Effects Of Microbiological Agents On Heavy Metal Passivation,reduction Of Antibiotic And Resistance Genes In Aerobic Composting Of Pig Manure

In recent years,heavy metals such as copper（Cu）and zinc（Zn）and various antibiotics have been added to poultry breeding to promote animal growth and reduce the risk of disease.However,these elements cannot be fully absorbed by animals,and most of them are excreted in the feces and still be highly active.As a result,antibiotic resistance genes in pig manure are frequently detected,causing huge environmental risks.At present,the best way to dispose livestock manure is composting.However,the traditional composting method not only has a long cycle and serious nitrogen loss,but also produces a foul smell and weakens the agricultural value of composting products.Based on this,this study was conducted to determine the effects of microbial inoculants（i.e.Phanerochaete chrysosporium（p）,Aspergillus niger（a）and Bacillus licheniformis（b））and their different combination on the composting process,antibiotics,heavy metal removal,antibiotic resistant gene passivation,and nitrogen transformation during pig manure composting.And the mechanism of removing antibiotics and reducing drug resistance genes in livestock feces by aerobic compost inoculation with compound bacteria was discussed.In this experiment,20 strains of thermophilic bacteria were screened from fresh pig manure by high temperature screening method,and then their heat resistance was tested.Seven heat-resistant strains,including LB1,LB5,GS1,GS2,GS3,GS4 and GS6,were observed morphologically.Combined with the results of heat resistance experiment and morphological observation,LB1 was selected as the thermophilic bacteria added in compost.It was identified as Bacillus licheniformis by molecular biology.This study was conducted to determine the effects of microbial inoculants（i.e.Phanerochaete chrysosporium（p）,Aspergillus niger（a）and Bacillus licheniformis（b））and their different combination on the composting process,heavy metal passivation and nitrogen transformation during pig manure composting.The results indicated the promoting efficiency of composting process and maturity ranked in the following order:the inoculation treatment（b+p+a）>the inoculation treatment（b+p）>the inoculation treatment（p）.The germination index（GI）reached 80%and C/N ratio decreased to 11.4 on day 16 in the inoculation treatment D（b:p:a was 1:5:5）,indicating that the inoculation strategy achieved the best composting effect.The highest passivation efficiency for heavy metals（Cu of 69.7%,and Zn of 59.4%）was obtained in the inoculation treatment（p）,followed by the inoculation treatment（b+p+a）and the inoculation treatment（b+p）.The best nitrogen retention was achieved in the inoculation treatment（b+p）,followed by the inoculation treatment（b+p+a）and the inoculation treatment（p）.Compared to control treatment（CK）,the loss and total nitrogen and organic nitrogen in inoculation treatment（b:p:a=1:8:0）were reduced by17.3%and 18.5%,respectively.On this basis,the effects of different strains of the compound bacteria on the removal of antibiotics and ARGs and the evolution of bacterial communities were further investigated.The results showed that the highest removal extent of tetracycline and oxytetracycline occurred in pile D（p:b:a=1:5:5,biomass）reaching 89.2%and 87.8%,respectively,while the highest removal extent of doxycycline and enrofloxacin occurred in pile A（p:b:a=1:0:0,biomass）reaching 98.6%and 89%,respectively.Compared with the levels in B（control check）,in D,antibiotic resistance genes（ARGs）,except those for sulfonamides,decreased by 1.059×10^-3–6.68×10^-2gene copies/16S r RNA copies.Inoculation with p alone effectively reduced int I1 and int I2.Pearson correlation analysis（PCA）showed that the physical and chemical properties,microbial community structure,antibiotic resistance genes and mobile genetic factors of compost were strongly correlated with antibiotic residues.Antibiotic residues and Treponema2 Rikenellaceae RC9gutgroup,Streptococcus,Ruminococcaceae TCG-005,Lachnospiraceae XPB1014group,Ruminococcaceae TCG-002,Ruminococcaceae TCG-014 and Acinetobacter present a strong positive correlation,which indicates that these bacteria may have the ability to degrade these antibiotics.Treponema2,Ruminococcaceae TCG-005,Prevotellaceae TCG-003 and Lactobacillus were positively correlated with most resistant gene,shows that these species may be linked to the spread of drug-resistant genes are closely related.From what has been discussed above,we discussed the mechanism of removing antibiotic and drug resistance genes by adding bacterial agent D（p+b+a）in aerobic compost.The addition of bacterial agent to the reactor accelerated the biochemical reaction of fermentation,and the physicochemical properties of the reactor changed significantly.The change of these physical and chemical factors was the main reason for the change of microbial community structure.Bacteria that are closely related to the removal of antibiotics and the reduction of drug resistance genes,proliferate in the dramatic evolution of microbial community structure.They rapidly promoted the composting process while degrading antibiotics and reducing resistance genes,and finally achieved a good composting effect in a short time.