Nanopore sequencing reveals the genomic diversity of the variants of concern of SARS-CoV-2 during 2021 disease outbreak in Pakistan
Abstract The global spread of SARS-CoV-2 was significantly impacted by the emergence of Variants of Concern (VOC), with Pakistan experiencing similar trends to other countries. To gain a comprehensive understanding of the genomic epidemiology of SARS-CoV-2 strains circulating during Pakistan’s third...
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Nature Portfolio
2025-08-01
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| Online Access: | https://doi.org/10.1038/s41598-025-12774-1 |
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| author | Zohaib Ul Hassan Minkyu Park Dongju Park Humaira Amin Ibrar Ahmed Nazish Badar Asif Ali Muhammad Salman Massab Umair Hamza Ahmed Mirza Abdul Ahad Maryam Muzaffar Falak Fazal Sana Bibi Madiha Islam Yasar Mehmood Yousafzai Sher Shah Muhammad Zakria Zia Ul Haq Seil Kim |
| author_facet | Zohaib Ul Hassan Minkyu Park Dongju Park Humaira Amin Ibrar Ahmed Nazish Badar Asif Ali Muhammad Salman Massab Umair Hamza Ahmed Mirza Abdul Ahad Maryam Muzaffar Falak Fazal Sana Bibi Madiha Islam Yasar Mehmood Yousafzai Sher Shah Muhammad Zakria Zia Ul Haq Seil Kim |
| author_sort | Zohaib Ul Hassan |
| collection | DOAJ |
| description | Abstract The global spread of SARS-CoV-2 was significantly impacted by the emergence of Variants of Concern (VOC), with Pakistan experiencing similar trends to other countries. To gain a comprehensive understanding of the genomic epidemiology of SARS-CoV-2 strains circulating during Pakistan’s third and fourth pandemic waves, we conducted the largest single sequencing effort in the country to date. Using the GridION platform (Oxford Nanopore Technologies), we performed whole genome sequencing on 1052 confirmed COVID-19 patient samples collected from multiple cities across Pakistan between March and October 2021. Our analysis revealed a clear temporal shift in variant dominance. The Alpha variant (B.1.1.7 lineage) predominated in the first half of 2021, while the Delta variant (B.1.617.2) became most prevalent in the latter half. This transition reflects global trends and provides crucial insights into the timing and dynamics of this shift within Pakistan. Mutational analysis revealed that the most frequent nucleotide mutations in Pakistani SARS-CoV-2 samples were A23403G (associated with the D614G mutation in the spike protein), C3037T, C14408T, and C241T, potentially contributing to increased disease transmission and evasion of host immune responses. The rapid evolution and spread of these circulating variants highlight the possibility of novel variants emerging with enhanced mutational fitness. The AY.108 lineage, which has been reported at relatively low frequencies globally including in Europe and North America, with Pakistan accounting for approximately 34% of global cases, suggesting potential regional evolution or specific introduction events. Our findings underscore the dynamic nature of SARS-CoV-2 and emphasize the critical importance of ongoing, large-scale genomic surveillance in Pakistan. This study demonstrates the feasibility and utility of using nanopore sequencing for comprehensive viral monitoring in Pakistan’s public health context. While we utilized the higher-throughput GridION platform for centralized processing, the Oxford Nanopore technology still provided distinct advantages through its simplified library preparation protocol, flexible run capacities, and reduced computational requirements for base-calling compared to traditional high-throughput sequencers. These benefits enabled efficient processing of our large sample batches collected from distributed sites across the country, demonstrating a scalable approach for genomic surveillance that balances throughput needs with resource availability. This could inform future public health strategies, including vaccine updates and targeted interventions. Continued monitoring and adaptive strategies are essential to keep pace with the evolving viral ecology and to enhance preparedness for future outbreaks. |
| format | Article |
| id | doaj-art-64ad4fdb6b2c42038076a878463e7f7d |
| institution | Kabale University |
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| language | English |
| publishDate | 2025-08-01 |
| publisher | Nature Portfolio |
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| spelling | doaj-art-64ad4fdb6b2c42038076a878463e7f7d2025-08-20T03:42:45ZengNature PortfolioScientific Reports2045-23222025-08-0115111110.1038/s41598-025-12774-1Nanopore sequencing reveals the genomic diversity of the variants of concern of SARS-CoV-2 during 2021 disease outbreak in PakistanZohaib Ul Hassan0Minkyu Park1Dongju Park2Humaira Amin3Ibrar Ahmed4Nazish Badar5Asif Ali6Muhammad Salman7Massab Umair8Hamza Ahmed Mirza9Abdul Ahad10Maryam Muzaffar11Falak Fazal12Sana Bibi13Madiha Islam14Yasar Mehmood Yousafzai15Sher Shah16Muhammad Zakria17Zia Ul Haq18Seil Kim19Microbiological Analysis Team, Korea Research Institute of Standards and Science (KRISS)Microbiological Analysis Team, Korea Research Institute of Standards and Science (KRISS)Microbiological Analysis Team, Korea Research Institute of Standards and Science (KRISS)Alpha Genomics Private LimitedMicrobiological Analysis Team, Korea Research Institute of Standards and Science (KRISS)Public Health