monitoring and controlling viral contamination of shellfish · monitoring and controlling viral...

Monitoring and controlling viral

contamination of shellfish

Bill Doré

Marine Institute -National Reference Laboratory

Food safety challenges: Controlling Foodborne Viruses. Dublin Dec1st 2015

1

Presentation Overview

Food safety challenges: Controlling Foodborne Viruses. Dublin Dec1st 2015 2

Why do we have a problem with viruses in bivalve

molluscan shellfish?

e.g. Mussels, cockles, & oysters

Current control measures

Progress and challenges for setting an international

regulatory standards for viruses in bivalve shellfish

Method standardisation

Balancing Industry Impact and health benefits

Risk management procedures used by the shellfish industry.

3

Shellfish, Sewage and Public Health

Shellfish growing areas may also be impacted by sewage

Shellfish feed by filtration

Sewage-derived pathogens are concentrated

Shellfish are eaten raw or lightly cooked

Outbreaks of disease can occur

4

Shellfish European Control Measures

Control of sewage pollution in

shellfisheries

Classification and monitoring

of harvesting areas

Commercial processing

(depuration, relaying, cooking)

End-product controls

(quality tests, traceability)

EU

reg.

2004/8

53

EU

reg.

2004/8

54

WF

Directive

2000/6

0/E

C

EU

reg.

2004/8

53

5

Controls in Europe have virtually

eliminated bacterial illness associated

with shellfish consumption. However

the risk of virus illness remains and

outbreaks of illness continue to

occur throughout Europe……….


Virus illness outbreaks caused by oysters

6

Scandinavia, Jan 1997 (J. of Shell. Res. 17(5):1633-1635)

– Gastro-enteritis associated with oysters

– Denmark: 356 cases recorded (Sweden, Netherlands and Germany)

– Approx. 35,000 contaminated oysters sold in Denmark alone

Europe Jan/March 2010 (Euro Surveill. 15(12): 1-5)

– 334 cases in 65 clusters in Sweden, UK, Norway, France and Denmark

– Oysters produced in France, UK and Ireland

– Winter 2010 cold and high gastro-enteritis activity in community


Virus illness outbreaks caused by oysters

7

Scandinavia, Jan 1997 (J. of Shell. Res. 17(5):1633-1635)

– Gastro-enteritis associated with oysters

– Denmark: 356 cases recorded (Sweden, Netherlands and Germany)

– Approx. 35,000 contaminated oysters sold in Denmark alone

Europe Jan/March 2010 (Euro Surveill. 15(12): 1-5)

– 334 cases in 65 clusters in Sweden, UK, Norway, France and Denmark

– Oysters produced in France, UK and Ireland

– Winter 2010 cold and high gastro-enteritis activity in community


Norovirus

Relatively mild gastroenteritis (2-4 days)

nausea, diarrhoea, vomiting, fever and abdominal pain

The most common cause of infectious intestinal disease in the community

Seasonal distribution “Winter Vomiting Disease”

Coincides with peak periods for oyster consumption

Genetically diverse virus with two main genogroups (GI, GII) and many genotypes that infect humans.

Detection methods

8


Norovirus

Food safety challenges: Controlling

Foodborne Viruses. Dublin Dec1st 2015

9

Person to person spread

Highly infectious (<10 virus particles)

Easily spread in closed communities

Cruise ships, hospitals, care homes, schools…..

Vomiting Larry

Role of shellfish in the spread of NoV?

Global trade contaminated with multiple strains

Shellfish (oysters)

Discharge to the environment

person-to-person spread

Virus shedding in feaces:

104-1010 viral particles/g

new variant

recombination

10



11

Virus Controls in shellfish

12

Despite the recognised risk there are currently no

EU regulatory standards for viruses in shellfish

Until recently there has been a lack of reliable

methods for the detection and quantification of

viruses in shellfish (and other foods)

NoV not culturable

Sensitivty (low target numbers)

Strain diversity


NoV detection in shellfish ISO/TS 15216-1: published March 2013

Quantitative real-time RT-PCR method for norovirus (GI & GII) and hepatitis A in shellfish

Developed by European working group (CEN) lead by EURL and including MI

Also bottled water, hard surfaces, soft fruit and salad vegetables

Marine Institute -accredited to ISO17025 standard in 2010 for the real-time RT-PCR method

An internationally recognised standard method is the first step towards establishing a standard for Nov in shellfish?

