1. Antibiotic Resistant Escherichia Coli in Non-Tidal Wetlands of Maryland
Speakers: Neil Agarwal & Robert Vocke III
University of Maryland, College Park

2. Establishing a Baseline
MDE
Clean Water Act
Antibiotics

3. Site Overview
Non-tidal, mitigation sites
3 plots per site

4. Data Collection Vegetation Soils Land Use Water Quality
Antibiotic Resistance

5. Our focus today… Water quality data Why Antibiotic Resistance?
The Concept of Measuring Resistance
Preliminary Results

6. Water Quality Results
Nitrate (mg/L)

7. Water Quality (continued)
Ammonia (mg/L)

8. Water Quality (continued)
Total N (mg/L)

9. Water Quality (continued)
Total P (mg/L)

10. Why look at antibiotic resistance?
The pharmaceutical problem
Antibiotics and resistance
Testing for antibiotic resistance is cost effective
Questions to be asked:
Prevalence and temporal changes
Correlation to other aspects of health
Resistance ‘sinks’
Nutrient Cycling

11. Disk Diffusion: The Concept
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12. Disk Diffusion: The Concept
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13. Fleming’s Discovery
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14. Disk Diffusion: The Procedure
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15. Disk Diffusion: The Procedure
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16. Disk Diffusion: The Procedure
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17. Disk Diffusion: The Procedure
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18. Disk Diffusion: The Procedure
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19. Disk Diffusion: The Procedure
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20. Our specific procedure
Collected surface samples
Serial dilutions
Culture on ColiBlue

21. ColiBlue media
Red = Coliforms
Blue = E. Coli

22. Our specific procedure
Collected surface samples
Serial dilutions
Culture on ColiBlue
Isolate a single E. coli colony
Disk diffusion using 5 antibiotics:
Ampicillin
Erythromycin
Tetracycline
Sulfisoxazole
Ciprofloxacin
Negative Control: Vancomycin

23. Converting Inhibition Zones
CLSI standard
Example:
Ampicillin with 7mm zone of inhibition
<13 mm = Resistant mm = Intermediate >17 mm = Susceptible

24. Converting Inhibition Zones
CLSI standard
Example:
Ampicillin with 7mm zone of inhibition
<13 mm = Resistant mm = Intermediate >17 mm = Susceptible

25. Results of Testing Ampicillin Tetracycline Ciprofloxacin Sulfisoxazole
Susceptible 1 2 32 11
Intermediate 5 13 12
Resistant 27 18
Total 33 34
% Resistant
82%
55% 0%
32%

26. What’s next? Find correlations to health Test upstream & downstream
Effect on nutrient cycling

27. Antibiotic Resistant Escherichia Coli in Non-Tidal Wetlands of Maryland
Speakers: Neil Agarwal & Robert Vocke III
University of Maryland, College Park