Antimicrobial Resistance Gene Transfer Dynamics in Wastewater Treatment Systems
DOI:
https://doi.org/10.5281/zenodo.22687952Keywords:
antimicrobial resistance, horizontal gene transfer, wastewater treatment plants, mobile genetic elements, metagenomics, systematic reviewAbstract
Wastewater treatment plants receive antibiotic residues, resistant bacteria and mobile genetic elements from
hospitals, households, farms and industry in a single mixed stream, and the conditions inside a treatment train --
high cell density, sub-inhibitory antibiotic concentrations, and prolonged retention time -- are widely suspected to
favour horizontal transfer of resistance genes rather than simply diluting them away. We conducted a systematic
review of studies quantifying antimicrobial resistance gene abundance or horizontal gene transfer frequency
across sequential stages of conventional activated-sludge wastewater treatment, searching five databases from
January 2010 to March 2023 and identifying 1,842 unique records, of which 61 met pre-specified inclusion criteria
after full-text screening. Across the 61 included studies, absolute gene copy numbers per millilitre for the most
commonly reported resistance determinants (sul1, sul2, tetA, blaCTX-M and intI1, a class 1 integron-integrase
marker of anthropogenic gene-transfer potential) declined by a median of 1.6-2.3 log10 units between raw influent
and final effluent, driven mainly by the substantial reduction in total bacterial biomass achieved by secondary
treatment. Relative abundance, normalised to 16S rRNA gene copies, showed a markedly different and more
heterogeneous pattern: 34 of 61 studies reported no significant change or a net increase in relative resistance
gene abundance across the activated-sludge stage specifically, and conjugation-based transfer frequency,
measured directly in 14 of the 61 studies using filter-mating or fluorescence-based assays, was consistently one to
three orders of magnitude higher in activated-sludge mixed liquor than in raw influent or treated effluent. UV and
chlorine disinfection reduced culturable resistant bacteria substantially but had inconsistent and generally smaller
effects on extracellular and intracellular resistance gene DNA detectable by qPCR. We conclude that conventional
wastewater treatment achieves large absolute reductions in resistance gene load primarily through biomass
removal, while the activated-sludge process itself appears to be a zone of enhanced, rather than suppressed,
horizontal gene transfer potential, a distinction with direct implications for how treatment plant performance with
respect to antimicrobial resistance should be measured and reported.
References
Rizzo L, Manaia C, Merlin C, Schwartz T, Dagot C, Ploy MC, Michael I, Fatta-Kassinos D (2013) Urban wastewater treatment plants as hotspots for antibiotic resistant bacteria and genes spread into the environment. Science of the Total Environment 447:345-360. https://doi.org/10.1016/j.scitotenv.2013.01.032
Karkman A, Do TT, Walsh F, Virta MPJ (2018) Antibiotic-resistance genes in waste water. Trends in Microbiology 26(3):220-228. https://doi.org/10.1016/j.tim.2017.09.005
Manaia CM, Rocha J, Scaccia N, Marano R, Radu E, Biancullo F, Cerqueira F, Fortunato G, Iakovides IC, Zammit I (2018) Antibiotic resistance in wastewater treatment plants: tackling the black box. Environment International 115:312-324. https://doi.org/10.1016/j.envint.2018.03.044
Berendonk TU, Manaia CM, Merlin C, Fatta-Kassinos D, Cytryn E, Walsh F, Burgmann H, Sorum H, Norstrom M, Pons MN (2015) Tackling antibiotic resistance: the environmental framework. Nature Reviews Microbiology 13(5):310-317. https://doi.org/10.1038/nrmicro3439
