichoderma guizhouese NJAU 4742 bio-organic fertilizer(NJAU 4742 BIO)produced in the first part on the biomass and yield of cucumber,as well as the microbial communities of cucumber bulk and rhizosphere soil were evaluated.Moreover,we studied the effects of soil microbial community influenced by NJAU 4742 BIO on biomass accumulation and microbial communites of cucumber and Arabidopsis through soil slurry experiment.The main results were summarized as follows:1.All the proteins of the dead pigs could be hydrolyzed into amino acids under the acid hydrolyzed reaction conditions:5 M sulfuric acid,80 ? reaction temperature,1:1.5(Ig fresh lean meat:1.5 ml acid solution)materials ratio and 6 h reaction time.After three times of dead pigs amendment,the concentration of free amino acids was 12.4%.After pre-compost for 4 days before Bacillus or Trichoderma inoculation,treatments of matured chicken or pig manure added with 0.2 ml/g(DW)of acidolysis amino acids resulted in a maximum cell density of functional strains.Results showed that the additional acidolysis amino acids decreased the pH value and increased the total nitrogen(TN)content of matured manure.MiSeq sequencing of the 16S rRNA and internal transcribed spacer(ITS)genes showed that the additional acidolysis amino acids reduced the bacterial richness and increased the relative abundance of Bacillus,providing a suitable environment for SQR9 colonization,which greatly influenced the bacterial community.The relative abundance of Trichoderma increased significantly when inoculated NJAU 4742 in the materials after pre-compost,and the fungal community was greatly influenced.2.Results showed that different treatments resulted in different disease incidences(DI)of tomato in the field experiment,and soil amended with organic fertilizer(OF,DI = 30%),amino acid organic fertilizer(AOF,DI = 16%),and SQR9 BIO(DI = 6%)were progressively and significantly suppressed the tomato disease in comparison with chemical fertilizer(CF,DI = 59%)in the three seasons field experiment.Interestingly,the rhizospheric soil bacterial and fungal communities of the different fertilization managements were well separated from each other,as the same trends were observed for bulk soil bacterial and fungal communities.Moreover,we found that 6 bacterial and 10 fungal rhizospheric genus correlated significantly with the tomato disease incidence.Furthermore,the relative abundances of 8 of 16 genus in the rhizosphere were significantly(P<0.01)and positively correlated with the same genus in bulk soil.Additionally,CF-treated bulk soil contained high relative abundances of Ralstonia and Fusarium.Compared to the CF,soil amended with SQR9 BIO showed significant lower relative abundances of Ralstonia and Fusarium,but only the bulk soils in OF and AOF treatments showed significant lower relative abundance of Ralstonia.3.Compared to the CF,OF and AOF,application of NJAU 4742 BIO significantly promoted cucumber growth and increased cucumber yield.Compared to the AOF,soil amend with NJAU 4742 BIO significantly shifted the bulk soil fungal community,and influenced the rhizospheric fungal community similarly.In addition,application of NJAU 4742 BIO significantly increased the relative abundance of Trichederma and decrased the relative abundance of Fusarium in both cucumber bulk and rhizospheric soils.Results of pot experiments showed that it was not the Trichoderma guizhouese NJAU 4742 playing the most important role in promoting cucumber growth.Actually,compared to the AOF,the soil microbial community regulated by NJAU 4742 BIO significantly promoted the growth and influenced the fungal community of cucumber and Arabidopisis.Moreover,the soil suspension without fungi regulated by NJAU 4742 BIO and AOF showed no significant difference in promotion of the growth of cucumber and Arabidopisis.In conclusion,this work successfully applied acidolysis amino acids from dead pigs in production of novel SQR9 and NJAU 4742 BIOs.Application of the novel SQR9 BIO significantly decreased the disease incidence of tomato,reduced the relative abundance of Ralstonia and Fusarium in the rhizosphere,and altered the rhizospheric bacterial and fungal microbial communities through shifting the bacterial and fungal microbial communities in the bulk soil.In addition,application of the novel NJAU 4742 BIO significantly promoted the growth of cucumber,maybe through regulating the fungal microbial community in bulk soil.