Development of a reverse transcription-loop-mediated isothermal amplification assay for the detection of parrot bornavirus 4

Ryo Komorizono; Keizo Tomonaga; Akiko Makino

doi:10.1016/j.jviromet.2019.113749

Development of a reverse transcription-loop-mediated isothermal amplification assay for the detection of parrot bornavirus 4

J Virol Methods. 2020 Jan:275:113749. doi: 10.1016/j.jviromet.2019.113749. Epub 2019 Oct 14.

Authors

Ryo Komorizono¹, Keizo Tomonaga², Akiko Makino¹

Affiliations

¹ Laboratory of RNA Viruses, Department of Virus Research, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan; Laboratory of RNA Viruses, Department of Mammalian Regulatory Network, Graduate School of Biostudies, Kyoto University, Kyoto, Japan.
² Laboratory of RNA Viruses, Department of Virus Research, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan; Laboratory of RNA Viruses, Department of Mammalian Regulatory Network, Graduate School of Biostudies, Kyoto University, Kyoto, Japan; Department of Molecular Virology, Graduate School of Medicine, Kyoto University, Kyoto, Japan.

PMID: 31622637
DOI: 10.1016/j.jviromet.2019.113749

Abstract

Avian bornavirus (ABV) is the causative agent of proventricular dilatation disease, which is fatal in psittacine birds. ABVs have spread worldwide, and outbreaks have led to mass deaths of captive birds in commercial and breeding facilities. The segregation of infected birds is a countermeasure to prevent ABV spread in aviaries. However, this approach requires a highly sensitive detection method for the screening of infected birds before virus transmission. In this study, we developed a reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay for the diagnosis of parrot bornavirus 4 (PaBV-4), a dominant ABV genotype. Using this assay, we successfully detected PaBV-4 RNA in cell cultures, brain tissues, and feces. We also developed methods for simple RNA extraction and visual detection without electrophoresis. The sensitivity of the newly established RT-LAMP assay was 100-fold higher than that of the real-time PCR (RT-qPCR) assay. Accordingly, the RT-LAMP assay developed in this study is suitable for the rapid and sensitive diagnosis of PaBV-4 without specialized equipment and will contribute to virus control in aviaries.

Keywords: Avian bornavirus; Captive birds; Parrot bornavirus 4; RT-LAMP.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Bird Diseases / diagnosis*
Bird Diseases / virology
Bornaviridae / genetics
Bornaviridae / isolation & purification*
Feces / virology
Genotype
Molecular Diagnostic Techniques / methods*
Mononegavirales Infections / diagnosis
Mononegavirales Infections / veterinary*
Nucleic Acid Amplification Techniques / methods*
Parrots / virology*
Phylogeny
RNA, Viral / genetics
Reverse Transcription*

Substances

RNA, Viral

Supplementary concepts

LAMP assay