Overview of Project

Objective: Evaluate trans-Pacific survival of FADs and endemic diseases through the use of surrogate viruses (when necessary), select feed ingredients and the transboundary model.

Hypothesis: Survival will depend on the correct combination of virus and ingredient.

Literature Review: Feed as a vehicle for PEDV transmission

PEDV was identified in the US during April 2013◦ Chen 2014

The US isolate is 99.5% similar to a Chinese variant◦ Huang 2013

PEDV can live in feed◦ Goyal 2014

PEDV can be transmitted by feed◦ Dee 2014

The MID in feed is 1 log◦ Schumacher 2015

PEDV survival is dependent upon the ingredient◦ Dee 2015

PEDV risk in feed can be reduced via chemical mitigation◦ Dee 2014, Cochrane 2015

Methods Virus Selection: Target Pathogen* (Surrogate)

1. Foot & Mouth Disease Virus (Seneca Valley Virus)

2.Classical Swine Fever Virus (Bovine Virus Diarrhea Virus)

3.Pseudorabies Virus (Bovine HerpesVirus-1)

4.Vesicular Exanthema of Swine Virus (Feline Calici Virus)

5. Nipah Virus (Canine Distemper Virus)

6. Swine Vesicular Disease Virus (Porcine Sapelovirus)

7. Vesicular Stomatitis Virus

8. PCV2

9. PRRS Virus 174

10. Influenza A Virus-Swine

11. African Swine Fever Virus

12. PEDV

* = SHIC swine disease matrix project

Transboundary Model

1. IngredientsUS Govt Harmonized Tariff ScheduleStandard quantities of ingredient (5g) & virus (1 x 105 TCID50)

2. Environmental chamberHistorical Temperature & % Relative Humidity data

3. Timetable: SeaRates.com37 day (Trans-Pacific)30 day (Trans-Atlantic, ASFV)

4. Sampling: 4 batches of ingredients = key cities & ports

5. Multiple metrics: PCR, VI, bioassay, T ½ (Bryan et al)

6. Controls: CF (+), CF (-), stock virus (no feed matrix)

Ingredients Imported: China to San Francisco*

Ingredients 2012 (Kg) 2013 (Kg) 2014 (Kg) 2015 (Kg) 2016 (Kg)Soy oil cake 15,126,647 7,977,560 13,545,880 24,201,390 36,962,316

DDGS 4,008,000 2,640,000 2,808,000 2,416,363 1,738,182

Pet food 4,075,353 3,068,722 623,734 51,587 1,412,165

Soybean meal 1,832,561 1,816,100 1,340,270 979,627 185,400

Pork sausage casings 129,365 216,845 457,427 420,005 582,093

Lysine 33,000 95,000 19,764 2,325,236 2,393,915

Choline 19,000 400 0 0 0

Vitamin D 26,000 21,000 14,000 0 0

TOTAL (Kg) 25,249,926 7,198,012 18,809,075 30,394,208 43,274,071

* US Govt. Harmonized Tariff schedule

Relationship of Trans-Pacific route and sampling points

Batch 1 (d 1 PI)

Batch 2 (d 8 PI)

Batch 3 (d 25 PI)

Batch 4 (d 37 PI)

Mean daily temperature & % RH data utilized during the Trans-Pacific model

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1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37

China Pacific USADec 23-30, 2012 Dec 31, 2012- Jan 15, 2013 Jan 16-28, 2013

C0%

RH

ASFV: Relationship of Trans-Atlantic route and sampling points

Batch 1 (d 1 PI)

Batch 2 (d 9 PI)

Batch 3 (d 19 PI)

Batch 4 (d 30 PI)

Mean daily temperature & % RH data utilized during the Trans-Atlantic model

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Atlantic Ocean New York City CLE: CHI: DSM

European segment Oceanic segment US segment

Warsaw-Hannover-Le Harve

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Study Period

Results: Summary of virus survival from batch 4 samples (37 DPI or 30 DPI) across both models.

IngredientSVA

(FMDV) ASFVPSV

(SVDV) PEDVFCV

(VESV) PCV2BHV-1(PRV)

PRRSV174

BVDV(CSFV) VSV

CDV(NiV) IAV-S

Soybean meal-Conventional (+) (+) (+) (+) (+) (-) (+) (+) (-) (-) (-) (-)Soybean meal-Organic (-) (+) (+) (+) (-) (-) (-) (-) (-) (-) (-) (-)Soy oil cake (+) (+) (+) NT (-) (-) (+) (-) (-) (-) (-) (-)DDGS (+) (-) (-) NT (-) (-) (-) (+) (-) (-) (-) (-)Lysine (+) (-) (+) (+) (+) (+) (-) (-) (-) (-) (-) (-)Choline (+) (+) (-) (+) (-) (+) (-) (-) (-) (-) (-) (-)

Vitamin D (+) (-) (+) (+) (-) (+) (-) (-) (-) (-) (-) (-)Moist cat food (+) (+) (+) NT (-) (-) (-) (-) (-) (-) (-) (-)Moist dog food (+) (+) (+) NT (-) (-) (-) (-) (-) (-) (-) (-)

