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表达外源抗原的重组绵羊痘病毒的构建

Engineering of Recombinant Sheep Pox Viruses Expressing Foreign Antigens.

作者信息

Chervyakova Olga, Tailakova Elmira, Kozhabergenov Nurlan, Sadikaliyeva Sandugash, Sultankulova Kulyaisan, Zakarya Kunsulu, Maksyutov Rinat A, Strochkov Vitaliy, Sandybayev Nurlan

机构信息

Research Institute for Biological Safety Problems, RK ME&S-Science Committee, Gvardeiskiy 080409, Kazakhstan.

State Research Center of Virology and Biotechnology "Vector", Koltsovo, 630559 Novosibirsk Region, Russia.

出版信息

Microorganisms. 2021 May 7;9(5):1005. doi: 10.3390/microorganisms9051005.

DOI:10.3390/microorganisms9051005
PMID:34067124
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8150597/
Abstract

Capripoxviruses with a host range limited to ruminants have the great potential to be used as vaccine vectors. The aim of this work was to evaluate attenuated sheep pox virus (SPPV) vaccine strain NISKHI as a vector expressing several genes. Open reading frames SPPV020 (ribonucleotide kinase) and SPPV066 (thymidine kinase) were selected as sites for the insertion of foreign genes. Two integration plasmids with expression cassette were designed and constructed. Recombinant SPPVs expressing an enhanced green fluorescent protein (EGFP) (rSPPV(RRΔ)EGFP and rSPPV(TKΔ)EGFP), Foot-and-mouth disease virus capsid protein (VP1), and spp. outer membrane protein 25 (OMP25) (rSPPV(RRΔ)VP1A-(TKΔ)OMP25) were generated under the transient dominant selection method. The insertion of foreign genes into the SPPV020 and SPPV066 open reading frames did not influence the replication of the recombinant viruses in the cells. Successful foreign gene expression in vitro was assessed by luminescent microscopy (EGFP) and Western blot (VP1 and OMP25). Our results have shown that foreign genes were expressed by rSPPV both in permissive (lamb testicles) and non-permissive (bovine kidney, saiga kidney, porcine kidney) cells. Mice immunized with rSPPV(RRΔ)VP1A-(TKΔ)OMP25 elicited specific antibodies to both SPPV and foreign genes VP1 and OMP25. Thus, SPPV NISKHI may be used as a potential safe immunogenic viral vector for the development of polyvalent vaccines.

摘要

宿主范围仅限于反刍动物的山羊痘病毒具有用作疫苗载体的巨大潜力。本研究的目的是评估减毒绵羊痘病毒（SPPV）疫苗株NISKHI作为表达多个基因的载体。选择开放阅读框SPPV020（核糖核苷酸激酶）和SPPV066（胸苷激酶）作为插入外源基因的位点。设计并构建了两个带有表达盒的整合质粒。在瞬时显性选择方法下，产生了表达增强型绿色荧光蛋白（EGFP）的重组SPPV（rSPPV(RRΔ)EGFP和rSPPV(TKΔ)EGFP）、口蹄疫病毒衣壳蛋白（VP1）和 种外膜蛋白25（OMP25）（rSPPV(RRΔ)VP1A-(TKΔ)OMP25）。将外源基因插入SPPV020和SPPV066开放阅读框并不影响重组病毒在细胞中的复制。通过荧光显微镜（EGFP）和蛋白质印迹（VP1和OMP25）评估了外源基因在体外的成功表达。我们的结果表明，rSPPV在允许细胞（羔羊睾丸）和非允许细胞（牛肾、赛加羚羊肾、猪肾）中均表达了外源基因。用rSPPV(RRΔ)VP1A-(TKΔ)OMP25免疫的小鼠产生了针对SPPV以及外源基因VP1和OMP25的特异性抗体。因此，SPPV NISKHI可作为开发多价疫苗的潜在安全免疫原性病毒载体。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/a14ec9a44fad/microorganisms-09-01005-g008.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/aaa707779823/microorganisms-09-01005-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/27add201ee5d/microorganisms-09-01005-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/1ed80038597e/microorganisms-09-01005-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/8d5eae592473/microorganisms-09-01005-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/f4a4264f3be4/microorganisms-09-01005-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/f31a405e92a3/microorganisms-09-01005-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/0cd641053ff4/microorganisms-09-01005-g007.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/a14ec9a44fad/microorganisms-09-01005-g008.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/aaa707779823/microorganisms-09-01005-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/27add201ee5d/microorganisms-09-01005-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/1ed80038597e/microorganisms-09-01005-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/8d5eae592473/microorganisms-09-01005-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/f4a4264f3be4/microorganisms-09-01005-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/f31a405e92a3/microorganisms-09-01005-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/0cd641053ff4/microorganisms-09-01005-g007.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/48bc/8150597/a14ec9a44fad/microorganisms-09-01005-g008.jpg

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