Despite low viral titer in saliva samples, Sanger-based SARS-CoV-2 spike gene sequencing is highly applicable for the variant identification
Abstract Background This study aimed to compare the performance of Sanger-based SARS-CoV-2 spike gene sequencing and Next Generation Sequencing (NGS)-based full-genome sequencing for variant identification in saliva samples with low viral titer. Methods Using 241 stocked saliva samples collected fro...
| Main Authors: | , , , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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BMC
2023-08-01
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| Series: | BMC Medical Genomics |
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| Online Access: | https://doi.org/10.1186/s12920-023-01633-5 |
| _version_ | 1797451154412535808 |
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| author | Ko Ko Kazuaki Takahashi Noriaki Ito Aya Sugiyama Shintaro Nagashima Kei Miwata Yoshihiro Kitahara Mafumi Okimoto Serge Ouoba Golda Ataa Akuffo Bunthen E Tomoyuki Akita Toshiro Takafuta Junko Tanaka |
| author_facet | Ko Ko Kazuaki Takahashi Noriaki Ito Aya Sugiyama Shintaro Nagashima Kei Miwata Yoshihiro Kitahara Mafumi Okimoto Serge Ouoba Golda Ataa Akuffo Bunthen E Tomoyuki Akita Toshiro Takafuta Junko Tanaka |
| author_sort | Ko Ko |
| collection | DOAJ |
| description | Abstract Background This study aimed to compare the performance of Sanger-based SARS-CoV-2 spike gene sequencing and Next Generation Sequencing (NGS)-based full-genome sequencing for variant identification in saliva samples with low viral titer. Methods Using 241 stocked saliva samples collected from confirmed COVID-19 patients between November 2020 and March 2022 in Hiroshima, SARS-CoV-2 spike gene sequencing (nt22735-nt23532) was performed by nested RT-PCR and Sanger platform using in-house primers. The same samples underwent full-genome sequencing by NGS using Illumina NextSeq2000. Results Among 241 samples, 147 were amplified by both the Sanger and the Illumina NextSeq2000 NGS, 86 by Sanger only, and 8 were not amplified at all. The overall amplification rates of Illumina NextSeq2000 NGS and Sanger were 61% and 96.7%, respectively. At low viral titer (< 103 copies/mL), Illumina NextSeq2000 NGS provided 19.2% amplification, while Sanger was 89.7% (p < 0.0001). Both platforms identified 38 wild type, 54 Alpha variants, 84 Delta variants, and 57 Omicron variants. Conclusions Our study provided evidence to expand the capacity of Sanger-based SARS-CoV-2 spike gene sequencing for variants identification over full-genome by Illumina NextSeq2000 NGS for mass screening. Therefore, the feasible and simple Sanger-based SARS-CoV-2 spike gene sequencing is practical for the initial variants screening, which might reduce the gap between the rapid evolution of SARS-CoV-2 and its molecular surveillance. |
| first_indexed | 2024-03-09T14:50:44Z |
| format | Article |
| id | doaj.art-d1ff00131ff147de8e0b12f2d80561b0 |
| institution | Directory Open Access Journal |
| issn | 1755-8794 |
| language | English |
| last_indexed | 2024-03-09T14:50:44Z |
| publishDate | 2023-08-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Medical Genomics |
| spelling | doaj.art-d1ff00131ff147de8e0b12f2d80561b02023-11-26T14:30:26ZengBMCBMC Medical Genomics1755-87942023-08-011611910.1186/s12920-023-01633-5Despite low viral titer in saliva samples, Sanger-based SARS-CoV-2 spike gene sequencing is highly applicable for the variant identificationKo Ko0Kazuaki Takahashi1Noriaki Ito2Aya Sugiyama3Shintaro Nagashima4Kei Miwata5Yoshihiro Kitahara6Mafumi Okimoto7Serge Ouoba8Golda Ataa Akuffo9Bunthen E10Tomoyuki Akita11Toshiro Takafuta12Junko Tanaka13Department of Epidemiology, Infectious Disease Control and Prevention, Graduate School of Biomedical and Health Sciences, Hiroshima UniversityDepartment of Epidemiology, Infectious Disease Control and Prevention, Graduate School of Biomedical and Health Sciences, Hiroshima UniversityHiroshima City Funairi Citizens HospitalDepartment of Epidemiology, Infectious Disease Control and Prevention, Graduate School of Biomedical and Health Sciences, Hiroshima UniversityDepartment of Epidemiology, Infectious Disease