Antibiotics in Wastewater Treatment Plants in Tangshan: Perspectives on Temporal Variation, Residents’ Use and Ecological Risk Assessment

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

Antibiotics in Waste Water Treatment Plants in Tangshan: Per-1 spective on temporal variation, residents' use and ecological 2 risk assessment

Journal

Water (ISSN 2073-4441)

Manuscript ID:  water-3009836

Comments

In the manuscript submitted by Dong and co-workers, various type of antibiotics in the influent and effluent of Waste Water Treatment Plants (WWTPs) in the urban and suburban of Tangshan was reported. For each investigated antibiotic, their risk quotient (RQ) values were calculated (spring, summer, autumn and winter). Based on the risk quotient (RQ) ecological risk of each antibiotics was evaluated and reported. In my opinion, the topic addressed is interesting and important. In conclusion, the manuscript needs minor amendments to be accepted for publication in Journal “Water”.

The following suggestion must be added before the publication of article in Journal “Water”

1)     Rewrite line no 67-69 for better understanding

2)     Provide chronic toxicity data (ChV) of antimicrobial drugs (i.e. for this study) in the text.

3)     Table S1: change to Table 1. Accordingly change Table no. in manuscript.

4)     The quality of the discussion section must be improved. In so doing, the authors must organize the discussion from the general to the specific and link their findings to the literature.

 

5)     Limitations of the study must be presented in the conclusion section.

Comments on the Quality of English Language

Minor English grammar editing required 

Author Response

Reviewers' comments:

Reviewer #1

1) Rewrite line no 67-69 for better understanding

Thank you for your suggestion. We have reflected this comment in line 99-102 by “Seasonal sampling campaigns were conducted in 2023 [January to March (winter), April to May (spring), July to August (summer), and October (autumn) ] in Tangshan. Samples were collected from influent and effluent of WWTPs in the urban (n=2) and suburban (n=2).” 

 

2) Provide chronic toxicity data (ChV) of antimicrobial drugs (i.e. for this study) in the text.

We have reflected this comment in line 150-153 by “In this study, the ChV values for roxithromycin, tetracycline, chlortetracycline, oxytetracycline, ciprofloxacin, norfloxacin, ofloxacin, sulfadiazine, and sulfamethoxazole were 0.6, 20, 20, 20, 116, 114, 116, 0.101, and 0.068 mg/L, respectively.”

 

3) Table S1: change to Table 1. Accordingly change Table no. in manuscript.

Thank you for your suggestion. We have reflected this comment in the manuscript.

 

4) The quality of the discussion section must be improved. In so doing, the authors must organize the discussion from the general to the specific and link their findings to the literature.

Thank you for your suggestion.

We have supplied the content in line 226-241 by “In the influent of WWTPst, (1) the concentration ranges of the nine antibiotics selected in this study, except for Aureomycin, Oxytetracycline, and Ciprofloxacin, are much lower than those in Beijing (2018) in winter [32]; (2) the concentration of tetracycline antibiotics in summer is lower than existing research data, while the concentration of Aureomycin and Oxytetracycline in summer is higher than existing research data (except for Jiulongjiang River Basin); the concentration of Ciprofloxacin in quinolone antibiotics in summer and winter is higher than existing research data (except for Urumqi and Shihezi). In summer, the concentration of norfloxacin surpasses that of the Jiulongjiang River Basin, yet remains lower than that of Urumqi and Shihezi at its peak. In summer, the concentration of ofloxacin is lower than that of the Jiulongjiang River Basin, yet surpasses that of Yibin, Urumqi, and Shihezi[33-35]. (3) Among the sulfa antibiotics, sulfadiazine's concentration in summer and autumn surpasses that of the Jiulongjiang River Basin, yet remains lower than that of Urumqi and Shihezi. The concentration of sulfamethoxazole in summer exceeds that of the Jiulongjiang River Basin, though its peak value is lower than that observed in Beijing (2019) [34-36](Table S2).” 

