Analysis of Newcastle disease virus quasispecies and factors affecting the emergence of virulent virus.

Genome sequence analysis of a number of avirulent field isolates of Newcastle disease virus revealed the presence of viruses (within their quasispecies) that contained virulent F0 sequences. Detection of these virulent sequences below the ~1% level, using standard cloning and sequence analysis, proved difficult, and thus a more sensitive reverse-transcription real-time PCR procedure was developed to detect both virulent and avirulent NDV F0 sequences. Reverse-transcription real-time PCR analysis of the quasispecies of a number of Newcastle disease virus field isolates, revealed variable ratios (approximately 1:4-1:4,000) of virulent to avirulent viral F0 sequences.

A simple approach for preparing real-time PCR positive reaction controls for rare or emerging viruses.

Background: Laboratories often have difficulties obtaining positive control material for polymerase chain reaction (PCR) diagnosis of rare or emerging viruses. This is particularly problematic during outbreaks caused by emerging infectious diseases, when delays can impede the public health response.

Objectives: The aim of this study was to develop a simple approach for preparing real-time PCR positive reaction controls for rare or emerging viruses.

Detection of novel influenza A(H1N1) virus by real-time RT-PCR.

Accurate and rapid diagnosis of novel influenza A(H1N1) infection is critical for minimising further spread through timely implementation of antiviral treatment and other public health based measures. In this study we developed two TaqMan-based reverse transcription PCR (RT-PCR) methods for the detection of novel influenza A(H1N1) virus targeting the haemagglutinin and neuraminidase genes. The assays were validated using 152 clinical respiratory samples, including 61 Influenza A positive samples, collected in Queenland, Australia during the years 2008 to 2009 and a further 12 seasonal H1N1 and H3N2 influenza A isolates collected from years 2000 to 2002.

Sequence variation in the Newcastle disease virus genome.

Full-length genome sequences of five virulent and five avirulent strains of Newcastle disease virus isolated between 1998 and 2002 in Victoria and New South Wales, Australia were determined. Comparisons between these strains revealed that coding sequence variability in the haemagglutinin-neuraminidase (HN), matrix (M) and phosphoprotein (P) gene sequences appeared to be more variable than in the fusion (F), nucleocapsid (N) and RNA dependent-RNA replicase (L) genes. Sequence analysis of a number of other isolates made during the recent virulent NDV outbreaks, also identified the presence of a number of variants with altered F gene cleavage sites, which resulted in altered biological properties of those viruses.

Characterisation of an Australian bat lyssavirus variant isolated from an insectivorous bat.

In 1996 a variant lyssavirus was isolated from an insectivorous bat (yellow bellied, sheath tail bat-Saccolaimus flaviventris) in Australia. The nucleocapsid protein (N), matrix protein (M), phosphoprotein (P), glycoprotein (G) and polymerase (L) genes of the Australian bat lyssavirus (ABL) insectivorous isolate were compared with that previously described from a frugivorous bat (Pteropus sp.), and showed sequence divergence at both the nucleotide and amino acid sequence level of 20% and 4-12%, respectively.