Generation and Characterization of a Replication-Competent Human Adenovirus Type 55 Encoding EGFP
Human adenovirus 55 (HAdV-55) has recently caused outbreaks of acute respiratory disease (ARD), posing a significant public threat to civilians and military trainees. Efforts to develop antiviral inhibitors and quantify neutralizing antibodies require an experimental system to rapidly monitor viral...
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MDPI AG
2023-05-01
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| Online Access: | https://www.mdpi.com/1999-4915/15/5/1192 |
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| author | Wei Li Yuehong Chen Ye Feng Jing Li Xiaoping Kang Sen Zhang Yuchang Li Zhiyan Zhao Wenguang Yang Lu Zhao Huiyao Wang Tao Jiang |
| author_facet | Wei Li Yuehong Chen Ye Feng Jing Li Xiaoping Kang Sen Zhang Yuchang Li Zhiyan Zhao Wenguang Yang Lu Zhao Huiyao Wang Tao Jiang |
| author_sort | Wei Li |
| collection | DOAJ |
| description | Human adenovirus 55 (HAdV-55) has recently caused outbreaks of acute respiratory disease (ARD), posing a significant public threat to civilians and military trainees. Efforts to develop antiviral inhibitors and quantify neutralizing antibodies require an experimental system to rapidly monitor viral infections, which can be achieved through the use of a plasmid that can produce an infectious virus. Here, we used a bacteria-mediated recombination approach to construct a full-length infectious cDNA clone, pAd55-FL, containing the whole genome of HadV-55. Then, the green fluorescent protein expression cassette was assembled into pAd55-FL to replace the E3 region to obtain a recombinant plasmid of pAd55-dE3-EGFP. The rescued recombinant virus rAdv55-dE3-EGFP is genetically stable and replicates similarly to the wild-type virus in cell culture. The virus rAdv55-dE3-EGFP can be used to quantify neutralizing antibody activity in sera samples, producing results in concordance with the cytopathic effect (CPE)-based microneutralization assay. Using an rAdv55-dE3-EGFP infection of A549 cells, we showed that the assay could be used for antiviral screening. Our findings suggest that the rAdv55-dE3-EGFP-based high-throughput assay provides a reliable tool for rapid neutralization testing and antiviral screening for HAdV-55. |
| first_indexed | 2024-03-11T03:14:08Z |
| format | Article |
| id | doaj.art-1ba1a939a15f4619853d9e536f0f7ac7 |
| institution | Directory Open Access Journal |
| issn | 1999-4915 |
| language | English |
| last_indexed | 2024-03-11T03:14:08Z |
| publishDate | 2023-05-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Viruses |
| spelling | doaj.art-1ba1a939a15f4619853d9e536f0f7ac72023-11-18T03:40:17ZengMDPI AGViruses1999-49152023-05-01155119210.3390/v15051192Generation and Characterization of a Replication-Competent Human Adenovirus Type 55 Encoding EGFPWei Li0Yuehong Chen1Ye Feng2Jing Li3Xiaoping Kang4Sen Zhang5Yuchang Li6Zhiyan Zhao7Wenguang Yang8Lu Zhao9Huiyao Wang10Tao Jiang11State Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaState Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaState Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaState Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaState Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaState Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaState Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaState Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaState Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaState Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaState Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaState Key Laboratory of Pathogen and Biosecurity, Institute of Microbiology and Epidemiology, Academy of Military Medical Sciences, Beijing 100071, ChinaHuman adenovirus 55 (HAdV-55) has recently caused outbreaks of acute respiratory disease (ARD), posing a significant public threat to civilians and military trainees. Efforts to develop antiviral inhibitors and quantify neutralizing antibodies require an experimental system to rapidly monitor viral infections, which can be achieved through the use of a plasmid that can produce an infectious virus. Here, we used a bacteria-mediated recombination approach to construct a full-length infectious cDNA clone, pAd55-FL, containing the whole genome of HadV-55. Then, the green fluorescent protein expression cassette was assembled into pAd55-FL to replace the E3 region to obtain a recombinant plasmid of pAd55-dE3-EGFP. The rescued recombinant virus rAdv55-dE3-EGFP is genetically stable and replicates similarly to the wild-type virus in cell culture. The virus rAdv55-dE3-EGFP can be used to quantify neutralizing antibody activity in sera samples, producing results in concordance with the cytopathic effect (CPE)-based microneutralization assay. Using an rAdv55-dE3-EGFP infection of A549 cells, we showed that the assay could be used for antiviral screening. Our findings suggest that the rAdv55-dE3-EGFP-based high-throughput assay provides a reliable tool for rapid neutralization testing and antiviral screening for HAdV-55.https://www.mdpi.com/1999-4915/15/5/1192human adenovirus type 55infectious clonereporter virusenhanced GFPreplication-competenthomologous recombination |
| spellingShingle | Wei Li Yuehong Chen Ye Feng Jing Li Xiaoping Kang Sen Zhang Yuchang Li Zhiyan Zhao Wenguang Yang Lu Zhao Huiyao Wang Tao Jiang Generation and Characterization of a Replication-Competent Human Adenovirus Type 55 Encoding EGFP Viruses human adenovirus type 55 infectious clone reporter virus enhanced GFP replication-competent homologous recombination |
| title | Generation and Characterization of a Replication-Competent Human Adenovirus Type 55 Encoding EGFP |
| title_full | Generation and Characterization of a Replication-Competent Human Adenovirus Type 55 Encoding EGFP |
| title_fullStr | Generation and Characterization of a Replication-Competent Human Adenovirus Type 55 Encoding EGFP |
| title_full_unstemmed | Generation and Characterization of a Replication-Competent Human Adenovirus Type 55 Encoding EGFP |
| title_short | Generation and Characterization of a Replication-Competent Human Adenovirus Type 55 Encoding EGFP |
| title_sort | generation and characterization of a replication competent human adenovirus type 55 encoding egfp |
| topic | human adenovirus type 55 infectious clone reporter virus enhanced GFP replication-competent homologous recombination |
| url | https://www.mdpi.com/1999-4915/15/5/1192 |
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