Designing one-step reverse transcriptase loop-mediated isothermal amplification for serotype O foot-and-mouth disease virus detection during the 2022 outbreak in East Java, Indonesia

Vet World Vol.16 September-2023 Article-15

Research Article

Veterinary World, 16(9): 1889-1896

https://doi.org/10.14202/vetworld.2023.1889-1896

Eduardus Bimo Aksono^1,2, Mirni Lamid¹, Rimayanti Rimayanti¹, Iwan Sahrial Hamid¹, Mustofa Helmi Effendi¹, Fedik Abdul Rantam¹, Widjiati Widjiati¹, Mufasirin Mufasirin^1,3, Heni Puspitasari³, Munawaroh Fitria⁴, Nur Syamsiatul Fajar³, Lucia Tri Suwanti¹, Nusdianto Nusdianto¹, Andi Hamim Zaidan², Yuta Kanai⁵, and Teguh Hari Sucipto³

1. Department of Veterinary Medicine, Faculty of Veterinary Medicine, Universitas Airlangga, Surabaya, 60115, Indonesia.
2. Institute of Life Science, Technology and Engineering, Universitas Airlangga, Surabaya, 60115, Indonesia.
3. Institute of Tropical Disease, Universitas Airlangga, Surabaya, 60115, Indonesia.
4. Department of Clinical Pathology, Faculty of Medicine, Universitas Airlangga, Surabaya, 60132, Indonesia.
5. Research Institute for Microbial Diseases, Osaka University, Suita, Osaka, 565-0871, Japan.

Background and Aim: Various methods can detect foot-and-mouth disease (FMD) in cows, but they necessitate resources, time, costs, laboratory facilities, and specific clinical specimen submission, often leading to FMD virus (FMDV) diagnosis delays. The 2022 FMD outbreak in East Java, Indonesia, highlighted the need for an easy, inexpensive, rapid, and accurate detection approach. This study aims to devise a one-step reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) technique and phylogenetic analysis to detect the serotype O FMDV outbreak in East Java.

Materials and Methods: Swab samples were collected from the foot vesicles, nasal secretions, and saliva of five suspected FMDV-infected cows in East Java between June and July 2022. The RT-LAMP design used hydroxy naphthol blue dye or SYBR Green I dye, with confirmatory analysis through reverse transcriptase polymerase chain reaction (RT-PCR) targeting 249 base pairs. PCR products underwent purification, sequencing, and nucleotide alignment, followed by phylogenetic analysis.

Results: The RT-LAMP method using hydroxy naphthol blue dye displayed a positive reaction through a color shift from purple to blue in the tube. Naked-eye observation in standard light or ultraviolet (UV) light at 365 nm, with SYBR Green I stain, also revealed color change. Specifically, using SYBR Green I dye, UV light at 365 nm revealed a color shift from yellow to green, signifying a positive reaction. Nucleotide alignment revealed mutations and deletion at the 15th sequence in the JT-INDO-K3 isolate from the East Java FMDV outbreak. Despite differing branches, the phylogenetic tree placed it in the same cluster as serotype O FMDV from Malaysia and Mongolia.

Conclusion: JT-INDO-K3 exhibited distinctions from Indonesian serotype O FMDV isolates and those documented in GenBank. Then, the RT-LAMP method used in this study has a detection limit 10 times higher latter than the conventional RT-PCR limit, without any cross-reactivity among strains. Keywords: cow, East Java, foot-and-mouth disease virus, reverse transcriptase loop-mediated isothermal amplification, reverse transcriptase polymerase chain reaction, serotype O.

Keywords: cow, East Java, foot-and-mouth disease virus, reverse transcriptase loop-mediated isothermal amplification, reverse transcriptase polymerase chain reaction, serotype O.

How to cite this article: Aksono EB, Lamid M, Rimayanti R, Hamid IS, Effendi MH, Rantam FA, Widjiati W, Mufasirin M, Puspitasari H, Fitria M, Fajar NS, Suwanti LT, Nusdianto N, Zaidan AH, Kanai Y, and Sucipto TH (2023) Designing one-step reverse transcriptase loop-mediated isothermal amplification for serotype O foot-and-mouth disease virus detection during the 2022 outbreak in East Java, Indonesia, Veterinary World, 16(9): 1889-1896.

Received: 28-03-2023 Accepted: 23-08-2023 Published online: 17-09-2023

Corresponding author: Eduardus Bimo Aksono E-mail: eduardus-b-a-h@fkh.unair.ac.id

DOI: 10.14202/vetworld.2023.1889-1896

Copyright: Aksono, et al. This article is an open access article distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http:// creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.