Baquero F, Martinez JL, Canton R (2008) Antibiotics and antibiotic resistance in water environments. Current Opinion in Biotechnology 19(3):260-265. https://doi.org/10.1016/j.copbio.2008.05.006
Rizzo L, Sannino D, Vaiano V, Sacco O, Scarpa A, Pietrogiacomi D (2014) Effect of solar simulated N-doped TiO2 photocatalysis on the inactivation and antibiotic resistance of an E. coli strain in biologically treated urban wastewater. Applied Catalysis B 144:369-378. https://doi.org/10.1016/j.apcatb.2013.07.033
Zhang XX, Zhang T, Fang HHP (2009) Antibiotic resistance genes in water environment. Applied Microbiology and Biotechnology 82(3):397-414. https://doi.org/10.1007/s00253-008-1829-z
Munck C, Albertsen M, Telke A, Ellabaan M, Nielsen PH, Sommer MOA (2015) Limited dissemination of the wastewater treatment plant core resistome. Nature Communications 6:8452. https://doi.org/10.1038/ncomms9452
Alexander J, Hembach N, Schwartz T (2020) Evaluation of antibiotic resistance spread during wastewater treatment and chlorination disinfection. Scientific Reports 10:8952. https://doi.org/10.1038/s41598-020-65635-4
Barancheshme F, Munir M (2018) Strategies to combat antibiotic resistance in the wastewater treatment plants. Frontiers in Microbiology 8:2603. https://doi.org/10.3389/fmicb.2017.02603
Rice LB (2012) Mechanisms of resistance and clinical relevance of resistance to beta-lactams, glycopeptides, and fluoroquinolones. Mayo Clinic Proceedings 87(2):198-208. https://doi.org/10.1016/j.mayocp.2011.12.003
von Wintersdorff CJH, Penders J, van Niekerk JM, Mills ND, Majumder S, van Alphen LB, Savelkoul PHM, Wolffs PFG (2016) Dissemination of antimicrobial resistance in microbial ecosystems through horizontal gene transfer. Frontiers in Microbiology 7:173. https://doi.org/10.3389/fmicb.2016.00173
Beaber JW, Hochhut B, Waldor MK (2004) SOS response promotes horizontal dissemination of antibiotic resistance genes. Nature 427(6969):72-74. https://doi.org/10.1038/nature02241
Lu J, Wang Y, Li J, Mao L, Nguyen SH, Duarte T, Coin L, Bond P, Yuan Z, Guo J (2018) Triclosan at environmentally relevant concentrations promotes horizontal transfer of multidrug resistance genes within and across bacterial genera. Environment International 121:1217-1226. https://doi.org/10.1016/j.envint.2018.10.040
Lin H, Sun W, Zhang Z, Chapman SJ, Freitag TE, Fu J, Zhang X, Ma J (2016) Effects of manure and mineral fertilization strategies on soil antibiotic resistance gene levels and microbial community in a paddy-upland rotation system. Environmental Pollution 211:332-337. https://doi.org/10.1016/j.envpol.2016.01.007
Guo J, Li J, Chen H, Bond PL, Yuan Z (2017) Metagenomic analysis reveals wastewater treatment plants as hotspots of antibiotic resistance genes and mobile genetic elements. Water Research 123:468-478. https://doi.org/10.1016/j.watres.2017.07.002
Pruden A, Pei R, Storteboom H, Carlson KH (2006) Antibiotic resistance genes as emerging contaminants: studies in northern Colorado. Environmental Science & Technology 40(23):7445-7450. https://doi.org/10.1021/es060413l
Michael-Kordatou I, Michael C, Duan X, He X, Dionysiou DD, Mills MA, Fatta-Kassinos D (2015) Dissolved effluent organic matter: characteristics and potential implications in wastewater treatment and reuse applications. Water Research 77:213-248. https://doi.org/10.1016/j.watres.2015.03.011
Gillings MR (2014) Integrons: past, present, and future. Microbiology and Molecular Biology Reviews 78(2):257-277. https://doi.org/10.1128/MMBR.00056-13
