and Antibiotic Resistance Is there a Connection Jean E McLain Associate Director University of Arizona Water Resources Research Center Statewide Water Quality Management Working Group October 11 2016 ID: 1032111
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1. Recycled Municipal Wastewater and Antibiotic Resistance: Is there a Connection?Jean E. McLain, Associate DirectorUniversity of Arizona Water Resources Research CenterStatewide Water Quality Management Working GroupOctober 11, 2016
2. OutlineWhat is antibiotic resistance?Antibiotic resistance in the news
3. OutlineWhat is antibiotic resistance?Antibiotic resistance in the newsCurrent studies examining effects of anthropogenic chemical pollutants on environmental resistanceEstablishing guidelines for future resistance studies
4. The ability of a bacterium to prevent an antibiotic from adversely affecting that isolate, strain, or group.Antibiotic Resistant Bacteria
5. The ability of a bacterium to prevent an antibiotic from adversely affecting that isolate, strain, or group.Horizontal Gene Transfer confers antibiotic resistance in response to selective pressure Clinical settings High antibiotic dosages Antibiotic Resistant Bacteria
6. The ability of a bacterium to prevent an antibiotic from adversely affecting that isolate, strain, or group.Horizontal Gene Transfer confers antibiotic resistance in response to selective pressure Clinical settings High antibiotic dosages WWTPsAntibiotic Resistant Bacteria
7. Environmental Antibiotic ResistanceAgricultural and clinical use of antibioticsUp to 75% excreted unaltered or as metabolites
8. Environmental Antibiotic ResistanceAgricultural and clinical use of antibioticsUp to 75% excreted unaltered or as metabolitesMany WWTPs not designed for the removal of these chemicals Recycled wastewater BiosolidsProposed that this is a “key source of resistance to the environment”
9. Earliest antibiotics: naturally produced (e.g., Penicillin)ABR bacteria isolated from deep terrestrial subsurface (>250 m deep caves) and prehistoric (120,000+ YBP) ice coresAntibiotic Resistant Bacteria
10. Earliest antibiotics: naturally produced (e.g., Penicillin)ABR bacteria isolated from deep terrestrial subsurface (>250 m deep caves) and prehistoric (120,000+ YBP) ice coresABR genes evolved in absence of selective pressure Competition for scarce resourcesAntibiotic Resistant Bacteria
11. Broth microdilution methodsAgar disc diffusionClinical and Laboratory Standards InstituteResistance Analysis Methods: Bacterial Isolation/Culturing
12. Broth microdilution methodsAgar disc diffusionClinical and Laboratory Standards InstitutePROS:Highly reproducible, robustCan assess at clinical levelsCONS:Analyze a single isolate in each plateResistance Analysis Methods: Bacterial Isolation/Culturing
13. Resistance Analysis Methods: Molecular ID of GenesPolymerase Chain Reaction (PCR), Quantitative PCR
14. Resistance Analysis Methods: Molecular ID of GenesPolymerase Chain Reaction (PCR), Quantitative PCRPROS:Can identify/quantify resistance genes in entire DNA sampleAbility to quantify to single gene marker levelCONS:No discrimination between live and dead bacteria
15. Resistance Analysis Methods: Molecular ID of GenesPolymerase Chain Reaction (PCR), Quantitative PCRPROS:Can identify/quantify resistance genes in entire DNA sampleAbility to quantify to single gene marker levelCONS:No discrimination between live and dead bacteria
16. 2012, Funded by private citizen15 Recycled water samples; collected without guidance or permissionShipped to Virginia Tech University for analysisHow Did ABR Become Connected to Recycled Water Use?The Case of Flagstaff, Arizona
17. Aug 13: Antibiotic resistance genes in pipe outflows of recycled water systemHow Did ABR Become Connected to Recycled Water Use?The Case of Flagstaff, Arizona
18. Aug 13: Antibiotic resistance genes in pipe outflows of recycled water systemSept 26: “New Study Finds Antibiotic-Resistant Bacteria Genes in Flagstaff’s Reclaimed Water”How Did ABR Become Connected to Recycled Water Use?The Case of Flagstaff, Arizona
19. Aug 13: Antibiotic resistance genes in pipe outflows of recycled water systemSept 26: “New Study Finds Antibiotic-Resistant Bacteria Genes in Flagstaff’s Reclaimed Water”Oct 11: “Wastewater Snow-Making Could Breed Super-Bacteria”Oct 15: “Manmade Snow From Recycled Water May Contain Antibiotic Resistant Bacteria”How Did ABR Become Connected to Recycled Water Use?The Case of Flagstaff, Arizona
