By: Alyssa Ruiz Mentor: Dr. Tesfa Yacob PI: Dr. Karl Linden REINVENT THE TOILET CHALLENGE: URINE TREATMENT ENVIRONMENTAL SUSTAINABILITY REU

 2.5 billion people practice open defecation or lack adequate sanitation facilities  Poor sanitation contributes to 1.5 million child deaths from diarrhea each year  Main Objectives  radically change our world’s current sanitation state through innovative discoveries that convert our waste into viable resources  decrease the statistic of child mortality that societies around the globe are facing and to improve the quality of life in these communities ABOUT THE CHALLENGE 2

BACKGROUND: URINE hydrolysis 3

 Source separated toilet  utilize the properties found in urine  create biochar  energy efficient  Main source of pathogens in source separated urine comes from misplaced feces  Source separated urine high potential for regrowth  Less water compared to latrines  Daily sanitation  End Product = Fertilizer ABOUT THE SYSTEM 4

 Experiments have shown disinfection over long periods of time (weeks) at low temperature and varying ammonia concentrations  Assess short term (hours) disinfection with these variables 1.the optimum concentration of ammonia 2.the intensity of heat treatment 3.the duration of the application of heat treatment  Assess the potential for regrowth in urine 0BJECTIVES 5

 Two surrogates used from frozen cultures  E. coli Famp  MS2  Enumeration Methods  Membrane Filtration - E. coli Famp  Agar Plating - MS2 MATERIALS AND METHODS 6

 Initial Experiment: 40°C & 60°C over 1 hr  Surrogates: E. coli & MS2  Total Ammonia Concentration: 2 g/L EXPERIMENTS: PRELIMINARY DISINFECTION 7

RESULTS: PRELIMINARY DISINFECTION 8

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 Previous experiment shows 60°C more than enough while literature says must be above 45°C  Maximize efficiency  Chose 50°C because previous data implied should take less than one day - optimal  Curious about a synergistic effect  3 treatment methods  AH: Ammonia + Heat  H: Heat Only  A: Ammonia Only  Tested over 24 hours  2.3 g/L NH3  Two surrogates used: E. coli Famp & MS2 EXPERIMENTS: 50°C DISINFECTION 10

RESULTS: 50°C DISINFECTION 11

RESULTS: 50°C DISINFECTION 12

 Factors that could support regrowth  Contamination  High abundance of nutrients found in urine  Tested 3 urine dilutions  1:0 (2.4 g/L NH3)  1:1 (1.2 g/L NH3)  1:5 (0.4 g/L NH3)  Tested 2 controls  Positive: TSB growth media  Negative: PBS  Designed to run for 21 days  10^4 starting E. coli concentration EXPERIMENTS: REGROWTH 13

SampleDay 0 (avg log CFU/ 1 mL) Day 2 (avg log CFU/ 1 mL) Day 24 (avg log CFU/ 1 mL) 1: ± : ± : ± TSB (pos) 4.38 ± ± 0.13NA PBS (neg) 4.07 ± ± ± 0.03 RESULTS: REGROWTH 14

 First of these types of experiments  Synergistic effect proven  significant in short term urine disinfection  implications for other avenues: passive solar heating  Further test synergistic effect with varying ammonia concentrations  Regrowth experiments with spiking before heat  Short, low heat + storage  Other surrogates – Enterococcus, ascaris, eggs CONCLUSIONS & FURTHER EXPERIMENTS 15

 NSF REU Program  CU Boulder – Dr. Montoya  Bill & Melinda Gates Foundation  Dr. Karl Linden  Dr. Tesfa Yacob  Ryan Mahoney  Sara Beck  Cori Oversby  Linden/Gates Team ACKNOWLEDGEMENTS 16

 "Bill & Melinda Gates Foundation." Water, Sanitation & Hygiene. N.p., n.d. Web. 8 June .  "Hach – Manufactures Water Quality Testing and Analytical Instruments & Reagents." Hach –  Manufactures Water Quality Testing and Analytical Instruments & Reagents. N.p., n.d. Web. 8 June  Höglund, C. et al. "Evaluation of Faecal Contamination and Microbial Die-Off in Urine  Separating Sewage Systems." Water Science & Technology 38.6 (1998): Print.  Putnam, David F. “Composition and Concentrative Properties of Human Urine.” (1971): n.  pag. Google Scholar. Web.  McCartney, Daryl, and Kristine M Wichuk. “A Review of the Effectiveness of Current Time–  Temperature Regulations on Pathogen Inactivation During Composting.” Journal of Environmental Engineering and Science6.5 (2007): 573–586. CrossRef. Web.  Udert, K. M. et al. “Fate of Major Compounds in Source-separated Urine.” Water Science &  Technology 54.11–12 (2006): Print.  Vinnerås, Björn et al. “Inactivation of Bacteria and Viruses in Human Urine Depending on  Temperature and Dilution Rate.” Water Research (2008): 4067–4074. CrossRef. Web. REFERENCES 17

“OUR GOAL: to enable universal access to sustainable sanitation services by supporting the development of radically new sanitation technologies as well as markets for new sanitation products and services.” – Gates Foundation QUESTIONS? 18