Characterization and evaluation of the pathogenicity of a natural recombinant transmissible gastroenteritis virus in China

Abstract

Porcine transmissible gastroenteritis virus (TGEV) is one of the major etiological agents of viral enteritis and fetal diarrhea in suckling piglets. In this study, a TGEV JS2012 strain was isolated from the feces of piglets in Jiangsu Province, China. The phylogenetic analysis showed that TGEV JS2012 was placed between the Purdue and the Miller clusters. Analysis of recombination confirmed that TGEV JS2012 is a natural recombinant strain between Miller M6 and Purdue 115. Similar to Miller M6, virulent Purdue and China strain TS, in S gene the JS2012 maintained genetic integrity and the characteristics of the TGEV virulent strains. In vivo, TGEV JS2012 caused 100% mortality in newborn piglets, indicating the strong pathogenicity of this isolate. These results reveal that the JS2012 is a novel natural recombinant TGEV with high virulence. Our findings provide valuable information about genetic diversity and infection mechanism of the coronavirus family.

Keywords: Pathogenicity; Phylogenetic; Recombination; TGEV.

Fig. 1

Virus isolation and identification of TGEV strain JS2012. (a) The cytopathic effect of JS2012 on ST cells. (b) Fluorescence microscopy of JS2012 infection on ST cells. (c) Electron microscopy observation of JS2012. Scale bars = 50 nm. (d) Growth kinetics of JS2012 with different passages in ST cells.

Fig. 2

Phylogenetic analysis based on the nucleotide sequence of the complete genome and S gene of TGEV strains. (a) Complete genome. (b) S gene. The trees were created using the neighbor-joining method in MEGA version 6.0, with 1000 bootstrap replicates to measure confidence of the groupings. The black origin indicates the new JS2012 reported in this study.

Fig. 3

Visualization of genomic insertion and deletion regions of the 22 TGEV strains. The nucleotide sequences of 22 TGEV strains were analyzed using the CLUSTAL W program by MEGA version 6.06.

Fig. 4

Mutation in S protein and S gene of TGEV JS2012. (a) Mutation in S protein of TGEV JS2012. (b) T residue at nucleotide 1753 of JS2012 strains Virulent M6, Virulent Purdue, and TS, and G in the other strains. The amino acid and nucleotide sequences of the complete S gene of TGEV JS2012 and other TGEV strains were analyzed using the CLUSTAL W program by MEGA version 6.06.

Fig. 5

Recombination analysis of TGEV JS2012 with other TGEV strains. (a) TGEV genome organization. (b) Recombination of Miller M6 and Purdue P115 located at nucleotides 23240–24324 of S gene. (c) Recombination score = 0.638.

Fig. 6

Transmissible gastroenteritis virus JS2012 infection in newborn piglets. (a) Mean fecal scores after viral inoculation. (b) Mean quantitative reverse transcription polymerase chain reaction titers of fecal samples. (c) Survival curves of JS2012-infected clusters.

Fig. 7

Histology and IHC staining of TGEV JS2012-infected pig intestines. (a) HE-stained small intestines of piglets inoculated with JS2012 strains. (b) For IHC assays, the intestinal tissue sections were stained with a TGEV N monoclonal antibody (1:500 dilution). Positive cells presented in villous epithelial cells and the crypt enterocytes of Jejunum.