Abstract
In this study, we develop a reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay for rapid and easy detection of goose astroviruses (GAstVs) in clinical samples. The specific LAMP primer sets were designed targeting the ORF2 gene of GAstV. The conditions of LAMP amplification were optimized in terms of reaction time and temperature. The optimal conditions are 60 min in a 60 °C water bath. No cross-reactivity was noted with fowl adenovirus serotype 4 (FAdV-4), duck tembusu virus (DTMUV), goose parvovirus (GPV), avian infectious bronchitis virus (IBV), or chicken anemia virus (CAV). The proposed RT-LAMP method was compared with conventional RT polymerase chain reaction (RT-PCR) and with nested RT-PCR. The results showed that the sensitivity of the proposed method was comparable to that of nested RT-PCR and tenfold higher than that of the conventional RT-PCR. Clinical samples (N = 129) of the liver and kidney from sick geese collected from six commercial goose farms were tested. The positive rate was 39.5% (51/129), 38.8% (50/129), and 34.9% (45/129) using RT-LAMP, nested RT-PCR and conventional RT-PCR, respectively. The developed RT-LAMP diagnostic method is not only simple, rapid, and highly specific, but also portable for use on the field. It may be used in epidemiological investigation to detect GAstVs.
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The authors express gratitude to editage for the help with revising this manuscript.
Funding
This work was supported financially by the National Natural Science Foundation of China (no. 31602063), National key research and development program (no. 2018YFD0502006), Anhui Key Research and Development Program (no. 201904f06020030). Accession number: A new GAstV strain (no. MN099162), FAdV-4 strain (no. KY379035), DTMUV strain (no. KY623435), GPV strain (no. MK333463), IBV strain (no. MH020185) and CAV strain (no. KU645509), was uploaded to GenBanK database.
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ZY and DZ conceived of the study, carried out the experiment and drafted the manuscript, contributed equally to this work. KY and CB participated in the data collection and analysis. YL involved in drafting of the manuscript. JL and SJ participated in statistical analysis. YW conceived of the study, revising the manuscript critically. All authors have read and approved the final manuscript.
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Yu, Z., Zhang, D., Yang, K. et al. A simple and rapid diagnostic method to detect new goose astrovirus using reverse-transcription loop-mediated isothermal amplification. 3 Biotech 10, 20 (2020). https://doi.org/10.1007/s13205-019-2006-z
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DOI: https://doi.org/10.1007/s13205-019-2006-z