Dry dog food (+) (+) (+) NT (-) (-) (-) (-) (-) (-) (-) (-)Pork sausage casings (+) (+) (+) NT (+) (-) (-) (-) (-) (-) (-) (-)Complete feed (+ control) (+) (+) (+) NT (+) (+) (-) (-) (-) (-) (-) (-)Complete feed (- control) (-) (-) (-) (-) (-) (-) (-) (-) (-) (-) (-) (-)Stock virus control (-) (+) (-) (-) (-) (-) (-) (-) (-) (-) (-) (-)

SVA (FMDV): D 37 PI (Batch 4 = Beijing to Des Moines )

Ingredient VI MEAN TITER T ½ (days)Soybean meal-Conventional Pos/Pos 10*4.5 9.7

Soybean meal-Organic Neg/Neg (NEG BIOASSAY) NT NT

Soy oil cake Pos/Pos 10*3.5 3.4

DDGS Pos/Pos 10*4.25 7.1

Lysine Pos/Pos 10*3.25 2.6

Choline Neg/Neg (POS BIOASSAY) NT NT

Vitamin D Pos/Pos 10*2.25 1.7

Moist cat food Pos/Pos 10*4.25 6.5

Moist dog food Pos/Pos 10*3.25 4.1

Dry dog food Pos/Pos 10*3.25 2.7

Pork sausage casings Pos/Pos 10*4.25 5.6

Complete feed (positive control) Pos/Pos 10*3.75 4.1

Complete feed (negative control)

Neg/Neg NT NT

Stock virus (positive control) Neg/Neg NT NT

FCV (VESV): D 37 PI (Batch 4 = Beijing to Des Moines )

Ingredient VI Mean titer T ½ (days)Soybean meal-Conventional Pos/Pos 10*4.8 26.6

Soybean meal-Organic Neg/Neg NT NT

Soy oil cake Neg/Neg NT NT

DDGS Neg/Neg NT NT

Lysine Pos/Pos 10*3.3 2.8

Choline Neg/Neg NT NT

Vitamin D Neg/Neg NT NT

Moist cat food Neg/Neg NT NT

Moist dog food Neg/Neg NT NT

Dry dog food Neg/Neg NT NT

Pork sausage casings Pos/Neg 10*3.7 3.5

Complete feed (positive control) Pos/Neg 10*2.3 1.9

Complete feed (negative control) Neg/Neg NT NT

Stock virus (positive control) Neg/Neg NT NT

PSV (SVDV): D 37 PI (Batch 4 = Beijing to Des Moines )Ingredient VI MEAN TITER T ½ (days)Soybean meal-Conventional Pos/Pos 10*3.2 2.7

Soybean meal-Organic Pos/Pos 10*3.2 2.7

Soy oil cake Pos/Pos 10*3.5 3.2

DDGS Neg/Neg NT NT

Lysine Neg/Neg NT NT

Choline Neg/Neg NT NT

Vitamin D Pos/Pos 10*3.5 3.5

Moist cat food Pos/Pos 10*3.3 3.1

Moist dog food Pos/Pos 10*3.7 3.8

Dry dog food Pos/Pos 10*3.3 2.8

Pork sausage casings Pos/Pos 10*2.8 2.2

Complete feed (positive control) Pos/Pos 10*3.2 2.7

Complete feed (negative control)

Neg/Neg NT NT

Stock virus (positive control) Neg/Neg NT NT

ASFV: D 30 PI (Batch 4 = Warsaw to Des Moines )Ingredient VI Mean titer T ½ (days)Soybean meal-Conventional Pos/Pos 10*3.0 1.3

Soybean meal-Organic Pos/Pos 10*3.1 1.8

Soy oil cake Pos/Pos 10*3.2 2.1

DDGS Neg/Neg (NEG BIOASSAY) NT NT

Lysine Neg/Neg (NEG BIOASSAY) NT NT

Choline Pos/Pos 10*3.2 2.2

Vitamin D Neg/Neg (NEG BIOASSAY) NT NT

Moist cat food Pos/Pos 10*3.0 1.3

Moist dog food Pos/Pos 10*2.8 1.8

Dry dog food Pos/Pos 10*2.8 1.8

Pork sausage casings Pos/Pos 10*2.9 1.9

Complete feed (positive control) Pos/Pos 10*2.9 1.9

Complete feed (negative control) Neg/Neg (NEG BIOASSAY) NT NT

Stock virus (positive control) Pos/Pos 10*3.0 1.8

PRRSV 174: D 37 PI (Batch 4 = Beijing to Des Moines )Ingredient VI Mean CtSoybean meal-Conventional Neg/Neg (POS BIOASSAY) 22.2

