USE OF BIOFERTILIZERS IN BERRY FIELD AND FOOD SAFETY

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1 USE OF BIOFERTILIZERS IN BERRY FIELD AND FOOD SAFETY Meijun Zhu, Benedict Chris, Chad Eugene, Margaret Drennan Washington State University

2 FOODBORNE ILLNESS IS A SIGNIFICANT BURDEN There are 1400 foodborne outbreaks reported annually. These outbreaks result in 47.8 million illnesses 128,000 hospitalizations 3,000 deaths 2011 Fact Sheets

3 FOODBORNE OUTBREAKS Outbreaks and contamination incidents are global and wide reaching. Involve in different food products. Meat, dairy and egg products Low moisture foods Minimally processed foods, fresh produce

4 EPIDEMIOLOGY 2011 Fact Sheets

5 LISTERIOSIS IS RARE, BUT DEADLY Pathogen Illnesses Deaths Case-fatality rate L. monocytogenes 1, % Campylobacter 1,300, % Salmonella 1,230, % Shiga toxin- producing E. coli O157 96, % Scallan E et al, Emerging Infectious Diseases Has a high mortality rate (~30%)

6 THREE IMPORTANT FOODBORNE PATHOGENS Listeria monocytogenes High mortality Environmental species Is listed by the FDA as a pathogen of concern Salmonella All strains and serotypes are pathogenic Implicated in diverse food outbreaks.

7 THREE IMPORTANT FOODBORNE PATHOGENS Shiga toxin producing E. coli (STEC) Hemorrhagic colitis and Hemolytic Uremic Syndrome (HUS) >100 STEC associated with human disease. E. coli O157:H7 Non-O157 STEC Numerous outbreaks have been traced to leafy greens E. coli O157:H7 outbreaks linked to romaine lettuce The fecal shedding is the main source of contamination Law, Allison, 2007;

8 FOODBORNE PATHOGENS - UNIQUE CHALLENGES Grown in an open environment Pathogens can be persistent in the environment Contamination can happen during growing, harvesting, packing and through-out the supply chain No Kill step

9 FSMA (FOOD SAFETY MODERNIZATION ACT) FSMA targets the safety of fresh and minimally processed foods because of recent outbreaks. Proposed Produce Rule Focus on the growing, harvesting, and post-harvest handling of produce Focus is on the prevention.

10 FIVE AREAS OF FOCUS IN PRODUCE RULE Agricultural water Biological soil amendments of animal origin Domesticated and wild animals Personnel qualifications, training, and health and hygiene Equipment, tools, buildings and sanitation

11 BIOLOGICAL SOIL AMENDMENTS OF ANIMAL ORIGIN Untreated animal waste may be a source of pathogens. Proper manure management is crucial to reduce risk billion tons of manure are produced annually Majority is from cattle Valuable nutrients and organic matters for soil enrichment and crop production

12 MANURE IMPLICATED IN E. COLI OUTBREAK 2006 spinach E. coli O157:H7 outbreak cause ~205 cases and 3 deaths. Jay et al., Emerging Infectious Diseases, 13: FDA traced sources 4 California ranches near spinach fields Case Counts by State The outbreak strains Cattle farm Feral Swine farm or higher

13 OBJECTIVE To evaluate dairy manure-derived fertilizer application on microbial safety of soil, leaf, and fruits in berry fields. Salmonella E. coli O157:H7 and STEC L. monocytogenes

14 EXPERIMENTAL DESIGN FOR RASPBERRY FIELD AS: PS: Ammonium sulfate Phosphorous solid MA2 AS DLE: Digested liquid effluent CON SL: Straight lagoon DLE COM: Compost CON: Standard fertilization COM PS o 4 plots per treatment. o 2 soil samples, 2-3 foliar samples and 3 fruit samples per plot

15 EXPERIMENTAL DESIGN FOR BLUEBERRY FIELD AS1: Ammonium sulfate AS2: Ammonium sulfate slurry split CON: Standard fertilization AS1 CON AS2 o 4 plots per treatment o 2 soil samples, 2-3 foliar samples and 3 fruit samples per plot

16 TIMELINE AND SAMPLING PLAN Soil sample (~500 g) subsamples ~20 g/subsample Pre-application Post- application Before fruit harvest After fruit harvest Feb March April May Day June 0 July August Fertilizer sampling Fertilizer sample (~500 g) 8-10 subsamples 50 g/subsample Foliar sampling Foliar sample (~100 g) 200~250 subsamples Fruit sampling Fruit sample (~100 g) 10 subsamples 10 g/subsample