Bengtsson-Palme J, Milakovic M, Svecova H, Ganjto M, Jonsson V, Grabic R, Udikovic-Kolic N (2019) Industrial wastewater treatment plant enriches antibiotic resistance genes and alters the structure of microbial communities. Water Research 162:437-445. https://doi.org/10.1016/j.watres.2019.06.073
Auerbach EA, Seyfried EE, McMahon KD (2007) Tetracycline resistance genes in activated sludge wastewater treatment plants. Water Research 41(5):1143-1151. https://doi.org/10.1016/j.watres.2006.11.045
LaPara TM, Burch TR, McNamara PJ, Tan DT, Yan M, Eichmiller JJ (2011) Tertiary-treated municipal wastewater is a significant point source of antibiotic resistance genes into Duluth-Superior Harbor. Environmental Science & Technology 45(22):9543-9549. https://doi.org/10.1021/es202775r
Page MJ, McKenzie JE, Bossuyt PM, Boutron I, Hoffmann TC, Mulrow CD, Shamseer L, Tetzlaff JM, Akl EA, Brennan SE (2021) The PRISMA 2020 statement: an updated guideline for reporting systematic reviews. BMJ 372:n71. https://doi.org/10.1136/bmj.n71
Sterne JA, Hernan MA, Reeves BC, Savovic J, Berkman ND, Viswanathan M, Henry D, Altman DG, Ansari MT, Boutron I (2016) ROBINS-I: a tool for assessing risk of bias in non-randomised studies of interventions. BMJ 355:i4919. https://doi.org/10.1136/bmj.i4919
Yuan QB, Guo MT, Yang J (2015) Fate of antibiotic resistant bacteria and genes during wastewater chlorination: implication for antibiotic resistance control. PLOS ONE 10(3):e0119403. https://doi.org/10.1371/journal.pone.0119403
Czekalski N, Diez EG, Burgmann H (2014) Wastewater as a point source of antibiotic-resistance genes in the sediment of a freshwater lake. ISME Journal 8(7):1381-1390. https://doi.org/10.1038/ismej.2014.8
Grenni P, Ancona V, Barra Caracciolo A (2018) Ecological effects of antibiotics on natural ecosystems: a review. Microchemical Journal 136:25-39. https://doi.org/10.1016/j.microc.2017.02.006
Forsberg KJ, Reyes A, Wang B, Selleck EM, Sommer MOA, Dantas G (2012) The shared antibiotic resistome of soil bacteria and human pathogens. Science 337(6098):1107-1111. https://doi.org/10.1126/science.1220761
Su JQ, Wei B, Ou-Yang WY, Huang FY, Zhao Y, Xu HJ, Zhu YG (2015) Antibiotic resistome and its association with bacterial communities during sewage sludge composting. Environmental Science & Technology 49(12):7356-7363. https://doi.org/10.1021/acs.est.5b01012
Zainab SM, Junaid M, Xu N, Malik RN (2020) Antibiotics and antibiotic resistant genes (ARGs) in groundwater: a global review on dissemination, sources, interactions, environmental and human health risks. Water Research 187:116455. https://doi.org/10.1016/j.watres.2020.116455
Bezuidenhout CC, Mthembu N, Puckree T, Lin J (2002) Microbiological evaluation of the Mhlathuze River, KwaZulu-Natal (RSA). Water SA 28(3):281-286. https://doi.org/10.4314/wsa.v28i3.4900
Martinez JL (2009) Environmental pollution by antibiotics and by antibiotic resistance determinants. Environmental Pollution 157(11):2893-2902. https://doi.org/10.1016/j.envpol.2009.05.051
Marti E, Variatza E, Balcazar JL (2014) The role of aquatic ecosystems as reservoirs of antibiotic resistance. Trends in Microbiology 22(1):36-41. https://doi.org/10.1016/j.tim.2013.11.001
Aminov RI (2011) Horizontal gene exchange in environmental microbiota. Frontiers in Microbiology 2:158. https://doi.org/10.3389/fmicb.2011.00158
Nesme J, Simonet P (2015) The soil resistome: a critical review on antibiotic resistance origins, ecology and dissemination potential in telluric bacteria. Environmental Microbiology 17(4):913-930. https://doi.org/10.1111/1462-2920.12631
Stalder T, Barraud O, Casellas M, Dagot C, Ploy MC (2012) Integron involvement in environmental spread of antibiotic resistance. Frontiers in Microbiology 3:119. https://doi.org/10.3389/fmicb.2012.00119
