Biochar-induced migration of tetracycline and the alteration of microbial community in agricultural soils

Journal Pre-proof Biochar-induced migration of tetracycline and the alteration of microbial community in agricultural soils

Hua-Yu Liu, Chao Song, Shan Zhao, Shu-Guang Wang PII:

S0048-9697(19)36082-6

DOI:

https://doi.org/10.1016/j.scitotenv.2019.136086

Reference:

STOTEN 136086

To appear in:

Science of the Total Environment

Received date:

2 September 2019

Revised date:

1 December 2019

Accepted date:

10 December 2019

Please cite this article as: H.-Y. Liu, C. Song, S. Zhao, et al., Biochar-induced migration of tetracycline and the alteration of microbial community in agricultural soils, Science of the Total Environment (2018), https://doi.org/10.1016/j.scitotenv.2019.136086

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© 2018 Published by Elsevier.

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Biochar-induced migration of tetracycline and the alteration of microbial community in agricultural soils Hua-Yu Liu, Chao Song*, Shan Zhao, Shu-Guang Wang* Shandong Key Laboratory of Water Pollution Control and Resource Reuse, School of Environmental Science and Engineering, Shandong University, Qingdao 266237, China

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Corresponding author. Tel.: +86 532 58630936; Fax: +86 532 58630907.

E-mail address: [email protected] (Chao Song); [email protected] (Shu-Guang Wang) 1

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Abstract Recently, biochar is widely used as a soil amendment to improve soil properties, which might affect the fate and behavior of contaminants in soil. In this study, we investigated the effect of biochar on the migration of tetracycline (TC) in soil and their combined impacts on microbiome. Due to the strong interaction between soil and

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TC, adsorption, rather than photolysis or biodegradation, was the dominating dissipation way of TC in soil. Moreover, biochar could promote the vertical migration

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of TC through the decreased soil bulk density and its lower adsorption capacity. After

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90-day incubation, only slight impact of TC on soil bacterial community was

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observed due to the rapid dissipation of TC in soil, whereas more available C supply

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induced by biochar significantly altered bacterial community via the enhancement of

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copiotrophic bacteria. Besides, biochar could decrease the soil pH and thus change the composition of fungal community. The effect of TC on fungal community was

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partially counteracted by biochar, which could adsorb part of TC and thus decrease the contact of TC with microorganisms. This work will improve our understanding of the fate of organic pollutants and evolution of microbiome in soil where biochar servers as soil amendment.

Keywords: Tetracycline; Biochar; Soil; Microbiome; Migration

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1. Introduction Nowadays, tetracycline antibiotics are widely used in the treatment and prevention of infections for livestock (Zhou et al., 2013). Unfortunately, 30%-90% of intake antibiotics can not be absorbed by body, and they are excreted as parent compounds or metabolites through urine and feces into the soil (Erşan et al., 2013; Ji

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et al., 2012; Zhao et al., 2010). Besides, the manure contaminated with antibiotics are also applied as fertilizers onto arable lands, leading to the enrichment of antibiotics in

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the soil (Duan et al., 2017). Residues of tetracycline have already been detected at

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concentrations ranging from ng/g to g/g in soil (Song et al., 2017; Thiele-Bruhn and

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Beck, 2005). As a broad-spectrum antibiotic, tetracycline could exhibit adverse

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influences on soil ecosystem, such as inhibiting bacteria activity, disturbing microbial

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metabolism and even changing soil microbiome (Jechalke et al., 2014; Kong et al., 2006). The residues would also induce the evolution of antibiotic resistance genes,

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which are considered as a potential threat to both ecosystem and human health (Hong et al., 2013). Therefore, understanding the fate and behavior of tetracycline in soil environment becomes indispensable. Biochar is a carbon-rich material which is produced by pyrolysis of various forms of biomass (De la Rosa et al., 2019; Duku et al., 2011). In recent years, biochar is commonly used as a soil amendment to improve soil properties (Srinivasan et al., 2015). Moreover, biochar could increase the microbial biomass and activity, and even reshape the structure of microbiome (Ahmad et al., 2014). It is worth noticing that biochar is a porous material with high surface area and exhibits excellent adsorption 3