Laboratories Division, National Institute of HealthDepartment of Pathology, College of Medicine, Qassim UniversityPublic Health Laboratories Division, National Institute of HealthPublic Health Laboratories Division, National Institute of HealthPublic Health Laboratories Division, National Institute of HealthPublic Health Laboratories Division, National Institute of HealthAlpha Genomics Private LimitedAlpha Genomics Private LimitedAlpha Genomics Private LimitedDepartment of Biotechnology & Genetics Engineering, Hazara UniversityInstitute of Pathology and Diagnostic Medicine, Khyber Medical UniversityPublic Health Reference Lab, Khyber Medical UniversityPublic Health Reference Lab, Khyber Medical UniversityInstitute of Public Health and Social Sciences, Khyber Medical UniversityMicrobiological Analysis Team, Korea Research Institute of Standards and Science (KRISS)Abstract The global spread of SARS-CoV-2 was significantly impacted by the emergence of Variants of Concern (VOC), with Pakistan experiencing similar trends to other countries. To gain a comprehensive understanding of the genomic epidemiology of SARS-CoV-2 strains circulating during Pakistan’s third and fourth pandemic waves, we conducted the largest single sequencing effort in the country to date. Using the GridION platform (Oxford Nanopore Technologies), we performed whole genome sequencing on 1052 confirmed COVID-19 patient samples collected from multiple cities across Pakistan between March and October 2021. Our analysis revealed a clear temporal shift in variant dominance. The Alpha variant (B.1.1.7 lineage) predominated in the first half of 2021, while the Delta variant (B.1.617.2) became most prevalent in the latter half. This transition reflects global trends and provides crucial insights into the timing and dynamics of this shift within Pakistan. Mutational analysis revealed that the most frequent nucleotide mutations in Pakistani SARS-CoV-2 samples were A23403G (associated with the D614G mutation in the spike protein), C3037T, C14408T, and C241T, potentially contributing to increased disease transmission and evasion of host immune responses. The rapid evolution and spread of these circulating variants highlight the possibility of novel variants emerging with enhanced mutational fitness. The AY.108 lineage, which has been reported at relatively low frequencies globally including in Europe and North America, with Pakistan accounting for approximately 34% of global cases, suggesting potential regional evolution or specific introduction events. Our findings underscore the dynamic nature of SARS-CoV-2 and emphasize the critical importance of ongoing, large-scale genomic surveillance in Pakistan. This study demonstrates the feasibility and utility of using nanopore sequencing for comprehensive viral monitoring in Pakistan’s public health context. While we utilized the higher-throughput GridION platform for centralized processing, the Oxford Nanopore technology still provided distinct advantages through its simplified library preparation protocol, flexible run capacities, and reduced computational requirements for base-calling compared to traditional high-throughput sequencers. These benefits enabled efficient processing of our large sample batches collected from distributed sites across the country, demonstrating a scalable approach for genomic surveillance that balances throughput needs with resource availability. This could inform future public health strategies, including vaccine updates and targeted interventions. Continued monitoring and adaptive strategies are essential to keep pace with the evolving viral ecology and to enhance preparedness for future outbreaks.https://doi.org/10.1038/s41598-025-12774-1SARS-CoV-2Genomic diversityVariant of concernSequencingOxford nanopore |
| spellingShingle | Zohaib Ul Hassan Minkyu Park Dongju Park Humaira Amin Ibrar Ahmed Nazish Badar Asif Ali Muhammad Salman Massab Umair Hamza Ahmed Mirza Abdul Ahad Maryam Muzaffar Falak Fazal Sana Bibi Madiha Islam Yasar Mehmood Yousafzai Sher Shah Muhammad Zakria Zia Ul Haq Seil Kim Nanopore sequencing reveals the genomic diversity of the variants of concern of SARS-CoV-2 during 2021 disease outbreak in Pakistan Scientific Reports SARS-CoV-2 Genomic diversity Variant of concern Sequencing Oxford nanopore |
| title | Nanopore sequencing reveals the genomic diversity of the variants of concern of SARS-CoV-2 during 2021 disease outbreak in Pakistan |
| title_full | Nanopore sequencing reveals the genomic diversity of the variants of concern of SARS-CoV-2 during 2021 disease outbreak in Pakistan |
| title_fullStr | Nanopore sequencing reveals the genomic diversity of the variants of concern of SARS-CoV-2 during 2021 disease outbreak in Pakistan |
| title_full_unstemmed | Nanopore sequencing reveals the genomic diversity of the variants of concern of SARS-CoV-2 during 2021 disease outbreak in Pakistan |
| title_short | Nanopore sequencing reveals the genomic diversity of the variants of concern of SARS-CoV-2 during 2021 disease outbreak in Pakistan |
| title_sort | nanopore sequencing reveals the genomic diversity of the variants of concern of sars cov 2 during 2021 disease outbreak in pakistan |
| topic | SARS-CoV-2 Genomic diversity Variant of concern Sequencing Oxford nanopore |
| url | https://doi.org/10.1038/s41598-025-12774-1 |
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