Novel genes identified in the nonreference sequences including 14 genes in the MHC. (A) Geographic distributions of pig breeds used for the construction of pig pangenome. Northern Chinese pigs (dark red) are as follows: Hetaodaer (HTDE), Min (MIN), Laiwu black (LW), and Erhualian (EHL). Southern Chinese pigs (pink) are as follows: Wuzhishan (WZS), luchuan (LC), bamaxiang (BMX), Baoshan (BS), Debao (DB), Diannan small-ear (DNS), Gaoligongshan (GLGS), Guangdong small-ear spotted (GDS), Guanzhuang spotted (GZ), Guizhong spotted (GZS), Huai (HUAI), Lantang (LT), large black-white (LBW), Longlin (LONG), Minbei spotted (MBS), Putian (PT), Saba (SB), Tunchang (TCH), and Yuedong black (YDB). European pigs are as follows: Duroc (DR), Yorkshire (YS), landrace (LR), Pietrain (PTR), Hampshire (HSH), Berkshire (BSH), Landrace × Yorkshire (LRYS), and Duroc × (Landrace × Yorkshire) (DLY). (B) Venn diagram displaying the distribution of 3438 genes from the pangenome not found in the reference genome, including genes shared among or unique to either the Chinese or European breeds. (C) Enriched GO terms of nonreference genes. (D) The pipeline of the PRRSV challenge trial: (1) Piglets from the PRRSV-infected group and noninfected group were injected intramuscularly with PRRSV and 2% DMEM, respectively; (2) lungs of piglets were collected at 7, 14, and 21 days post infection (dpi) and tissue slices examined microscopically by hematoxylin-eosin staining; (3) extraction of bronchoalveolar lavage fluid (BALF); (4) collection of porcine alveolar macrophages (PAMs); and (5) RNA extraction and sequencing. (E) Fourteen nonreference SLA genes suffered from PAV among 103 individuals from 22 pig breeds. The seven nonreference SLA genes in the gray box were differentially expressed after PRRSV infection. (F) Volcano plots show DEGs under PRRSV infection at three time points (7, 14, and 21 dpi). The black circles represent the 60 DEGs identified in the nonreference sequences. Orange and blue points denote the up-regulated and down-regulated DEGs, respectively. The line chart describes that the number of DEGs increases as dpi increase.