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capacity on organic contaminants due to its surface aromaticity and functionality (Vithanage et al., 2016). Several studies have reported the biochar based removal of antibiotics such as tetracycline, ciprofloxacin and sulfonamides (Afzal et al., 2018; Ahmad et al., 2014; Rajapaksha et al., 2016). Thus, biochar may exhibit a significant impact on the migration of tetracycline in contaminated soils, which is highly related

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to the behaviors of tetracycline in soils (Thiele-Bruhn, 2003). However, the effects of biochar application on the migration and behaviors of tetracycline in soils, as well as

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its underlying mechanism, are still unclear.

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In this study, we investigated the impacts of biochar on the migration and

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behaviors of tetracycline in soils, and the objectives of this study were as follows: (i)

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to quantify the temporal dissipation and vertical migration of tetracycline in natural

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soils with/without biochar amendment; (ii) to explore the roles of tetracycline and biochar on soil microbial community; and (ⅲ) to elucidate the combined effects of

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tetracycline and biochar on soil microbiome. This work will extend our understanding of the fate and behavior of antibiotics in soils, which is vitally important to control the persistence and negative effects of antibiotics in agricultural environment. 2. Materials and methods 2.1 Chemicals and reagents Tetracycline hydrochloride (TC) were of USP grade and purchased from Aladdin Industrial Corporation, USA. Acetonitrile, methanol and other chemicals used for mobile phase were of HPLC grade and obtained from Sinopharm Chemical Reagent Company, China. The biochar was purchased from Huansheng Carbon Production 4

Journal Pre-proof Company, China, which was produced from coconut shell at 300-500oC in a vertical kiln and its particle size was 1-3 mm. All other chemicals were of analytical reagent grade and used without further purification. 2.2 Soil samples and biochar characterization Fresh soils were collected at a depth of 0-20 cm from Shandong University in

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Qingdao, China on June 2018. The samples were mixed and sieved with a 2 mm-mesh screen to remove large particles. Meanwhile, the initial soil was analyzed, and no

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tetracycline was detected.

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The surface area and pore characteristics of biochar were measured by N2

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adsorption using a SSA-4300 surface area analyzer. The basic characteristics of

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biochar were: special surface area (SSA) =1116.16 m2/g, the total pore volume (TPV)

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=1.65 cm3/g, the average pore radius (APR)= 4.17 nm. The isotherm curve (Figure S1) showed hysteresis loops in the relative pressure (p/p0) ranging from 0.2 to 1.0,

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suggesting uniform mesopores in biochar structure. 2.3 Temporal dissipation of TC The sieved soils were divided into four group: control group, dark group, sterilized group, and dark-sterilized group. In each group, thirty petri dishes were prepared with 10 g soil (dry weight) and TC at 50 g/g soil. In control group, all dishes were placed on laboratory table at room temperature without further operation. In dark group, aluminum foil was used to cover the dishes, which were then kept in dark place. For sterilized group, the dishes with 10 g soil were autoclaved at 121oC for 30 min before adding TC, and then they were placed in the same condition with 5

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control group. In dark-sterilized group, all dishes were covered with aluminum foil and placed in dark after autoclaved. At predetermined time intervals, three soil samples in one dish of each group were taken for further analysis to determine the amount of TC. Each dish was used for one-time sample collection. 2.3 Vertical transport of TC

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The sieved soils (60 g) were filled into a tube (3.5 cm inner diameter), where they were designated as Layer 1 to Layer 6 from top to bottom equally based on their

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depth.

