Hepatitis E virus (HEV) infection in haemodialysis patients

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Dialysis. Transplantation. Hepatitis £ virus (HEV) infection in haemodialysis patients ... Laiko General Hospital;4Blue Cross Clinic;5Athinaiki Clinic;6Renal Unit,.
Nephrol Dial Transplant (1996) 11: 1093-1095

Nephrology Dialysis Transplantation

Original Article

Hepatitis £ virus (HEV) infection in haemodialysis patients M. Psichogiou1, E. Vaindirli1, E. Tzala1, S. Voudiclari2, J. Boletis3, G. Vosnidis3, S. Moutafis4, G. Skoutelis5, V. Hadjiconstantinou6, H. Troonen7, A. Hatzakis1, for the Multicentre Haemodialysis Cohort Study on Viral Hepatitis* 'Department, of Hygiene and Epidemiology, Athens University Medical School; 2Division of Nephrology, Hippokration General Hospital; 'Division of Nephrology, Laiko General Hospital; 4Blue Cross Clinic; 5Athinaiki Clinic; 6Renal Unit, Amalia Fleming Hospital, Athens, Greece; 7Abbott Laboratories, Abbott Park IL, USA and Wiesbaden, Germany

Abstract

Background. The aim of this study was to determine the prevalence of hepatitis E virus (HEV) infection among patients undergoing haemodialysis (HD) and to evaluate whether chronic haemodialysis is associated with an increased risk of HEV infection. Methods. Serum samples from 420 HD patients and 316 healthy volunteers were tested for IgG and IgM antibodies to HEV (anti-HEV). Anti-HEV testing was done by an enzyme immunoassay (ElA) based on recombinant proteins of HEV (Abbott Labs). All antiHEV IgG positive sera were confirmed using synthetic peptides. Results. Anti-HEV IgG was confirmed in 27/420 (6.4%) of the HD patients and in 7/316 (2.2%) of the reference group (P=0.007). However, multiple logistic regression analysis showed that the prevalence of anti-HEV IgG was not significantly higher in HD patients compared with the reference group, after controlling for age and sex. No patient was found positive for antiHEV IgM. The presence of anti-HEV was associated with sex in HD patients (/> = 0.04). No significant association was found between anti-HEV and underlying renal disease, anti-HCV, anti-HBc, blood transfusions, history of elevated transaminases, history of clinical hepatitis and renal transplantation. A marginal association, which was observed with the duration of haemodialysis in univariate analysis (P=0.07), was not confirmed in multivariate analysis. Conclusions. Chronic haemodialysis is not associated with an increased risk of exposure to HEV, and the

high prevalence of anti-HEV IgG in HD patients reported in uncontrolled studies is possibly due to the confounding effect of age and sex. Key words: haemodialysis; hepatitis E virus; IgG antiHEV Introduction

Hepatitis E virus (HEV) is considered to be the main aetiological agent of the enterically transmitted non-A, non-B hepatitis (ET-NANBH) [1]. It is generally believed that in developed countries only travellers in endemic areas face the threat of ET-NANBH. Since the cloning of HEV [2] and the development of serological assays for the detection of antibodies to recombinant expressed HEV antigens [3], our knowledge of the epidemiology of this infection has been enhanced. Seroprevalence studies conducted in developed countries suggest that among blood donors the prevalence of HEV infection ranges between 0.4 and 3.2% [4-6]. We recently identified two acute non-A, non-B hepatitis cases caused by HEV, in Greece, without an apparent association with endemic areas, suggesting that HEV infection may be endemic in a European population (J Hepatol (in press)). The possibility of parenteral transmission of HEV has been proposed [7]. An association with hepatitis C virus infection has been reported, pointing to similar or overlapping routes of transmission [8,9]. As a transient viraemic stage occurs during the infection, Correspondence and offprint requests to: Angelos Hatzakis, Nationalthe possibility of parenteral transmission cannot be Retrovirus Reference Centre, Dept of Hygiene and Epidemiology, ignored [10]. Athens University Medical School, Athens 11527, Greece. Patients undergoing chronic haemodialysis (HD) * Investigators and Institutions in the Multicentre Haemodialysis Cohort Study on Viral Hepatitis: A. Hatzakis, A. Katsoulidou, M. potentially have an increased risk of exposure to parenPsichogiou, E. Vaindirli (Athens Univ. Medical School); G. terally transmitted agents. A higher prevalence of IgG Vosnidis, J. Boletis, D. Stamatiadis (Laiko General Hospital); S. antibodies to HEV has already been reported among Voudiclari, Ch. Priftis, M. Koulousiou (Hippokration General this group of patients [11]. Our study aimed to deterHospital); S. Moutafis, K. Ifandi, I. Thanou (Kianous Stavros Clinic); G. Skoutelis, K. Triandafilou, M. Matzaropoulou (Athinaiki mine the prevalence of HEV infection among HD Clinic); V. Hadjiconstantinou, D. Arvanitis, E. Logothetis (A. patients and to evaluate whether chronic haemodialysis Fleming Hospital). is associated with an increased risk of HEV infection. © 1996 European Dialysis and Transplant Association-European Renal Association

