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Review
. 2023 Oct 7;10(10):609.
doi: 10.3390/vetsci10100609.

Emergent Molecular Techniques Applied to the Detection of Porcine Viruses

Elda A Flores-Contreras  1   2 , Jorge Alberto Carrasco-González  3 , Daniel C L Linhares  4 , Cesar A Corzo  5 , J Israel Campos-Villalobos  6 , Alexandra Henao-Díaz  7 , Elda M Melchor-Martínez  1   2 , Hafiz M N Iqbal  1   2 , Reyna Berenice González-González  1   2 , Roberto Parra-Saldívar  1   2 , Everardo González-González  1
Affiliations
Review

Emergent Molecular Techniques Applied to the Detection of Porcine Viruses

Elda A Flores-Contreras et al. Vet Sci. .

Abstract

Molecular diagnostic tests have evolved very rapidly in the field of human health, especially with the arrival of the recent pandemic caused by the SARS-CoV-2 virus. However, the animal sector is constantly neglected, even though accurate detection by molecular tools could represent economic advantages by preventing the spread of viruses. In this regard, the swine industry is of great interest. The main viruses that affect the swine industry are described in this review, including African swine fever virus (ASFV), porcine reproductive and respiratory syndrome virus (PRRSV), porcine epidemic diarrhea virus (PEDV), and porcine circovirus (PCV), which have been effectively detected by different molecular tools in recent times. Here, we describe the rationale of molecular techniques such as multiplex PCR, isothermal methods (LAMP, NASBA, RPA, and PSR) and novel methods such as CRISPR-Cas and microfluidics platforms. Successful molecular diagnostic developments are presented by highlighting their most important findings. Finally, we describe the barriers that hinder the large-scale development of affordable, accessible, rapid, and easy-to-use molecular diagnostic tests. The evolution of diagnostic techniques is critical to prevent the spread of viruses and the development of viral reservoirs in the swine industry that impact the possible development of future pandemics and the world economy.

Keywords: ASFV; LAMP; NASBA; PCR; PCV; PEDV; PRRSV; PSR; RPA.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Viruses with the most significant impact in the pig industry, highlighting the main characteristics. Pig population and its geographical distribution, referenced from the Our World in Data database [2]. Note: African swine fever virus (ASFV), porcine reproductive and respiratory syndrome virus (PRRSV), porcine epidemic diarrhea virus (PEDV), porcine circovirus (PCV).
Figure 2
Figure 2
The proportion of isothermal tests developed to identify porcine viruses that have been published in scientific papers using the NCBI database. In addition, displayed are the most extensively studied viruses in the cases of LAMP and RPA. Note: African swine fever virus (ASFV), Porcine reproductive and respiratory syndrome virus (PRRSV), Porcine epidemic diarrhea virus (PEDV), Porcine circovirus (PCV), Loop-mediated isothermal amplification (LAMP), Nucleic acid sequence-based amplification (NASBA), Recombinase polymerase amplification (RPA), Polymerase spiral reaction (PSR).
Figure 3
Figure 3
Schematic illustration of the CRISPR-Cas system’s mechanism of action using ssDNA (single-stranded DNA) as a reporter for the detection of ASFV. (a) The ASFV DNA target sequence is amplified by the RPA or LAMP isothermal assays. (b) Components required for ASFV detection by the CRISPR-Cas system, which are CRISPR-RNA = crRNA, the enzyme Cas12a, and the ssDNA reporter labeled with a fluorophore (F) and a quencher (Q). (c) The Cas12a enzyme is guided by the crRNA to the target viral sequence. (d) The ASFV DNA sequence is cut by the CRIPR-Cas system. (e) After cutting the target sequence, the Cas12a enzyme is activated to perform non-specific ssDNase activity. (f) The non-specific cut in the ssDNA reporter sequence results in the release of the quencher, with a fluorescence signal indicating the presence of ASFV.
See this image and copyright information in PMC

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