| Doxycycline is a kind of therapeutic veterinary antibiotic commonly used in pig breeding.Antibiotic resistance genes(ARGs)induced by doxycyline would be excreted with feces into the surrounding environment.This paper studied the change rule and factors of tetracyclines resistance genes in swine manure after doxycycline feeding,to accurately access the ecotoxicological effects by feeding doxycycline and provide a theoretical basis to decrease the discharge of ARGs from animals to environment.A total of 27 Duroc × Landrace × Large White pigs of 33.5 ± 0.72 kg were divided into 3groups with 9 pigs in each group.Based on "Veterinary Pharmacopoeia of the People’s Republic of China",doxycycline of 3 mg/kg body weight and 5 mg/kg body weight were added in feed in group L and group H,respectively.The blank group(group CK)did not add with any veterinary antibiotics.The medication and withdraw periods were set for 5 days and 28 days.Pig manure was regularly collected to determine the concentration of doxycycline,ARGs,mobile genetic genes(MGEs),microbial community structure and physicochemical indicators.The results obtained in this article are as follows:(1)After feeding with doxycycline,the concentration of doxycycline in group L peaked at157.05 ± 16.43 mg/kg DM on day 5,while that in group H peaked at 272.49 ± 28.24 mg/kg DM on day 4.The average concentration of group L and group H during the medication period were 117.63 ± 13.54 mg/kg DM and 202.03 ± 24.91 mg/kg DM,respectively.The average concentration during the drug withdrawal period were 23.45 ± 10.31 mg/kg DM and 37.89 ±16.02 mg/ kg DM,respectively.On day 20,doxycycline was lower than the detection limit.(2)The total relative abundance of tetracycline ARGs in pig manure ranged from 0.35 to0.41 copies/16 S r RNA copies,accounting for 38.00% to 46.47% of the total ARGs.Doxycycline had no significant effect on the total relative abundance of tetracycline ARGs.On day 5,the absolute abundance of tet A in pig manure in group H was significantly lower than that in group L,and the absolute abundance of tet W was significantly lower than that in groups CK and L(P < 0.05).Meanwhile,the absolute abundance of tol C,tnp A and int1 in group H were significantly lower than those in group L(P < 0.05).(3)Compared with CK group,in the medication period,the relative abundance of Blautia,Dorea,Parabacteroides and Oscillibacter in pig manure in group L was significantly decreased(P < 0.05),and the relative abundance of unclassified clostridium was significantly increased(P < 0.05).In the withdrawal period,the relative abundance of Streptococcus in pig manure in group L increased significantly,while that of Enterorhabdus in group H decreased significantly(P < 0.05).At the species level,a significant reduction of Marinobacter adhaerens resulted in a significant reduction of tet A and tol C,and the significant reduction of 8 species such as Bacillus toyonensis,Ruminococcus albus and Lachnoclostridium phocaeense resulted in a significant decline of tet W.(4)The microbial was an important factor affecting the abundance of tetracycline ARGs in pig manure,followed by the doxycycline,MGEs and physicochemical properties.Tetracycline resistance genes were significantly and positively correlated with a variety of MGEs(P < 0.05),for example,tet S and int1,tet O and tni A and Inc Q1.tet L were significantly and positively correlated with 28 MGEs,which may be one of the tetracycline reisstance genes most susceptible affected by MGEs.In summary,feeding doxycycline did not significantly affect the total relative abundance of tetracycline resistance genes,but the high concentration of doxycycline did change the microbial community structure of pig manure and lead to the decrease of potential host bacteria of tet W,tet A and tol C,thus leading to a significant decrease in the absolute abundance of tetW,tetA and tolC. |
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