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Surveillance and Risk Assessment of Antibiotic Resistance in the Urban Water Cycle, Le Thai Hoang
- 1. Surveillance and Risk Assessment of Antibiotic Resistance in Urban Water Cycle Le Thai Hoang, PhD Lecturer, Environmental Engineering International University - Vietnam National University HCMC ProSPER.Net Young Researchers’ School 6 to 15 March, 2017
- 3. Annual death by Antimicrobial resistance (AMR) Tetanus 60,000 Car traffic accidences 1,200,000 Cholera 100,000-120,000 Diarrhea diseases 1,300,000 Measles 130,000 Diabetes 1,500,000 AMR 700,000 AMR in 2050 10,000,000 Cancers 8,200,000 WHO. Review on Antimicrobial Resistance. 2014
- 4. Timeline of antibiotic resistance 1930 1935 1940 1945 1950 1955 1960 1965 1970 1975 1980 1985 1990 1995 2000 2005 Sulfonamides Penicillin Streptomycin chloramphenicol Tetracycline Erythromycin Vancomycin Methicillin Ampicillin Cephalosporins Linezolid Daptomycin Antibiotic deployment Sulfonamides Penicillin Streptomycin Tetracycline Erythromycin Vancomycin Methicillin Chloramphenicol Ampicillin Cephalosporins Linezolid Daptomycin Antibiotic resistance observed Nature Chemical Biology 3, 541 - 548 (2007)
- 5. Antimicrobial uses by category ACS Infectious Diseases, 2015
- 6. Antibiotic in environment Antibiotics Livestock uses Human uses Excretion Treatment of manure Land application Excretion Flushing unused Topical application Municipal WW Runoff Treated discharge Leach to ground water Contaminate surface water Potential drinking water sources
- 7. Why is resistance monitoring important? Baseline Spread Trend Source tracking Veterinary/ human use Risk factors Education Policy
- 8. ANTIBIOTIC RESISTANCE 16 ARGs & class 1 integrons 20 Antibiotic residues Pathogenic ARBs Diversity of ARB & ARG Risk assessment Quantitative PCR LC-MS/MS Culture-based method Integrated approach Metagenomics Quantitative microbial risk assessment
- 9. Application Baseline A. Reservoirs & catchments B. Hospital Wastewater C. Domestic wastewater
- 10. 10.0 km Reservoirs & Catchments Catchments & Reservoirs K PKR1 (R) WQ4A (C) WQ5 (C) WQ6 (C) M RmbA (R) CmbB (C) CmbH (C) U RUSA (R) RUSF (R) RUSH (R) FRESHWATER Mc RmcA (R) K U Mc M
- 11. Hospital Wastewater Hospital wastewater S6 S7 H1 H2 • Reservoirs for pathogenic bacteria • High usage of antibiotics • Concern with transmission and long term survival in the environment • Discharged into domestic sewage system without any treatments routes of dissemination to environment
- 12. Domestic wastewater Influent Effluent Primary Clarifiers Anoxic/Aerobic tank Secondary Clarifiers Return Activated Sludge Effluent Primary Clarifiers Anoxic/Aerobic tank Membrane Bioreactor Return Activated Sludge INF A1 A2 B1 B2 CAS treatment process MBR treatment process
- 13. Occurrences of AMR in reservoirs, hospital wastewater, and domestic wastewater
- 14. Target analytes Min (ppb) (n=8) Median (ppb) Max (ppb) (n=8) MDL (ppt) CFZ <DL <DL <DL 268 MER 0.10 0.82 0.94 15 CAP <DL Only one detection 0.44 19 CIPX <DL 2.44 71.94 70 LIN <DL <DL <DL 1.5 CLI <DL 0.83 1.14 28 ERY <DL <DL <DL 17 AZT 0.12 0.30 1.23 2 CLAR 1.02 2.63 56.77 2.4 TYL <DL <DL <DL 243 SMZ <DL <DL <DL 8 SMX 1.00 14.34 24.72 22 TMP 0.81 9.51 61.18 5 TET <DL <DL <DL 50 MIN <DL <DL <DL 94 CTC <DL <DL <DL 12 OXY <DL <DL <DL 74 VAN 0.15 6.94 42.59 5 Concentrations of AB in hospital wastewaters
