Wastewater Treatment Plants & Bacteria Strategies for Compliance

1. Wastewater Treatment Plants & Bacteria Strategies for Compliance Best Practices for Effluent Sampling TANNY BUSBY & LAURA BONJONIA ENVIRODYNE LABORATORIES, INC. April 26, 2011

2. Where Does the Analytical Testing Begin? • Container Preparation • Sample Collection • Chain of Custody • Preservatives • Field QC

3. Best Practices for Effluent Sampling What is a Representative Sample? Definition Every Member of the Population has an “Equal Chance” of being Selected Typically a Population is very large A Sample is a Subset of the Population

5. Best Practices for Effluent Sampling The sample reflect characteristics of the Effluent

6. Best Practices for Effluent Sampling Identification of Sampling Points • Effluents • Influents • Process Control • Collection Systems

7. Grab e-Coli Fecal Coliform Oil & Grease *DO *pH Cyanide VOC *Chlorine Residual Composite Metals BNA Pesticides TPH Minerals Nutrients (Nitrogen) BOD TSS (Solids) DEFENSIBLE DATASample Collection

8. DEFENSIBLE DATAQUALITY CONTROL • Calibrate Field Equipment - (Document in field log) ~ pH, DO & Cl2 Meters • Field Blanks • Trip Blanks • Rinseates • Field and Lab Duplicates • Field and Lab Matrix Spikes (MS) • Laboratory control samples (LCS) • Method Numbers

9. Gram stain Gram negative (turns red) Do not produce spores Bacilli (rod shaped) Aerobic (require oxygen) or Facultative Anaerobic WHAT ARE COLIFORM

10. WHERE ARE COLIFORM(s) FOUND? • Surface Water • Decaying Plants • Soil • Intestinal Tract of Warm-blooded Animals

11. TYPES OF COLIFORM BACTERIA

12. ~ Microbes ~ Commonly Regulated in Wastewater Effluents • Fecal Coliform • 200 CFU/100ml ~ Monthly Geo-Mean (Avg.) • 400 CFU/100ml ~ 7 Day Geo-Mean (Avg.) • 800 CFU/100ml ~ Single Grab Max. • E. coli ~ (Escherichia coli) • 126 CFU/100ml ~ Monthly Geo-Mean (Avg.) • 394 CFU/100ml ~ Single Grab Max. • Enterococci • 36 CFU/100ml ~ Monthly Geo-Mean (Avg.)

13. COLIFORM LAB TESTING METHODS • Multiple Tube Fermentation (Quantifies) • Reported as MPN (most probable number) • Membrane Filtration (Quantifies) • Reported as CFU’s/100 mL (colony forming units) • Quanti-tray (Quantifies) • Colilert – 18-24 hours • Reported as MPN • Positive sample turns yellow; E.coli fluorescence • Presence/Absence (Qualifies) • Colilert, Colisure • Positive sample turns yellow (Colilert); turns magenta (Colisure) • E.coli fluorescence

14. E. coli • Membrane filtration • m-ColiBlue 24 • Total coliform and • E. coli simultaneously • E. coli – Blue colonies • Total coliform – Red or blue colonies • Other methods are approved

15. ENTEROCOCCI PLATE • Membrane filtration • SM 9230 C • CFUs/100 ml • Blue colonies • Other methods • Enterolert • MPN (Most Probable Number)

16. COLILERT- QUANTITRAY Incubate 18-24 hours- Most Probable Number TOTAL COLIFORMS E.Coli (UV Light)

17. SAMPLING HOLD TIMES • Fecal Coliform, E. coli and Enterococci • Grab (Not Composite) • Sterile container • Hold Time ~ Not to Exceed a Total of 8Hrs • 6 hours transport + 2 hours set up

18. Micro Samples How to Collect Effluent • Directly into Sterile Testing Container

19. How to Collect and Split Samples

20. Total Coliform Qualitative Analysis (PRESENCE~ABSENCE) E.coli positive (UV Light) • COLILERT-18 • Incubate 18-22 hours • COLILERT • -Incubate 24-28 hours COLISURE -Incubate 24-48 hours

21. QUESTIONS