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TC was spiked to Layer 1 at the dosage of 100 g TC/g soil. In addition, biochar was

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mixed in Layer 2 at 2% w/w, which was considered as the most suitable proportion

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(Huang et al., 2018). The tube without biochar was also prepared as control. All the

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tubes were covered with aluminum foil and kept in dark. Deionized water was sprayed to soils as fine droplets using a plastic nebulizer. The spray was very slow

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with suitable time interval to make the water equally distributed. The total volume of sprayed water was 2.4 mL, which was amount to the rainfall of a heavy rain in China (Day et al., 2018). Then, amounts of TC in each layer was measured to investigate its acute vertical migration in soils. All experiments were repeated in triplicate, and the error bars represent the standard deviations calculated for each independent experiment. 2.4 Amount of TC in soil TC in soils was extracted using the method described in previous study (Pan and Chu, 2016) with minor modifications. Briefly, soil sample (1 g) was transferred to a 6

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50 mL centrifuge tubes with 10 mL of 1/1(v/v) EDTA-Mcllvaine buffer (pH=4.0). Then, the sample was vortexed for 1 min, ultrasonically extracted for 30 min, and centrifuged at 4500 min for 5 min. The supernatants were all decanted into a 50 mL centrifuge tube, and the sediment was treated twice again with the same procedure to achieve a high extraction efficiency. All supernatants were collected together, filtered with 0.45 μm fiber filter (purchased fromShanghai Xinya purification device factory),

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and then pumped into an Oasis HLB cartridge (6 mL, 500 mg, Waters) at a flow of 3 –

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5 mL/min. Then, the cartridge was rinsed with 15 mL ultrapure water and eluted with

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8 mL methanol (containing 0.1% formic acid, v/v), followed by collecting the eluent

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for HPLC analysis.

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The quantification of TC was carried out using a high-performance liquid

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chromatography (HPLC) system equipped with an ODS C18 column (4.6×150 mm, 5-m). The wavelength of UV detector was set at 360 nm. The mobile phase (v/v)

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consisted of 0.01M anhydrous oxalic acid (80%), acetonitrile (16%) and methanol (4%) at a flow rate of 1.0 mL/min. Each sample was filtered through a 0.22 μm membrane filter before analysis. 2.5 Soil microbiome analysis Batch experiments were designed to evaluate the alteration of soil microbiome. As shown in Fig. 1, glass column reactors (internal diameter 7.5 cm, height 13 cm) were designed with porous PVC plates and 1cm-diameter pipes at the bottom. A piece of filter paper covered on the PVC plate was arranged to prevent the leakage of soil. The sieved soils (480 g) were added into reactors and designated as Layer 1 to Layer 7

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6 (2 cm for every Layer) from top to bottom. Based on the amounts of TC and biochar, all reactors were divided into four groups: control group with only sieved soils (CK group), control group with 2% (w/w) biochar in Layer 2 (BC group), experimental group with 50 μg/g TC in Layer 1 (TC group), experimental group with both TC (50 μg/g in Layer 1) and biochar (2% in Layer 2) (BT group). Artificial rain (11.55 mL,

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equivalent to 2.5 mm) was sprayed to the soils by a plastic nebulizer within 1h to simulate rainfall and maintain moisture. The rain was conducted every 3 days and the

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entire process lasted for 90 days. Three parallel reactors were set in identical

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operation for each group. Soil samples were collected from each reactor after 90-day

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operation for further analysis.

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DNA was extracted from 500 mg of soil using PowerSoil DNA Isolation Kit

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(MoBio Laboratories, Carlsbad, CA) according to the manufacturer's instructions. The extracted DNA was amplified via PCR on a Mastercycler Gradient (Eppendorf,

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Germany), and the products were purified using a QIA quick Gel Extraction Kit (QIAGEN, Germany). Then, the DNA samples were analyzed with 16S rRNA and 18S rRNA high-throughput pyrosequencing. The raw data and sequences were first quality-filtered and clustered into operational taxonomic units (OTUs) at a similarity level of 97% using Illumina Analysis Pipeline Version 2.6 and QIIME (Edgar, 2013). And then the sequences were classified into different taxonomic groups by Ribosomal Database Project (RDP) Classifier tool (Cole et al., 2009). For bacterial analysis, a total of 18649 reads belonging to 2733, 2564, 2664, 2651 operational taxonomic units (OTUs) of CK, BC, TC and BT treatment were found, respectively. For fungal 8

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analysis, 17560 reads belonging to 681, 850, 585, 836 operational taxonomic units (OTUs) of CK, BC, TC and BT treatment were found, respectively. The richness and diversity indices were calculated and PCA were used using R (Wang et al., 2012). And an Unweighted Pair Group Method with Arithmetic Mean (UPGMA) was used to describe the dissimilarity between multiple samples. (Jiang et al., 2013).

