Microplastics combined with tetracycline in soils facilitate the formation of antibiotic resistance in the Enchytraeus crypticus microbiome☆
Graphical abstract
Introduction
Microplastics have increasingly becoming appreciated as an environmental pollutant in world since the concept was proposed in 2004 (Rillig and Ingraffia, 2017; Thompson et al., 2004). Studies have primarily focused on the pollution and eco-toxicological effects of microplastics in aquatic ecosystems (Peiponen et al., 2019; Wang et al., 2019). In the past few years, the pollution and risks of microplastics in terrestrial ecosystems have received increasing attention (da Costa et al., 2019; Duis and Coors, 2016). In terrestrial ecosystems, microplastics may be an emerging threat through influence on soil biota at different trophic levels (He et al., 2018; Machado et al., 2018). Evidence shows that microplastics can act as a vector for the transfer of pollutants including plastic additives and other pollutants (such as heavy metals and persistent organic pollutants) absorbed from soil matrices, and influence the transport of pollutants, thus posing potential risks for soil biota (He et al., 2018; Hodson et al., 2017; Kwon et al., 2017; Zhang et al., 2018a). The coexistence of microplastics and pollutants inevitably induces interactions between them, and can affect the toxicity of individual pollutants (Seidensticker et al., 2019; Zhang et al., 2018b). A small number of studies have demonstrated that hydrophobic organic pollutants can affect the eco-toxicity of microplastics (Duis and Coors, 2016). For example, the adsorption capacity of triclosan on microplastics can directly affect the toxicity of different microplastics (Zhu et al., 2019b). A previous study showed that microplastics can enhance the effects of an organo-phosphorous flame retardant on mice to induce oxidative stress, neurotoxicity, amino acid metabolism and energy metabolism (Deng et al., 2018). Accordingly, more attention should be paid to the effects of combined pollution from microplastics and other pollutants on terrestrial organisms.
Polyamide (PA) and polyvinyl chloride (PVC) are two types of most common microplastics in the environment. PA (the basis of nylon) has been widely used in domesticity and industry, such as clothes, belts and automobile tires, and has been widely detected in field studies (de Sa et al., 2018; Khosrovyan and Kahru, 2020). However, it has received the rarest attention. As a synthetic resin made by polymerization of vinyl chloride monomer, PVC is the third-most widely-produced plastic in the world, after polyethylene and polypropylene (Tang et al., 2018b). It is estimated that approximately 50.27 million tons will be consumed globally in 2020 (Tang et al., 2018b). PVC plays a crucial role in our daily lives such as building, transport, packaging, electrical and healthcare applications, as a result, many PVC products or debris are discharged into environment during their utilization (Tang et al., 2018b). In addition, previous studies have verified that PA and PVC can effectively adsorb antibiotics, especially for tetracycline (Li et al., 2018; Xu et al., 2018).
In order to improve growth and prevent diseases in production animals, veterinary antibiotics have been widely used in conventional animal husbandry over the past few decades (Dibner and Richards, 2005). However, approximately 30–90% of the antibiotics cannot be assimilated by animals, and enter the environment via feces or urine (Fang et al., 2015). Tetracycline (TC, C22H24N2O8), as a kind of polar, ionizable and broad-spectrum antibiotic, it is widely applied in livestock farming to prevent and treat diseases, but most of it is not absorbed by farm animals’ bodies and therefore large amount are excreted into the environment (Sarmah et al., 2006; Xu et al., 2018). Consequently, tetracycline residues have been widely detected in water and soil, where microplastics co-occur (Xu et al., 2018). In addition, various antibiotic residues (such as tetracycline) induce bacteria to develop resistance, and genes inside the resistant bacterial cells become antibiotic resistance genes (ARGs) (Wright, 2007). ARGs are recognized as one of the most important emerging contaminants of the 21st century, posing a severe threat to public health (Chen et al., 2019; Pruden et al., 2006; Sanderson et al., 2016). Tetracycline in the environment can be adsorbed by microplastics (Li et al., 2018; Xu et al., 2018). Although a small number of studies have focused on the eco-toxicity of combined pollution from microplastics and antibiotics (Fonte et al., 2016; Zhang et al., 2019a; Zhang et al., 2019c), these studies have not targeted terrestrial systems. Recent evidence has shown that microplastics can act as a vector for the spread of ARGs in the marine environment (Lagana et al., 2019; Yang et al., 2019). However, it is not clear whether the combined pollution from microplastics and antibiotics can affect the incidence and pattern of ARGs in the terrestrial environment, especially in soil fauna.
