Isolation and Identification of a Rare Spike Gene Double-Deletion SARS-CoV-2 Variant From the Patient With High Cycle Threshold Value
Coronavirus disease 2019 (COVID-19) is an emerging life-threatening pulmonary disease caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which originated in Wuhan, Hubei Province, China, in December 2019. COVID-19 develops after close contact via inhalation of res...
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Frontiers Media S.A.
2022-01-01
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| Series: | Frontiers in Medicine |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fmed.2021.822633/full |
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| author | Li-Teh Liu Jih-Jin Tsai Jih-Jin Tsai Jih-Jin Tsai Chun-Hong Chen Chun-Hong Chen Ping-Chang Lin Ching-Yi Tsai Yan-Yi Tsai Miao-Chen Hsu Wan-Long Chuang Wan-Long Chuang Jer-Ming Chang Jer-Ming Chang Jer-Ming Chang Shang-Jyh Hwang Shang-Jyh Hwang Inn-Wen Chong Inn-Wen Chong |
| author_facet | Li-Teh Liu Jih-Jin Tsai Jih-Jin Tsai Jih-Jin Tsai Chun-Hong Chen Chun-Hong Chen Ping-Chang Lin Ching-Yi Tsai Yan-Yi Tsai Miao-Chen Hsu Wan-Long Chuang Wan-Long Chuang Jer-Ming Chang Jer-Ming Chang Jer-Ming Chang Shang-Jyh Hwang Shang-Jyh Hwang Inn-Wen Chong Inn-Wen Chong |
| author_sort | Li-Teh Liu |
| collection | DOAJ |
| description | Coronavirus disease 2019 (COVID-19) is an emerging life-threatening pulmonary disease caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which originated in Wuhan, Hubei Province, China, in December 2019. COVID-19 develops after close contact via inhalation of respiratory droplets containing SARS-CoV-2 during talking, coughing, or sneezing by asymptomatic, presymptomatic, and symptomatic carriers. This virus evolved over time, and numerous genetic variants have been reported to have increased disease severity, mortality, and transmissibility. Variants have also developed resistance to antivirals and vaccination and can escape the immune response of humans. Reverse transcription polymerase chain reaction (RT–PCR) is the method of choice among diagnostic techniques, including nucleic acid amplification tests (NAATs), serological tests, and diagnostic imaging, such as computed tomography (CT). The limitation of RT–PCR is that it cannot distinguish fragmented RNA genomes from live transmissible viruses. Thus, SARS-CoV-2 isolation by using cell culture has been developed and makes important contributions in the field of diagnosis, development of antivirals, vaccines, and SARS-CoV-2 virology research. In this research, two SARS-CoV-2 strains were isolated from four RT–PCR-positive nasopharyngeal swabs using VERO E6 cell culture. One isolate was cultured successfully with a blind passage on day 3 post inoculation from a swab with a Ct > 35, while the cells did not develop cytopathic effects without a blind passage until day 14 post inoculation. Our results indicated that infectious SARS-CoV-2 virus particles existed, even with a Ct > 35. Cultivable viruses could provide additional consideration for releasing the patient from quarantine. The results of the whole genome sequencing and bioinformatic analysis suggested that these two isolates contain a spike 68-76del+spike 675-679del double-deletion variation. The double deletion was confirmed by amplification of the regions spanning the spike gene deletion using Sanger sequencing. Phylogenetic analysis revealed that this double-deletion variant was rare (one per million in public databases, including GenBank and GISAID). The impact of this double deletion in the spike gene on the SARS-CoV-2 virus itself as well as on cultured cells and/or humans remains to be further elucidated. |
| first_indexed | 2024-04-11T20:29:24Z |
| format | Article |
| id | doaj.art-845a20f862a84bce86a681a1d42cb578 |
| institution | Directory Open Access Journal |
| issn | 2296-858X |
| language | English |
| last_indexed | 2024-04-11T20:29:24Z |
| publishDate | 2022-01-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Medicine |
| spelling | doaj.art-845a20f862a84bce86a681a1d42cb5782022-12-22T04:04:33ZengFrontiers Media S.A.Frontiers in Medicine2296-858X2022-01-01810.3389/fmed.2021.822633822633Isolation and Identification of a Rare Spike Gene Double-Deletion SARS-CoV-2 Variant From the Patient With High Cycle Threshold ValueLi-Teh Liu0Jih-Jin Tsai1Jih-Jin Tsai2Jih-Jin Tsai3Chun-Hong Chen4Chun-Hong Chen5Ping-Chang Lin6Ching-Yi Tsai7Yan-Yi Tsai8Miao-Chen Hsu9Wan-Long Chuang10Wan-Long Chuang11Jer-Ming Chang12Jer-Ming Chang13Jer-Ming Chang14Shang-Jyh Hwang15Shang-Jyh Hwang16Inn-Wen Chong17Inn-Wen Chong18Department of Medical Laboratory Science and Biotechnology, College of Medical Technology, Chung-Hwa University of Medical Technology, Tainan, TaiwanTropical Medicine Center, Kaohsiung Medical University Hospital, Kaohsiung, TaiwanDivision of Infectious Diseases, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, TaiwanSchool of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, TaiwanNational Mosquito-Borne Diseases Control Research Center, National Health Research Institutes, Zhunan, TaiwanNational Institute of Infectious Diseases and Vaccinology, National Health Research