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3. Results and discussion 3.1 The short-term fate of TC in soil

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The temporal dissipation of TC was first investigated in natural soils. As shown

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in Fig. 2, the concentration of TC decreased from 50 g/g to 9.98 ± 0.89 g/g after 30

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days in control group, indicating that most TC (80.0% ± 1.77%) could be removed

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under natural conditions. Similar results were also obtained in the dark group with

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final concentration of 9.54 ± 2.01 g/g, suggesting that little TC was eliminated via photodegradation. Previous study monitored dissipation of antibiotics in manure piles

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and found no apparent effects of light on tetracycline (Storteboom et al., 2007). Similarly, photodegradation of antibiotics is assumed to be limited due to poor light penetration in realistic soils (Ozaki et al., 2011). The role of hydrolysis was also evaluated in this study, and there was no obvious change in Fig. S2, indicating the little role of hydrolysis. Meanwhile, more TC residuals were detected in sterilized group, resulting from the role of microorganism. The loss via biodegradation was calculated to be 9.34% ± 0.15% in this study. Compared to 30% oxytetracycline loss due to biodegradation in soil under aerobic conditions (Yang et al., 2009), the contribution of biodegradation is quite small in our study. This could be explained by 9

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two reasons: first, microorganisms need time to be screened out to degrade TC which didn’t exist in soil before. Second, the stronger adsorption property of TC (kd: 1093 L/kg) makes it more difficult to desorb from soil than oxytetracycline (kd: 1026 L/kg)(Wszolek and Alexander, 1979). Furthermore, there was no significant difference (RMSE=1.69) between the TC amounts on the 30th day in sterilized group

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and those in dark-sterilized group, which confirmed the limited contribution of photolysis. After 30-day operation, 64.01% ± 6.63% TC was removed from sterilized

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soil in dark condition, which was attributed to the strong adsorption of TC by soils.

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The adsorption of antibiotics to soil include reversible equilibrium process,

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sequestration and non-extractable residues (Jechalke et al., 2014). TC dissipation in

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soils could be divided into two phases: the faster phase before the 8th day and the

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slower phase after (Fig. 2). It is consistent with the two dynamic processes proposed to characterize TC adsorption to soils: a rapid initial adsorption on the outer surface

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and then the slower intercalation to microscopes (Chang et al., 2009). Most TC was sequestered into micro- and nanopores or formed non-extractable residues with soil via complexation or other mechanisms, decreasing its bioavailability and making it difficult to be extracted or detected by the method used here. Moreover, pH is an important factor that affects complexation. TC mainly exists as zwitterion in common soil with pH 3.3 – 8.0 (pH 6.7 in this study), where surface complexation between TC and soil is the dominant mechanism (Sassman and Lee, 2005). However, this part of TC will be released with slow rates and subsequently degraded by microorganisms. The incubation period (30 days) was too short to reveal this process. Moreover, the 10

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removal data was fitted with first-order kinetic model, and the estimates and 95% confidence intervals of TC half-life were listed in Table 1. The half-life of TC in soils varied from 13 to 16 days, which were consistent with that at 6 to 15 days in other study (Storteboom et al., 2007). 3.2 Vertical migration of TC in soil

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To evaluate the effect of biochar on vertical migration of TC in soil, we analyzed TC concentration in different depth of soil after TC spiking on soils with and without

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biochar, respectively. As shown in Fig. 3, TC was only found in Layer 1 in the control

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group and undetectable in deeper soils, resulting from the strong adsorption of soil on

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TC (Pan and Chu, 2016). However, TC was detected in all layers with the addition of

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biochar. Most TC (81.68 ± 3.72 g/g) was retained in Layer 1 while 7.56 ± 1.25 g/g

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in Layer 2 and 11.32 ± 12.94 g/g in other layers. These results suggest that biochar amendment promotes the migration of TC to deeper soils. The adsorption isotherms

