ABSTRACT
OBJECTIVE: To isolate and characterise avian paramyxoviruses and other haemagglutinating viruses amongst Victorian wild bird populations. PROCEDURE: Tracheal and cloacal material was collected from wild duck, pigeon, quail and other wild birds throughout Victoria. Samples were processed and cultured in embryonating eggs. Viral isolates were characterised based on their haemagglutination and haemagglutination-inhibition activity using a panel of specific antisera. Reverse transcriptase polymerase chain reaction and DNA sequencing were used to characterise Newcastle disease virus isolates. RESULTS: Twenty-five nonpathogenic haemagglutinating viruses were isolated from 605 wild bird samples. The majority were characterised as APMV-6 or influenza A virus, H3N2. Two isolates were identified and characterised as APMV-1 (avirulent NDV) based on nucleotide and deduced amino acid sequence analysis at the F0 cleavage site. CONCLUSIONS: Twenty-five viruses were isolated, none of which resembled progenitor or virulent genotypes. This study provides valuable epidemiological information against which to compare future isolates from outbreaks of disease to determine their origin.
Subject(s)
Avulavirus Infections/veterinary , Avulavirus/isolation & purification , Bird Diseases/epidemiology , Birds , Influenza A virus/isolation & purification , Animals , Animals, Wild , Avulavirus Infections/epidemiology , Bird Diseases/etiology , Bird Diseases/virology , Cloaca/virology , Columbidae , DNA Primers , DNA, Viral/analysis , Ducks , Newcastle Disease/epidemiology , Quail , RNA, Viral/analysis , Reverse Transcriptase Polymerase Chain Reaction/veterinary , Trachea/virology , Victoria/epidemiologyABSTRACT
Respiratory syncytial virus (RSV) causes severe lower respiratory tract infection in children and calves. Antibodies to ovine RSV (ORSV) are common in sheep, but the clinical disease is not well defined. There is no report of ORSV infection in Australian sheep although respiratory distress syndrome has been described. This discrepancy may be due to the lack of a suitable diagnostic test. In this report, we have characterised the ORSV G protein in an attempt to study its relatedness to human and bovine RSV (HRSV, BRSV) and for use in the development of a suitable diagnostic assay. Full length and a truncated variant of ORSV G protein were expressed in mammalian cells and the expressed proteins characterised by indirect immunofluorescence and radioimmunoprecipitation assays. Our results indicate that like HRSV, the ORSV G protein is heavily glycosylated. The expressed protein was membrane bound as well as secreted and could be purified from culture supernatants and may be suitable for use in development of a diagnostic assay.
Subject(s)
HN Protein , Respiratory Syncytial Virus Infections/veterinary , Respiratory Syncytial Viruses/isolation & purification , Sheep Diseases/diagnosis , Animals , Australia , Base Sequence , Cattle , Cell Line , Child , Cloning, Molecular , Diagnosis, Differential , Fluorescent Antibody Technique, Indirect , Humans , Molecular Sequence Data , Radioimmunoassay , Respiratory Syncytial Virus Infections/diagnosis , Respiratory Syncytial Virus, Bovine , Respiratory Syncytial Virus, Human , Respiratory Syncytial Viruses/genetics , Sheep , Sheep Diseases/virology , Transfection , Viral Envelope Proteins , Viral Proteins/analysis , Viral Proteins/geneticsABSTRACT
The respiratory syncytial virus (RSV) G glycoprotein mediates attachment of RSV to cells via an unknown receptor. To study G glycoprotein function we have cloned two variants of the RSV G gene into a Semliki Forest virus (SFV) expression vector, a full length (rG) and soluble (srG) G glycoprotein variant. By immunofluorescence microscopy, rG was found to be predominantly membrane associated, while srG was mostly cytoplasmic. The rG (80-85 kDa) and srG (75-80 kDa) constructs produced heavily glycosylated proteins, however they were slightly smaller than the G glycoprotein expressed in RSV infected HEp-2 cells (85-90 kDa). The biological activity of purified srG was tested by its ability to bind to RSV permissive cells. Purified srG bound to HEp-2 cells and the amount bound increased linearly with the quantity added. Binding was not saturable with the small quantities of protein available. Binding of srG to HEp-2 cells was inhibited (67-68%) by MAb 30 and neutralising anti-G MAb 29. Nonpermissive SF9 insect cells bound 20-50 times less srG than HEp-2 cells. SFV expressed recombinant RSV G glycoprotein should be useful for studying interactions between the RSV G glycoprotein and cells.
