Manli Duan

Variable effects of oxytetracycline on antibiotic resistance gene abundance and the bacterial community during aerobic composting of cow manure

Livestock manure is often subjected to aerobic composting but little is known about the variation in antibiotic resistance genes (ARGs) during the composting process under different concentrations of antibiotics. This study compared the effects of three concentrations of oxytetracycline (OTC; 10, 60, and 200mg/kg) on ARGs and the succession of the bacterial community during composting. Very similar trends were observed in the relative abundances (RAs) of each ARG among the OTC treatments and the control during composting. After composting, the RAs of tetC, tetX, sul1, sul2, and intI1 increased 2-43 times, whereas those of tetQ, tetM, and tetW declined by 44-99%. OTC addition significantly increased the absolute abundances and RAs of tetC and intI1, while 200mg/kg OTC also enhanced those of tetM, tetQ, and drfA7. The bacterial community could be grouped according to the composting time under different treatments. The highest concentration of OTC had a more persistent effect on the bacterial community. In the present study, the succession of the bacterial community appeared to have a greater influence on the variation of ARGs during composting than the presence of antibiotics. Aerobic composting was not effective in reducing most of the ARGs, and thus the compost product should be considered as an important reservoir for ARGs.

Reducing antibiotic resistance genes, integrons, and pathogens in dairy manure by continuous thermophilic composting

This study explored the effects of composting using three temperature regimes, namely, insufficient thermophilic composting (ITC), normal thermophilic composting (NTC), and continuous thermophilic composting (CTC), on antibiotic resistance genes (ARGs), integrons, and human pathogenic bacteria (HPB), as well as the mechanisms involved. The NTC and CTC treatments led to greater decreases in 5/10 ARGs and two integrons than ITC, and the abundances of ARGs (tetC, tetG, and tetQ) and int1 only declined in the NTC and CTC treatments. The abundances of HPB decreased by 82.8%, 76.9%, and 96.9% under ITC, NTC, CTC, respectively. Redundancy analysis showed that both bacterial succession and horizontal gene transfer play important roles in the variation of ARGs, and the changes in different ARGs were due to diverse mechanisms. CTC performed significantly better at reducing ARGs, integrons, and HPB, thus it may be used to manage the public health risks of ARGs in animal manure.

Mechanism and Effect of Temperature on Variations in Antibiotic Resistance Genes during Anaerobic Digestion of Dairy Manure

Animal manure comprises an important reservoir for antibiotic resistance genes (ARGs), but the variation in ARGs during anaerobic digestion at various temperatures and its underlying mechanism remain unclear. Thus, we performed anaerobic digestion using dairy manure at three temperature levels (moderate: 20 °C, mesophilic: 35 °C, and thermophilic: 55 °C), to analyze the dynamics of ARGs and bacterial communities by quantitative PCR and 16S rRNA gene sequencing. We found that 8/10 detected ARGs declined and 5/10 decreased more than 1.0 log during thermophilic digestion, whereas only four and five ARGs decreased during moderate and mesophilic digestion, respectively. The changes in ARGs and bacterial communities were similar under the moderate and mesophilic treatments, but distinct from those in the thermophilic system. Potential pathogens such as Bacteroidetes, Proteobacteria, and Corynebacterium were removed by thermophilic digestion but not by moderate and mesophilic digestion. The bacterial community succession was the dominant mechanism that influenced the variation in ARGs and integrons during anaerobic digestion. Thermophilic digestion decreased the amount of mesophilic bacteria (Bacteroidetes and Proteobacteria) carrying ARGs. Anaerobic digestion generally decreased the abundance of integrons by eliminating the aerobic hosts of integrons (Actinomycetales and Bacilli). Thermophilic anaerobic digestion is recommended for the treatment and reuse of animal manure.