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for tetracycline on biochar and soil also indicated the slightly lower adsorption capacity of biochar (Fig. S3). Similarly, the adsorption affinity of TC on biochar was lower than that on soil, largely due to the strong interaction between soil clay and TC (Zhang et al., 2013). Meanwhile, biochar can decrease the bulk density of soil through creating large void spaces (Jones et al., 2011), making it easier for TC to spread. However, some studies found contrary result between antibiotics and biochar/soil. For instance, the application of burcucumber-derived biochar showed an enhancement on the adsorption of sulfamethoxazole in soil (Vithanage et al., 2014). This discrepancy mainly results from the different properties of both biochar and molecular structures 11

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of pollutants. The larger molecular size of TC (MW=444.43) causes stronger steric hindrance when approaching biochar, which weaken its adsorption on biochar (Zhang et al., 2013). 3.3 Evolution of bacterial community in soils Soil samples in the four groups were collected after 90-day incubation, and both and

fungal

community were

investigated

using

high-throughput

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bacterial

pyrosequencing. The differences in bacterial communities were analyzed with Partial

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least squares Discriminant (PLS-DA) plot. As shown in Fig. 4, there was significant

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difference between samples in CK, BC, and BT group whereas slight difference was

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observed between samples in CK and TC group. It has been reported that biochar

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could increase bacterial diversity and richness in rice paddy soil from south China

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(Chen et al., 2015). In contrast, TC exhibited limited impacts on bacterial community, as no significant effect was found on dominant microbial species in soil during

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120-day incubation (Ma et al., 2016). Therefore, biochar exhibited more profound shaping influence on soil microbial community than that of TC, which might be attributed to the rapid dissipation of TC in soil. The phylogenetic classification at phylum and genus level was summarized in Fig. 5. After 90-day incubation, the most dominant phyla were Acidobacteria and Proteobacteria in all samples with relative abundance at 32.2%~36.5% and 28.4~36.4%, which is similar to other studies on bacterial communities with carbon materials (Xu et al., 2014). Other prevalent phyla included Nitrospirae, Bacteroidetes, Gemmatimonadetes, and Verrucomicrobia with relative abundance above 1%. At the 12

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genus level, RB41 was found to be the dominant genus (relative abundance = 10%). Other highly abundant genus including Sphingomonas, Nitrospira, 11-24, and H16 were also detected with relative abundance above 2% of the total community. Furthermore, Bacterial community in biochar-amended soil was different from that in CK. The addition of biochar increased the relative abundance of Actinobacteria

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and decreased that of Acidobacteria and Nitrospirae. Biochar addition to soil commonly increases soil C/N ratio (Chan et al., 2007). Meanwhile, micro- and

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macropores in biochar hold more oxygen and air content which has been reported to

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stimulate more soil organic matter (SOM) decomposition (Parsons and Smith, 1989).

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The resulting available C leads to the promotion of copiotrophic taxa (i.e.

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Actinobacteria and Proteobacteria), which was consistent with other studies (Jiang et

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al., 2016; Xu et al., 2016). In this study, biochar might be utilized by microorganisms such as Actinobacteria, which could degrade recalcitrant polymers (Jones et al., 2011;

also

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Kirby, 2006), causing the shift of community composition. At genus level, biochar enhanced

the

growth

of

Sphingomonas,

which

is

classified

in

alphaproteobacteria class and able to degrade refractory contaminants (Leys et al., 2005). In contrast, no obvious change was observed on the relative abundance of Acidobacteria, implying little impact of biochar on these bacteria. Acidobacteria prefers oligotrophic soils with low carbon availability (Mao et al., 2012). Moreover, the relative abundance of Nitrospira at genus level decreased slightly with biochar amendment, which is because that Nitrospira prefers to live under oligotrophic conditions (Shen et al., 2013). 13

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3.4 Evolution of fungal community in soils All samples were roughly clustered into four regions in the PLS-DA plot (Fig. S4), suggesting the alteration of fungal community induced by biochar or tetracycline. It is in agreement with other studies (Jechalke et al., 2014; Yao et al., 2017). The effect of biochar on fungal community was first evaluated at the phylum level. As