Soil fauna is an important part of soil ecosystem, harboring various soil microorganisms (van den Hoogen et al., 2019). As a widespread soil invertebrate, Enchytraeus crypticus (E. crypticus, Oligochaeta, Enchytraediae) plays an important role in maintaining soil quality through mineralization, decomposition and redistribution of soil organic matter (Castro-Ferreira et al., 2012). Due to its higher sensitivity to pollutants and broader tolerance to soil environments, E. crypticus has been widely used as a model species to investigate the toxicity of pollutants such as heavy metals, nanoparticles, and organic pollutants in soil environments (Arp et al., 2014; Castro-Ferreira et al., 2012; Gomes et al., 2015; He et al., 2017). Previous studies have paid attention to the microbiome and ARGs in E. crypticus (Ma et al., 2019a; Ma et al., 2019b; Zhang et al., 2019b; Zhu et al., 2018a). However, it is still not clear how and why the combination of microplastic and antibiotic pollutants in soils induce species-specific responses in the microbiome and the incidence of ARGs in E. crypticus.
In the present study, we chose TC, and PA and PVC to construct a microcosm simulating the combined soil pollution from antibiotics and microplastics. Our aim was to exam the effects of the combined pollutants on the microbiome and ARGs in the non-target organism E. crypticus. We hypothesis that, 1) Single microplastics and their combination with TC have different effects on the alteration of the microbial community in E. crypticus; 2) The effects on ARGs abundance in E. crypticus from combinations of microplastics and TC are greater than for single microplastic treatments. The study aimed to further extend our knowledge of the combined effects of microplastics and antibiotics on the ARGs and microbiome of soil invertebrates.
Section snippets
Soil, microplastics, tetracycline and test organisms
Approximately 5 kg soil used in this study was sampled from the top layer (0–20 cm) of a vegetable field located in Xiamen city, Fujian province, China (24°38′ N, 118°01′E). Soil had a saturated water holding capacity of 16.56 g water per 100 g moisture-saturated soil. Soil had a pH of 6.55, and electric conductivity (EC) of 127.6 μs cm−1. Soil total nitrogen, carbon, sulfur accounted for 0.10%, 1.23% and 0.04%, respectively. Soil was air-dried, ground, and sieved using a plastic soil sieve
Mortality, growth, reproduction, and TC concentration in E. crypticus
After single or combined exposure to PVC, PA and TC for 21 days, the mortality of E. crypticus was below 5% in all microcosms. The body weights of individuals in the TC, PVC, and their combined exposure treatments were significantly higher than in the Control (Fig. S2, ANOVA, P < 0.05). Reproduction in the TC, PVC, and their combined exposure treatments were significantly lower than in the Control (Fig. S2, ANOVA, P < 0.05). However, PA treatment was significantly higher than in the Control,
Bioaccumulation of TC in E. crypticus
The accumulation of TC in E. crypticus was detected after either exposure to TC alone or when TC was combined with PA or PVC in soils. Many types of microplastics including PA and PVC can effectively adsorb various antibiotics including but not limited to TC (Li et al., 2018; Wang et al., 2018; Xu et al., 2018). Although the adsorption capacities of PA and PVC for TC are generally different (Li et al., 2018), the difference in accumulation of TC by PA+TC and PVC+TC treated E. crypticus showed
Conclusions
Microplastics and TC exposure significantly disturbed the microbial community of the E. crypticus microbiome, single microplastics and their combination with TC have different effects on the alteration of the microbial community in E. crypticus. Microplastics and TC exposure significantly decreased the microbial alpha diversity of the E. crypticus microbiome, while no significant toxic synergies on the diversity of E. crypticus microbiome between tetracycline and microplastics in soil
CRediT authorship contribution statement
Jun Ma: Conceptualization, Methodology, Writing - original draft. G. Daniel Sheng: Conceptualization, Data curation, Writing - review & editing. Patrick O’Connor: Writing - review & editing.
Declaration of competing interest
The authors declare no competing financial interest.
Acknowledgements
This research was funded by the National Key Research and Development Program of China (2017YFE0107300), and the National Natural Science Foundation of China (21437004).
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This paper has been recommended for acceptance by Eddy Y. Zeng.