Elsevier

Water Research

Volume 45, Issue 2, January 2011, Pages 681-693
Water Research

Release of antibiotic resistant bacteria and genes in the effluent and biosolids of five wastewater utilities in Michigan

https://doi.org/10.1016/j.watres.2010.08.033Get rights and content

Abstract

The purpose of this study was to quantify the occurrence and release of antibiotic resistant genes (ARGs) and antibiotic resistant bacteria (ARB) into the environment through the effluent and biosolids of different wastewater treatment utilities including an MBR (Membrane Biological Reactor) utility, conventional utilities (Activated Sludge, Oxidative Ditch and Rotatory Biological Contactors-RBCs) and multiple sludge treatment processes (Dewatering, Gravity Thickening, Anaerobic Digestion and Lime Stabilization). Samples of raw wastewater, pre- and post-disinfected effluents, and biosolids were monitored for tetracycline resistant genes (tetW and tetO) and sulfonamide resistant gene (Sul-I) and tetracycline and sulfonamide resistant bacteria. ARGs and ARB concentrations in the final effluent were found to be in the range of ND(non-detectable)-2.33 × 106 copies/100 mL and 5.00 × 102–6.10 × 105 CFU/100 mL respectively. Concentrations of ARGs (tetW and tetO) and 16s rRNA gene in the MBR effluent were observed to be 1–3 log less, compared to conventional treatment utilities. Significantly higher removals of ARGs and ARB were observed in the MBR facility (range of removal: 2.57–7.06 logs) compared to that in conventional treatment plants (range of removal: 2.37–4.56 logs) (p < 0.05). Disinfection (Chlorination and UV) processes did not contribute in significant reduction of ARGs and ARB (p > 0.05). In biosolids, ARGs and ARB concentrations were found to be in the range of 5.61 × 106–4.32 × 109 copies/g and 3.17 × 104–1.85 × 109 CFU/g, respectively. Significant differences (p < 0.05) were observed in concentrations of ARGs (except tetW) and ARB between the advanced biosolid treatment methods (i.e., anaerobic digestion and lime stabilization) and the conventional dewatering and gravity thickening methods.

Introduction

The escalating problem of emergence of antibiotic resistant bacteria and their resistant genes is becoming a major global health issue (Levy, 2002, Chee-Sanford et al., 2001). The use of numerous antimicrobial agents as treatments in animal, human, and plant health maintenance, is a worldwide practice providing both desirable and undesirable consequences. Links have been found to exist between antibiotic use and the emergence of antibiotic resistant bacterial pathogens (Aminov et al., 2001, Levy, 2002, Peak et al., 2007, Séveno et al., 2002). Studies have proven increase in antibiotic resistance strains that belong to pathogenic bacteria (Blasco et al., 2008) and over the years, nearly every bacterial pathogen has developed resistance to one or more clinical antibiotics (Todar, 2008).

The general observation published in different studies is that the environmental compartments which are most directly impacted by human or agricultural activities showed higher concentrations of antibiotic resistant bacteria and antibiotic resistant genes (Pruden et al., 2006, Chee-Sanford et al., 2001). Large amounts of antibiotics are released into municipal wastewater due to incomplete metabolism in humans or due to disposal of unused antibiotics (Nagulapally et al., 2009), which finally find their ways into different natural environmental compartments. Antibiotic resistant genes and antibiotic resistant bacteria have been detected in wastewater samples (Zhang et al., 2009a, Zhang et al., 2009b, Auerbach et al., 2007, Brooks et al., 2007, Pruden et al., 2006, Reinthaler et al., 2003). Also, the release of antibiotic resistant organisms through wastewater effluents into streams has been previously reported (Gallert et al., 2005, Iwane et al., 2001). Iwane and their colleagues reported approximately 8% and 6.7% of tetracycline resistant bacteria to be found in the pre- and post-chlorinated samples of a wastewater treatment plant respectively and then close to discharge location in the river water, similar percentages of bacteria were found to be resistant to tetracycline (Iwane et al., 2001). In addition, biosolids samples were reported to contain a high concentration of antibiotic resistant bacteria (Brooks et al., 2007). Also, the role of wastewater treatment plants in reducing the load of antibiotic resistant bacteria present in raw sewage is not well known (Rijal et al., 2009). However, it has been suggested that certain conditions within the wastewater treatment plants might increase the number of antibiotic resistant bacteria during the treatment process (Silva et al., 2006, Reinthaler et al., 2003). To the best of our knowledge, comparisons between different wastewater and biosolids treatment processes have not been studied so far.

