Hepatitis E is (re-)emerging in the Netherlands. HEV RNA is present in domestic pigs, wild boars, meat products, sewage, surface water, and drinking water in the Netherlands. The prevalence of HEV among blood donors and patients is increasing. However, the reasons for the increasing human cases and the relative importance of different transmission pathways such as the environment or food are still unknown. Upcoming patient-control studies aim to identify risk factors for HEV infection and clinical disease.

1. Hepatitis E
(Re-)Emerging in the Netherlands
Presented by Wilfrid van Pelt

2. CIb-RIVM
EPI Agnetha Hofhuis, Wilfrid van Pelt, Roan Pijnacker,
Rody Zuidema, Susan Hahné, Sofie Mooij
IDS Erwin Duizer, Harry Vennema
Z&O Saskia Rutjes, Martijn Bouwknegt
LCI Madelief Mollers, Jim van Steenbergen
NVWA (Dutch FSA) Aloys Tijsma, Ingeborg Boxman
Sanquin Hans Zaaijer, Boris Hogema
Anses (FR) Nicole Pavio
VVVL (BE) Isabel De Boosere
Bfr (DE) Reimar Johne
Acknowledgements
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

3. ● Considered a travelers disease until the end of the last century
● 1 serotype, genotypes 1-4, with different transmissionroutes
Kamar et al. Lancet 2012; 379: 2477–88
Hepatitis E

4. Hepatitis E

5. Kamar et al. Lancet 2012; 379: 2477–88
● Liverinfection (O/B)
● Mostly asymptomatic, or
very mild complaints.
● Symptoms look like
hepatitis A:
jaundice, fever, abdominal
pain, fatigue…
● Underlying disease increases
risk symptomatic infection,
serious course and chronic
infection (GB, NA/PTS)
Guillain-Barré, Neuralgic amyotrophy/Parsonage-Turner S
●
Hepatitis E
(A) Acute icteric hepatitis
(B) Chronic infection
(C) Drug-induced injury
(D) Neurological injury
(E) Miscellaneous clinical
syndromes
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

6. Sanquin bloeddonor tests, 18-21 jaar, Hogema, 2014, Transfusion
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015
HEV Inreasing in NL?

7. Virological laboratories 2012-2015 (patients)
Viremie Sanquin donoren
1:1800 (‘13) 1-800 (‘14,’15)
Viraemic Sanquin donors
1:1800 (‘13) 1-800 (‘14,’15)
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015
HEV Inreasing in NL?

8. All large lab’s show
increase 2014
Many labs started in
2015 after joining
the Ca-Co study
0 50 100
Erasmus MC Rotterdam
Izore Friesland
LUMC
UMC Groningen
Streeklab Hengelo
UMC Utrecht
Vir Infectieziekten Groningen
HagaZiekenhuis
Veldhoven
Diakonessenhuis Utrecht
Streeklab Zeeland
Rijnstate Ziekenhuis Arnhem
VieCuri Venlo
UMC Nijmegen
Vumc
Maasstad Ziekenhuis Rdam
AZM Maastricht
MCH
GGD Amsterdam
AMC Adam
Streeklab Tilburg
SSDZ Delft
Laurentius Ziekenhuis Roermond 2012
2013
2014
2015
RIVM: all type 3
(3a,3c(mainly),3e,3f)
No regional clusters
No molecular clusters
HEV Inreasing in NL?

9. Seroprevalence
● Nederland
– Bloeddonors Sanquin 2012: 26,7% (23,6%-30,5%)
› 5239 sera
› Wantai anti-HEV IgG
– Livestock Farming & Resident Health Study (VGO), 2013-2014
› ~2500 sera residents
› Wantai anti-HEV IgG
› Preliminary seroprevalence 28,7%
● Europe
› Large differences, also dependent on assay
● France (october 2011)
› Dependent geography 20% - 71,3% Ariège Zuid Frankrijk
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015
No association proximity of pig livestock farms
Age: OR 20-30yrs vs ≥60 yrs 5.43 [2.14-13.78]
Sex: OR Males vs Females 1.36 [1.16-1.64]

10. % HEV RNA positive donations in Europe
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

11. HEV RNA positive blood donors
a) Baylis,Vox 2012; b) Vollmer, JCM 2012; c) Slot, EuroSurv. 2013; d) Cleland, Vox 2013;
e) Hewitt, Lancet 2014; f) Gallian, EID 2014; g) Sauleda, Transfusion 2014; h) Fischer, PlosOne 2015;
i) Ikeda, ISS 2009; j) CBS Surveilance Report 2014;
k) pers. communication J.S. Epstein; l) Hogema, Transfusion 2015

12. What we know about sources in the Netherlands
● HEV RNA in animals
– Pigs (>90% seropositive; 50% RNA positive)
– Wild game, Veluwe (wild boar 25% seropositive, 8% RNA positive, deer lower)
– HEV RNA shown in oysters and mussels, Oosterschelde (5%)
› Other countries in oysters, mussels, cockles
– Not shown in faeces from muskrat (2007), veals (1999), dairy cattle (1999) and
poultry (1999)
● HEV RNA in food
– Pigmeat (6% in livers, 50% meats tested infectious, 2% meats in EU-study)
– Wildgame (prevalence in liver and meats <5% in NL, higher abroad)
– Raspberries (frozen) EU study (not NL) 3%
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

