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A previously unrecognized virus belonging to the subfamily Pneumovirinae and most closely related to murine pneumovirus (MPV) was identified in domestic dogs in 2 related animal shelters. Additional diagnostic testing yielded 3 new viral isolates and identified 6 additional PCR positive dogs from other USA locations indicating that its distribution is not geographically limited. Nucleotide sequences encompassing 9 of the 10 genes were compared to the only 2 available MPV strains, 15 and J3666. Several features distinguished the canine pneumovirus (CnPnV) from the murine strains. Two regions of diversity were identified in the amino-proximal region of P and the overlapping P2 ORF was only 54 amino acids (aa) compared to 137aa in MPV. The G protein had an amino-terminal cytoplasmic tail 18aa longer than in the MPV strains. The CnPnV SH protein showed the highest divergence with only 90.2% aa identity when compared to MPV strain 15. Like strain 15, the CnPnV SH ORF coded for a protein of 92aa while J3666 has a 114aa variant. Comparison of CnPnV isolates at culture passages 4 and 17 revealed 7nt differences within the 8598nt sequenced. Of note was a substitution at nt 364 in G resulting in a termination codon that would produce a truncated G protein of 122aa. Analysis of early passage and ex vivo samples showed the termination codon in G to be predominant after 6 days in culture indicating rapid selection of the mutation in A72 cells.

The objective of the current study was to determine the capability of 3 recently described one-step TaqMan real-time reverse transcription polymerase chain reaction (real-time RT-PCR) assays targeting the nucleoprotein (NP), matrix (M), and hemagglutinin (HA) genes of H3N8 Equine influenza virus (EIV NP, EIV M, and EIV HA3 assays, respectively) to detect Canine influenza virus (CIV). The assays were initially evaluated with nucleic acid extracted from tissue culture fluid (TCF) containing the A/canine/FL/43/04 strain of Influenza A virus associated with the 2004 canine influenza outbreak in Florida. The EIV NP, EIV M, and EIV HA3 assays could detect CIV nucleic acid at threshold cycle (Ct) values of 16.31, 23.71, and 15.28, respectively. Three assays using TCF or allantoic fluid (AF) samples containing CIV ( n = 13) and archived canine nasal swab samples ( n = 20) originally submitted for laboratory diagnosis of CIV were further evaluated. All TCF and AF samples, together with 10 nasal swab samples that previously tested positive for virus by attempted isolation in embryonated hens' eggs or Madin–Darby canine kidney cells, were positive in all 3 real-time RT-PCR assays. None of the 3 assays detected the H1N1 Swine influenza virus strain in current circulation. These findings demonstrate that previously described real-time RT-PCR assays targeting NP, M, and H3 HA gene segments of H3N8 EIV are also valuable for the diagnosis of CIV infection in dogs. The assays could expedite the detection and identification of CIV.

To determine which respiratory viruses circulate among confined dogs, we analyzed nasal and pharyngeal swab specimens from shelter dogs with acute respiratory disease. An unknown virus was isolated. Monoclonal antibody testing indicated that it was probably a pneumovirus. PCR and sequence analysis indicated that it was closely related to murine pneumovirus.