A novel RNA-based in situ hybridization to detect Seneca Valley virus in neonatal piglets and sows affected with vesicular disease

Abstract

Seneca Valley virus (SVV) is the causative agent of an emerging vesicular disease in swine, which is clinically indistinguishable from other vesicular diseases such as foot-and-mouth disease. In addition, SVV has been associated with neonatal mortality in piglets. While a commercial SVV qRT-PCR is available, commercial antibodies are lacking to diagnose SVV infections by immunohistochemistry (IHC). Thus, a novel in situ hybridization technique-RNAscope (ISH) was developed to detect SVVRNA in infected tissues. From a total of 78 samples evaluated, 30 were positive by qRT-PCR and ISH-RNA, including vesicular lesions of affected sows, ulcerative lesions in the tongue of piglets and various other tissues with no evidence of histological lesions. Nineteen samples were negative for SVV by qRT-PCR and ISH-RNA. The Ct values of the qRT-PCR from ISH-RNA positive tissues varied from 12.0 to 32.6 (5.12 x 106 to 5.31 RNA copies/g, respectively). The ISH-RNA technique is an important tool in diagnosing and investigating the pathogenesis of SVV and other emerging pathogens.

Fig 1. SVV distribution in vesicular lesions in the snout of an affected sow and in necrotizing lesions in tongue of an affected piglet.

a) Skin from infected sow. Ballooning degeneration (intracellular edema) of keratinocytes in the stratum spinosum with formation of intraepidermal vesicles. H&E, 20x; b) Skin from infected sow. Intraepidermal vesicle showing strong SVV positive staining within the cytoplasm of keratinocytes. ISH-RNA, 40x; c) Tongue from infected piglet. Necrotizing glossitis. H&E, 20x; d) Tongue from the infected piglet shown in (c). Red dots and clusters represent the presence of SVV mRNA within an erosive lesion. ISH-RNA, 40x.

Fig 2. SVV distribution in tissues without evidence of histological lesions.

Swine, ISH-RNA. a) Piglet, spleen (central arteriole). Strong SVV positive staining diffusely distributed throughout the splenic parenchyma. ISH-RNA, 20x.; b) Piglet, spleen. Negative control. ISH-RNA, 20x; c) Piglet, small intestine. SVV mRNA was multifocally distributed within enterocytes (black arrows) and lamina propria. ISH-RNA, 20x; d) Piglet, lung, SVV positive signals were found in alveolar septum. ISH-RNA, 20x.