Effects of soil habitat changes on antibiotic resistance genes and related microbiomes in paddy fields.

The changes of paddy soil habitat profoundly affect the structure and function of soil microorganisms, but how this process drives the growth and spread of manure- derived antibiotic resistance genes (ARGs) after entering the soil is unclear. Herein, this study explored the environmental fate and behavior of various ARGs in the paddy soil during rice growth period. Results showed that most ARG abundances in flooded soil was lower than that in non-flooded soil during rice growth (decreased by 33.4 %). And soil dry-wet alternation altered microbial community structure in paddy field (P < 0.05), showing that Actinobacteria and Firmicutes increased in proportion under non-flooded conditions, and Chloroflexi, Proteobacteria and Acidobacteria evolved into the dominant groups in flooded soil. Meanwhile, the correlation between ARGs and bacterial communities was stronger than that with mobile genetic elements (MGEs) in both flooded and non-flooded paddy soils. Furthermore, soil properties, especially oxidation reduction potential (ORP), were proved to be an essential factor in regulating the variability of ARGs in the whole rice growth stage by structural equation model, with a direct influence (λ = 0.38, P < 0.05), following by similar effects of bacterial communities and MGEs (λ = 0.36, P < 0.05; λ = 0.29, P < 0.05). This study demonstrated that soil dry-wet alternation effectively reduced the proliferation and dissemination of most ARGs in paddy fields, providing a novel agronomic measure for pollution control of antibiotic resistance in farmland ecosystem.

Exploring antibiotic resistance load in paddy-upland rotation fields amended with commercial organic and chemical/slow release fertilizer.

Agricultural fertilization caused the dissemination of antibiotic resistance genes (ARGs) in agro-ecological environment, which poses a global threat to crop-food safety and human health. However, few studies are known about the influence of different agricultural fertilization modes on antibiotic resistome in the paddy-upland rotation soils. Therefore, we conducted a field experiment to compare the effect of different fertilization (chemical fertilizer, slow release fertilizer and commercial organic fertilizer replacement at various rates) on soil antibiotic resistome in paddy-upland rotation fields. Results revealed that a total of 100 ARG subtypes and 9 mobile genetic elements (MGEs) occurred in paddy-upland rotation soil, among which MDR-ARGs, MLSB-ARGs and tet-ARGs were the dominant resistance determinants. Long-term agricultural fertilization remarkably facilitated the vertical accumulation of ARGs, in particular that bla ampC and tetO in relative abundance showed significant enrichment with increasing depth. It's worth noting that slow release fertilizer significantly increased soil ARGs, when comparable to manure with 20% replacing amount, but chemical fertilizer had only slight impact on soil ARGs. Fertilization modes affected soil microbial communities, mainly concentrated in the surface layer, while the proportion of Proteobacteria with the highest abundance decreased gradually with increasing depth. Furthermore, microbial community and MGEs were further proved to be essential factors in regulating the variability of ARGs of different fertilization modes by structural equation model, and had strong direct influence (λ = 0.61, p < 0.05; λ = 0. 55, p < 0.01). The results provided scientific guidance for reducing the spreading risk of ARGs and control ARG dissemination in agricultural fertilization.

Study on the Quality of Mixed Silage of Rapeseed with Alfalfa or Myriophyllum.

The objective of this study was to improve the comprehensive rate of utilization of rapeseed (Brassica napus subsp. napus L.), Myriophyllum (Myriophyllum spicatum L.) spicatum and alfalfa (Medicago sativa L.), reduce resource waste and environmental pollution. In this experiment, the effects of different proportions of the mixed silage of rapeseed and alfalfa or M. spicatum on the fermentation and nutritional quality were analyzed and further improved the quality of mixed silage using molasses and urea. Rapeseed was separately silaged with alfalfa and M. spicatum based on the ratios of 3:7, 5:5 and 7:3. After 60 days of mixed silage, the fermentation index and nutrient contents were measured to explore the appropriate ratio of mixed silage. The mixing ratio of rapeseed and alfalfa was better at 3:7: The contents of NH3-N/TN (4.61%), lactic acid (96.46 g·kg-1 dry matter [DM]) were significantly higher (p < 0.05). The crude protein content (118.20 g·kg-1 DM) was the highest (p < 0.05), while the pH (4.56) was the lowest when the mixing ratio of rapeseed and M. spicatum was 7:3. Considering the fermentation and nutrition quality, it is suggested that rapeseed and alfalfa should be mixed as silage at a ratio of 3:7 with 3% molasses and 0.3% urea, and rapeseed and M. spicatum should be mixed as silage at a ratio of 7:3 with 3% molasses.