- 15. 1.00E+00 1.00E+01 1.00E+02 1.00E+03 1.00E+04 1.00E+05 1.00E+06 1.00E+07 1.00E+08 1.00E+09 MPN100ml E.coli Enterococci Pseudomonas aeruginosa Reservoirs and catchments: - Enterococci: between 1.76 x 101 and 2.54 x 103 MPN/100mL - E.coli: between 2.11 x 101 and 4.28 x 103 MPN/100mL Reservoirs water quality all below thresholds recommended by USEPA. Biological indicators for WQ
- 16. Concentrations of ARB ARB concentrations (geometric means): Hospital wastewaters - (1.40 x 105 CFU/mL) Domestic wastewaters – (5.94 x 105 CFU/mL) Freshwaters – (5.14 x 102 CFU/mL)
- 17. Relative abundance of ARGs Relative abundance of ARGs (geometric means): Hospital wastewaters – Average (8.91x10-2) Domestic watewaters – Average (3.62x10-2 ) Freshwaters – Average (8.67x10-4) 1. ARG abundance in freshwaters 2 magnitudes lower 2. All 4 bla-gene targets found in freshwaters (10-5-10-7) , however at least a magnitude lower than in wastewaters (10-3-10-5)
- 18. Phylogenetic composition of ARB Dominant AR bacteria: Wastewaters: Aeromonas, Enterobacteriaceae (Klebsiella, Enterobacter, E.coli), Pseudomonas, Acinetobacter Freshwaters: Flectobacillus, Pseudomonas, Acinetobacter, Flavobacterium, Aeromonas
- 19. Risk assessment of Antibiotic resistant E. coli O157H7 in Recreational Health Risks
- 20. Hazard Identification Dose Response Assessment Exposure Assessment DALYs Probability of infection/illness Reservoirs water Treated water • Sewage • Hospital effluent ARB at MIC Indicator organism Antibiotics ARGs/Integr ons ARB pathogens (e.g., E. coli, K. pneumoniae, etc.) Library of ARB • Frequency • Severity (e.g., last resort AB, pathogen, virulence factor) Risk Risk Controll QMRA approach for Antibiotic resistance MIC/MDR ARGs Virulence genes
- 21. Occurrence of Antibiotic resistant E. coli <100 CFU/100ml <10,000 CFU/100ml • Prevalence of E. coli in agricultural and urbanized area > 100 times in reservoirs • Among 4 reservoirs, Marina is the highest prevalence of AMR E. coli. • CIP and SXT are the most prevalent. AMK was the least. • Average concentration of E. coli in reservoirs < EPA guideline (200 EC/100ml).
- 22. Concentration of E. coli O157H7 Eco CEFT-Eco CIP-Eco SXT-Eco MEM-Eco Average 108.58 0.02 0.34 1.01 0.05 Median 1.6 0 0.01 0.01 0 Mode 0.04 0 0 0 0 SD 7,537.19 0.23 12.74 91.98 1.08 Distribution lnorm lnorm lnorm lnorm lnorm E. coli : E. coli O157H7 = 1: 0.08 Reference: Haas et al., 1999; Howard et al., 2006; Assumption AR E. coli : AR E. coli O157H7 = 1:0.08
- 23. Exposure and dose-response parameters Distribution Parameters References Exposure duration (h) PERT(minimum, likeliest, maximum) (0.25, 0.5, 2) Mcbridge 2013 Ingestion rate (ml/h) PERT(minimum, likeliest, maximum) (2,10, 20) Dorevitch 2010, 2011 Dose-response model Beta-poison model: 𝑃𝑃 = 1 − (1 + 𝐷𝐷𝐷𝐷𝐷𝐷𝐷𝐷 × 2 1 𝛼𝛼−1 𝑁𝑁50 )−𝛼𝛼 Exposure for 2nd contact activities (Rowing, canoeing, kayaking) alpha N50 illness/infection rate Reference E.coli O157H7 2.10E-01 1.12E+03 0.35 Hass 1999, Horward and Pedley 2004 Assumption Susceptible and resistant E. coli O157H7 have the same ability to infect to human.