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shown in Fig. 6a, the addition of biochar decreased the relative abundance of Ascomycota from 40.4% in CK group to 26.6% in BC group, which was attributed to

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the low pH in soil with biochar. Soil pH has been considered as a key factor on soil

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fungal community (Zhang et al., 2016). In this study, soil pH decreased from 6.72 ±

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0.05 to 4.22 ± 0.51 in BC group and 4.38 ± 0.07 in BT group with the addition of

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biochar (Table 2), which was due to the acidic property of biochar (Lehmann et al.,

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2011). Then, the pH increased to 5.58 ± 0.04 in BC group and 5.47 ± 0.16 in BT group after 30-day incubation, which was similar to another study (Xu et al., 2014). It

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has been reported that higher pH (7.29 ± 0.02) was beneficial to the growth of Ascomycota (Li et al., 2019). The genus Staurothele from Ascomycota was also adapted to neutral to alkaline environments (Gueidan et al., 2014). These could explain the decreased relative abundance of Ascomycota in BC and BT group. Moreover, the relative abundance of Basidiomycota slightly increased from 6.5% in CK group to 8.0% in BC group and 7.5% in BT group. Similarly, biochar also slightly increased the relative abundance of Basidiomycota in a forest soil conducted for 96 days (Hu et al., 2014). Basidiomycota could degrade organic materials such as lignin, hemicellulose and cellulose (Nie et al., 2018), and thus the carbon-rich conditions 14

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induced by biochar promoted its growth in BC and BT group in this study. At genus level, Trinema in groups with biochar was detected three times more than that in CK group. The amount of Trinema is generally dependent on the moisture content of the soil (Wilkinson and Mitchell, 2010), and the presence of biochar could promote the migration of rainwater to deeper soil in this work. Tausonia, a genus of Basidiomycota,

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was also observed at higher abundance in BC and BT group due to the carbon-rich conditions induced by biochar (Nie et al., 2018). Moreover, the genus of Mortierella,

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saprotrophs in soil, was also enhanced by biochar amendment, implying that biochar

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could provide a better survival habitat for Mortierella (Liu et al., 2015; Zheng et al.,

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2016).

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Tetracycline exhibited no significant effect on fungal diversity (Fig. S5) but it

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obviously changed the composition of fungal community in soil (Fig. 6). The most frequent fungal phyla such as Ascomycota, Nematoda and Basidiomycota were found

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less abundant in TC group. Several studies have also highlighted the adverse impact of veterinary antibiotics on soil microbial biomass (Jechalke et al., 2014; Kong et al., 2006). However, there were some fungal genus with higher abundant in TC group than others, such as Thermothelomyces, a kind of fungi able to degrade aromatic compounds (Périgon et al., 2019). Hence, the presence of TC could also enrich the microorganism which can degrade antibiotics. Compared to TC group, fungal community was less impacted in BT group, and the abundance of some dominant phyla recovered to normal levels with Basidiomycota at 7.5% and Nematoda at 6.3%, which was attributed to the combined effect of TC and biochar. Liu et al. (2014) also 15

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found that the diversity of soil microbial community stimulated by chlortetracycline could be partially counteracted by organic matter. Besides, TC would be partly adsorbed by biochar due to its large surface area (Quilliam et al., 2013). Therefore, biochar could adsorb some quantities of tetracycline, which limited the contact between TC and microorganisms, decreasing its bioavailability and mitigating its

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effect on fungal community in soil. 4. Conclusion

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This study evaluated the effect of biochar on the migration of tetracycline in soil

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and their combined impacts on microbiome. TC was mainly dissipated in soil via

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adsorption with little fraction being photolyzed or biodegraded. Moreover, the vertical

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migration of TC was accelerated in the presence of biochar, mainly due to the

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decreased soil bulk density and lower adsorption capacity of TC on biochar. Biochar could stimulate available C to promote the growth of copiotrophic bacteria and thus

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significantly change the composition of bacterial community in soil, whereas TC played a minor role due to its rapid dissipation in soil. Besides, biochar changed the composition of fungal community due to its acidic impact on soil and partially counteracted the role of TC through limiting its contact with microbiome. Acknowledgement The research was supported by the China Major Science and Technology Program for Water Pollution Control and Treatment (2017ZX07101003) and National Natural Science Foundation of China (21676161 and 21808127). The authors wish to thank Project funded by China Postdoctoral Science Foundation (2019M652389) and 16

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Young Scholars Program of Shandong University.

