Interfering factors in the diagnosis of Senecavirus A

• Published in: Molecular biology reports 2024-12, Vol.51 (1), p.777, Article 777
• Main Authors: Fonseca Júnior, Antônio Augusto, Laguardia-Nascimento, Mateus, Barbosa, Aline Aparecida Silva, da Silva Gonçalves, Valdenia Lopes, Camargos, Marcelo Fernandes
• Format: Article
• Language: English
• Subjects: Animal Anatomy; Animal Biochemistry; Animals; Automation; Biomedical and Life Sciences; Brazil; Diagnosis; Disease; Foot & mouth disease; Foot-and-Mouth Disease - diagnosis; Foot-and-Mouth Disease - virology; Histology; Life Sciences; Morphology; Oligonucleotides; Original Article; Picornaviridae - genetics; Picornaviridae - isolation & purification; Picornaviridae Infections - diagnosis; Picornaviridae Infections - veterinary; Picornaviridae Infections - virology; Real-Time Polymerase Chain Reaction - methods; Reproducibility of Results; RNA viruses; RNA, Viral - genetics; Sensitivity and Specificity; Stomatitis; Swine; Swine Diseases - diagnosis; Swine Diseases - virology; Swine vesicular disease; Swine Vesicular Disease - diagnosis; Swine Vesicular Disease - virology; Vesicular stomatitis
• ISSN: 0301-4851; 1573-4978; 1573-4978
• DOI: 10.1007/s11033-024-09692-2

Background Senecavirus A (SV-A) is an RNA virus that belongs to the genus Senecavirus within the family Picornaviridae. This study aimed to analyze factors that can influence the molecular diagnosis of Senecavirus A, such as oligonucleotides, RNA extraction methods, and RT-qPCR kits. Methods Samples from suspected cases of vesicular disease in Brazilian pigs were analyzed for foot-and-mouth disease, swine vesicular disease, and vesicular stomatitis. All tested negative for these diseases but positive for SV-A. RT-qPCR tests were used, comparing different reagent kits and RNA extraction methods. Sensitivity and repeatability were evaluated, demonstrating efficacy in detecting SV-A in clinical samples. Results In RNA extraction, significant reduction in Cq values was observed with initial dilutions, particularly with larger supernatant volumes. Trizol and Maxwell showed greater sensitivity in automated equipment protocols, though results varied in tissue tests. RT-qPCR kit comparison revealed differences in amplification using viral RNA but minimal differences with plasmid DNA. Sensitivity among methods was comparable, with slight variations in non-amplified samples. Repeatability tests showed consistent results among RT-qPCRs, demonstrating similarity between methods despite minor discrepancies in Cq values. Conclusions Trizol, silica columns, and semi-automated extraction were compared, as well as different RT-qPCR kits. The study found significant variations that could impact the final diagnosis.