We have supplied the content in line 293-309 by “ In the effluent of WWTPs, (1) the nine antibiotics selected in this study (excluding Tetracycline, Aureomycin, Oxytetracycline, and Ciprofloxacin) exhibited lower winter concentration ranges in comparison to Beijing (2018)[32]; (2) Roxithromycin concentrations in summer were lower than those in the Zijiang River Basin of central Hunan, while in spring and autumn, they were an order of magnitude higher than those in Shenyang[32,35,42,43]; (3) Quinolone and tetracycline concentrations in spring and autumn were an order of magnitude higher than those in Shenyang, while in summer, they were an order of magnitude lower than those in Yibin; (4) Sulfa antibiotic concentrations in summer and autumn were higher than those reported by Beijing (2019) and Shenyang, and in summer, they were an order of magnitude higher than those in Yibin [32,35,42,43] (Table S3). The variation in the spatial and temporal distribution of antibiotics concentrations is significant. The variation in the spatial and temporal distribution of antimicrobial drug concentrations is significant. This variation is primarily attributed to a complex interplay of factors, including the treatment processes and surface temperatures employed by sewage treatment plants across different regions, the sources and composition of sewage within the service area, and the size of the population served by these WWTPs.”

 

5) Limitations of the study must be presented in the conclusion section.

We have incorporated your comments in line 382-399 by “This study focuses solely on the temporal distribution of target antibiotics in the influent and effluent of WWTPs. Certain antibiotics are susceptible to hydrolysis and removal in aquatic environments. Research indicates that macrolide antibiotics are prone to hydrolysis. Tetracycline antibiotics are not stable in water; for instance, the hydrolysis rate of oxytetracycline increases with deviations from neutral pH (pH =7) and rising temperatures, whereas sulfonamides and fluoroquinolones are resistant to hydrolysis. pH and temperature are significant factors influencing hydrolysis. Consequently, it is imperative to further investigate the impact of seasonal variations in pH and temperature at the ends of the drainage systems and within the treatment units of WWTPs on the temporal distribution of antibiotics, to elucidate the driving factors behind any temporal trends observed in these agents' distribution in the influent and effluent.Moreover, the adsorption of antibiotics by sewage plant sludge is a significant factor in enhancing the removal rate of these antibiotics. For antibiotics primarily removed through sludge adsorption (such as fluoroquinolones and sulfonamides), an extension in sludge retention time concurrently enhances their removal efficiency. However, the removal of certain antibiotics may not be impacted by sludge retention time. Hence, there is an urgent need for a comprehensive assessment of the physical adsorption and biodegradation of antibiotics in sewage plants.”

 

Comments on the Quality of English Language

Minor English grammar editing required

We agree with you and have incorporated this suggestion throughout the manuscript.

 

 

Author Response File: Author Response.pdf

Reviewer 2 Report

Comments and Suggestions for Authors

It is an interesting work that assesses the temporal variation of four types of nine antibiotics in the influent and effluent of urban and suburban-WWTPs, and estimates the usage and annual discharge of antibiotics in the urban and suburban based on per capita pollution load.

 

1. Introduction: L. 40-46.

It remains to be said that there are technologies installed in the WWTP that remove antibiotics (e.g., chemical precipitation, chemical oxidation, UF, NF and IO), but these are perhaps only applied in the treatment of industrial effluents and are not common in the treatment of suburban effluents, where they prevail. biological treatments.

 

2. Section 2.1. Experimental reagents and instruments.

It remains to be said what led to the selection of antibiotics. The prevalence of these molecules in urban wastewater in this region? In the majority use of these antibiotics by the population.

 

3. Section 3.2. Effluent

Information is missing on how WWTP could be technologically improved to remove antibiotic molecules and what costs they could add to the operation. it would have to be a pragmatic and realistic analysis., but important to evaluate how the problem could be solved or its impact could be minimized.

This section lacks comparison with other similar studies. Make a comparison with other studies.

