Identification of a truncated nucleoprotein in avian metapneumovirus-infected cells encoded by a second AUG, in-frame to the full-length gene

Rene Alvarez,Bruce S Seal

doi:10.1186/1743-422x-2-31

Abstract

BackgroundAvian metapneumoviruses (aMPV) cause an upper respiratory disease with low mortality, but high morbidity primarily in commercial turkeys. There are three types of aMPV (A, B, C) of which the C type is found only in the United States. Viruses related to aMPV include human, bovine, ovine, and caprine respiratory syncytial viruses and pneumonia virus of mice, as well as the recently identified human metapneumovirus (hMPV). The aMPV and hMPV have become the type viruses of a new genus within the Metapneumovirus. The aMPV nucleoprotein (N) amino acid sequences of serotypes A, B, and C were aligned for comparative analysis. Based on predicted antigenicity of consensus protein sequences, five aMPV-specific N peptides were synthesized for development of peptide-antigens and antisera.ResultsThe presence of two aMPV nucleoprotein (N) gene encoded polypeptides was detected in aMPV/C/US/Co and aMPV/A/UK/3b infected Vero cells. Nucleoprotein 1 (N1) encoded from the first open reading frame (ORF) was predicted to be 394 amino acids in length for aMPV/C/US/Co and 391 amino acids in length for aMPV/A/UK/3b with approximate molecular weights of 43.3 kilodaltons and 42.7 kilodaltons, respectively. Nucleoprotein 2 (N2) was hypothesized to be encoded by a second downstream ORF in-frame with ORF1 and encoded a protein predicted to contain 328 amino acids for aMPV/C/US/Co or 259 amino acids for aMPV/A/UK/3b with approximate molecular weights of 36 kilodaltons and 28.3 kilodaltons, respectively. Peptide antibodies to the N-terminal and C-terminal portions of the aMPV N protein confirmed presence of these products in both aMPV/C/US/Co- and aMPV/A/UK/3b-infected Vero cells. N1 and N2 for aMPV/C/US/Co ORFs were molecularly cloned and expressed in Vero cells utilizing eukaryotic expression vectors to confirm identity of the aMPV encoded proteins.ConclusionThis is the first reported identification of potential, accessory in-frame N2 ORF gene products among members of the Paramyxoviridae. Genomic sequence analyses of related members of the Pneumovirinae other than aMPV, including human respiratory syncytial virus and bovine respiratory syncytial virus demonstrated the presence of this second potential ORF among these agents.

Highlights

Avian metapneumoviruses cause an upper respiratory disease with low mortality, but high morbidity primarily in commercial turkeys
We present evidence for utilization of a secondary open reading frame, within the N gene encoding a truncated nucleoprotein (N2) among Avian metapneumoviruses (aMPV)/C/Co and aMPV/A/UK/3b infected cells
Avian metapneumovirus N gene possess several putative AUG start sites The aMPV/C/US/Co nucleoprotein is encoded by the N gene with a predicted molecular weight of 42–45 kD [7,19]

Summary

Introduction

Avian metapneumoviruses (aMPV) cause an upper respiratory disease with low mortality, but high morbidity primarily in commercial turkeys. During 1997, mortality due to aMPV infections among commercial turkeys in the U.S ranged from zero, to 30% when accompanied by bacterial infections, with condemnations due to air sacculitis. This was the first reported outbreak of aMPV infections in the U.S which was previously considered exotic to North America. The virus causing disease was designated a new aMPV type C genetically different from European counterparts [3,4,5] and was subsequently demonstrated to be most closely related to human metapneumovirus (hMPV) from diverse geographic locations [6,7]. Infections among commercial turkeys with aMPV/C continue in the north-central U.S resulting in substantial economic loss to the poultry industry [6,8,9]

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