Soybean meal-Organic Neg/Neg (NEG BIOASSAY) 26.7

Soy oil cake Neg/Neg (NEG BIOASSY) 24.5

DDGS Neg/Neg (POS BIOASSAY) 24.3

Lysine Neg/Neg (NEG BIOASSY) 22.0

Choline Neg/Neg (NEG BIOASSY) 25.6

Vitamin D Neg/Neg (NEG BIOASSY) 24.5

Moist cat food Neg/Neg (NEG BIOASSY) 28.6

Moist dog food Neg/Neg (NEG BIOASSY) 28.7

Dry dog food Neg/Neg (NEG BIOASSY) 25.0

Pork sausage casings Neg/Neg (NEG BIOASSY) 24.5

Complete feed (positive control) Neg/Neg (NEG BIOASSY) 26.8

Complete feed (negative control) Neg/Neg (NEG BIOASSY) 38

Stock virus (positive control) Neg/Neg (NEG BIOASSY) 25.5

PCV2: D 37 PI (Batch 4 = Beijing to Des Moines )Ingredient VI Mean CtSoybean meal-Conventional NEG BIOASSAY 15.6

Soybean meal-Organic NEG BIOASSAY 15.9

Soy oil cake NEG BIOASSAY 15.7

DDGS NEG BIOASSAY 16.3

Lysine POS BIOASSAY 16.0

Choline POS BIOASSAY 18.1

Vitamin D POS BIOASSAY 17.1

Moist cat food NEG BIOASSAY 19.6

Moist dog food NEG BIOASSAY 17.4

Dry dog food NEG BIOASSAY 16.9

Pork sausage casings NEG BIOASSAY 16.4

Complete feed (positive control) POS BIOASSAY 16.6

Complete feed (negative control) NEG BIOASSAY 38

Stock virus (positive control) NEG BIOASSAY 16.2

BHV-1 (PRV): D 37 PI (Batch 4 = Beijing to Des Moines )Ingredient VI Mean titer T ½ (days)Soybean meal-Conventional Pos/Pos 10*2.7 2.2

Soybean meal-Organic Neg/Neg NT NT

Soy oil cake Pos/Pos 10*2.7 2.2

DDGS Neg/Neg NT NT

Lysine Neg/Neg NT NT

Choline Neg/Neg NT NT

Vitamin D Neg/Neg NT NT

Moist cat food Neg/Neg NT NT

Moist dog food Neg/Neg NT NT

Dry dog food Neg/Neg NT NT

Pork sausage casings Neg/Neg NT NT

Complete feed (positive control) Neg/Neg NT NT

Complete feed (negative control) Neg/Neg NT NT

Stock virus (positive control) Neg/Neg NT NT

BVDV (CSFV): D 37 PI (Batch 4 = Beijing to Des Moines )Ingredient VI Mean CtSoybean meal-Conventional Neg/Neg 23.4

Soybean meal-Organic Neg/Neg 26.6

Soy oil cake Neg/Neg 21.5

DDGS Neg/Neg 23.9

Lysine Neg/Neg 23.4

Choline Neg/Neg 24.4

Vitamin D Neg/Neg 26.6

Moist cat food Neg/Neg 21.7

Moist dog food Neg/Neg 21.6

Dry dog food Neg/Neg 23.1

Pork sausage casings Neg/Neg 21.1

Complete feed (positive control) Neg/Neg 29.5

Complete feed (negative control) Neg/Neg 38

Stock virus (positive control) Neg/Neg 27.6

IAV-S: D 37 PI (Batch 4 = Beijing to Des Moines )Ingredient VI Mean CtSoybean meal-Conventional Neg/Neg (NEG BIOASSAY) 25.8

Soybean meal-Organic Neg/Neg (NEG BIOASSAY) 26.2

Soy oil cake Neg/Neg (NEG BIOASSAY) 28.0

DDGS Neg/Neg (NEG BIOASSAY) 28.7

Lysine Neg/Neg (NEG BIOASSAY) 26

Choline Neg/Neg (NEG BIOASSAY) 30.1

Vitamin D Neg/Neg (NEG BIOASSAY) 29.7

Moist cat food Neg/Neg (NEG BIOASSAY) 35.3

Moist dog food Neg/Neg (NEG BIOASSAY) 29.4

Dry dog food Neg/Neg (NEG BIOASSAY) 28.6

Pork sausage casings Neg/Neg (NEG BIOASSAY) 26.6

Complete feed (positive control) Neg/Neg (NEG BIOASSAY) 31.6

Complete feed (negative control) Neg/Neg (NEG BIOASSAY) 38

Stock virus (positive control) Neg/Neg (NEG BIOASSAY) 31.5

ConclusionsUnder the conditions of this study:

1. Certain viruses survive in certain feed ingredients.

2. This affects multiple species.

3. Viral transport through feed may be a frequent event.

4. This work suggests the need for “feed biosecurity”, both domestically and globally.

CritiqueStrengths: Replication.Experimental design.Actual pathogen (n = 6).Proper controls.

Limitations:Model.Surrogates (n = 6).Actual contamination level in feed is unknown.

Discussion and Next Steps

We now propose a new hypothesis for the transboundary movement of pathogens: FEED.

◦ Missing information:◦ Oral infectious dose◦ Contamination level in field

◦ Responsible Importer concept (L. Minion):◦ Mitigation◦ Extended storage…. “Quarantine”◦ Sourcing decisions