 

4. Section 3.4. Estimation of usage and sewage discharge

This section lacks comparison with other similar studies. Make a comparison with other studies.

 

5. Paragraphs too long

Paragraphs are too long in several sections of the text. They should be smaller, for example L. 181-225, L. 232-257, and L. 259-282.

Comments on the Quality of English Language

None

Author Response

Reviewers' comments:

Reviewer #2

It is an interesting work that assesses the temporal variation of four types of nine antibiotics in the influent and effluent of urban and suburban-WWTPs, and estimates the usage and annual discharge of antibiotics in the urban and suburban based on per capita pollution load.

 

1. Introduction: L. 40-46.

It remains to be said that there are technologies installed in the WWTP that remove antibiotics (e.g., chemical precipitation, chemical oxidation, UF, NF and IO), but these are perhaps only applied in the treatment of industrial effluents and are not common in the treatment of suburban effluents, where they prevail biological treatments.

We agree with you and have supplied the content in line 67-73 by “It should be noted that sewage treatment technologies like advanced chemical oxidation, chemical precipitation, ultrafiltration, nanofiltration, and ion exchange are not effective at removing antibiotics and may be better suited for industrial wastewater treatment. In contrast, suburban wastewater treatment primarily relies on biological processes, with advanced oxidation and other sewage treatment techniques being less common, resulting in a lower capacity for treating antibiotics in wastewater.”

 

2. Section 2.1. Experimental reagents and instruments.

Thank you for your suggestion. We have supplied the content in line 87-91 by “Ultra-high purity compounds (>99%) of nine antibiotics, including roxithromycin (macrolides), ofloxacin (quinolones), norfloxacin(quinolones), ciprofloxacin (quinolones), tetracycline (tetracyclines), chlortetracycline (tetracyclines), oxytetracycline (tetracyclines), sulfadiazine(sulfonamides) and sulfamethoxazole (sulfonamides), were bought from Sigma-Aldrich (St. Louis, MO, USA).”

 

Thank you for your suggestion. We have supplied the content in line 121-126 by“

The pretreated samples were analyzed using UPLC-MS/MS (QTRAPTM5500 LC/MS/MS system, SCIEX, USA), employing a Waters Cortecs T3 column (2.1 mm × 100 mm, 2.7 μm). The injection volume for liquid chromatography was 2 μL, with a flow rate of 0.3 mL·min-1 and a column temperature of 40 °C. The mobile phase was a gradient elution of 0.1% formic acid aqueous solution and acetonitrile.” 

 

It remains to be said what led to the selection of antibiotics. The prevalence of these molecules in urban wastewater in this region? In the majority use of these antibiotics by the population.

We have reflected this comment in line 55-59 by“As veterinary antibacterial drugs, tetracycline antibiotics are the most commonly used, accounting for 40.5% of the total, followed by sulfonamides and macrolides. Quinolones are highly used in hospitals due to the high incidence of respiratory tract infections and mycoplasmal pneumonia in spring, autumn, and winter.”

 

3. Section 3.2. Effluent

Information is missing on how WWTP could be technologically improved to remove antibiotic molecules and what costs they could add to the operation. it would have to be a pragmatic and realistic analysis., but important to evaluate how the problem could be solved or its impact could be minimized.

Thank you for your suggestion. We have reflected this comment in line 245-251 by “Currently, most WWTPs employ biological treatment processes to degrade organic matter, including antibacterial drugs. These processes primarily involve microorganisms in activated sludge attaching to the cell surface through adsorption and absorption. Different types of microorganisms utilize their metabolic capabilities to decompose and transform antibacterial drugs. Ultimately, these microorganisms break down the molecular structure of the antibacterial drugs into smaller organic compounds or CO₂ through enzyme production and oxidation, releasing corresponding metabolites.”