13 Food safety challenges: Controlling Foodborne Viruses. Dublin Dec1st 2015



14

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2g of digestive glands, add equal

vol. of proteinase K, 37°C and

60 and centrifuge

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Norovirus Controls-European Context

15

EFSA scientific opinion on Norovirus in oysters:

methods, limits and control options 2012

www.efsa.europa.eu/en/efsajournal

“….risk managers should consider establishing an acceptable

limit for NoV in oysters to be harvested and placed on the

market”

European Commission DG-Sanco Working Group on

Live Bivalve Molluscs

Includes discussions on virus control measures (standards)

NoV detection by PCR–what does it mean?

16

Detects short genome fragments from both infectious and non-infectious virus particles

NoV often detected in oysters in the absence of any apparent illness (outbreaks)

UK study 76% of oysters contain NoV but 52% of these oysters contained concentrations <LOQ of the assay

Lowther et al (2012) Appl. Environ Microbiol. 2012 Aug; 78(16):5812-7

This makes setting a standard (acceptable limit) for NoV in oysters difficult

Impact on industry vs public health benefit


17

Cost

(indust

ry im

pac

t)

Benefit

(food s

afety

)

Calculating the balance

Benefit (difficult) Cost (relatively easy)

Dose Response NoV concentration in oysters

Outbreak data European baseline survey

EU wide Baseline Study

18

EFSA & EURL are currently establishing protocols for European wide survey across shellfish areas to establish the distribution of NoV in oysters across European shellfish.

Aim

To gather information on the extent of NoV contamination in European oysters

Establish the impact for industry of the introduction of a NoV standard in Europe

Will not establish a safe or acceptable limit for NoV in shellfish

will not gather illness data

will not establish a threshold of infection

will not determine exposure levels for consumers


Survey of Market Ready Oysters

19

Project with UCD (part funded by Food for Health Research Initiative

Contract 07FHRIT AFRC5)

8 companies/suppliers

C. gigas (1 site both C. gigas and O. edulis)

Depurated or Category A

Monthly sampling between Jan 2012 and March

2013

Total of 114 samples


NoV (GI & GII) concentration in market ready oysters

20

203 348 150 1548 120

Overall Percentage detection rates (n=114)

Not detected Positive >200 >500 >1000

61.5 38.5 22.0 7.9 5.3


21

Cost

(indust

ry im

pac

t)

Benefit

(food s

afety

)

Calculating the balance

Benefit (difficult) Cost (relatively easy)

Dose Response NoV concentration in oysters

Outbreak data European baseline survey

Dose response (published information)

22

A few published human volunteer studies with a range of

50% human infectious dose reported

18-1,015 genome copies (Teunis et al 2008)

1,320-2,800 genome copies (Atmar et al 2014)

Published outbreak data

Host susceptibility and immunity

Secretor positive individuals

Genotype Differences?


Outbreak Information

Date No. ill NoV Levels (GI+GII) Comments

Feb

2010

>70 Restaurants 2,350 & 2040 A number of illnesses clusters in

Ireland and UK Harvest area 2,890 &1,920

Jan

2012

18 Restaurant 2,380 Dublin Restaurant

Harvest area 4,000

Feb

2012

20 Restaurant 1,446- 4,413 Illness in Denmark

Restaurant results from Danish NRL.

RASSF Harvest area 1,074

Nov

2012

<10 Restaurant 433 Galway Restaurant.