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Author contributions: Hua-Yu Liu: Conceptualization, Methodology, Validation, Investigation, Formal analysis, Writing - Original Draft, Writing – Review & Editing Chao Song: Conceptualization, Investigation, Writing - Original Draft, Writing – Review & Editing Shan Zhao: Investigation, Formal analysis, Writing – Review & Editing

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Shu-Guang Wang: Conceptualization, Supervision, Project administration, Writing –

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Review & Editing

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√ The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

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may be considered as potential competing interests:

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☐The authors declare the following financial interests/personal relationships which

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Journal Pre-proof Figure Caption Fig. 1 Schematic design of the soil column experiments among different groups. CK: control group; BC: biochar group; TC: tetracycline group; BT: biochar and tetracycline group. Fig. 2 Dynamic dissipation and removal rate of TC under different conditions during 30 days incubation. Error bars represent the mean±standard deviations (n=3).

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Fig. 3 Acute vertical migration of TC from Layer 1 to Layer 6 in soils with/without biochar. Error bars represent the mean±standard deviations (n=3). CK: control

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group; BC: biochar group.

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Fig. 4 Bacterial community changes: Partial least squares Discriminant (PLS-DA)

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analysis of bacterial community at OTUs level. CK: control group; BC: biochar

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group; TC: tetracycline group; BT: biochar and tetracycline group. Fig. 5 Distribution of partial sequences of bacterial 16S rRNA genes from different

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groups at the phylum (a) and genus (b) level on day 90. Phylogenetic groups accounting for 1% or less of the classified sequences in all samples were

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summarized in the group “others”. CK: control group; BC: biochar group; TC: tetracycline group; BT: biochar and tetracycline group. Fig. 6 Distribution of partial sequences of fungal 18S rRNA genes from different groups at the phylum (a) and genus (b) level on day 90. Phylogenetic groups accounting for 1% or less of the classified sequences in all samples were summarized in the group “others”. CK: control group; BC: biochar group; TC: tetracycline group; BT: biochar and tetracycline group.

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Table. 1 First-order kinetic parameters for the dissipation of tetracycline in soils among control, dark, sterilized and dark-sterilized groups. Treatments

k (mg kg-1d-1) a

R2

t1/2 (days) b

Control Dark Sterilized Dark-sterilized

0.05484 0.05064 0.04318 0.04594

0.992 0.991 0.986 0.993

13 14 16 15

k, rate constant of the first-order reaction kinetics; R2, coefficient of determination. b t1/2, half-life of dissipation (ln 2/k).

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Journal Pre-proof Table. 2 pH values of soils in different groups during 30 days incubation. CK BC TC BT

Day 1

Day 10

Day 20

Day 30

6.72±0.05a 4.22±0.51b 6.57±0.08a 4.38±0.07b

6.12±0.21a 4.88±0.25b 6.54±0.05a 5.21±0.21b

6.90±0.03a 5.08±0.05b 6.80±0.04a 5.04±0.17b

7.11±0.20a 5.58±0.04b 7.09±0.04a 5.47±0.16b

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CK: control group, BC: biochar group, TC: tetracycline group, BT: biochar and tetracycline group. Different letters in a row demonstrate significant difference among the treatments at p<0.05.

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Highlights 

Adsorption is the dominant dissipation way for tetracycline in soils.

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Biochar promoted vertical migration of tetracycline.

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Biochar and tetracycline greatly shifted bacterial and fungal community

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composition in 90 days.

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Figure 1

Figure 2

Figure 3

Figure 4

Figure 5

Figure 6