- 24. Probability of Gastrointestinal illness EPA guideline (2012): 36 illnesses/ 1000 cases
- 25. Number of GI cases per 1000 recreators 2.9% 0.02% Statistics Eco157 CAZ-Eco157 CIP-Eco157 SXT-Eco157 MEM-Eco157 Mean 4.04 0.00397 0.0317 0.0953 0.00626 Median 0.167 0.000185 0.000817 0.00109 0.000164 Minimum 0.0006 0 0 0 0 Maximum 219 5.49 15.9 69.3 6.49 EPA guideline (2012): 36 illnesses/ 1000 cases Frequency of exceeding the EPA guideline 2012
- 26. Removal of Antibiotic Resistance in Domestic Wastewater by The Membrane Bioreactor Treatment Influent Effluent Primary Clarifiers Anoxic/Aerobic tank Secondary Clarifiers Return Activated Sludge Effluent Primary Clarifiers Anoxic/Aerobic tank Membrane Bioreactor Return Activated Sludge INF A1 A2 B1 B2 CAS treatment process MBR treatment process
- 27. Membrane bioreactor treatment 27 • Introduced in late 1960s • Is the combination of a membrane process with a suspended growth bioreactor • is now widely used for wastewater treatment. • Advantages over the activated sludge treatment: • high quality of effluent: low turbidity, bacteria, TSS, BOD • can operate at high concentration of MLSS, low reactor volume OBJECTIVE: To evaluate the removal efficiency of AB, ARB, and ARG in the MBR process compared to the CAS process.
- 28. 28 𝑹𝑹𝑹𝑹𝑹𝑹𝑹𝑹𝑹𝑹𝑹𝑹𝑹𝑹 𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆𝒆 𝒆𝒆𝒆𝒆𝒆𝒆 % = 𝑪𝑪𝑰𝑰𝑰𝑰𝑰𝑰 − 𝑪𝑪 × 𝟏𝟏𝟏𝟏𝟏𝟏 𝑪𝑪𝑰𝑰𝑰𝑰𝑰𝑰 Removal of Antibiotic residues • On average, about 75% and 80% AB were removed in CAS and MBR processes. • Both Secondary clarifier and MBR treatment did not efficiently remove AB. ng/l CAS MBR High >200 Chlotetracycline Chlotetracycline Oxytetracycline Amoxicilin Tetracycline Oxytetracycline Azithromycin Clarithromycin Clarithromycin Sulfamethaxazole Ciprofloxacin Tetracycline Sulfamethaxazole Ciprofloxacin Medium 10-200 Trimethoprim Azithromycin Sulfamethazine Erythromycin Erythromycin Sulfamethazine Meropenem Trimethoprim Lincomycin Meropenem Vancomycin Lincomycin Vancomycin Low <10 Clindamycin Clindamycin Minocycline Minocycline Chloramphenicol Chloramphenicol Ceftazidime Ceftazidime Tylosin Tylosin Amoxicilin
- 29. 29 Removal of Antibiotic resistant bacteria • Prevalence of ARB in the effluent were from 102 to 104 CFU/ml in CAS, and under detection limit in MBR. • Average log removal of ARB in final effluent were about 2.3 in CAS, and 5.5 in MBR. • MBR treatment was highly efficient in removal of ARB. 𝑳𝑳𝑳𝑳𝑳𝑳 𝒓𝒓𝒓𝒓𝒓𝒓𝒓𝒓𝒓𝒓𝒓𝒓𝒓𝒓 (𝑨𝑨𝑨𝑨𝑨𝑨) = −log𝟏𝟏𝟏𝟏 𝑪𝑪 𝑪𝑪𝑰𝑰𝑰𝑰𝑰𝑰 * * P=0.016 P=2.5x10-9
- 30. 30 • Average log removal of ARG were approximately 1.5 in CAS, and 3.0 in MBR. • Compared to the CAS, MBR showed a better efficiency in removal of ARG genes. CFU/ml CAS MBR High >1000 16S 16S sul1 sul1 tetO tetO aac6 int1 ermB Medium <1000 qnrB ermB blaCTX-M qnrB tetM blaCTX-M blaSHV tetM blaKPC Low <100 qnrA qnrA vanA int1 dfrA vanA sul2 dfrA cfr blaKPC blaNDM1 aac6 sul2 cfr blaSHV blaNDM1 * Removal of Antibiotic resistant genes
- 31. Overall summary ∗ Antibiotic resistance (AB, ARB, ARG) is already a global concern threatening environmental and community health, not something in future. ∗ Surveillance effort, especially on aquatic environment, need to be raised worldwide to understand the current status, baseline, and guideline for further management. ∗ Culture-based method, qPCR, LC-MSMS, and metagenomics are demonstrated a good method to detect and analyze AR. ∗ There need to be a specific treatment of AR in WWTP to increase removal of AR factors (AB, ARB, ARG) ∗ Burden of disease for AR pathogen needs to evaluate.
- 32. Acknowledgements A/Prof. Karina Gin Dr. Ng Charmaine Dr. Laurence Haller National Research Foundation (NRF) International University HCMC RCE ESD Southern Vietnam