Dredged Oysters Harvest area 926; 2,775


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0

500

1000

1500

2000

2500

3000

3500

4000

NoV copies g-1

Sample (n=162)

NoV (GI & GII) Concentration in Market Ready

Oysters and Oysters Associated with Outbreaks

• Threshold of infection

remains undetermined

• Dose response –Increasing

concentration of NoV =

increasing risk of apparent

infection (outbreaks)


FIRM Funded Project

NORORISK -developing a risk

assessment framework for NoV in Irish oyster areas

25

Risk Assessment Project

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Three year project funded under FIRM with UCD and

Marine Institute

Establish a Risk Assessment Framework for Norovirus in

Irish Oyster Production Areas

Use a sample area to develop a framework

Develop exposure model using data from sample area

Assess impact of management interventions (e.g. depuration)

on NoV concentrations

Run published dose response models to estimate disease

Framework will provide a model that can be applied to

other areas


NoV concentrations in individual

oysters • Measured NoV (GII) concentration in 30 individual oysters

from one growing bag

• Repeated at low (~LOQ), medium and high NoV

concentrations

• Estimate potential range of consumer exposures based on

individual oyster concentrations estimate

27

28 0

500

1000

1500

2000

2500

3000

3500

4000

0 10 20 30

0

100

200

300

400

500

600

700

800

900

1000

0 10 20 30

0

50

100

150

200

250

300

350

400

0 10 20 30

Concentration of NoV GII per g in individual shellfish

Mean=88.2

Range=0-355

Mean=328.8

Range=89-892

Mean=1248.6

Range =220-3741


Consumer Exposure

Oysters

eaten

Copies/g = 85 Copies/g = 325 Copies/g = 1250

2 146

(0 - 428)

598

(238 - 1138)

2516

(855 - 4821)

4 293

(47 - 661)

1196

(627 - 1941)

5062

(2521 - 8184)

6 442

(130 - 885)

1794

(1076 - 2712)

7605

(4374 – 11341)

8 589

(214 - 1089)

2388

(1544 - 3430)

10116

(6362 - 14422)

10 734

(308 - 1273)

2988

(2057 - 4121)

12612

(8417 - 17466)

95% interval for Norovirus genome copies consumed per meal size

Food safety challenges: Controlling Foodborne Viruses. Dublin Dec1st 2015 29

30

Drivers for NoV standards in shellfish

Food Safety (regulation)

outbreaks

EFSA opinion

DG Sanco

WG


31

Drivers for NoV standards in shellfish

Food Safety (regulation)

outbreaks

EFSA opinion

DG Sanco

WG

Business

outbreaks

retailers

importing countries


Risk Management

32

MI worked with a number of producers to implement risk management procedures

1. Characterisation of harvest areas Identification of pollution risks in production areas

NoV concentrations (monitoring)

2. Identification of high risk periods Season, community outbreaks and high rainfall events

Nov monitoring (increase in NoV concentrations)

3. Determine effectiveness of post harvest treatment depuration (elevated temp and extended time)

Relay in areas of clean seawater?


Dep.Area 2Area 1

5000

4000

3000

2000

1000

0

Ge

no

me

co

ncn

etra

tio

ns p

er g

Boxplot of of NoV concentrations in oysters


Viruses & Shellfish –Summary (1)

34

Sewage contamination of bivalve shellfish is a significant public health concern

EU regulation controls bacterial risks but there remains a risk from viruses

In Ireland the most clearly identified risk is NoV in oysters

A standardised quantitative test method for the detection of NoV is available (ISO/TS 15216-1)

Robust and reliable method to quantify NoV in oysters (and other foods)

does not distinguish between non-infectious and infectious virus particles


Viruses & Shellfish –Summary (2)

35

A recognised need to set a regulatory standard for NoV in oysters Threshold of infection is not known but a dose response exists

(increasing genome copies increasing risk)

Balance of cost (industry impact) against benefit (food safety gains)

EU wide baseline study to establish industry impact

UCD/MI risk assessment project

Industry risk management practice Identify control points points and develop risk management

plans

NoV monitoring is of major risk management tool

Export market key driver


Acknowledgements

36

Marine Institute Sinead Keaveney

Agnieszka Rupnik

Leon Devilly

UCD (risk assessment project) Francis Butler

Kevin Hunt


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