Hepatitis E virus surveillance in Italy

1. Hepatitis E surveillance in Italy
Maria Elena Tosti
1st ECDC Hepatitis E virus expert panel meeting
ECDC Stockholm. 9th-10th December 2015
Centre for Epidemiology, Surveillance and Health Promotion
National Institute of Health - ISS, Rome - Italy

2. 1. routine surveillance in Italy
2. an integrated epidemiological, virological,
and environmental surveillance
3. results from a recent prevalence study
4. conclusions
Schedule

3. classification: A
B
nonA-nonB
unknown
Mandatory surveillance in Italy
Viral hepatitis is included in the national notifiable
infectious diseases surveillance system
By Ministry of Health
Diseases at high frequency and/or liable of control
interventions

4. classification: A
B
C
Delta
E
nonA-nonE
Voluntary surveillance in Italy
Since 1984
SEIEVA surveillance
Sistema Epidemiologico Integrato dell’Epatite Virale Acuta
(Integrated Epidemiological System for Acute Viral Hepatitis)
By National Institute of Health
Introduced in
2007

5. target cases: acute hepatitis
network: voluntary Local Health Units
interview by a standardized questionnaire
ascertainment of markers
data flow to coordination centre (ISS)
SEIEVA Methodology

6. 2/11
2/9
5/8
12/15
4/11
5/6
2/5
Local Health Units collaborating with SEIEVA
by December 31st 2014
N° of participating LHUs
151/189 = 79.9%
Covered population
43,951,734 = 77.2%
complete
partial
not participating

7. Acute hepatitis E: case definition
Based on clinical and serological criteria
Clinical criteria:
acute illness compatible with hepatitis
ALT > 10 times the upper limit of the normal range
Serological criteria:
IgM anti-HEV positive
IgM anti-HBc negative
IgM anti-HAV negative
Cases IgM anti-HEV positive, in absence of clinical signs, are
included among “acute hepatitis E cases”

8. Collected information
socio-demographic characteristics
exposure to faecal-oral risk factors (within 6 weeks
before disease onset)
laboratory parameters (ALT, AST, bilirubin, INR)
serological markers of infection
clinical course

9. Hepatitis N° cases %
A 5.174 50.3
B 3.514 34.2
C 876 8.5
E 144 1.4
nonA-nonE* 45 0.4
nonA-nonC** 153 1.5
Unknown 378 3.7
Total 10.285 100.0
* cases negative for hepatitis A, B, C and E
viruses
** cases negative for hepatitis A, B and C,
and not tested for IgM anti-HEV
Distribution of notified cases by hepatitis type.
SEIEVA 2007-2014
Hepatitis A Hepatitis B
Hepatitis C Hepatitis E
nonA-nonE nonA-nonC
Unknown
Hepatitis E

10. Distribution of notified hepatitis E cases by region.
SEIEVA 2007-2014
3
2 18
3
2
6
21
18
1717
37

11. acute hepatitis cases
where the presence of A, B
or C hepatitis viruses has
already been excluded
Geographical area
Test IgM anti-HEV
yes Total
N° (%) N° cases
North 83 (27.3) 304
Centre 123 (35.7) 345
South/Islands 4 (5.6) 71
Total 210 (29.2) 720
Distribution of IgM anti-HEV tests by geographical area and year
among possible acute hepatitis E cases. SEIEVA 2007-2014
0%
10%
20%
30%
40%
50%
60%
2007 2008 2009 2010 2011 2012 2013 2014
Year
Mean = 29.2%

12. Distribution of notified hepatitis E cases by age,
gender and nationality. SEIEVA 2007-2014
Characteristics N° (%)
Age
0-14 3 (2.1)
15-24 16 (11.1)
25-34 35 (24.3)
35-54 54 (37.5)
≥ 55 36 (25.0)
median (range) 40.5 (12-87)
Gender
Male 122 (84.7)
Female 22 (15.3)
Nationality
Italian 81 (56.6)
Other 62 (43.4)
Total n° of cases 144
Nationality N° (%)
Bangladesh 29 (46.8)
India 17 (27.4)
Pakistan 10 (16.1)
Bolivia, China,
Croatia, Morocco,
Peru, Romania
6 (9.7)
Total n° of cases 62 (100.0)

