Hygiene considerations for source-separated urine collection, storage and processing into a marketable fertilizer. Heather N.

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1 Hygiene considerations for source-separated urine collection, storage and processing into a marketable fertilizer Heather N. Bischel Postdoctoral Research Scientist Laboratory of Environmental Chemistry Swiss Federal Institute of Technology, Lausanne (EPFL) Simon Schindelholz, Manfred Schoger, Tamar Kohn September 15, 2016

2 Urine separation & Struvite production Kwazulu- Natal province Population: 3.5M Durban >80K Urine-Diverting Dry Toilets (UDDTs) in rural/peri-urban areas Struvite production: Add Mg 2+, gentle mix, Cotton bag Filters MgNH 4 PO 4 6H 2 O Drying outdoors under cover Valorisation of Urine Nutrients in Africa (VUNA) 2

3 Hygiene Considerations 1g of fresh feces from an infected person can contain: ~10 6 viral pathogens bacterial pathogens 10 4 protozoan cysts or oocysts helminth eggs Feachem et al, 1983 Source-separated urine is not sterile! Human pathogens detected in source-separated urine in Durban Diarrheal Bacteria E.g.,: Shigella spp., Vibrio spp., E. coli 0157:H7 Human viruses E.g.,: Rotavirus, Adenovirus, Hepatitis A Virus Bischel et al, Wat. Res

4 4 Overall Objectives Characterize and optimize hygiene of urine storage and recycling Pathogens: Excretion in feces / urine Urine Storage Struvite Production & Drying Fertilizer application 1. Which hazards are there? 2. How can we mitigate them? Inactivation Kinetics and Mechanisms 3. Health risk implications? Challenges Anticipate short-term urine storage for process scale-up or in dense locations Which microbial targets to evaluate? Pathogen indicators and in situ bacteria Manual struvite production in the field: Variable temperature and humidity

5 Inactivation Potential During Urine Storage BACTERIA: e.g., E. coli and Salmonella e.g., C. perfringens spores: VIRUSES: e.g., Rotavirus Rapid die-off T 90 < 5 d at ~20 C Little to no reduction Slower inactivation T 90 ~ 35 d at 20 C, 1:2 dilution (Höglund, Stenström et al. 1998; Vinneras et al. 2008; Schonning 2001; Höglund, Ashbolt et al Decrey et al. 2015, 2016) and others Inactivation depends on: Organism (e.g., gram negative/positive bacterium; viral genome type) Urine storage temperature Ammonia concentration (urine dilution and storage conditions) ph (ammonia speciation) Viable bacteria present after long-term storage And even urine treatment (Lahr et al, 2016) Inactivation during fertilizer production important after short-term urine storage

6 Inactivation during Struvite Precipitation & Drying Laboratory-made Struvite Cake Inactivation of virus and helminths during controlled struvite drying inactivation with moisture content inactivation with temperature (Decrey et al. Wat. Res. 2011) Evaluate: Retention of bacteria in struvite cake Bacteria inactivation in lab (controlled) and field Degradation of struvite with T > 55 C How to increase inactivation without heat treatment? 6

7 Methods: Bacteria Inactivation during Struvite Precipitation & Drying Targeted health-relevant bacteria: Heterotrophic plate counts: Salmonella typhimurium Enterococcus spp. Total viable bacteria by flow cytometry: File: Sample_003_struvite_SYBR+PI_t0_1a_10^3_diluted.FCS R2 R1 Date: FL3 (Red) Struvite (Dead) (Live) counts ~10 10 viable bacteria per gram of 40 struvite! FL1 (Green) SS

8 FL3 (Red) Log C/C 0 FL3 (Red) FL3 (Red) Total Live Cells: Log C (Counts/g ww) In situ measurements by Flow Cytometry 0,5 0-0,5-1 -1,5-2 -2, C/40% RH Dead Live Time (hrs) R² = 0, Heterotrophs: Log C (CFU/g ww) (Dead) (Live) 0 hrs 24 hrs 48 hrs FL1 (Green) FL1 (Green) FL1 (Green) 8

9 Isothermal Struvite Drying S. typhimurium Log C/C Low Relative Humidity Low Humidity p < 1 (Tailing) Time (hrs) Weibull Model: Distribution of resistances to inactivation log10(c) = log10(c 0 ) ( t δ )p Shape param. (p) captures tail or shoulder δ is the time for first-decimal reduction Log C/C High Relative Humidity High Humidity p > 1 (Shoulder) Time (hrs) Tailing can result from, e.g.: Reduced probability of lethal hit Evaporative cooling at low humidity Dehydration tolerant subpopulation Retention of moisture for continued inactivation (followed by desiccation) 9

10 Log(C/C 0 ) Log(C/C 0 ) 2 1 In situ Heterotrophic Bacteria Inactivation Controlled Lab Conditions ethekwini Batch Swiss Batch Field Conditions Nylon Filters Cotton Filters Relative Moisture Content, Log(θ/θ 0 ) Log(θ/θ 0 ) Dynamic Field conditions: Oscillating temperature and relative humidity 10

11 Recommendations for inactivation of pathogens during struvite production Wet-heating for enhanced inactivation of bacteria Soil Solarization Inactivation dependent on relative humidity Addabbo et al 2010 Minimum temperature (35 C) required for helminth inactivation struvite degrades T > 55 C Desiccation: reduce moisture content for virus, helminth, & bacteria inactivation 11

12 Summary & Next steps Characterize and optimize hygiene of urine storage and recycling Pathogens: Excretion in feces / urine Urine Storage Struvite Production & Drying Fertilizer application 1. Which hazards are there? Source-separated urine is not sterile! Contains fecal pathogens including persistent viruses Recommend: Personal protective equipment; sealed storage 2. How can we mitigate them? Struvite Production Bacteria retained in filtered struvite Field inactivation consistent with lab Recommend: initial retention of moisture followed by desiccation Quantitative microbial risks assessment: during urine collection, handling and struvite production 12

13 Acknowledgements Tamar Kohn, Laboratory of Environmental Chemistry Kai Udert & Bastian Etter, Eawag/VUNA Teddy Gounden, EWS Field staff Chris Buckley, Sara Rhoton, Nicola Rodda, UKZN Simon Schindelholz, Manfred Schoger, Loic Decrey Funding: US and Swiss NSF Bill & Melinda Gates Foundation Rotary Foundation EPFL, EWS and Eawag Contact Info After April 2017: Heather Bischel, Asst Prof University of California, Davis