Effects of bamboo charcoal on antibiotic resistance genes during chicken manure composting

Composting is widely used for animal waste disposal, and bamboo charcoal (BC) can be used for nitrogen conservation during composting. However, the effects of BC on antibiotic resistance genes (ARGs) during chicken manure composting are still unclear. This study investigated the effects on ARGs of adding different proportions of BC (0%, 5%, 10%, and 20% w/w) to chicken manure compost. After 26 days, the relative abundances (RAs) of most ARGs (tetC, tetG, tetW, tetX, sul2, drfA1, drfA7, ermB, ermF, ermQ, and ermX) and intI1 declined by 21.6-99.5%, whereas sul1 increased by 7.5-17.7 times. The average RAs reductions with 0%, 5%, 10%, and 20% BC were 0.85, 1.05, 1.08, and 1.15 logs, respectively. The most important environmental factor for the ARG profiles was temperature according to redundancy analysis. Furthermore, BC significantly decreased the bio-Cu and bio-Zn levels, thereby reducing the co-selection pressure from heavy metals. Different proportions of BC had no significant effects on the removal of tetG, tetW, tetX, sul2, drfA1, and ermB. Supplementation with 10% BC was more effective at removing tetC and drfA7 compared with the other treatments. The results suggested that 10% BC supplementation is appropriate for reducing ARGs in chicken manure compost.

Relationships between sulfachloropyridazine sodium, zinc, and sulfonamide resistance genes during the anaerobic digestion of swine manure.

In this study, swine manure containing sulfachloropyridazine sodium (SCPS) and zinc was subjected to mesophilic (37°C) anaerobic digestion (AD). The absolute abundances (AAs) of antibiotic resistance genes (ARGs) were evaluated, as well as intI1 and intI2, and the degradation of SCPS according to variation in the amount of bio-available zinc (bio-Zn). In digester that only contained SCPS, the concentrations of SCPS were lower than that digesters both contain SCPS and Zn. Compared with the control digester, the addition of SCPS increased the AAs of sul1, sul3, drfA1, and drfA7 by 1.3-13.1 times. However, compared with the digester with SCPS but no added Zn, the AAs of sul3, drfA1, and drfA7 were decreased by 21.4-70.3% in the presence of SCPS and Zn, whereas sul1 and sul2 increased 1.3-10.7 times. There were significant positive correlations (P<0.05) between the concentrations of SCPS with several ARGs and bio-Zn.

Effects of genetically modified cotton stalks on antibiotic resistance genes, intI1, and intI2 during pig manure composting

Genetically modified (GM) cotton production generates a large yield of stalks and their disposal is difficult. In order to study the feasibility of using GM cotton stalks for composting and the changes that occur in antibiotic resistance genes (ARGs) during composting, we supplemented pig manure with GM or non-GM cotton stalks during composting and we compared their effects on the absolute abundances (AA) of intI1, intI2, and ARGs under the two treatments. The compost was mature after processing based on the germination index and C/N ratio. After composting, the AAs of ARGs, intI1, and intI2 were reduced by 41.7% and 45.0% in the non-GM and GM treatments, respectively. The ARG profiles were affected significantly by temperature and ammonia nitrogen. In addition, excluding tetC, GM cotton stalks had no significant effects on ARGs, intI1, and intI2 compared with the non-GM treatment (p < 0.05). Thus, similar to non-GM cotton stalks, GM cotton stalks can be used for aerobic composting with livestock manure, and the AAs of ARGs can be reduced. Furthermore, the results of this study provide a theoretical basis for the harmless utilization of GM cotton stalks.

Effects of sulphamethazine and zinc on the functional diversity of microbial communities during composting

ABSTRACT The changes in the functional diversity of the microbial community in a compost matrix with a single or compound addition of zinc (Zn; 0, 600, and 1800 mg/kg) and sulphamethazine (SM2; 0, 1, and 25 mg/kg) were studied with the Biolog method during composting. The microbial community was extracted from the compost matrix comprising swine manure and wheat straw at day 6 (themophilic period) and day 25 (mature period) of composting. Results proved that the Shannon index, average well-colour development, and substrate utilization significantly decreased as the concentrations of SM2 and Zn increased on day 6. The negative effect of the combined addition of SM2 and Zn was lower than that of the individual addition of SM2 and Zn. On day 25, the inhibition effect disappeared, and microbial metabolic activities were higher than those on day 6. The effects of SM2 and Zn could be further differentiated via the principal component analysis (PCA) and cluster analysis. On day 6, the treatments were divided into three groups by PC1 and PC2. The separation of the different treatments in the PCA plots became increasingly apparent on day 25. In conclusion, the effects of SM2 and Zn on the microbial community during composting became evident in the themophilic period and that the microbial activity recovered in the mature period. The combination of SM2 and Zn decreased the inhibition with the addition of individual additive. GRAPHICAL ABSTRACT

