Serological Immunoassay for Detection of Hepatitis E Virus on the Basis of Genotype 3 Open Reading Frame 2 Recombinant Proteins Produced in Trichoplusia ni Larvae

Abstract

Hepatitis E virus (HEV) is a major cause of acute hepatitis in humans, and strains of genotypes 1 and 2 are endemic in many regions with suboptimal sanitary conditions. In many industrialized countries, HEV strains of genotype 3 are highly endemic in swine, and an increased number of autochthonous infections with HEV genotype 3 strains have been reported lately. Serological studies of HEV infection are often conducted with commercial assays based on peptides and recombinant proteins of HEV genotype 1 and 2 strains. For some patients with proven HEV genotype 3 infections, these assays failed to detect specific antibodies, and they are not applicable or validated for the detection of anti-HEV antibodies in swine. To elucidate the incidence of hepatitis E in regions where HEV genotype 3 infections can be expected, and to study the seroprevalence of HEV in swine, new tools with broad specificity for all genotypes of HEV are needed. We present the expression and partial characterization of recombinant HEV genotype 3 open reading frame 2 (ORF-2) proteins and their usefulness as diagnostic antigens in detecting anti-HEV antibodies in humans and swine with proven HEV genotype 3 infections. The recombinant antigens were produced at relatively high yields and at low cost upon infection of Trichoplusia ni larvae with recombinant baculoviruses expressing recombinant HEV genotype 3 ORF-2 proteins. The enzyme-linked immunosorbent assay based on the recombinant proteins showed good specificity and sensitivity for anti-HEV genotype 3 immunoglobulin G detection in human and swine sera. These recombinant HEV genotype 3 ORF-2 proteins might be added to diagnostic kits containing HEV genotype 1 and 2 antigens in order to develop a broadly sensitive new tool for the diagnosis of hepatitis E.

Head-to-Head Comparison of Three Vaccination Strategies Based on DNA and Raw Insect-Derived Recombinant Proteins against Leishmania

“…We have shown that insect-derived recombinant proteins produced in insect larvae used as living biofactories, even in its raw form, are useful for diagnosis [25], [26], [27], [28], [29], [30], [31], including that of leishmaniasis [32], [33], and vaccination [34], [35], [36], [37].…”

Restricted Enzooticity of Hepatitis E Virus Genotypes 1 to 4 in the United States

“…(29). Most of these EIAs employ for antibody capture antigens that are derived from HEV genotypes 1 and 2 and so may not be sufficiently sensitive to detect heterotypic antibodies generated in human and animal hosts infected by genotype 3 (19,24,45,48) or genotype 4 (2,23,62). Moreover, some indirect EIAs tend to generate false-positive reactivities (52,65) and therefore require, in order to validate the specificity of their reactivities, the extra steps of neutralization, immunoblotting, or prior production of genus-specific positive controls (1,12,35,45).…”

“…Moreover, in most assays, the antigens used for antibody capture are derived from HEV genotype 1 or genotypes 1 and 2. Because the HEV genotypes thus far identified to infect animals belong instead to genotype 3, genotype 4, and the newer putative genotypes, the possibility that low antibody detection rates reflect weak binding of antibody to heterotypic antigens rather than true antibody absence (2,19,24,44,45,48,62) cannot be excluded.…”

First Serological Study of Hepatitis E Virus Infection in Backyard Pigs from Serbia

“…However, most of these studies have been conducted using HEV ELISA that are based on genotype 1 or 2 peptides and are designed for humans, but not for swine (Pintó and Saiz 2007). Actually, these assays sometimes failed to detect specific antibodies in sera from individuals with proven HEV gt3 infections Jiménez de Oya et al 2009b). …”