Construction of enterovirus G expressing reporter genes for antiviral drug screening assays
Abstract The emergence of Enterovirus G strains harboring recombinant papain-like protease (EV-G-PLP) poses a significant threat to the global swine health with zoonotic potential. Addressing the critical lack of targeted treatments, we developed a reverse genetics system for the CH/20GXNN/PLP2020 s...
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| Format: | Article |
| Language: | English |
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BMC
2025-08-01
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| Series: | BMC Veterinary Research |
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| Online Access: | https://doi.org/10.1186/s12917-025-04960-0 |
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| author | Kaige Chen Dalin Hong Lingyou Zeng Jinni Bian Shiting Huang Yifeng Qin Yeshi Yin Weijian Huang Ying Chen Zuzhang Wei Kang Ouyang |
| author_facet | Kaige Chen Dalin Hong Lingyou Zeng Jinni Bian Shiting Huang Yifeng Qin Yeshi Yin Weijian Huang Ying Chen Zuzhang Wei Kang Ouyang |
| author_sort | Kaige Chen |
| collection | DOAJ |
| description | Abstract The emergence of Enterovirus G strains harboring recombinant papain-like protease (EV-G-PLP) poses a significant threat to the global swine health with zoonotic potential. Addressing the critical lack of targeted treatments, we developed a reverse genetics system for the CH/20GXNN/PLP2020 strain and generated three isogenic reporter recombinants (GFP/RFP/iLOV) through precise PLP gene substitution. Viral progeny exhibited parental-like replication kinetics, with r20GXNN-iLOV demonstrating genetic stability (> 10 passages) compared to GFP/RFP variants. Plaque phenotyping revealed an inverse correlation between plaque size and insert length at the 2C/3A junction. Leveraging this system, we engineered a high-throughput screening platform identifying four anti-EV-G compounds: niclosamide (0.02 µM), salinomycin (2 µM), chloroquine phosphate (12 µM), and ribavirin (400 µM). This study establishes a reverse genetics platform enabling picornavirus research advancement and reveals structural plasticity in the 2C/3A junction that facilitates antiviral screening. Furthermore, it identifies EV-G-specific drug candidates while providing a framework for pan-picornaviral (including EV-A71) compound discovery. |
| format | Article |
| id | doaj-art-74ca79397e204d3b91fe76150f92e947 |
| institution | Kabale University |
| issn | 1746-6148 |
| language | English |
| publishDate | 2025-08-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Veterinary Research |
| spelling | doaj-art-74ca79397e204d3b91fe76150f92e9472025-08-20T03:46:03ZengBMCBMC Veterinary Research1746-61482025-08-0121111110.1186/s12917-025-04960-0Construction of enterovirus G expressing reporter genes for antiviral drug screening assaysKaige Chen0Dalin Hong1Lingyou Zeng2Jinni Bian3Shiting Huang4Yifeng Qin5Yeshi Yin6Weijian Huang7Ying Chen8Zuzhang Wei9Kang Ouyang10College of Animal Science and Technology, Guangxi UniversityCollege of Animal Science and Technology, Guangxi UniversityCollege of Animal Science and Technology, Guangxi UniversityCollege of Animal Science and Technology, Guangxi UniversityCollege of Animal Science and Technology, Guangxi UniversityCollege of Animal Science and Technology, Guangxi UniversityCollege of Animal Science and Technology, Guangxi UniversityCollege of Animal Science and Technology, Guangxi UniversityCollege of Animal Science and Technology, Guangxi UniversityCollege of Animal Science and Technology, Guangxi UniversityCollege of Animal Science and Technology, Guangxi UniversityAbstract The emergence of Enterovirus G strains harboring recombinant papain-like protease (EV-G-PLP) poses a significant threat to the global swine health with zoonotic potential. Addressing the critical lack of targeted treatments, we developed a reverse genetics system for the CH/20GXNN/PLP2020 strain and generated three isogenic reporter recombinants (GFP/RFP/iLOV) through precise PLP gene substitution. Viral progeny exhibited parental-like replication kinetics, with r20GXNN-iLOV demonstrating genetic stability (> 10 passages) compared to GFP/RFP variants. Plaque phenotyping revealed an inverse correlation between plaque size and insert length at the 2C/3A junction. Leveraging this system, we engineered a high-throughput screening platform identifying four anti-EV-G compounds: niclosamide (0.02 µM), salinomycin (2 µM), chloroquine phosphate (12 µM), and ribavirin (400 µM). This study establishes a reverse genetics platform enabling picornavirus research advancement and reveals structural plasticity in the 2C/3A junction that facilitates antiviral screening. Furthermore, it identifies EV-G-specific drug candidates while providing a framework for pan-picornaviral (including EV-A71) compound discovery.https://doi.org/10.1186/s12917-025-04960-0Enterovirus GInfectious cloneReporter geneAntiviral drug screening |
| spellingShingle | Kaige Chen Dalin Hong Lingyou Zeng Jinni Bian Shiting Huang Yifeng Qin Yeshi Yin Weijian Huang Ying Chen Zuzhang Wei Kang Ouyang Construction of enterovirus G expressing reporter genes for antiviral drug screening assays BMC Veterinary Research Enterovirus G Infectious clone Reporter gene Antiviral drug screening |
| title | Construction of enterovirus G expressing reporter genes for antiviral drug screening assays |
| title_full | Construction of enterovirus G expressing reporter genes for antiviral drug screening assays |
| title_fullStr | Construction of enterovirus G expressing reporter genes for antiviral drug screening assays |
| title_full_unstemmed | Construction of enterovirus G expressing reporter genes for antiviral drug screening assays |
| title_short | Construction of enterovirus G expressing reporter genes for antiviral drug screening assays |
| title_sort | construction of enterovirus g expressing reporter genes for antiviral drug screening assays |
| topic | Enterovirus G Infectious clone Reporter gene Antiviral drug screening |
| url | https://doi.org/10.1186/s12917-025-04960-0 |
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