Deciphering discriminative antibiotic resistance genes and pathogens in agricultural soil following chemical and organic fertilizer.

Fertilizers containing rich nutrients can change the profiles of antibiotic resistant pathogens (ARPs) and antibiotic resistance genes (ARGs) in receiving soils; however, the discriminative ARGs and ARPs in agricultural soil following different fertilizer applications remain unknown. Using metagenomic sequencing combined with binning approach, the present study investigated the discriminative ARGs and ARPs under various fertilizer applications (chemical and organic fertilizer) in a 8-year field experiment. VanR, multidrug ARG transporter, vanS, ermA, and arnA were the discriminative ARGs in the chemical fertilizer group, whereas rosB, multidrug transporter, mexW, and aac(3)-I were enhanced in the organic fertilizer group. The metagenomic binning approach revealed that both fertilizer applications caused pathogen proliferation. Chemical fertilizer caused the increase in the pathogenic genus Luteimonas, and organic fertilizer facilitated the proliferation of the pathogenic genera Dokdonella and Pseudomonas. The pathogenic species Pseudomonas_H sp014836765, carrying mexW and multidrug transporter, was enriched only in the organic fertilizer group, indicating that it was a discriminative ARP in the organic fertilizer group. Our results demonstrated that fertilizer application, particularly organic fertilizer application, can facilitate the proliferation of ARGs and ARPs in the receiving soil, posing the risk of the development and spread of soil-borne ARPs.

Adsorption of Phosphate in Aqueous Phase by Biochar Prepared from Sheep Manure and Modified by Oyster Shells.

Sheep manure and oyster shells as C and Ca sources, respectively, were used to obtain Ca-enriched biochar materials with a high dephosphorization efficiency. This approach is helpful for the utilization of livestock manure and shell solid waste as well as for creating highly adsorbent materials. The results show that as the Ca content in biochar was increased, the material's phosphate adsorption capacity increased. The maximum adsorption efficiency reached 94%. The highest adsorption capacity (calculated using Langmuir fitting) of the material containing 1:1 biochar/oyster shell weight ratio reached 146.3 mg P/g. With the increase of the pH value of phosphate solution, the adsorption capacity of the sample gradually increased to 89.5-93.3 mg P/g. The adsorption occurred mainly by complexation. The results of this work provide insights into livestock manure and shell solid waste utilization, which yields a material with useful adsorption properties that can be applied for the removal of phosphate and other inorganics from water.

[Effects of Different Fertilization Patterns on Nitrogen Leaching Loss from Paddy Fields Under Reduced Nitrogen].

Nitrogen leaching loss in paddy fields is one of the main ways of farmland non-point source pollution. To explore the suitable fertilization of rice fields in the Erhai Lake Basin and reduce the nitrogen loss from paddy fields, a field experiment was conducted by setting single applications of chemical or organic fertilizer, combined organic and inorganic application, and single application of controlled release fertilizer under reduced nitrogen conditions. The results showed that, compared with the conventional fertilization treatment(CF), there was no significant difference in rice grain and straw yield between the single chemical fertilizer treatment(T1) and the organic-inorganic combined treatment(T3); the single organic fertilizer treatment(T2) decreased the rice grain yield by 13.0%, and decreased straw yield by 17.1%; single application of controlled-release fertilizer(T4) increased rice grain and straw yield by 15.7% and 21.0%, respectively. Further, compared with CF, the single application of chemical fertilizer(T1), organic fertilizer(T2), and organic-inorganic combined application(T3) reduced the total nitrogen leaching loss at 30 cm depths by 26.9%, 18.0%, and 33.9%, respectively. The loss of ammonia nitrogen leaching with T1, T2, and T3 decreased by 24.4%, 36.9%, and 36.6%, respectively, and the loss of nitrate nitrogen leaching decreased by 40.2%, 4.8% and 46.4%. The total nitrogen leaching at 60 cm soil depths was reduced by 34.2%, 26.3%, and 42.1%, the loss of ammonia nitrogen leaching was reduced by 31.4%, 35.7%, and 46.6%, and the loss of nitrate nitrogen leaching was reduced by 8.0%, 10.1%, and 23.9% for T1, T2, and T3, respectively. The total nitrogen loss at 30 and 60 cm depths increased by 41.6% and 14.0% in the single application of controlled release fertilizer(T4) treatment. Considering factors such as agronomic and environmental benefits of different fertilization modes, T1 and T3 are suitable environmentally friendly alternative fertilization modes.

Systematic analysis of occurrence, density and ecological risks of 45 veterinary antibiotics: Focused on family livestock farms in Erhai Lake basin, Yunnan, China.