Prevalence of quinolone resistance genes, copper resistance genes, and the bacterial communities in a soil-ryegrass system co-polluted with copper and ciprofloxacin

The presence of high concentrations of residual antibiotics and antibiotic resistance genes (ARGs) in soil may pose potential health and environmental risks. This study investigated the prevalence of plasmid-mediated quinolone resistance (PMQR) genes, copper resistance genes (CRGs), and the bacterial communities in a soil-ryegrass pot system co-polluted with copper and ciprofloxacin (CIP; 0, 20, or 80 mg kg-1 dry soil). Compared with the samples on day 0, the total relative abundances of the PMQR genes and mobile genetic elements (MGEs) were reduced significantly by 80-89% in the ryegrass and soil by the cutting stage (after 75 days). The abundances of PMQR genes and MGEs were reduced by 63-81% in soil treated with 20 mg kg-1 CIP compared with the other treatments, but the abundances of CRGs increased by 18-42%. The presence of 80 mg kg-1 CIP affected the microbial community structure in the soil by increasing the abundances of Acidobacteria and Thaumarchaeota, but decreasing those of Firmicutes. Redundancy analysis indicated that the pH and microbial composition were the main factors that affected the variations in PMQR genes, MGEs, and CRGs, where they could explain 42.2% and 33.3% of the variation, respectively. Furthermore, intI2 may play an important role in the transfer of ARGs. We found that 80 mg kg-1 CIP could increase the abundances of ARGs and CRGs in a soil-ryegrass pot system.

Combined effects of compost containing Sulfamethazine and zinc on pakchoi (Brassica chinensis L.) growth, soil sulfonamide resistance genes, and microbial communities

ABSTRACT To understand the ecotoxicity of antibiotics and heavy metals in soil, 5% (w/w) composts containing different concentrations of sulfamethazine (SMZ) and/or zinc (Zn) were applied to soil to investigate their effects on pakchoi (Brassica chinensis L.) growth, soil sulfonamide resistance genes (SRGs: sul1, sul2, and dfrA7), and soil microbial communities. Composts containing less than 1.0 mg SMZ kg–1 or less than 2.8 g Zn kg–1 promoted pakchoi growth and the metabolic activity of soil microbial communities. Compared with the control, the absolute abundances (AAs) of soil SRGs significantly increased by 0.85–4.54 times with 50.6 μg kg–1 SMZ treatment (P < 0.05), the AA of sul2 increased by 166% with 248.8 mg Zn kg–1 treatment. The combination treatments with 19.8 μg SMZ kg–1 and 179.9 mg Zn kg–1 in soil had synergistic stimulatory effects on pakchoi growth, soil SRGs and microbial metabolism, whereas 53.5 μg SMZ kg–1 and 262.1 mg Zn kg–1 had an opposite effect. Pearson’s correlation analysis showed that carbon metabolism by soil microorganisms had significant positive correlations with shoot height of pakchoi (r = 0.84, P < 0.05) and AAs of SRGs (r > 0.80, P < 0.05).

Effects of inoculation with organic-phosphorus-mineralizing bacteria on soybean (Glycine max) growth and indigenous bacterial community diversity

Three different organic-phosphorus-mineralizing bacteria (OPMB) strains were inoculated to soil planted with soybean (Glycine max), and their effects on soybean growth and indigenous bacterial community diversity were investigated. Inoculation with Pseudomonas fluorescens Z4-1 and Brevibacillus agri L7-1 increased organic phosphorus degradation by 22% and 30%, respectively, compared with the control at the mature stage. Strains P. fluorescens Z4-1 and B. agri L7-1 significantly improved the soil alkaline phosphatase activity, average well color development, and the soybean root activity. Terminal restriction fragment length polymorphism analysis demonstrated that P. fluorescens Z4-1 and B. agri L7-1 could persist in the soil at relative abundances of 2.0%-6.4% throughout soybean growth. Thus, P. fluorescens Z4-1 and B. agri L7-1 could potentially be used in organic-phosphorus-mineralizing biofertilizers. OPMB inoculation altered the genetic structure of the soil bacterial communities but had no apparent influence on the carbon source utilization profiles of the soil bacterial communities. Principal components analysis showed that the changes in the carbon source utilization profiles of bacterial community depended mainly on the plant growth stages rather than inoculation with OPMB. The results help to understand the evolution of the soil bacterial community after OPMB inoculation.