Submitted to: Clinical and Diagnostic Laboratory Immunology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: November 21, 2003
Publication Date: March 15, 2004
Citation: ALVAREZ, R., NJENGA, M., SCOTT, M.A., SEAL, B.S. DEVELOPMENT OF A NUCLEOPROTEIN-BASED ENZYME-LINKED IMMUNOSORBENT ASSAY UTILIZING A SYNTHETIC PEPTIDE ANTIGEN FOR DETECTION OF AVIAN METAPNEUMOVIRUS ANTIBODIES IN TURKEY SERA. CLINICAL AND DIAGNOSTIC LABORATORY IMMUNOLOGY. MAR 2004, P 245-249. Interpretive Summary: Avian metapneumoviruses (aMPV) cause an upper respiratory tract disease designated turkey rhinotracheitis (TRT). The disease in commercial turkeys does not usually cause death, but symptoms can be more severe when accompanied by secondary bacterial infections. Prior to 1996 the disease was not found in the United States. However, a new viral subtype different from European viruses is now present among commercial United States turkey flocks. Reagents utilized to detect these viruses are currently not standardized. Also, the United States virus is different enough from European varieties that new reagents must be created to more consistently detect presence of that agent. Based on physical characteristics of the aMPV nucleoprotein, a structural component of the virus, reagents were created to detect antibodies in sera from commercial turkeys. Portions of the nucleoprotein that were the same physically for all aMPV subtypes were chemically synthesized as small peptides. These peptides were used in a diagnostic assay to detect exposure of turkeys to any aMPV subtype and can be used to more reliably monitor presence of these viruses among commercial turkey flocks.
Technical Abstract: Avian metapneumoviruses (aMPV) cause an upper respiratory disease with low mortality, but high morbidity primarily in commercial turkeys that can be exacerbated by secondary infections. There are three aMPV serotypes of which the C type is found only in the United States. The aMPV nucleoprotein (N) amino acid sequences of serotypes A, B, and C were aligned for comparative sequence analysis. Based on predicted antigenicity of consensus sequences, five aMPV-specific N peptides were synthesized for development of a peptide-antigen, enzyme-linked immunosorbent assay (aMPV/N-peptide ELISA) to detect aMPV-specific antibodies among turkeys. Sera from naturally and experimentally infected turkeys were utilized to demonstrate the presence of antibodies reactive to the chemically synthesized aMPV N-peptides. Subsequently, the aMPV N-peptide 1 that had the sequence 10-DLSYKHAILKESQYTIKRDV-29 with variations at only three amino acids among aMPV serotypes was evaluated as a universal aMPV ELISA antigen. Data obtained with the peptide-based ELISA correlated positively with total aMPV viral antigen based ELISAs and the peptide ELISA provided higher optical density readings with less background. These results indicated that aMPV N-peptide 1 can be utilized as a universal ELISA antigen to detect antibodies for all aMPV serotypes.