13. USA (Feagins et al., 2007) 11% RNA+; 2/3 infectious
The Netherlands (Bouwknegt et al., 2007) 6.5%
Japan (Yazaki et al., 2003) 1.9%
Korea (Jung et al., 2007) 10.8%
India (Kulkarni et al., 2008) 0.83%
France (Rose et al 2011) 4%
Germany (Wenzel JJ, 2011) 4%
Italy (Di Bartolo et al 2012) 6%
Canada (Wilhelm et al 2014) 3,5%
Pork Liver contaminated with HEV : grocery stores, slaughterhouse
USA 26% (Meng et al 2002)
Sweden 13% (Olsen et al 2006)
Moldavia 51.1% (Drobeniuc et al 2001)
The Netherlands 11% (Bouwknegt et al 2007)
France 44% (Chaussade et al 2013)
Germany 32 % (Krumbholz et al 2014)
Direct contact with infected animals. High HEV seroprevalence in personnel with
swine occupation (veterinarians, butchers, slaughterhouse).
Pork Liver sausages contaminated with HEV
France (Pavio et al 2014) 30%
Germany (Johne R communication) 25%
Italy (Di Bartolo et al 2014) 22%
Sources abroad
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

14. HEV RNA in the environment in the Netherlands
● Drinking water
HEV RNA shown in 2 water reservoirs with partly cleaned drinking water
● Sewage
– HEV RNA shown in slaughterhouse sewage
› Other countries in city sewages and sewage of pig farms
● Surfacewater
– In NL shown in the river Maas (17%)
› Other countries in irrigation water
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

15. Genetic relationship of HEV in the Netherlands
3c
3a
3f
3e
Prototype
Pig
Pig liver
Wild boar
Water
Human
Prototype
Pig
Pig liver
Wild boar
Water
Human
0 - 3 nt
4 - 11 nt
12 - 15 nt
16 - 20 nt
0 - 3 nt
4 - 11 nt
12 - 15 nt
16 - 20 nt
2004-2006
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

16. ● HEV-infection is caused by consumption and/or direct or indirect
contact with specific (pig-)(meat-)products
Patient-Control studies
Working Hypothesis
Pilot patient-control comparison
(n=50 patients, n=147 age-sex matched control person (continuous survey))
● Patients predominantly middle aged man
● People with underlying disease more often get a symptomatic HEV
infection (Diabetes, Cardio-vascular disease, Immunodeficiency, Rheumatism)
● Medication, Meat and Food, Animal Contact: unclear
Riskfactor study, ideal
● Difference HEV IgM/PCR positive……IgG negatives
● Difference patients……asymptomatic viremic cases (medicatie-mod/cnt)
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015
Vegetarians (bd 400) seroprevalence 15% lower at all ages (Sanquin p.c.)

17. Patients from 21 virological lab’s:
– Cases: HEV positive IgM or RNA Controls: HEV IgG negative
– June 2015 - june 2017
– Questionnair: from Laboratory –to- Treating Physician –to- Patient
– Sample: to RIVM for IgG testing and genotyping op IDS/VIR
– ~15 invitations / month Cases; ~30 invitations / month Controls
– ~50% expected respons
Participants from Livestock Farming & Resident Health Study (VGO)
– Cases: HEV positive IgG Controls: HEV IgG negative
– June 2013 - june 2014; 2500sera and questionnairs
Blood donors Sanquin (predominantly middle aged man)
– Cases: HEV positive IgG Controls: HEV IgG negative
– November 2015 (Februari 2016)
– 2500sera and questionnairs; 60% expected respons
Patient-Control studies
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

18. ● Questionnair: Complaints, Medication (pig derived) Medical history, General
health, SES, Education, Age, Residence
– Exposure (2month before illness):
 travel history  fruit & vegetables profession
 meat consumption  eating outside outdoor activities
 shellfish/crustacean  water contact
 rawmilk products  animal contact
Extra Controls from the general population
– No serological testing
– 100 invitations / month, general control study
Pig meat products
in high detail
Patient-Control studies
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

19. “Hide(meat) Glue”
●Used for glueing pieces of meat:
› Steaks, Ham, Beefsteak, Pork tenderloin
●Thrombine and Fibrinogen derived from Cattle/Pig blood.
Dutch FSA approved animals (not-tested for HEV)
●No HEV-inactivating production step
●Endproduct not tested for HEV
●Risk-control: now (erronously) retailer/consumer
●Mandatory on productwrapper “Pig Protein” “Varkenseiwit”
●Production 2014: 175.000 kg
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

20. Spray Dried Porcine Plasma (SDPP)
● Hide (meat) Glue ≠ SDPP
● SDPP production (spray drying ~15 seconds 220 – 80 C):
Inactivation HEV?
● SDPP is used for:
● binding fats and water
● colouring food products
● Animal feed (piglets)
● Total production 2014 SDPP (1 big producer):
13.000.000 kg
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

21. Spray Dried Porcine Plasma (SDPP)
● Pujols et al. Virology Journal 2014, 11:232:
› 100 % tested SDPP batches HEV AB present
› 22,4% HEV RNA +
› HEV RNA in SDPP batches fed to pigs: no HEV seroconversion…….
– Meaning for Hide/Meat Glue and other pig blood derived
products?
Action plan Dutch FSA, Hide Glue:
●HEV incorporating in the “foodsafetyplan”
●Monitoring-project, starting in 2016
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015

22. Recent (re-)emerging of Hepatitis E cases in the Netherlands,
among blood donors as well
HEV RNA is present / found in the Netherlands in:
› domestic pigs and wild boars in NL
› meat products: pigmeat and wildgame in NL
› Sewage, surface water and drinking water in NL
Unknown:
› Why HEV is increasing in humans
› Role Hide/Meat Glue and SDPP in foods
› Relative importance of different transmission pathways (env/food)
› Risk factors for developing clinical disease (also dose-resp)
› Inactivation of HEV in foods
› Stability/Infectivity of HEV in food. Infectivity testing is difficult
Conclusions
ECDC- HEV-Exp meeting , Stockholm, 9,10 Dec 2015