Development of real time reverse transcription loop-mediated isothermal amplification assay for rapid detection of genotype VII of Newcastle disease viruses

ABSTRACT

1. This paper details the establishment of a diagnostic system based on the real-time reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for the rapid, simple and sensitive detection of genotype VII of Newcastle disease virus (NDV) directly from field samples. One specific set of six primers were designed which targeted the fusion protein gene of G-VII viruses. The target gene can be amplified and the results recorded within 40 min.

2. The merit of this technique was the feasibility of reading results either by examining turbidity by the naked eye or via the amplification curve generated by real-time PCR. This study tested the sensitivity and specificity of this system against NDV-G-VII and other avian viruses. The real-time RT-LAMP has been found to be more sensitive than real-time RT-PCR. Moreover, 24 out of 35 suspected field samples were positive for genotype VII by real-time RT-LAMP within 30 min in comparison to the real-time RT-PCR for detection of universal NDV.

3. Accordingly, real-time RT-LAMP revealed higher sensitivity than real-time RT-PCR and had specificity only for the NDV-G-VII genotype. Additionally, this system was more rapid and had lower cost than real-time RT-PCR. Based on the results, the RT-LAMP-based assay is a useful tool for the rapid and sensitive diagnosis of NDV-G-VII infection.

KEYWORDS:

• RT-LAMP
• RT-qPCR
• NDV-genotype VII
• sensitivity
• specificity

Acknowledgements

The authors would like to thank Dr Naglaa Hagag and Dr Nahed Yahia in the Gene Analysis Unit at Reference Laboratory for Veterinary Quality Control on Poultry Production (RLQP).

Disclosure statement

No potential conflict of interest was reported by the authors.

Authors’ contributions

KS, designed research; AA, performed research; AA and KA, wrote the draft of the manuscript; and SE and MS, revision of the manuscript and supervision of the workflow. All authors read and approved the final manuscript.

Availability of data and materials

The datasets supporting the conclusions of this article are included within the article.

Supplementary material

Supplemental data for this article can be accessed online at https://doi.org/10.1080/00071668.2022.2094219

Additional information

Funding

Science, Technology & Innovation Funding Authority (STDF STDF project no. 25406) – grant no. 25406.