Control and Prevention, Graduate School of Biomedical and Health Sciences, Hiroshima UniversityHiroshima City Funairi Citizens HospitalHiroshima City Funairi Citizens HospitalHiroshima City Funairi Citizens HospitalDepartment of Epidemiology, Infectious Disease Control and Prevention, Graduate School of Biomedical and Health Sciences, Hiroshima UniversityDepartment of Epidemiology, Infectious Disease Control and Prevention, Graduate School of Biomedical and Health Sciences, Hiroshima UniversityDepartment of Epidemiology, Infectious Disease Control and Prevention, Graduate School of Biomedical and Health Sciences, Hiroshima UniversityDepartment of Epidemiology, Infectious Disease Control and Prevention, Graduate School of Biomedical and Health Sciences, Hiroshima UniversityHiroshima City Funairi Citizens HospitalDepartment of Epidemiology, Infectious Disease Control and Prevention, Graduate School of Biomedical and Health Sciences, Hiroshima UniversityAbstract Background This study aimed to compare the performance of Sanger-based SARS-CoV-2 spike gene sequencing and Next Generation Sequencing (NGS)-based full-genome sequencing for variant identification in saliva samples with low viral titer. Methods Using 241 stocked saliva samples collected from confirmed COVID-19 patients between November 2020 and March 2022 in Hiroshima, SARS-CoV-2 spike gene sequencing (nt22735-nt23532) was performed by nested RT-PCR and Sanger platform using in-house primers. The same samples underwent full-genome sequencing by NGS using Illumina NextSeq2000. Results Among 241 samples, 147 were amplified by both the Sanger and the Illumina NextSeq2000 NGS, 86 by Sanger only, and 8 were not amplified at all. The overall amplification rates of Illumina NextSeq2000 NGS and Sanger were 61% and 96.7%, respectively. At low viral titer (< 103 copies/mL), Illumina NextSeq2000 NGS provided 19.2% amplification, while Sanger was 89.7% (p < 0.0001). Both platforms identified 38 wild type, 54 Alpha variants, 84 Delta variants, and 57 Omicron variants. Conclusions Our study provided evidence to expand the capacity of Sanger-based SARS-CoV-2 spike gene sequencing for variants identification over full-genome by Illumina NextSeq2000 NGS for mass screening. Therefore, the feasible and simple Sanger-based SARS-CoV-2 spike gene sequencing is practical for the initial variants screening, which might reduce the gap between the rapid evolution of SARS-CoV-2 and its molecular surveillance.https://doi.org/10.1186/s12920-023-01633-5SARS-CoV-2Sanger methodNext generation sequencingAmplificationVariantsScreening |
| spellingShingle | Ko Ko Kazuaki Takahashi Noriaki Ito Aya Sugiyama Shintaro Nagashima Kei Miwata Yoshihiro Kitahara Mafumi Okimoto Serge Ouoba Golda Ataa Akuffo Bunthen E Tomoyuki Akita Toshiro Takafuta Junko Tanaka Despite low viral titer in saliva samples, Sanger-based SARS-CoV-2 spike gene sequencing is highly applicable for the variant identification BMC Medical Genomics SARS-CoV-2 Sanger method Next generation sequencing Amplification Variants Screening |
| title | Despite low viral titer in saliva samples, Sanger-based SARS-CoV-2 spike gene sequencing is highly applicable for the variant identification |
| title_full | Despite low viral titer in saliva samples, Sanger-based SARS-CoV-2 spike gene sequencing is highly applicable for the variant identification |
| title_fullStr | Despite low viral titer in saliva samples, Sanger-based SARS-CoV-2 spike gene sequencing is highly applicable for the variant identification |
| title_full_unstemmed | Despite low viral titer in saliva samples, Sanger-based SARS-CoV-2 spike gene sequencing is highly applicable for the variant identification |
| title_short | Despite low viral titer in saliva samples, Sanger-based SARS-CoV-2 spike gene sequencing is highly applicable for the variant identification |
| title_sort | despite low viral titer in saliva samples sanger based sars cov 2 spike gene sequencing is highly applicable for the variant identification |
| topic | SARS-CoV-2 Sanger method Next generation sequencing Amplification Variants Screening |
| url | https://doi.org/10.1186/s12920-023-01633-5 |
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