Analysis on the genome of a teschovirus type 1 isolates with swine diarrhea

Abstract

Porcine Teschoviruses (PTVs) are associated with polioencephalomyelitis and various diseases, including reproductive and gastrointestinal disorders of pigs and wild boars, but rarely detected in the feces of pigs. In this study, a sample of swine diarrhea that tested positive for PTVs is subjected to high-throughput sequencing. The viral genome was 7221 nucleotides (nt) in length, which was consisted of twelve genes. Phylogenetic analysis showed and it was closely related to the PTV-HNMY(MG755212.1). The nucleotide homology of VP1 gene of PTVs JS2021 with PTV-1AF 296102.1 reached 82.97%, belonging to a branch of PTV-1 serotype. The nucleotide homology of VP1 protein with other serotypes of PTV is quite different from that of other serotypes of PTV. Bioinformatics analysis showed that PTVs have four capsid proteins, namely VP1, VP2, VP3 and VP4. The VP1 encodes a 29 kDa protein, which is the main protective antigen, a theoretical isoelectric point of 6.73, no transmembrane domain, no signal peptide and potential phosphorylation site. The VP1 protein is an unstable hydrophilic intracellular protein, which contains four secondary structures: irregular curl (c), extended chain (e), α-helix (h) and β-folded (t). The tertiary structure is heart-shaped and has multiple B cell epitopes. By analyzing the tertiary structure, we found that the amino acid at position 129 of VP1 mutated and reduction a larger alpha helix. This may lead to the main cause of piglet diarrhea. These findings enriched our knowledge of the viruses in the role of swine diarrhea, and help to develop an effective strategy for disease prevention and control.

Keywords: Diarrhea; High-throughput sequencing; PTVs; Phylogenetic analysis; VP1.

Fig. 1

Characterization of PTVs infection in ST cells. A. Morphological changes in ST cells at different time points after PTVs JS2021 infection. B. RT-PCR electrophoresis map of VP1 gene fragment of PTVs JS2021 isolate. C. The schematic diagram of genome structure of PTVs.

Fig. 2

(A) Neighbor-joining unrooted phylogenetic trees based on the nucleotide sequences of the full-length sequences of PTVs JS2021 and different genogroups. Our strain, PTVs JS 2021, is marked with a red triangle (◇). (B) Neighborjoining unrooted phylogenetic trees based on the VP1 gene sequences of PTVs. PTVs JS 2021, is marked with a red circle (Ο).

Fig. 3

Hydrophilic and hydrophobic analysis of VP1 protein. Verified by Protscale software. The peak value in the figure represents the hydrophilicity and hydrophobicity of amino acids, the highest score of Val (V) at position 213 was1.778, and the lowest score of Gln (Q) at position191 was −2.556. Figure A represents PTV JS2021. Figure B represents PTV-1 AF296102.1. Figure C represents PTV HNMY. Figure D represents PTV-8 JS2013.

Fig. 4

Prediction of signal peptide of VP1 protein. SP is used to distinguish whether the corresponding position is a signal peptide region. CS is used to distinguish whether it is a shear site or not, and the highest peak value is the first amino acid after the shear site (i.e., the first amino acid residue of mature protein), By software analysis, there is no fluctuation in sp value, so there is no signal peptide in the protein.

Fig. 5

Prediction of transmembrane domain of VP1 protein. The abscissa axis represents the serial number of amino acid residues corresponding to the submitted protein sequence. The value of ordinate axis is the probability value of each amino acid located in the medial membrane, lateral membrane and spiral region of crotch membrane on the transverse axis.

Fig. 6

Prediction of phosphorylation site specific to VP1 protein. The abscissa axis represents the serial number of amino acid residues corresponding to the submitted protein sequence. The value of ordinate axis is the score (0–1), and generally higher than 0.5 is the positive result. Figure A represents PTV JS2021. Figure B represents PTV-1 AF296102.1. Figure C represents PTV HNMY. Figure D represents PTV-8 JS2013.

Fig. 7

Prediction of B cell linear epitope of VP1 protein. The highest score was 2.200, the lowest was-0.005, and there were 12 linear epitopes by using IEDB software. Figure A represents PTV JS2021. Figure B represents PTV-1 AF296102.1. Figure C represents PTV HNMY. Figure D represents PTV-8 JS2013.

Fig. 8

Prediction of secondary structure of VP1 protein. Blue stands for Alpha helix, green stands for Beta turn, yellow stands for Random coil, and red stands for extended strand.

Fig. 9

Prediction of tertiary structure of VP1 protein. (A) Three-dimensional structure of VP1 protein after modeling. (B, C) Comparison of VP1 protein of PTV JS2021 with VP1 protein of classic strain PTV-8jinlin/2003, in which red represents amino acid at position 129. Among them, green stands for PTV JS 2021. Light blue represents PTV-8jinlin/2003.