Institutes, Zhunan, TaiwanTropical Medicine Center, Kaohsiung Medical University Hospital, Kaohsiung, TaiwanTropical Medicine Center, Kaohsiung Medical University Hospital, Kaohsiung, TaiwanTropical Medicine Center, Kaohsiung Medical University Hospital, Kaohsiung, TaiwanTropical Medicine Center, Kaohsiung Medical University Hospital, Kaohsiung, TaiwanSchool of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, TaiwanDivision of Hepatobiliary and Pancreatic, Kaohsiung Medical University Hospital, Kaohsiung, TaiwanSchool of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, TaiwanDivision of Nephrology, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, TaiwanDepartment of Medical Research, Kaohsiung Medical University Hospital, Kaohsiung, TaiwanSchool of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, TaiwanDivision of Nephrology, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan0Department of Internal Medicine and Graduate Institute of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan1Department of Pulmonary Medicine, Kaohsiung Medical University Hospital, Kaohsiung, TaiwanCoronavirus disease 2019 (COVID-19) is an emerging life-threatening pulmonary disease caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which originated in Wuhan, Hubei Province, China, in December 2019. COVID-19 develops after close contact via inhalation of respiratory droplets containing SARS-CoV-2 during talking, coughing, or sneezing by asymptomatic, presymptomatic, and symptomatic carriers. This virus evolved over time, and numerous genetic variants have been reported to have increased disease severity, mortality, and transmissibility. Variants have also developed resistance to antivirals and vaccination and can escape the immune response of humans. Reverse transcription polymerase chain reaction (RT–PCR) is the method of choice among diagnostic techniques, including nucleic acid amplification tests (NAATs), serological tests, and diagnostic imaging, such as computed tomography (CT). The limitation of RT–PCR is that it cannot distinguish fragmented RNA genomes from live transmissible viruses. Thus, SARS-CoV-2 isolation by using cell culture has been developed and makes important contributions in the field of diagnosis, development of antivirals, vaccines, and SARS-CoV-2 virology research. In this research, two SARS-CoV-2 strains were isolated from four RT–PCR-positive nasopharyngeal swabs using VERO E6 cell culture. One isolate was cultured successfully with a blind passage on day 3 post inoculation from a swab with a Ct > 35, while the cells did not develop cytopathic effects without a blind passage until day 14 post inoculation. Our results indicated that infectious SARS-CoV-2 virus particles existed, even with a Ct > 35. Cultivable viruses could provide additional consideration for releasing the patient from quarantine. The results of the whole genome sequencing and bioinformatic analysis suggested that these two isolates contain a spike 68-76del+spike 675-679del double-deletion variation. The double deletion was confirmed by amplification of the regions spanning the spike gene deletion using Sanger sequencing. Phylogenetic analysis revealed that this double-deletion variant was rare (one per million in public databases, including GenBank and GISAID). The impact of this double deletion in the spike gene on the SARS-CoV-2 virus itself as well as on cultured cells and/or humans remains to be further elucidated.https://www.frontiersin.org/articles/10.3389/fmed.2021.822633/fullCOVID-19SARS-CoV-2RT–PCRCtvirus culturespike gene |
| spellingShingle | Li-Teh Liu Jih-Jin Tsai Jih-Jin Tsai Jih-Jin Tsai Chun-Hong Chen Chun-Hong Chen Ping-Chang Lin Ching-Yi Tsai Yan-Yi Tsai Miao-Chen Hsu Wan-Long Chuang Wan-Long Chuang Jer-Ming Chang Jer-Ming Chang Jer-Ming Chang Shang-Jyh Hwang Shang-Jyh Hwang Inn-Wen Chong Inn-Wen Chong Isolation and Identification of a Rare Spike Gene Double-Deletion SARS-CoV-2 Variant From the Patient With High Cycle Threshold Value Frontiers in Medicine COVID-19 SARS-CoV-2 RT–PCR Ct virus culture spike gene |
| title | Isolation and Identification of a Rare Spike Gene Double-Deletion SARS-CoV-2 Variant From the Patient With High Cycle Threshold Value |
| title_full | Isolation and Identification of a Rare Spike Gene Double-Deletion SARS-CoV-2 Variant From the Patient With High Cycle Threshold Value |
| title_fullStr | Isolation and Identification of a Rare Spike Gene Double-Deletion SARS-CoV-2 Variant From the Patient With High Cycle Threshold Value |
| title_full_unstemmed | Isolation and Identification of a Rare Spike Gene Double-Deletion SARS-CoV-2 Variant From the Patient With High Cycle Threshold Value |
| title_short | Isolation and Identification of a Rare Spike Gene Double-Deletion SARS-CoV-2 Variant From the Patient With High Cycle Threshold Value |
| title_sort | isolation and identification of a rare spike gene double deletion sars cov 2 variant from the patient with high cycle threshold value |
| topic | COVID-19 SARS-CoV-2 RT–PCR Ct virus culture spike gene |
| url | https://www.frontiersin.org/articles/10.3389/fmed.2021.822633/full |
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