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Subjects and methods A total of 420 HD patients, followed up in five different haemodialysis units in Athens, Greece, were included in this study. Of these patients 235 were males and 185 females, with a mean age of 57.3 + 14.1 (SD) (range 16-84) years. The mean duration of haemodialysis was 61.4 + 52.9 (SD) (range 1.5-225.7) months. The most common causes of their renal failure were glomerulonephritis («=139), polycystic kidney disease (« = 54), diabetic nephropathy (n = 37), pyelonephritis (« = 27), undefined cause (n = 66), while other causes included nephrolithiasis, nephrosclerosis, lupus nephritis, etc. Among HD patients 64% had received blood transfusions, 5% reported a history of clinical hepatitis at least 6 months before the initiation of this study, while 18% had a history of elevated transaminases. Ten percent had a history of renal transplantation. As a reference group 316 healthy volunteers (266 males, 50 females) were recruited from occupational screening. Antibodies to HCV (anti-HCV) were detected by thirdgeneration enzyme immunoassay (Ortho Diagnostic Systems Inc, Raritan, NJ). Antibodies to hepatitis B virus core antigen (anti-HBc) were detected by Corzyme (Abbott). Antibodies to HEV (anti-HEV) were detected by an enzyme immunoassay based on two recombinant antigens (SG-3 and 8-5) derived from two different open reading frames (ORFs) of the HEV Burmese strain [12]. The recombinant antigen SG-3 is composed of 327 amino acids from the carboxy-terminus of ORF2, which encodes the major structural proteins of HEV [13-15]. The recombinant antigen 8-5 which consists of 123 amino acids represents the full length of ORF3. Each sample was tested for anti- HEV IgG. Specimens with absorbance values greater than or equal to the cut-off value were considered initially reactive and were then repeated in duplicate. Specimens that were repeatedly reactive were tested using synthetic peptides spB4-2, spB3-2, in order to confirm anti-HEV reactivity. All anti-HEV IgG positive sera were also tested for IgM antibodies by an enzyme immunoassay (EIA). Anti-HEV IgM reactivity was validated by DTT treatment for isotype and by synthetic peptide based EIA for antigenic specificity. Bivariate analysis was carried out by x2 a n d by Fisher's exact probability test. Multivariate analysis was done by logistic regression. A series of models were fitted with antiHEV as the dependent variable, and sex, age, unit, and duration of haemodialysis introduced into the models as independent variables.

Results Anti-HEV IgG was detected and confirmed in 27/420 (6.4%) of HD patients and in 7/316 (2.2%) of the healthy controls (i> = 0.007). Reactivity against spB4-2 (ORF3) was found in 29/34 (85%) anti-HEV IgG(+) individuals, and against spB3-2 (ORF2) in 12/34 (35%). None of the tested individuals was found positive for anti-HEV IgM. Multiple logistic regression analysis showed that the prevalence of anti-HEV IgG was not significantly higher in haemodialysis patients compared with the reference group (P=0.652), after controlling for age and sex. However, there was a statistically significant association with age (P = 0.024) (Table 1).