Given the multitude of antibiotics and their varying usage across regions, coupled with the diversity of urban sewage treatment processes, additional research in this area is still warranted. The objective of this study is to analyze temporal changes in the use of antibiotics in the influent and effluent of WWTPs, and to estimate the potential ecological risks associated with their consumption and discharge by residents. Improvements in sewage treatment processes are not within the scope of this study, but represent an important area of research.

 

This section lacks comparison with other similar studies. Make a comparison with other studies.

We agree with you and have supplied the content in line 226-241 by “In the influent of WWTPs, (1) the concentration ranges of the nine antibiotics selected in this study, except for Aureomycin, Oxytetracycline, and Ciprofloxacin, are much lower than those in Beijing (2018) in winter [32]; (2) the concentration of tetracycline antibiotics in summer is lower than existing research data, while the concentration of Aureomycin and Oxytetracycline in summer is higher than existing research data (except for Jiulongjiang River Basin); the concentration of Ciprofloxacin in quinolone antibiotics in summer and winter is higher than existing research data (except for Urumqi and Shihezi). In summer, the concentration of norfloxacin surpasses that of the Jiulongjiang River Basin, yet remains lower than that of Urumqi and Shihezi at its peak. In summer, the concentration of ofloxacin is lower than that of the Jiulongjiang River Basin, yet surpasses that of Yibin, Urumqi, and Shihezi[33-35]. (3) Among the sulfa antibiotics, sulfadiazine's concentration in summer and autumn surpasses that of the Jiulongjiang River Basin, yet remains lower than that of Urumqi and Shihezi. The concentration of sulfamethoxazole in summer exceeds that of the Jiulongjiang River Basin, though its peak value is lower than that observed in Beijing (2019) [34-36] (Table S2).” 

We have supplied the content in line 293-309 by “ In the effluent of WWTPs, (1) the nine antibiotics selected in this study (excluding Tetracycline, Aureomycin, Oxytetracycline, and Ciprofloxacin) exhibited lower winter concentration ranges in comparison to Beijing (2018)[32]; (2) Roxithromycin concentrations in summer were lower than those in the Zijiang River Basin of central Hunan, while in spring and autumn, they were an order of magnitude higher than those in Shenyang[32,35,42,43]; (3) Quinolone and tetracycline concentrations in spring and autumn were an order of magnitude higher than those in Shenyang, while in summer, they were an order of magnitude lower than those in Yibin; (4) Sulfa antibiotic concentrations in summer and autumn were higher than those reported by Beijing (2019) and Shenyang, and in summer, they were an order of magnitude higher than those in Yibin [32,35,42,43] (Table S3). The variation in the spatial and temporal distribution of antibiotics concentrations is significant. The variation in the spatial and temporal distribution of antibiotics concentrations is significant. This variation is primarily attributed to a complex interplay of factors, including the treatment processes and surface temperatures employed by WWTPs across different regions, the sources and composition of sewage within the service area, and the size of the population served by these WWTPs.”

 

4. Section 3.4. Estimation of usage and sewage discharge

This section lacks comparison with other similar studies. Make a comparison with other studies.

The per capita pollution load, annual usage, and annual emissions of antibiotics are determined based on the concentrations of these drugs in the influent and effluent of WWTPs, as well as the total population served by these plants. Upon reviewing the literature, we found that there is limited research that targets the same antibiotics as those in this study and that also includes the population of WWTP's service area. We understand that the reviewer's comments are aimed at improving the data mining process within this study, and to address this, we have included a section comparing the influent and effluent data of the antibiotics with historical research data.

 

 

5. Paragraphs too long

Paragraphs are too long in several sections of the text. They should be smaller, for example L. 181-225, L. 232-257, and L. 259-282.

Thank you for providing this insight. We have reflected this comment as follow:

Line 221-225: Despite the legitimate reasons for their use, the current standards for the dosage of various veterinary antibiotics are inconsistent and imprecise, leading to the potential overuse of these drugs in livestock farming. This in turn raises the concentration of antibiotics in the influent of wastewater treatment plants within the farming region.

