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Genetic diversity and viability of human norovirus strains detected in selected wastewater samples from Tshwane district, South Africa
Thesis (PhD (Medical Virology))--University of Pretoria, 2024.
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| Format: | Thesis |
| Language: | English |
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University of Pretoria
2025
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| _version_ | 1867613582127005696 |
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| access_status_str | Open Access |
| author2 | Mans, Janet |
| author_browse | Mans, Janet |
| author_facet | Mans, Janet |
| collection | Thesis |
| dc_rights_str_mv | © 2023 University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria. |
| description | Thesis (PhD (Medical Virology))--University of Pretoria, 2024. |
| format | Thesis |
| id | oai:repository.up.ac.za:2263/100994 |
| institution | University of Pretoria (South Africa) |
| language | English |
| last_indexed | 2026-06-10T12:38:26.101Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from UPSpace — University of Pretoria Institutional Repository |
| publishDate | 2025 |
| publishDateRange | 2025 |
| publishDateSort | 2025 |
| publisher | University of Pretoria |
| publisherStr | University of Pretoria |
| record_format | dspace |
| source_str | UPSpace — University of Pretoria Institutional Repository |
| spelling | oai:repository.up.ac.za:2263/100994 Genetic diversity and viability of human norovirus strains detected in selected wastewater samples from Tshwane district, South Africa Mans, Janet mabasa.victor@tuks.co.za Van Zyl, Walda B. Taylor, Maureen B. Mabasa, Victor Vusi UCTD Sustainable development goals (SDGs) Norovirus diversity Wastewater Viability PCR Next generation sequencing (NGS) Whole genome sequencing (WGS) Thesis (PhD (Medical Virology))--University of Pretoria, 2024. Norovirus, a leading cause of acute gastroenteritis worldwide, is characterised by its substantial genetic diversity and periodic emergence of novel variants, necessitating continuous monitoring to inform public health interventions. This study aimed to assess norovirus diversity and viability in wastewater from Pretoria, South Africa (SA), using next-generation sequencing (NGS) and advanced molecular assays. Between June 2018 and August 2020, 200 samples comprising raw sewage and final effluent were collected bi-weekly from two wastewater treatment works (WWTWs). Viruses were concentrated using skimmed milk flocculation and glass wool adsorption- elution methods. Noroviruses were detected in 81% of samples, with a higher positivity rate in raw sewage (89%) compared to final effluent (73%). Amplicon-based NGS targeting the norovirus BC genotyping region revealed 89 distinct genogroup (G) I and GII RNA-dependant RNA polymerase-capsid combinations, including 51 putative novel recombinants and one emerging novel strain, emphasizing the significant genetic variability present in wastewater. Complementing this, archived stool samples from paediatric gastroenteritis patients confirmed positive for GII norovirus underwent semi- nested PCR using in-house designed primers, followed by sequencing on the Illumina NextSeq 2000 platform. Complete genomes were successfully obtained for 56% of samples, uncovering different GII.4 variants and a GII.3[PNA] strain harbouring a putative novel P-type, which had 96.4% nucleotide similarity to a 2010 South African strain. Additionally, this study presented the first near-complete genome of GII.13[PNA8], which has only been reported in SA so far. Wastewater samples were analysed using both targeted and non-targeted NGS approaches. While the targeted method confirmed the presence of various GII strains, the non-targeted approach faced challenges with non-viral sequences predominating. The identification of strains such as GII.4 San Francisco[P31], previously observed in the United Kingdom, the United States of America, Gabon and India, underlines the global distribution and potential spread of this GII.4 variant. The detection of less common strains such as GII.7[P7] points to possible subclinical circulation in the population. A separate analysis focused on the viability of noroviruses in final effluent samples using viability RT-qPCR (vPCR) with PMAxx™- Triton X-100. While significant reductions in potentially viable noroviruses were observed post-treatment at both WWTWs, concerning levels of viable noroviruses persisted. At WWTW1, half of GI-positive and 64.9% of GII-positive final effluent samples showed no detectable RNA after vPCR, while WWTW2 achieved a 100% RNA reduction in 92.6% of GII-positive samples. These findings suggest that despite treatment processes, the discharge of viable noroviruses remains a public health concern due to their high initial load and low infectious dose. Overall, this comprehensive surveillance using NGS and vPCR highlights the genetic diversity, emergence of novel recombinants, and challenges in assessing viral viability in treated wastewater. Such data are crucial for enhancing wastewater-based epidemiology and guiding strategies to mitigate the spread of noroviruses in both environmental and clinical settings. NHLS Research Trust PhD Bursary and research funding National Research Foundation PhD Bursary Poliomyelitis Research Foundation PhD Bursary Medical Virology PhD (Medical Virology) Unrestricted Faculty of Health Sciences SDG-06: Clean water and sanitation 2025-02-17T13:06:43Z 2025-02-17T13:06:43Z 2025-04 2024-11 Thesis * A2025 http://hdl.handle.net/2263/100994 DOI: https://doi.org/10.25403/UPresearchdata.28429580.v1 https://doi.org/10.25403/UPresearchdata.28429580 en © 2023 University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria. application/pdf University of Pretoria |
| spellingShingle | UCTD Sustainable development goals (SDGs) Norovirus diversity Wastewater Viability PCR Next generation sequencing (NGS) Whole genome sequencing (WGS) Genetic diversity and viability of human norovirus strains detected in selected wastewater samples from Tshwane district, South Africa |
| title | Genetic diversity and viability of human norovirus strains detected in selected wastewater samples from Tshwane district, South Africa |
| title_full | Genetic diversity and viability of human norovirus strains detected in selected wastewater samples from Tshwane district, South Africa |
| title_fullStr | Genetic diversity and viability of human norovirus strains detected in selected wastewater samples from Tshwane district, South Africa |
| title_full_unstemmed | Genetic diversity and viability of human norovirus strains detected in selected wastewater samples from Tshwane district, South Africa |
| title_short | Genetic diversity and viability of human norovirus strains detected in selected wastewater samples from Tshwane district, South Africa |
| title_sort | genetic diversity and viability of human norovirus strains detected in selected wastewater samples from tshwane district south africa |
| topic | UCTD Sustainable development goals (SDGs) Norovirus diversity Wastewater Viability PCR Next generation sequencing (NGS) Whole genome sequencing (WGS) |
| url | http://hdl.handle.net/2263/100994 https://doi.org/10.25403/UPresearchdata.28429580 |