The objective of this study was to quantify the release of antibiotic resistant genes (ARGs) and antibiotic resistant bacteria (ARB) in the effluent and biosolids of wastewater treatment plants (WWTPs). This is the first study that surveys the release of ARGs and ARB into the environment through the effluent and biosolids of different wastewater treatment utilities including an MBR (Membrane Biological Reactor), conventional wastewater utilities and multiple sludge treatment processes. This study has attempted to provide comparisons between different wastewater treatment processes and biosolid treatment processes along with the comparison of release loads of ARGs and ARB in the environment through the effluent and biosolids. In this study, samples of raw wastewater, effluent and biosolids were monitored for tetracycline and sulfonamide resistant bacteria, tetracycline resistant genes (tetW and tetO) and sulfonamide resistant gene (SulI) using quantitative polymerase chain reaction (qPCR) assays and conventional heterotrophic plate count methods. Tetracycline and sulfonamide resistance genes (tetW, tetO and SulI) were chosen in this study because tetracycline and sulfonamide are the most commonly used antibiotics in human and veterinary medicine (Boxall et al., 2003, Chopra and Roberts, 2001). In addition, quantitative detection systems already exist for this class of genes (Pei et al., 2006, Aminov et al., 2001). TetW and tetO genes are common in intestinal and rumen environments (Aminov et al., 2001) and have been cited as being promiscuous in their ability to spread among and across populations (Pei et al., 2006, Smith et al., 2004, Billington et al., 2002). SulI gene is also one of the most commonly detected sulfonamide resistant genes in the environment (Pei et al., 2006).

Section snippets

Sample collection

Samples of raw wastewater, effluent prior to disinfection, and final effluent after disinfection were collected from five different WWTPs located in Michigan (U.S.A.). Biosolid samples were also collected from the same treatment plants. Characteristics of the different WWTPs based on wastewater treatment processes, disinfection methods and sludge treatment methods are given in Table 1a, Table 1b. Two or three sampling events were conducted from each of these treatment plants starting from

Overall concentrations of ARGs and ARB in wastewater treatment plants

Concentrations of ARGs and ARB found in this study are presented in Table 3a, Table 3b respectively. Variations among different WWTPs in the raw influent concentration for different genes are expected because of different locations and related human activities. Also wastewater treatment plants receive inflow from a wide variety of sources beyond human population including industrial, hospital and animal waste.

Overall, the trends observed in concentration ranges at different sampling points from

Discussion

This study documents the occurrence of ARGs and ARB at different points in multiple conventional WWTPs and an MBR facility in Michigan. Table 3a, Table 3b illustrate reported ranges of ARGs and ARB presented in different published studies along with a summary of concentration ranges detected in this study. We observed that even though the concentrations of ARGs and ARB in raw wastewater are significantly reduced with wastewater treatment, high concentration is discharged into the effluent.

Conclusions

Wastewater utilities seem to be a potential sources of emerging tetracycline and sulfonamide resistant genes and bacteria in our environment. All raw influent, effluent and biosolid samples analysed in this study were found to contain high concentrations of tetracycline and sulfonamide resistant genes and bacteria. The concentration levels of ARGs and ARB in raw sewage were found be much higher than their respective concentrations in treated effluent. The concentrations of these resistant

Acknowledgements

We would like to thank the managers of all the wastewater treatment plants for providing the samples and information needed for this study. Also, we would like to acknowledge sampling assistance and help provided by Frederick J. Simmons and Arun Kumar.

References (30)

  • J.P. Brooks et al.

    Occurrence of antibiotic-resistant bacteria and endotoxin associated with the land application of Biosolids

    Can. J. Microbiol.

    (2007)
  • J.C. Chee-Sanford et al.

    Occurrence and diversity of tetracycline resistance genes in lagoons and groundwater underlying two swine production facilities

    Appl. Environ. Microbiol.

    (2001)
  • I. Chopra et al.

    Tetracycline antibiotics: mode of action, applications, molecular biology, and epidemiology of bacterial resistance

    Microbiol. Mol. Biol. Rev.

    (2001)
  • C. Gallert et al.

    Antibiotic resistance of bacteria in raw and biologically treated sewage and in groundwater below leaking sewers

    Appl. Microbiol. Biotechnol.

    (2005)
  • T. Iwane et al.

    Possible impact of treated wastewater discharge on incidence of antibiotic resistant bacteria in river water

    Water Sci. Technol.

    (2001)
  • Cited by (733)

    View all citing articles on Scopus
    View full text