Line 242-292: 3.2. Effluent

The removal effect of antibiotics in WWTPs in different seasons is closely related to treatment processes, operating parameters, influent properties, and types of antibiotics. Currently, most WWTPs employ biological treatment processes to degrade organic matter, including antibiotics. These processes primarily involve microorganisms in activated sludge attaching to the cell surface through adsorption and absorption. Different types of microorganisms utilize their metabolic capabilities to decompose and transform antibiotics. Ultimately, these microorganisms break down the molecular structure of the antibiotics into smaller organic compounds or CO2 through enzyme production and oxidation, releasing corresponding metabolites. The seasonal variation characteristics of the detected concentration of antibiotics in the effluent of both urban- and suburban-WWTPs are shown in Figure 1. The total antibiotics concentration in the effluent of WWTPs in the winter is the highest, followed by spring, autumn, and summer. In winter, the concentration of antibiotics detected in the wastewater from the urban-WWTPs is higher than that from the suburban-WWTPs, while the trend is reversed in the other three seasons.(Figure 1). Currently, the WWTPs in Tangshan mainly use the A2O method for sewage treatment. The A2O method (anaerobic-anoxic-aerobic process), is widely used in the sewage treatment system of northern China [37]. However, the northern of China experience lengthy cold seasons, making it challenging for small-scale sewage biochemical treatment processes to operate stably [38]. Low temperatures decrease the activity of nitrifying and denitrifying bacteria, leading to a decline in the nitrogen removal efficiency of the A2O system and challenges in its stable operation[39-40]. Previous studies have found that temperature has a significant impact on the nitrogen removal efficiency of the AAO system. Nitrification reactions occur at 20-30°C, and almost stop at temperatures below 5°C; denitrification reactions occur at 20-40°C, and rapidly decrease at temperatures below 15°C. The winter temperatures in Tangshan and surrounding areas are low, with average temperatures below 0°C from January to February, which is not conducive to the degradation of antibiotics by microorganisms in activated sludge. Therefore, due to the impact of low temperatures on the efficiency of A2O sewage treatment, the total concentration of antibiotics in the effluent of sewage treatment plants in winter is one order of magnitude higher than that in spring, autumn, and summer. (Figure 1).

In the effluent of both urban- and suburban-WWTPs, macrolide antibiotics- roxithromycin has the highest detection concentration in spring,while correspond to the quinolone antibiotics in autumn and winter (Table 3). In the effluent samples of suburban-WWTPs in summer, the concentration of sulfa antibiotics in the effluent is higher than that in the influent (Table 3)[41]. Previous studies have also observed a similar phenomenon, where the concentration of sulfa antibiotics in the effluent after treatment by activated sludge processes has increased. This phenomenon may be due to the following reasons: (1) Antibiotics adsorbed in the activated sludge are released into the water, resulting in an increase in the concentration of these drugs in the effluent from WWTPs. (2) During the A2O sewage treatment process, sulfa antibiotics are converted into other substances in the aerobic stage, and these substances are converted back into sulfa antibiotics in the anaerobic stage, resulting in an increase in the concentration of sulfa antibiotics in the effluent. In this study, the removal rates of nine antibiotics in the urban- and suburban-WWTPs increased from 8.18% and 7.30% in winter to 70.14% and 66.82% in spring, and to 79.58% and 73.91% in autumn. The removal rates of tetracyclines, quinolones, and sulfonamides in the urban-WWTPs were higher than those of macrolides in all four seasons, while the suburban-WWTPs only followed the same trend in spring, autumn, and winter.