20. Aug 13: Antibiotic resistance genes in pipe outflows of recycled water systemSept 26: “New Study Finds Antibiotic-Resistant Bacteria Genes in Flagstaff’s Reclaimed Water”Oct 11: “Wastewater Snow-Making Could Breed Super-Bacteria”Oct 15: “Manmade Snow From Recycled Water May Contain Antibiotic Resistant Bacteria”Oct 17: “Antibiotic Resistance Racing Downriver”How Did ABR Become Connected to Recycled Water Use?The Case of Flagstaff, Arizona
21. December 2012: First meeting of Flagstaff City Manager’s Expert Panel on Antibiotic ResistanceHow Did ABR Become Connected to Recycled Water Use?The Case of Flagstaff, Arizona
22. December 2012: First meeting of Flagstaff City Manager’s Expert Panel on Antibiotic ResistancePanel Agrees on the Facts:1. Antibiotic resistance genes ≠ resistant bacteria (DNA can indicate living or dead)2. Recycled water pipe only: what about drinking/potable water pipes?How Did ABR Become Connected to Recycled Water Use?The Case of Flagstaff, Arizona
23. More Recently (March 2016)The Case of Los Angeles and CRECRE thrives in soil and water AND in the human gut; mortality rate of 50%
24. More Recently (March 2016)The Case of Los Angeles and CRECRE thrives in soil and water AND in the human gut; mortality rate of 50%Los Angeles Times“Deadly Superbugs Get Stronger in the Sewers…”WWTPs serve as a “luxury hotel” for superbugs“Chlorine is just not doing it”
25. More Recently (March 2016)The Case of Los Angeles and CRECRE thrives in soil and water AND in the human gut; mortality rate of 50%Los Angeles Times“Deadly Superbugs Get Stronger in the Sewers…”WWTPs serve as a “luxury hotel” for superbugs“Chlorine is just not doing it”The FactsEPA sampled “partially treated wastewater”Antibiotic resistance genes
26. Does Recycled Municipal Wastewater Induce Antibiotic Resistance?
27. Gilbert Riparian PreserveCreated in 1986, seven recharge basins receive tertiary-treated recycled water
28. Gilbert Riparian PreserveCreated in 1986, seven recharge basins receive tertiary-treated recycled waterControl site: irrigation retention pond at Maricopa Agricultural Center
29. Gram positive Enterococcus spp.GI tracts of humans and animals; environmental persistenceGreat capacity for gene transferField Sampling over Two Years
30. Gram positive Enterococcus spp.GI tracts of humans and animals; environmental persistenceGreat capacity for gene transferEmergence of multiple drug-resistant strains in clinical settingsIdeal bacterial group for investigating the ecology of AR developmentField Sampling over Two Years
31. Isolates tested for resistance to 16 widely used antibiotics using CLSI broth microdilution methodsLaboratory Methods
32. Isolates tested for resistance to 16 widely used antibiotics using CLSI broth microdilution methodsLaboratory Methods
33. High Level Antibiotic Resistance
34. High Level Antibiotic Resistance
35. High Level Antibiotic Resistance
36. Multi-Antibiotic Resistance
37. Multi-Antibiotic Resistance
38. Multi-Antibiotic Resistance?
39. Antibiotic resistance: “metabolically expensive”What Does This Mean?
40. Antibiotic resistance: “metabolically expensive”Recycled Water, Biosolids = Adding Soil CarbonDoes adding C at certain levels reduce bacterial competition and overcome the need to express resistance?What Does This Mean?
41. Antibiotic Resistance in the Environment: A Call to ActionPublished reports indicate that recycled water selects for resistant bacteria BUT many studies ignore natural antibiotic resistance and lack control sites
42. Antibiotic Resistance in the Environment: A Call to ActionPublished reports indicate that recycled water selects for resistant bacteria BUT many studies ignore natural antibiotic resistance and lack control sites We know that release of trace antibiotics and resistance genes is occurring, but propose that effects are unknown
43. Antibiotic Resistance in the Environment: A Call to ActionTop researchers around the world; meetings 2014 and 2015To reach consensus on methodsTo train next generation in new methodsTo address misinformationFive review papers in March/April issue of Journal of Environmental Quality
44. Antibiotic Resistance in the Environment: A Call to ActionTop researchers around the world; meetings 2014 and 2015To reach consensus on methodsTo train next generation in new methodsTo address misinformationWorld Health Organization: “one of the most critical human health challenges of the next 100 years”
45. AcknowledgementsUniversity of Arizona Channah Rock Leif AbrellUniversity of Nevada-Las Vegas Dan GerrityUSDA-NIFA Collaborators Lisa Durso Dan SnowUSDA-ARS Water Management and Conservation ProgramsUSGS 104b, WateReuse Foundation, Arizona Department of Agriculture
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