SummaryTwelve nasal swabs were collected from yearling horses with respiratory distress and tested for Equid herpesvirus 1 (EHV-1) and Equid herpesvirus 4 (EHV-4) by realtime PCR targeting the glycoprotein B gene. All samples were negative for EHV-1; however 3 were positive for EHV-4. When these samples were tested for EHV-2 and EHV-5 by PCR all samples were negative for EHV-2 and 11 were positive for EHV-5. All three samples that were positive for EHV-4 were also positive for EHV-5.These three samples gave a limited CPE in ED cells reminiscent of EHV-4 CPE.EHV-4 CPE was obvious after 3 days and was characterised by syncytia. None of the samples produced cytopathic effect (CPE) on African green monkey kidney (Vero) cells or hamster kidney (BSR) cells. Four of the samples, which were positive in the EHV-5 PCR, produced CPE on rabbit kidney (RK13) cells and equine dermis (ED) cells. EHV-5 CPE on both cell lines was slow and was apparent after four 7-day passages. On RK13 cells the CPE was characteristic of equid herpesvirus with the formation of syncytia. However in ED cells, the CPE was characterised by ringshaped syncytia.For the first time a case of equine respiratory disease involving dual infection with EHV-4 and EHV-5 has been reported in Queensland. This was shown by simultaneously isolating EHV-4 and EHV-5 from clinical samples. EHV5 was recovered from all samples except one, suggesting that EHV5 was more prevalent in young horses than EHV2.
High-throughput sequencing (HTS) provides the opportunity, once a diagnostic result is obtained, to extract additional information from a virus-containing sample. Hence, it offers advantages over established quantitative amplification technology, such as quantitative PCR, particularly in a public health environment. At this early stage of its clinical application, there have been limited studies comparing HTS performance to that of the more established quantitative PCR technology for direct detection of viruses. In this pilot-scale study, we tested HTS with a range of viruses and sample types routinely encountered in a public health virology laboratory. In comparison with quantitative PCR, our HTS method was able to sensitively (92%) detect all viruses in any sample type with the exception of certain tissues. Moreover, sufficient nucleotide sequence information was obtained to enable genotyping of strains detected, thus providing additional useful epidemiological information. While HTS sensitivity may not yet match that of PCR, the added value through enhanced epidemiological data has considerable potential to enable real-time surveillance of circulating strains so as to facilitate rapid and appropriate response to outbreaks and virus zoonotic spillover events.
A 5? Taq nuclease assay specific for Avibacterium paragallinarum was designed and optimized for use in diagnosing infectious coryza. The region chosen for assay design was one of known specificity for Av. paragallinarum. The assay detected Av. paragallinarum reference strains representing the three Page and the eight Kume serovars, and field isolates from diverse geographical locations. No cross-reactions were observed with other Avibacterium species, with other bacteria taxonomically related to Av. paragallinarum nor with bacteria and viruses likely to be present in swabs collected from suspected infectious coryza cases. The detection limit for the assay was 6 to 60 colony-forming units per reaction. Twenty-two out of 53 swabs collected from sick birds reacted in the 5? Taq nuclease assay, whereas Av. paragallinarum was not isolated from any of the swabs. All of the 22 swabs yielded other bacteria in culture. The presence of Av. paragallinarum in the swabs was also demonstrated by sequencing, thereby confirming the ability of the assay to detect Av. paragallinarum in the presence of other bacteria. The ability to quantify bacterial load in the swabs using the 5? Taq nuclease assay was demonstrated.
We sequenced the genomes of two chikungunya virus isolates obtained from viremic patients who had traveled to Australia. The first patient acquired the infection in Bangladesh in 2017, and the second was infected in Thailand in 2019. Phylogenetic sequence analysis demonstrated that both isolates belonged to the East/Central/South African genotype.
An outbreak of acute respiratory disease in layers was diagnosed as being of dual nature due to fowlpox and infectious laryngotracheitis using a multidisciplinary approach including virus isolation, histopathology, electron microscopy and polymerase chain reaction (PCR). The diagnosis was based on virus isolation of gallid herpesvirus 1 (GaHV-1) in chicken kidney cells and fowlpox virus (FWPV) in 9-day-old chicken embryonated eggs inoculated via the chorioallantoic membrane. The histopathology of tracheas from dead birds revealed intra-cytoplasmic and intra-nuclear inclusions suggestive of poxvirus and herpesvirus involvement. The presence of FWPV was further confirmed by electron microscopy, PCR and histology. All FWPV isolates contained the long terminal repeats of reticuloendotheliosis virus as demonstrated by PCR. GaHV-1 isolates were detected by PCR and were shown to have a different restriction fragment length polymorphism pattern when compared with the chicken embryo origin SA2 vaccine strain; however, they shared the same pattern with the Intervet chicken embryo origin vaccine strain. This is a first report of dual infection of chickens with GaHV-1 and naturally occurring FWPV with reticuloendotheliosis virus insertions. Further characterization of the viruses was carried out and the results are reported here.
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