Antigenic epitope composition and protectivity of avian hepatitis E virus (avian HEV) ORF2 protein and vertical transmission of avian HEV

Avian HEV was first isolated in chickens from the USA that had hepatitis-splenomegaly syndrome in 2001. Based on genetic identity and genomic organization, avian HEV has been classified into the genus Hepevirus, family Hepeviridae, which also includes human and swine HEVs. Avian HEV ORF2 protein was predicted to have common antigenic epitopes shared by avian, human and swine HEV, but its detailed antigenic epitope composition and protectivity have not been investigated. Vertical transmission of avian HEV in chickens also remains to be evaluated;To map HEV common and non-common epitopes on avian HEV ORF2 protein, nine synthetic peptides from the predicted four antigenic domains of the avian HEV ORF2 protein were synthesized and corresponding rabbit anti-peptide antisera were generated. With the use of recombinant ORF2 proteins, convalescent pig and chicken antisera, peptides and antipeptide rabbit sera, at least one epitope in domain II that is unique to avian HEV, one epitope in domain I that is common to avian, human and swine HEVs, and one epitope in domain IV that is shared between avian and human HEVs were identified;To determine if the capsid ORF2 protein of avian HEV can be protective and used as vaccine, twenty chickens were immunized with purified recombinant avian HEV ORF2 protein with aluminum as adjuvant, and another twenty chickens were mock immunized with KLH precipitated in aluminum. After challenge, all the tested mock-immunized control chickens developed typical avian HEV infection but not in the tested chickens immunized with avian HEV ORF2 protein;To identify neutralizing epitopes on avian HEV ORF2 protein, four Mabs (7B2, 1E11, 10A2, 5G10) against this protein were generated and characterized. 1E11, 10A2 and 5G10 were shown to bind to bona fide avian HEV particles in vitro, and partially neutralize virus in an animal based neutralization assay. The corresponding neutralization epiotpes were further localized by Western blot with the use of five avian HEV ORF2 recombinant proteins;Avian HEV was detected in egg white samples. Avian HEV contained in egg white was infectious as evidenced by viriema, fecal virus shedding and seroconversions in the chickens inoculated with avian HEV PCR positive egg white, but not in PCR negative egg white inoculated chickens. However, vertical transmission of avian HEV in chickens was not proved;The present studies pave the way for future avian HEV vaccine design and the development of differential immunoassays for the diagnosis of HEV cross-species infection. The results also implicate avian HEV infection in chicken could be a model system for studying HEV viral immune response, but not for studying HEV vertical transmission

Recovery Techniques For Finite Element Methods And Their Applications

Recovery techniques are important post-processing methods to obtain improved approximate solutions from primary data with reasonable cost. The practical us- age of recovery techniques is not only to improve the quality of approximation, but also to provide an asymptotically exact posteriori error estimators for adaptive meth- ods. This dissertation presents recovery techniques for nonconforming finite element methods and high order derivative as well as applications of gradient recovery. Our first target is to develop a systematic gradient recovery technique for Crouzeix- Raviart element. The proposed method uses finite element solution to build a better approximation of the exact gradient based on local least square fittings. Due to poly- nomial preserving property of least square fitting, it is easy to show that the new proposed method preserves quadratic polynomials. In addition, the proposed gra- dient recovery is linearly bounded. Numerical tests indicate the recovered gradient is superconvergent to the exact gradient for both second order elliptic equation and Stokes equation. The gradient recovery technique can be used in a posteriori error estimates for Crouzeix-Raviart element, which is relatively simple to implement and problem independent. Our second target is to propose and analyze a new effective Hessian recovery for continuous finite element of arbitrary order. The proposed Hessian recovery is based on polynomial preserving recovery. The proposed method preserves polynomials of degree (k + 1) on general unstructured meshes and polynomials of degree (k + 2) on translation invariant meshes. Based on it polynomial preserving property, we can able to prove superconvergence of the proposed method on mildly structured meshes. In addition, we establish the ultraconvergence result for the new Hessian recovery technique on translation invariant finite element space of arbitrary order. Our third target is to demonstrate application of gradient recovery in eigenvalue computation. We propose two superconvergent two-grid methods for elliptic eigen- value problems by taking advantage of two-gird method, two-space method, shifted- inverse power method, and gradient recovery enhancement. Theoretical and numer- ical results reveal that the proposed methods provide superconvergent eigenfunction approximation and ultraconvergent eigenvalue approximation. In addition, two mul- tilevel adaptive methods based recovery type a posterior error estimate are proposed