Prevalence of specific antibody to hepatitis E virus in the general population of the community of Madrid, Spain

INTRODUCTION

Hepatitis E virus (HEV) is an enterically transmitted agent causing acute hepatitis in humans. HEV strains cluster into four genotypes that might represent two separate evolutionary lineages, although all are considered as belonging to a single species because of lack of significant antigenic diversity. Genotypes 1 and 2 infect human beings exclusively and spread as waterborne epidemics by the oral-fecal route. Genotypes 3 and 4 infect mammals from different orders, and they can infect humans occasionally after consumption of contaminated liver meat or shellfish, and perhaps by other yet unknown routes [Lewis et al., 2010]. An additional HEV genotype has been found in rabbits [Zao et al., 2009], as well as a novel species, closely related to HEV, in Norwegian rats [Johne et al., 2010].

Although acute hepatitis E due to locally acquired infection by genotype 3 HEV strains has been reported from many European countries, the incidence of the disease in Europe is largely unknown. The prevalence of the HEV infection is also unknown for most countries of the region. Data obtained on samples representing the general population of the Lazio region of Italy, the Republic of San Marino and England and Wales reported frequency rates of antibody to HEV (anti-HEV) from 1.5 to 13% [Rapicetta et al., 1999; Vulcano et al., 2007; Ijaz et al., 2009]. The lowest prevalence was obtained in San Marino after retesting by a recombinant immunoblot test (RIBT) samples reactive for anti-HEV by enzyme-immunoassay (EIA) [Rapicetta et al., 1999]. In the British study, which was the largest study, the highest rate (13%) was recorded, and a significant increase of the frequency in relation to age was observed (25% in people aged over 50 years) [Ijaz et al., 2009].

Data obtained in Spain on samples representing the general population have been reported from Catalonia only. The prevalence observed ranged from 7.3% among adults to 4.6% among children aged 6–15 years [Buti et al., 2006, 2008]. Rates about 2% among unselected pregnant women [Suárez et al., 2004] and 2.5% among blood donors [Bernal et al., 1995; Mateos et al., 1999] were, in addition, recorded from other regions. RIBT was used to confirm samples reactive after EIA testing only in the study performed in blood donors in Granada [Bernal et al., 1995].

The prevalence rates found in Europe have been, therefore, moderate or low, but they do not fit well the scarcity of reports about patients displaying locally acquired acute hepatitis E in the region. The findings suggest that most acute HEV infections in Europe might be symptomless, but also that anti-HEV testing might be impaired by specificity issues [Shata and Navaneethan, 2008]. Cross-reactive antibody elicited by some unknown, non-pathogenic, HEV-related agent might be a potential cause, but other causes cannot be excluded. The wide experience obtained from screening antibody to hepatitis C virus in blood banks suggest that RIBT confirmation of samples reactive by EIA testing would mitigate the problem [Herremans et al., 2007].

A better understanding of the epidemiology of HEV in Europe requires an accurate estimation of the prevalence of the HEV infection among the general population. The objective of the present study was to record the frequency of anti-HEV among the inhabitants of the region of Madrid, as well as to assess the use of RIBT as a confirmatory test.

RESULTS

Fifty samples (2.17%) reacted for total anti-HEV after EIA testing (Table I). Anti-HEV IgG or IgM was detected by RIBT in 25 of these samples (50%; 1.08% of the 2,305 samples collected). Both EIA and RIBT reactivity rates did not show significant differences in regard to any of the parameters recorded from participants during the interview, but both showed a significant trend to increasing with age (linear χ² trend test: P < 0.001).

The frequency of samples reactive by EIA but negative by RIBT did not show a significant trend to increase with age observed for the total of samples reactive by EIA and for the samples confirmed by RIBT (linear χ² trend test: P < 0.05). The yield of RIBT confirmation of samples reactive in EIA was higher among the individuals aged above 20 years than among the younger of 21 years (19/31, 61.2% vs. 6/19, 31.6%; χ² = 4.16, P < 0.05). The yield of anti-HEV EIA testing was, in addition, significantly higher among the former (6/1,409, 0.4% vs. 19/896, 2.1%; χ² = 14.66, P < 0.001), and was less than 1% both among children aged less than 10 years (0.3%) and among adolescents (0.6%).

DISCUSSION

Confirmation by RIBT of samples reactive for anti-HEV after EIA testing has been recommended to improve diagnosis of HEV in settings of low endemicity [Herremans et al., 2007]. Following that recommendation, the overall prevalence of anti-HEV observed in the present study on a collection of samples from the general population of the Community of Madrid (1.08%) would be close to the prevalence recorded in San Marino (1.5%) under the same technical conditions [Rapicetta et al., 1999]. The highest prevalence was found among individuals older than 20 years (1.97–2.20%), and this figure was close to the frequency observed in San Marino among participants older than 40 years (2.1–2.5%). It represented, however, less than one tenth of the prevalence reported for England and Wales after testing the samples by EIA only [Ijaz et al., 2009].

Though these wide differences might in part be attributed to the exclusion of true anti-HEV positive samples after RIBT testing because of sensitivity issues, three findings from the present study argue against this interpretation. First, RIBT confirmation of EIA-reactive samples was twice more frequent among adults than among adolescents and children, and the difference was significant. Second, the yield of RIBT confirmation increased significantly with age, as did the overall prevalence of anti-HEV assessed by EIA. Third, the rate of EIA-reactive, RIBT-negative samples did not display a trend to increase with age, as should be expected if this pattern reflects the inability of RIBT to detect low amounts of anti-HEV after years of decay from the acute infection. The lowest number of confirmed results by RIBT was obtained, in addition, among individuals younger than 20 years (31.6%). Primary infections by Epstein–Barr virus and cytomegalovirus are frequent in this population group, and both conditions have been shown to interfere anti-HEV testing [Fogeda et al., 2009].

According to the results obtained after RIBT testing in the present study, contact with HEV seems very uncommon among children and adolescents of the Community of Madrid, and most infections seem to take place among persons older than 20 years. These observations agree with the data recorded from patients with locally acquired, acute hepatitis E in Spain [Echevarría et al., 2011], and are similar to the conclusions drawn from studies performed in the United Kingdom [Ijaz et al., 2009], the Bolivian Amazon [León et al., 1999], and Bangladesh [Labrique et al., 2009], though the conditions for HEV transmission would be expected to be very different in these three regions of the World. Results from the United Kingdom were interpreted as reflecting an increased spread of HEV under the exceptional circumstances of the Second World War, but such interpretation would not be valid for the present study, because sample collection was undertaken 17 years later. Specific factors favoring the spread of HEV among adults might, therefore, exist, and they might be working in regions of the World as diverse in culture, hygiene conditions, and economic development as Western Europe, Bangladesh, and the Amazon. Alimentary habits might account among them, but the participation of other factors cannot be excluded.

In conclusion, HEV infection is as prevalent in the Community of Madrid as in San Marino, less frequent than in Lazio and Catalonia, and much less frequent than in the United Kingdom. Contact with HEV seems very uncommon among children and adolescents, and the opportunity for infection would increase after the age of 20. Confirmation of EIA-reactive samples by RIBT reduces the final numbers of anti-HEV testing as much as 50%. Whether such testing protocol provides a better approach to the real prevalence of anti-HEV in settings of low endemicity such as Madrid remains to be demonstrated, though some findings of the present study would support it.