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. 2019 Jul 17;63(8):1240–1250. doi: 10.1007/s11427-019-9576-2

Next-generation sequencing and single-cell RT-PCR reveal a distinct variable gene usage of porcine antibody repertoire following PEDV vaccination

Ren Li 1, Fang Fu 1, Li Feng 1,✉, PingHuang Liu 1,✉
PMCID: PMC7088813  PMID: 31321668

Abstract

Porcine epidemic diarrhea virus (PEDV) is the most common diarrhea-causing pathogen in newborn piglets. The clarifications of the overall antibody repertoire and antigen-specific antibody repertoire are essential to provide important insights into the B-cell response and reshape new vaccines. Here, we applied next-generation sequencing (NGS) technology to investigate immunoglobulin (Ig) variable (V) gene segment usage of swine B-cells from peripheral blood lymphocytes (PBL) and mesenteric lymph node (MLN) cells following PEDV vaccination. We identified the transcripts of all functional Ig V-genes in antibody repertoire. IgHV1S2, IgKV1-11, and IgLV3-4 were the most prevalent gene segments for heavy, kappa, and lambda chains, respectively, in PBL and MLN. Unlike previous studies, IgKV1, instead of IgKV2, and IgLV3, instead of IgLV8, were the prevalent Ig V-gene families for kappa and lambda light chains, respectively. We further examined the antibody repertoire of PEDV spike-specific B cells by single-cell RT-PCR. In contrast to the overall antibody repertoire, Ig V-gene segments of PEDV spike-specific B cells preferentially adopted IgHV1-4 and IgHV1-14 for heavy chain, IgKV1-11 for kappa chain, and IgLV3-3 for lambda chain. These results represent a comprehensive analysis to characterize the Ig V-gene segment usage in the overall and PEDV spike-specific antibody repertoire in PBL and MLN.

Keywords: porcine epidemic diarrhea virus (PEDV), next-generation sequencing (NGS), single-cell RT-PCR, antibody repertoire

Acknowledgements

This work was supported by the National Natural Science Foundation of China (31772718) and the Open Research Fund of State Key Laboratory of Veterinary Biotechnology (SKLVBF2018XX). We thank Dr. Caixia Gao (Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, China) for assistance and advice in swine leukocyte antigens (SLA) types identification.

Footnotes

Compliance and ethics

The author(s) declare that they have no conflict of interest.

Contributor Information

Li Feng, Email: fengli@caas.cn.

PingHuang Liu, Email: liupinghuang@caas.cn.

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