In silico analyses of antigenicity and surface structure variation of an emerging porcine circovirus genotype 2b mutant, prevalent in southern China from 2013 to 2015

Abstract

Porcine circovirus type 2 (PCV2) is the pivotal pathogen causing porcine circovirus-associated diseases. In this study, 62 PCV2 isolates were identified from seven farms in southern China from 2013 to 2015 and phylogenetic trees were reconstructed based on whole-genome sequences or the cap gene. In this investigation, PCV2b was the main genotype in circulation throughout these farms. Furthermore, an emerging mutant (PCV2b-1C), isolated from PCV2-vaccinated farms, was the predominant strain prevalent on these farms. In addition, we isolated a new cluster that may represent evolution of the virus through recombination of PCV2b-1A/1B and PCV2b-1C. Finally, we discuss evidence that antigenicity and surface structure variation of the capsid resulted from mutation of the C-terminal loop (Loop CT) of the PCV2b-1C Cap in silico.

Fig. 1.

Phylogenetic tree of the 62 PCV2 isolates in our study with 28 reference strains from PCV1 (KJ408798), PCV2a (2A: AF109398, AY754017 and AY556474; 2B: AY180396, AY146993, AF166528; 2C: AF201308, AF201309, AF201310; 2D: AY322004, AY424403, AF381176; 2E: DQ104423, AJ223185, AB072301), PCV2b (1A: AY322003, AY424405, AF055393; 1B: AY484407, AY691169, HQ202970; 1C: HQ202973, HM038017, JX535296) and PCV2c (EU148503, EU148504, EU148505) deposited in GenBank. The phylogenetic tree was reconstructed based on complete genomic sequences of isolates described above. mega5.05 was used to reconstruct a neighbour-joining tree using a Kimura two-parameter model. Reliabilities of the tree were assessed (1000 bootstrap replications). ▴, 13 PCV2 isolates in 2013; ▪, 20 PCV2 isolates in 2014; ♦, 29 PCV2 isolates in 2015. Bar, 0.02 substitutions per site.

Fig. 2.

Phylogenetic trees of the 62 PCV2 isolates with 28 reference strains were reconstructed based on either the (a) cap or (b) rep genes. Bars, 0.05 (a) and 0.02 (b) substitutions per site.

Fig. 2.

Phylogenetic trees of the 62 PCV2 isolates with 28 reference strains were reconstructed based on either the (a) cap or (b) rep genes. Bars, 0.05 (a) and 0.02 (b) substitutions per site.

Fig. 3.

The new cluster of YiY-3-2-2 was a product of genome recombination between PCV2b-1A/1B and PCV2b-1C. The cap gene similarity was compared between the cap gene of the new cluster (YiY-3-2-2) and those of PCV2 reference strains of PCV2b-1A/1B (GenBank accession number FJ598044), PCV2b-1C (GenBank accession number HM038017) and PCV2a (GenBank accession number HM038034) using SimPlot molecular recombination software. The y-axis indicates identity within a sliding window (200 bp wide) centred on the position plotted, with a step size between plots of 20 bp. GapStrip, on; Kimura (two-parameter); T/t = 2.0. The red vertical line indicates a potential breakpoint.

Fig. 4.

Primary amino acid sequence alignment of PCV2b Cap and surface structure of PCV2 capsid. (a) Comparison of Cap amino acid residues amongst nine PCV2b reference isolates (GenBank accession numbers AY322003, AY424405, AF055393, AY484407, AY691169, HQ202970, HQ202973, HM038017 and JX535296). (b, c) The surface structure of a PCV2b capsid was generated based on the crystal structure of PCV2^CS (Protein Data Bank ID: 3R0R; Khayat et al., 2011) and displayed using PyMOL. The amino acid residues of Loop CT are labelled in blue (b) and the two amino acid residues (²³⁰PL²³¹) at the end of Loop CT are marked in magenta (c). Note that the last two amino acid residues (²³²NP²³³) of Cap were absent in the crystal structure.

Fig. 5.

Localization and orientation of β-strand B, Loop BC and Loop CT in the crystal structures of both the PCV2 Cap monomer and capsid. Localization of β-strand B (green), Loop BC (red) and Loop CT (blue) in the Cap monomer is displayed in a (a) surface model or (b) a backbone model. After 60 Cap monomers self-assembled into a capsid, the orientations of β-strand B, Loop BC and Loop CT on the surface are displayed on (c) a global scale or (d) an enlarged scale. Note that the last two amino acid residues (²³²NP²³³) of Cap were absent in the crystal structure.