M. Psichogiou el at. Table 1. Multiple-logistic-regression-derived relative risk (RR) and 95% confidence interval (CI) for anti-HEV IgG in HD patients vs reference group, adjusted for age and sex Variable Group Reference group Haemodialysis patients Sex Male Female Age (years)

RR

95% CI

P

1.00 1.28

— 0.43-3.82

— 0.652

1.00 1.40 1.03

0.87-2.23 1.00-1.06

0.162 0.024

Within the haemodialysis group a higher prevalence was observed in females (P = 0.04). No significant correlation was found between anti-HEV and antiHCV, anti-HBc, blood transfusions, history of elevated transaminases, history of clinical hepatitis, and the occurrence of renal transplantation, while a marginal association was observed with the duration of haemodialysis (P=0.07). The prevalence of antiHEV was higher in older haemodialysis patients but the difference did not reach statistical significance (Table 2). Also no association was found with renal disease (data not shown). In order to evaluate whether duration of haemodialysis and haemodialysis unit were associated with the prevalence of anti-HEV IgG, a logistic regression analysis was performed. No association was found with either variables (Table 3). Table 2. Univariate analysis of risk factors associated with HEV infection in 420 haemodialysis patients Characteristics

n

Age (years) 31 =£34 35-59 170 > 60 190 Sex Female 185 Male 235 Anti-HCV Negative 276 Positive 144 Anti-HBc Negative 231 Positive 189 Duration haemodialysis 100 < 20 months 21-40 81 >41 months 116 Blood transfusions None 153 1-9 198 >10 40 History of elevated transaminases Yes 77 No 314 History of clinical hepatitis Yes 21 No 371 Renal transplantatioi Yes 41 No 356

Anti-HEV (+) (%)

P

0.234 10(5.9) 15(7.9) 17(9.2) 10(4.3)

0.041

17 (6.2) 10 (6.9)

0.755

12(5.2) 15(7.9)

0.254

3 (3.0) 10(12.3) 7 (6.0)

0.074

11(7.2) 11 (5.6) 3(7.5)

0.741

2 (2.6) 23 (7.3)

0.129

1 (4.8) 24(6.5)

0.755

3(7.3) 22 (6.2)

0.777

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HEV in haemodialysis patients Table 3. Multiple-logistic-regression-derived relative risk (RR) and 95% confidence interval (CI) for anti-HEV IgG on HD patients Variable

RR

95% CI

Duration haemodialysis (months) Age (years) Sex Male Female Hospital Clinic 1 (reference) Clinic 3* Clinic 4 Clinic 5

1.00 1.03

0.99-1.01 0.99-1.06

0.603 0.166

1.00 2.11

— 0.90-4.97

— 0.085

1.00 2.36 0.42 0.45

— 0.75-7.80 0.09-2.04 0.14-1.52

— 0.140 0.286 0.202

of exposure to HEV, and the high prevalence of antiHEV IgG in HD patients found in uncontrolled studies is possibly due to the confounding effect of age and sex.

P

* Clinic 2 was not included since no anti-HEV IgG (+) patients were found.

Discussion The prevalence of anti-HEV IgG was considerably higher in haemodialysis patients (6.4%) than in the reference group (2.2%), although after controlling for age and sex the association was eliminated. The distribution of antibodies in the two different epitopes of the Burmese proteins, suggests that the B4-2 epitope, found in 85% of the tested individuals, is the most useful antigen in detecting anti-HEV [4]. According to our results a significant association was found between the presence of anti-HEV IgG and older age. This is in accordance with previous reports from Turkey that older age was significantly associated with anti-HEV IgG [8]. Similar findings were reported in other studies from Southern Italy [9] and the former Soviet Union [16]. The correlation of HEV with age may reflect a cohort phenomenon due to infection acquired some decades ago. Anti-HEV IgG can persist for a long time [17,18], and it is possible that some water-borne outbreaks that occurred earlier in the 20th century were actually hepatitis E and not A [19]. On the basis of the high prevalence of anti-HEV found in haemodialysis patients by Halfon and colleagues [11], it was suggested that orofaecal transmission may not be the only route of transmission of HEV and that patients with a high risk for B and C virus infections could also be infected by HEV. However, in our study the high prevalence of anti-HEV IgG in that group of patients was shown to be due to the confounding effect of age and sex. A logistic regression analysis showed that neither duration of haemodialysis, nor other variables related with haemodialysis were associated with HEV. In conclusion, this study suggests that chronic haemodialysis is not associated with an increased risk

Acknowledgements. The authors would like to thank Ms Aphrodite Leonditsi for typing the manuscript.

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