17 METHOD OVERVIEW Enrichment E. coli O157:H7 37 C, 42 C 25 g Salmonella 37 C 225 ml media Shake Listeria 30 C mbpwp E. coli O157:H7/STEC BPW BLEB Salmonella L. monocytogenes ml VRBA VRBA 25 g 225 ml media 1 ml 1 ml 1 ml PBS 10-2 VRBA 200 rpm/1 min 9 ml LST Coliform/generic E. coli

18 METHOD OVERVIEW E. coli O157:H7 STEC CT-SMAC CHROMSTEC CHROMO157 Immunomagnetic Separation L. monocytogenes MOX CHROMListeria Salmonella RV XLD HE BS CHROMSalmonella TT XLD HE BS CHROMSalmonella

19 E. coli O157: H7 Negative E. coli O157:H7 CT-SMAC CHROMagar O157/STEC E.coli O157:H7 mauve

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21 Listeria monocytogenes MOX Listeria CHROMListeria L. monocytogenes

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23 Salmonella spp. Salmonella RV XLD HE BS CHROMSalmonella TT XLD HE BS CHROMSalmonella BS HE XLD CHROMSalmonella

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26 a Results were confirmed by Latex Salmonella and PCR detection of inva gene; b Positive replicates/total replicates for each treatment. c RV and TT with the same result was only reported once. RV: Rappaport-Vassiliadis Salmonella enrichment broth; TT: Tetrathionate Salmonella enrichment broth; XLD: Xylose Lysine Desoxycholate agar; BS: Bismuth Sulfite agar; HE: Hektoen Enteric agar; CHROM: CHROMagar Salmonella agar; CON: standard fertilization; AS1: ammonium sulfate slurry; AS2: ammonium sulfate slurry split.

27 a Results were confirmed by Latex Salmonella and PCR detection of inva gene; b Positive replicates/total replicates for each treatment. c RV and TT with the same result was only reported once. RV: Rappaport-Vassiliadis Salmonella enrichment broth; TT: Tetrathionate Salmonella enrichment broth; XLD: Xylose Lysine Desoxycholate agar; BS: Bismuth Sulfite agar; HE: Hektoen Enteric agar; CH: CHROMagar Salmonella agar; CON: standard fertilization; AS1: ammonium sulfate slurry; AS2: ammonium sulfate slurry split.

28 Total coliform in fertilizer, 2017 (Log 10 CFU/g) TOTAL COLIFORM IN 2017 FERTILIZERS/SOILS IN RASPBERRY FIELD 5.0 b 4.0 c a a a CON AS COM DLE PS COM: Compost CON: Standard fertilization AS: PS: Ammonium sulfate Phosphorous solid DLE: Digested liquid effluent

29 Total count in fertilizer, 2018 (Log 10 CFU/g) TOTAL COLIFORM IN 2018 FERTILIZERS AND SOILS IN RASPBERRY FIELDS 5.0 Total coliform 4.0 b Generic E. coli 3.0 c c c e d a 0.0 a a a a a CON AS COM DLE PS SL COM: Compost AS: Ammonium sulfate DLE: Digested liquid effluent CON: Standard fertilization PS: Phosphorous solid SL: Straight lagoon

30 Total count in foliar, 2017 (Log 10 CFU/g) Total count in foliar, 2018 (Log 10 CFU/g) COLIFORM IN FOLIAR SAMPLES OF RASPBERRY FIELD Total coliform Generic E. coli 3.0 Total coliform Generic E. coli a a a a a a a a a a CON AS COM DLE PS a a a a a a a a a a a a CON AS COM DLE PS SL AS Ammonium sulfate. PS Phosphorous solid DLE Digested liquid effluent SL Straight lagoon COM Compost CON Standard fertilization

31 TOTAL COLIFORM IN SOILS IN BLUEBERRY FIELD AS1: Ammonium sulfate; AS2: Ammonium sulfate slurry split; CON: Standard fertilization

32 Total count in fertilizer, 2018 (Log 10 CFU/g) GENERIC E. COLI IN FERTILIZER/SOIL IN 2018 BLUEBERRY FIELD 5.0 Total coliform Generic E. coli 4.0 a a a b b b CON AS1 AS2 AS1: Ammonium sulfate; AS2: Ammonium sulfate slurry split; CON: Standard fertilization

33 CONCLUSIONS No E. coli O157:H7 or L. monocytogenes was detected from fertilizer, soil, foliar, or fruit sample. Salmonella was detected from some of fertilizer and early season soil samples. Coliform o Compost had the highest number of coliform. o Total coliform level in soil remains relatively stable over seasons Varied by years and berry fields o Coliform was very low in foliar sample and barely detectable in fruit samples Application of manure derived fertilizer had no effect on the soil coliform and food safety

34 ACKNOWLEDGEMENTS Dr. Margaret Drennan Randy Honcoop Farm Curt Maberry Farm

35 Go Cougs! Meijun Zhu School of Food Science (509)