Amarasiri M, Sano D, Suzuki S (2020) Understanding human health risks caused by antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARG) in water environments. Critical Reviews in Environmental Science and Technology 50(19):2016-2059. https://doi.org/10.1080/10643389.2019.1692611
Wellington EMH, Boxall ABA, Cross P, Feil EJ, Gaze WH, Hawkey PM, Johnson-Rollings AS, Jones DL, Lee NM, Otten W (2013) The role of the natural environment in the emergence of antibiotic resistance in Gram-negative bacteria. The Lancet Infectious Diseases 13(2):155-165. https://doi.org/10.1016/S1473-3099(12)70317-1
Ashbolt NJ, Amezquita A, Backhaus T, Borriello P, Brandt KK, Collignon P, Coors A, Finley R, Gaze WH, Heberer T (2013) Human health risk assessment (HHRA) for environmental development and transfer of antibiotic resistance. Environmental Health Perspectives 121(9):993-1001. https://doi.org/10.1289/ehp.1206316
Bengtsson-Palme J, Kristiansson E, Larsson DGJ (2018) Environmental factors influencing the development and spread of antibiotic resistance. FEMS Microbiology Reviews 42(1):fux053. https://doi.org/10.1093/femsre/fux053
Uluseker C, Kaster KM, Thorsen K, Basiry D, Shobana S, Jain M, Kumar G, Kommedal R, Pala-Ozkok I (2021) A review on occurrence and spread of antibiotic resistance in wastewaters and in wastewater treatment plants: mechanisms and perspectives. Frontiers in Microbiology 12:717809. https://doi.org/10.3389/fmicb.2021.717809
Rocha J, Cacace D, Kampouris I, Guilloteau H, Jager T, Marano RBM, Karaolia P, Berendonk TU, Fatta-Kassinos D, Manaia CM (2020) Inter-laboratory calibration of quantitative analyses of antibiotic resistance genes. Journal of Environmental Chemical Engineering 8(1):102214. https://doi.org/10.1016/j.jece.2018.02.022
Christgen B, Yang Y, Ahammad SZ, Li B, Rodriquez DC, Zhang T, Graham DW (2015) Metagenomics shows that low-energy anaerobic-aerobic treatment reactors reduce antibiotic resistance gene levels from domestic wastewater. Environmental Science & Technology 49(4):2577-2584. https://doi.org/10.1021/es505521w
Bengtsson-Palme J, Boulund F, Fick J, Kristiansson E, Larsson DGJ (2014) Shotgun metagenomics reveals a wide array of antibiotic resistance genes and mobile elements in a polluted lake in India. Frontiers in Microbiology 5:648. https://doi.org/10.3389/fmicb.2014.00648
Fahrenfeld N, Ma Y, O'Brien M, Pruden A (2013) Reclaimed water as a reservoir of antibiotic resistance genes: distribution system and irrigation implications. Frontiers in Microbiology 4:130. https://doi.org/10.3389/fmicb.2013.00130
Baker-Austin C, Wright MS, Stepanauskas R, McArthur JV (2006) Co-selection of antibiotic and metal resistance. Trends in Microbiology 14(4):176-182. https://doi.org/10.1016/j.tim.2006.02.006
Chagas TP, Seki LM, Cury JC, Oliveira JA, Davila AM, Silva DM, Asensi MD (2011) Multiresistance, beta-lactamase-encoding genes and bacterial diversity in hospital wastewater in Rio de Janeiro, Brazil. Journal of Applied Microbiology 111(3):572-581. https://doi.org/10.1111/j.1365-2672.2011.05072.x
Rizzo L (2011) Bioassays as a tool for evaluating advanced oxidation processes in water and wastewater treatment. Water Research 45(15):4311-4340. https://doi.org/10.1016/j.watres.2011.05.035
Karkman A, Parnanen K, Larsson DGJ (2019) Fecal pollution can explain antibiotic resistance gene abundances in anthropogenically impacted environments. Nature Communications 10:80. https://doi.org/10.1038/s41467-018-07992-3
Baquero F, Alvarez-Ortega C, Martinez JL (2009) Ecology and evolution of antibiotic resistance. Environmental Microbiology Reports 1(6):469-476. https://doi.org/10.1111/j.1758-2229.2009.00053.x