Line 310-349:3.3 Ecological risk assessment

In the effluent of WWTPs in winter, the RQ values of roxithromycin, tetracycline, chlortetracycline, and oxytetracycline in urban were 0.22, 0.1 (0.09), 0.1 (0.08), and 0.23, while correspond to roxithromycin and oxytetracycline were 0.17 and 0.1 (0.09) in suburban.. This indicating that these macrolides and tetracyclines were medium-risk antibiotics. In other three seasons, the four categories of nine antibiotics had RQ values of ≤0.1 in the effluent of WWTPs and shown low-risk antibiotics. It is worth noting that macrolides, including roxithromycin, are medium-risk antibiotics in the effluent of urban- and suburban-WWTPs in winter, and there is a possibility of overuse of these drugs by residents in Tangshan. Chen et al. used RQs to assess the ecological risks of antimicrobial drugs. The results showed that erythromycin, roxithromycin, tetracycline, chlortetracycline, sulfamethoxazole, and norfloxacin were high-risk pollutants in water bodies in China, accounting for 20.9% [44]. In winter, various respiratory diseases, including mycoplasma pneumoniae, influenza, adenovirus, and respiratory syncytial virus infections, are highly prevalent. The peak season for mycoplasma pneumoniae infection occurs from August to February of the subsequent year, with the highest incidence around December to January of the following year [45]. Macrolide antibiotics, such as roxithromycin and clarithromycin, are stable to acid, have a long half-life (35-48 hours), a broad antibacterial spectrum, high bioavailability, are widely distributed in the body, and have significant efficacy, with minimal gastrointestinal irritation [46]. They have become the first choice for treating mycoplasma pneumoniae infection[47,48]. At present, the resistance rate of Mycoplasma pneumoniae to macrolides has been on the rise worldwide [49]. East Asia is the region with the most serious resistance to macrolide drugs for Mycoplasma pneumoniae in the world. Studies have shown that the resistance rate in some areas of China has reached over 90%.

3.4 Estimation of usage and sewage discharge

The per capita pollution load, annual usage, and annual emissions of antibiotics in Tangshan are presented in Table 4 and 5. The per capita pollution load of antibiotics was highest in spring, summer, and autumn for sulfamethoxazole, while the highest in winter for ofloxacin. From spring to winter, the per capita pollution load of antibiotics for urban residents is 9.63-13.74 times that of suburban residents, suggesting that urban residents may be at risk of antibiotics abuse (Table 4). In urban, the usage of roxithromycin (5.87 kg/a in spring), sulfamethoxazole (8.96kg/a in summer), sulfamethoxazole (5.77 kg/a in autumn) and ofloxacin (17.76kg/a in winter) significantly surpasses that of other antimicrobial agents. In contrast, the usage levels in suburban are as follows: sulfamethoxazole (0.17 kg/ain spring), norfloxacin (0.10 kg/ain summer), sulfamethoxazole (0.12 kg/ain autumn) and ofloxacin (0.32kg/ain winter) (Table 4). It should be noted that the usage of antibioticss in the urban was 1 orders of magnitude higher than that in the suburban (Table 5).

 

Comments on the Quality of English Language

None

 

 

Author Response File: Author Response.pdf

Round 2

Reviewer 2 Report

Comments and Suggestions for Authors

The authors greatly improve the quality of the manuscript.

Comments on the Quality of English Language

none

Author Response

1. The authors should provide under the Materials chapter a flow scheme of the wastewater treatment plant

Thank you for your suggestion. We have added the Figure 1 (Schematic diagram of Waste Water Treatment Plants (WWTPs) in Tangshan.) in the manuscript.

2. The references for all equations should be provided and the same stands for the significance and the units of measures for each of their terms. There is a problem with the definition of pollutions loads (generally calculated by the multiplication of flows and concentrations, this is a mass flux, expressed usually as g/s or kg/day). But the authors express many loads such as mass load of antibiotics in the effluent of WWTP (M, g/a), what represents "a"?, without being consistent. Normally the units of measure should be those of the international system

We have reflected this comment in line 159-169.

3. The authors should spell out acronyms on first mention, both in the abstract and the main body of the paper. Even if you have defined an acronym in the abstract, it has to be defined again in the paper when first mentioned. For example COD, A2O are not explained when they are first used.

Thank you for your suggestion. We have reflected this comment in the whole manuscript.

 

Author Response File: Author Response.pdf