13. Most of the travel are
to the country of origin
Risk factors Italians Other p-value
Shellfish consumption 47.5 30.0 0.048
Raw 35.3 61.5 0.104
Drinking well water 9.1 38.8 <0.001
Travel to endemic areas 16.5 82.8 <0.001
Contact with a jaundice
case (within 6 weeks)
0.0 2.4 0.180
Total cases 81 62
Percent of notified hepatitis E cases who report exposure
to the listed risk factors by nationality. SEIEVA 2007-2014

14. Integrated epidemiological, virological, and environmental
surveillance of hepatitis E in Italy: emerging disease in
industrialized countries
Granted by MoH
ME Tosti ISS – National Centre for Epidemiology, Surveillance and Health Promotion
AR Ciccaglione ISS – Department of Infectious, Parasitic and Immune-Mediated Diseases
G La Rosa ISS – Department of Environment and Primary Prevention
L Romanò Milan University – Department of Biomedical Sciences for Health
Project conducted within:
the SEIEVA network
the Italian major reference center for viral hepatitis at ISS
the regional reference center for viral hepatitis at Milan University
Settings: 41 LHUs
Study population:
 all HEV cases from LHUs able to perform anti-HEV laboratory confirmation
 all nonA-nonC cases from LHUs not able to perform anti-HEV laboratory
confirmation
Years: 2012-2015

15. Study population Total cases
to be confirmed 126
E cases 30
Total 156
Study population
E cases 58 (37.2)
nonA-nonE cases 98 (62.8)
Total 156 (100.0)
IgM anti-HEV pos.
n (%)
28 (22.2)
30 (100.0)
58 (37.2)

16. Characteristics
Hepatitis
p-value
E nonA-nonE
Age % %
10-30 21.4 11.8
0.178
31-45 25.0 33.3
46-60 32.1 23.7
61-87 21.4 31.2
Gender
Male 84.5 55.7
<0.001
Female 15.5 44.3
Nationality
Italian 69.0 83.7
0.032
Other 31.0 16.3
Geogr. area of diagnosis
North 48.3 51.0
0.533Centre 48.3 41.8
South/Islands 3.4 7.1
Comparison between E and nonA-nonE cases: demographic characteristics

17. risk factors
Hepatitis
p-value
E nonA-nonE
% %
Travel 39.3 29.2 0.210
Shellfish consumption 56.8 40.4 0.119
Raw 38.9 0.0 0.095
Berries consumption 25.0 50.0 0.245
Drinking well water 12.9 9.5 0.708
Pork meat consumption 70.3 49.2 0.041
Sausage consumption 60.6 31.3 0.009
Wildfowl consumption 50.0 25.0 0.306
Comparison between E and nonA-nonE cases: risk factors

18. Study flow-chart with the results of molecular analysis
58 Hep. E
cases
45 tested for
RNA
40 HEV-RNA
positive
17
sequenced
16
G3
1
G4
23 not
sequenced
5 HEV-RNA
negative
13 not tested
for RNA
Croatian
travel to Ukraine
all Italians
no travels to endemic areas

19. Molecular analysis of nonA-nonC acute hepatitis cases sent
for diagnostic to the ISS laboratory. 2004-2015
124 cases
39 IgM anti-
HEV positive
(31.5%)
31 HEV-RNA
positive
8 HEV-RNA
negative
85 IgM anti-
HEV negative
2 HEV-RNA
positive
83 HEV-RNA
negative
all tested for
IgM anti-HEV and
HEV-RNA
33 HEV-RNA
positive
11
G1
6
G3
41
acute hepatitis E
on 124 nonA-nonC
33.1%

20. Geographic Information System (GIS) map of the
Wastewater Treatment Plants (WTPs) under study
 56 wastewater treatment plants
 717 sewage samples

21.  37 positive samples
5.2%
 21 Genotype 3
 16 Genotype 1
Geographic distribution of HEV-RNA positive samples

