Detection of HEV in Estonia Anna Ivanova, Irina Golovljova, Valentina Tefanova
National Institute for Health Development Department of Virology Zoonotic pathogens Tick-borne pathogens: TBEV, Borrelia sp., Rickettsia sp., Babesia sp., Ehrlichia sp. Rodent-borne pathogens: Hantaviruses Hepatitis viruses HAV, HBV, HCV, HEV
Hepatitis E virus (HEV) Family: Hepeviridae Genus: Hepevirus HEV genome Small, non-enveloped virus ssrna (+) Hepatitis E: An emerging disease M. Pérez-Gracia et al. Infection, Genetics and Evolution, Volume 22, 2014, 40-59
Geographical distribution of hepatitis E virus genotypes. S. Mirazo, et al Hepat Med. 2014; 6: 45 59.
Transmission routes of HEV Hepatitis E: An emerging disease M. Pérez-Gracia, et al. Infection, Genetics and Evolution, Volume 22, 2014, 40-59
Aims To assess HEV seroprevalence level in different human populations in Estonia To assess the presence of HEV infection in Estonian domestic pig and wild boar populations
Materials Human serum samples Retrospective samples Samples collected for the current study Ambulatory pt. without any known infectious desease (AP; 2003) 151 Acute nona-nonc (1994-2000) 762 Health care workers (2004) 163 Hemodialysis pat. (1996-2006) 176 IDU (2009-2010) 205 Veterinarians 158 Pig farm workers 67 Hunters 144 Samples from domestic pigs and wild boars Serum samples from adult pigs ( 1,5 y) 380 Fecal samples from pigs (2 4 months) 449 Meat juice from wild boars 471
Location of 17 farms where samples were collected 1 - Serum samples of pig farm workers 2 - Serum samples of domestic pigs 3 Fecal samples from pigs
Collection of serum samples from hunters and wild boars 21/68 7/48 7/4 3/53 15/16 1/37 9/13 3/23 30/46 2/30 10/41 13/30 1/28 4/34
Methods Serological assays Human samples: ELISA: Confirmatory IB test: recomwell IgG and IgM (Mikrogen GmbH, Germany) recomline HEV IgG/IgM (Mikrogen GmbH, Germany) Pig serum and wild boar meet juice samples ELISA: PrioCHECK HEV porcine kit (Prionics AG, Switzerland) Detection of viral RNA qpcr for samples screening (Jothikumar et al., 2006) Samples positive in qpcr amplified in the ORF2 region (1011 nt)
% of HEV IgG/IgM positive samples Detection of antibodies against HEV in Estonian human population 24 22 20 18 16 14 12 * 10 8 6 4 IgM IgG * * * * 2 0 Pig farm workers Ambulatory Healthy Hemodialysis Hunters Healthcare patients workers IDU nona-nonc Veterinarians
% of HEV IgG positive samples Anti-HEV IgG seroprevalence in different age groups 10 * 8 6 4 * 2 0 17-40 41-60 >61
HEV antibodies and HEV RNA in domestic pigs and wild boars Hiiumaa F12 (50%) Saaremaa 13% 25% F5 (100%) Läänemaa F11 (58,3%) 4% F10 (80,6%) F pig farm (seroprevalence %) 13% 30% F3 (100%) F2 (71,4%) F1 (27,3%) 26% Pärnumaa Harjumaa 19% Raplamaa Seroprevalence in wild boar HEV RNA in domestic swine 11% Järvamaa 33% Viljandimaa Lääne- Virumaa Valgamaa F17 (38,5%) Jõgevamaa 23% 4% Tartumaa 7% F6 (35,7%) F7 (50%) Ida-Virumaa 12% Põlvamaa 7% Võrumaa 21% HEV RNA in wild boar F14 (60%) F16 (75%) F15 (85%) F13 (61,2%)
Phylogenetic tree based of Estonia HEV sequences on the partial ORF2 HEV sequences from wild boar HEV sequences from domestic pigs 99 0.2 JF443726 DQ459342 95 98 66 KF691584 GENOTYPE 1 82 90 94 FJ998016 3e wild boar, Germany AB780453 3e wild boar, Japan JX912476 3e human, Russia HM055578 3e domestic pig, Hungary HQ380118 domestic pig, Russia Est343 Est91 99 Est187 Est346 Est90 Est112 F9 4-5 F2 3-30 II F14 4-25 F14 4-29 F14 3-18 99 F14 4-27 F14 3-19 F14 4-26 AB291961 human, Japan KC145143 3f domestic swine, Spain KC961258 3f human, Belgium EU514956 3f human, France HM641365 3f domestic swine, Sweden 99 F16 4-12 F16 4-26 HQ380077 domestic pig, Russia HQ399148 domestic pig, Russia Est442 F4 3-21 100 F4 3-30 100 F3 2-10 81 F3 3-16 F8 4-8 F8 4-23 88 F11 3-15 99 F11 4-6 F6 4-19 98 F6 2-4 94 F6 4-2 92 F6 2-15 IV JQ013794 3c human, France KJ701409 3i human, France KF152884 3b human, Brazil AF060668 3a human, USA AY115488 3j domestic pig, Canada AB220975 GENOTYPE 4 100 82 M74506 GENOTYPE 2 I III GENOTYPE 3
Geographical distribution of HEV lineages F2 F3 F6 Harjumaa Lääne- Virumaa Ida-Virumaa Hiiumaa Saaremaa F11 F8 Läänemaa Järvamaa Raplamaa Jõgevamaa Pärnumaa Viljandimaa Tartumaa Põlvamaa Valgamaa Võrumaa F4 F14 F16 F9 Lineage 1 Lineage 2 Lineage 3 Lineage 4 Sequences from wild boars Sequences from domestic pigs
Conclusions HEV circulates in Estonia Domestic pigs and wild boars are possible reservoirs and source of human HEV infection Seroprevalence in pig farm workers appears to be highest among the analysed groups All Estonian HEV strains from animals belong to genotype 3, but form 4 separate lineages
Acknowledgments Arvo Viltrop, Brian Lassen, Tiiu Saar, Department of Basic Veterinary Science and Population Medicine, Estonian University of Life Science Jevgenia Epstein, Kuulo Kutsar, Natalia Kerbo Public Health Board of Estonia The study was financially supported by the European Regional Development Fund, Programme TerVe, Projects ZoonRisk and EKZE