4.2.1. Pilot-Scale

A research group from China studied applying CWs to remove veterinary antibiotics and antibiotic resistant genes from swine wastewater for many years. In their studies, various CW types and their combinations as well as substrate medium and target contamination compounds were investigated. Their 2013 study results found that SSVF-CWs could efficiently remove target antibiotics and ARGs (68–95% and 50–90%, respectively) with the major mechanism being the physical sorption towards the wetland medium [38]. In another study, the results showed that removal performance ranged from high to low in the order of SSVF-Low water level > SSVF-High water level > SSHF > SF (based on average removal rates) indicating that the various design, flow path and water level led to different antibiotic removal rates through impacting the parameters, such as temperature, oxygen transfer, oxidation-reduction potential, sorption sites, etc. [89]. Another key finding from this study was that the seasonality might pose different impacts on different veterinary antibiotics (significant effect on sulfamethazine (SMZ) while no significant effect on TC) and different CW types (significant effect on SF while no significant effect on SSVF) [89]. Another long-term study indicated that high removal rates, ranging from 69.0% to 99.9%, were achieved for the target contaminants in all three treatments with different initial concentrations [86]. In another short-term study, flow direction showed no significant influence since they obtained comparable removal rates, but accumulation of antibiotics and ARGs in the surface soil was observed in down-flow treatments indicating a concern to the local environment due to likeliness of antibiotics enrichment and ARGs abundance [87].

Besides antibiotics, studies have also been conducted on removal of herbicides via application of CW systems, as Gikas et al. demonstrated up to 74% removal of terbuthylazine [92] and 60% removal of S-metolachlor [93] in horizontal subsurface flow CWs. Other researchers also investigated the removal performances of hybrid, SSHF and SF CW systems using various substrate and vegetations [62,63,88]. In a 2019 study, different combinations of CW units (SSHF-SSVF (up-flow); SSHF-SSVF (down-flow); SSFV (down-flow)-SSVF (up-flow)) were run for 84 days to treat antibiotics, ARGs and nutrients from goose wastewater. The researchers reported that the comparable antibiotic removal performance of different combinations of hybrid CWs was probably due to the highly spiked-up influent concentrations (2500 μg/L for tilmicosin (TMS) and 30 μg/L for doxycycline (DOC)), which likely concealed the differences on effluent concentrations among different treatments [63]. This may indicate the importance of conducting full-scale field studies receiving much lower antibiotic concentrations to simulate the real-world scenario, instead of pursuing the high removal efficiency results by dosing up the influent water

to an unrealistic level. Besides livestock and poultry, CW has also been applied to treat wastewater from aquaculture. For example, Huang et al. conducted a study using SSHF with different vegetations (single or mixture of *Iris pseudacorus* and *Phragmites australis*) to remove ENR, SMZ and AGRs from wastewater of a local fish farm achieving up to 80% removal performance [88].