Antibiotic pollution from family animal farms is often neglected, but the waste from these farms usually caused more harm to the surroundings because arbitrary discharge without effective disposal. The pollution status and ecological risks of 45 veterinary antibiotics on 33 family animal farms in Dali city, Erhai Lake basin of China, were firstly delivered. The results showed that antibiotic contamination was prevalent in different environmental mediums (feed, manure, wastewater and soil) on these family farms. Manure had highest antibiotic levels among all the environmental mediums. Tetracyclines (TCs) usually had higher concentrations (ND-404.95 mg/kg) than the other classes, among which chlorotetracycline (CTC) was the dominant type. Among different animal species, target 13 pig farms had the highest antibiotic concentrations, the most total types and unique types of antibiotics, which were followed by target 11 chicken farms then target 9 cattle farms. The antibiotic densities of animal waste were calculated by per animal, which showed that pig waste presented high density; and family chicken farms were characterized by quinolone antibiotics (QAs) and macrolide antibiotics (MAs) pollution. For the antibiotic ecological risks in effluent water, oxytetracycline (OTC), CTC, ofloxacin (OFL), enrofloxacin (ENR), ciprofloxacin (CIP) and sulfamethoxazole (SMX2) exhibited much more toxic effects on algae. OTC and doxycycline (DXC) posed high risk for invertebrate; while no antibiotic caused high ecological risk for fish. Some antibiotics were quantitatively detected in the soil but no antibiotic posed obvious ecological risks on soils. However, the interaction of synergistic or antagonistic effects between different antibiotics should be brought to the forefront. This study gave some information of antibiotic pollution on family livestock farms, which indicated that animal waste from family farms was indeed an important pollution source of antibiotics for the environment.

Family livestock waste: An ignored pollutant resource of antibiotic resistance genes.

The random discharge of livestock waste from family farms without utilization and treatment has caused great pressure on the rural ecological environment and gravely increased the environmental pollution. In this study, we targeted 26 family livestock farms to assess the occurrence characteristics of antibiotic resistance genes (ARGs) in livestock waste and its receiving farmland environment in Erhai Lake basin of China by real-time fluorescence quantitative PCR. The results showed that various common ARGs and some high-risk ARGs (i.e., blaampC, blaOXA-1 and blaTEM-1) were prevalent in family livestock waste, and the pollution of tetracycline resistance genes was the most serious in these family livestock farms. Meanwhile, we also found that the ARG levels were higher in family chicken farms than that in pig and cattle farms, and ARGs pollution in layer waste and sow waste was more severe than that in broiler waste and piglet/fattening pig waste, respectively. Troublesomely, significant ARGs levels could be discharged via manure application, further causing the increase of ARGs abundance in soil environment (approximately 11-36 times). This study demonstrated the high prevalence and severity of ARGs contamination in family livestock farms, also emphasizing that family livestock waste was a non-ignored important pollutant resource of ARGs in the environment.

Study on nitrogen removal from rice paddy field drainage by interaction of plant species and hydraulic conditions in eco-ditches.

Eco-ditches (ecological ditches) not only drain water from rice paddies, but also facilitate the removal of nitrogen (N). We established an experiment with both static and flowing water in 2017 to observe N removal from rice paddy drainage by eco-ditches containing three different types of monoculture vegetation: Zizania aquatica, Canna indica L., and Pontederia cordata. Results showed that ammonia volatilization and plant uptake contributed little to N removal. Harvest of Z. aquatica from the eco-ditch during the late growing season had an appreciable effect on N removal. However, harvest of C. indica L. and P. cordata had negligible effects. During static test, the concentration of total N (TN) and ammonium N (NH4+-N) and the pH all decreased from the surface to the bottom of water. The concentration of nitrate N (NO3--N) did not exhibit stratification. In a flowing water experiment, ditches with Z. aquatica, C. indica L., and P. cordata had the following average removal rates: TN 15.8%, 11.6%, and 27.9%; NO3--N 4.2%, 8.4%, and 17.8%; NH4+-N 22.8%, 16.4%, and 37.5%, respectively. The removal rates of TN and NH4+-N decreased with the increase of water level, while that of NO3--N increased significantly. Nitrogen removal rates decreased with the increase of influent TN concentration or flow rate. Nitrogen removal rate of P. cordata ditch was highly dependent on the influent TN concentration, but the flow rate was not as important due to the great drag caused by its large density. While the contrary was observed in the C. indica L. ditch. For Z. aquatica ditch, both the flow rate and the TN concentration had a strong negative correlation with the N removal rate.