22. Epidemiological study performed in 2014:
Prevalence of HEV infection among blood donors
resident in Abruzzo, central Italy
• cross-sectional study February-March
2014
• serum samples from 313 blood
donors*
• tested for anti-HEV IgM, anti-HEV IgG,
HEV RNA
* unpaid voluntary donors, 18-68 years old, median
age 48; 98.7% Italian subjects, 80.5% male
Lucarelli C, … and Ciccaglione AR. High prevalence of anti-hepatitis E virus
antibodies among blood donors in central Italy. Euro surveillance in press

23. Laboratory results:
313 blood donors tested:
309 Italians; 4 foreign citizens
80.5% male; median age 48
• 153 positive for anti-HEV IgG: 48.9%
(95% CI 0.4-0.5) (Wantai Elisa kit)
• 2 positive for anti-HEV IgM: 1.3%
(95% CI 0.1-4.6) (Wantai Elisa kit)
• 2 positive for HEV RNA:
 blood donor 784: IgM/IgG + ; HEV RNA 100 IU/mL; G3
 blood donor 771: IgM/IgG - ; HEV RNA 10,000 IU/mL; G3

24. Prevalence of anti-HEV IgG by age group
0%
10%
20%
30%
40%
50%
60%
70%
<35 35-44 45-54 ≥55
HEV IgG positive, %
age (years)
(n=82) (n=104) (n=75)(n=54)ANTI-HEV IgG PREVALENCE INCREASED WITH AGE
However, high values were also found in the first age groups
suggesting that viral transmission takes place even at young age

25. age (years)
Epidemiological study performed in 2014:
As a control: prevalence of HEV infection among
blood donors resident in Lazio, central Italy
9 anti-HEV IgG positive: 9% (95% CI 0.43-0.54)
100 subjects, all Italians
89% male; median age 40

26. Multivariate analysis
Univariate Analysis Multivariateanalysis
Risk factor PR 95% CI P value APRR 95% CI P value
Sex (M/F) 1.6 0.9-3 0.097
Birth rural area 1.6 1-2.7 0.042
Work with animals 1.7 0.7-4.1 0.189
Contacts with cattle 1.5 0.9-2.4 0,111
Contacts with dogs 0.5 0.3-1 0,028
Kitchen gardening 1.6 1-2.6 0,052
Raw dried pork liver
sausage
4.00 2-8.3 0 2.35 1.5-3.8 0
vegetable
consumption of
own kitchen garden
1.4 0.8-2.5 0,197
usually vegetable
consumption of
own kitchen garden
1.5 0.9-2.4 0,103
the only risk factor independently
associated with past and present infection
was the consumption of
raw dried pork liver sausage
(air-dried sausages )
APRR: adjusted prevalence rate ratios
A special food habit of that area
supporting the hypothesis of
zoonotic transmission

27. Blood donor 771_ISS
Blood donor 784_ISS
Phylogenetic analysis
of HEV
typing assay: ORF1 (broad-range MTase assay)
blood donor 784_ISS
clustered with strains from Italian ENVIRONMENTAL SAMPLES
wastewater (central Italy: WTP1496; northern Italy: WTP1929,
WTP1577)
Sequences from 2 blood donors: 771_ISS and 784_ISS
clustered with GENOTYPE 3 GROUP along with:
strains from swine
possible zoonotic origin.

28. Phylogenetic analysis
of HEV
subtyping assay: ORF2 (capsid region)
Sequence from blood donor 771_ISS
• clustered with SUB-GENOTYPE 3c GROUP
the group includes other 7 strains:
• 5 swine: 3 from The Netherland, 2 from Italy
• 1 wild boar: from Germany
• 1 human: from France

29. in Italy, hepatitis E surveillance is limited to a voluntary
system, as SEIEVA
SEIEVA, at present, covers 77% of the Italian population
surveillance is affected by underreporting
HEV is not routinely investigated, as only few laboratories
perform anti-HEV tests
laboratory tests performance seems to be increasing
underreporting is probably also attributable to a high
percentage of asymptomatic cases
Conclusions

30. Conclusions
When the specific tests are performed, about 30% acute
hepatitis negative for HAV, HBV and HCV are attributable
to hepatitis E virus
in all the presented data, G3 and G1 are prevalent in Italy,
in human cases as in the environment
microbiological and environmental surveillance is
currently performed in Italy only within granted projects,
but …
in 2015 started a integrated epidemiological and
microbiological surveillance of acute viral hepatitis,
supported by the MoH