Simple and Economical Extraction of Viral RNA and Storage at Ambient Temperature

Hernandez, Sarah; Cardozo, Fátima; Myers, David R.; Rojas, Alejandra; Waggoner, Jesse J.

doi:10.1128/spectrum.00859-22

Cited by 4 publications

(2 citation statements)

References 42 publications

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“…An alternative, low-cost RNA extraction method has been developed for resource-constrained settings. The method involves storing the lysed sample on an RNA-binding membrane for over a week, eliminating the need for specialized equipment, expensive materials, and cold chain preservation [18]. This makes it easier to perform dengue testing in resource-limited areas, which is critical for early detection and effective disease management.…”

Section: Rna Extraction From Clinical Samplesmentioning

confidence: 99%

Molecular Detection and Genotyping of Dengue Viruses: Current Techniques and Future Prospect

Lee,

Morshed

2024

Bangla. J. Microbiol.

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The Dengue virus is a prevalent mosquito-borne disease, and its global incidence has been steadily increasing due to the favorable environmental conditions that promote mosquito breeding, primarily influenced by rising temperatures. Bangladesh has been particularly hard-hit by an intense and ongoing outbreak, resulting in a surge of cases and fatalities. Effective management of this disease necessitates the implementation of robust public health measures, including rigorous surveillance and early diagnosis. While serological tests are commonly employed in clinical diagnosis, molecular methods hold a critical role in identifying the specific Dengue virus strain, thereby contributing to a more comprehensive understanding of disease severity. This article serves as an extensive review, delving into various molecular testing techniques employed for both surveillance and clinical diagnosis. It offers valuable insights for research and clinical laboratories engaged in the detection of Dengue virus RNA in mosquitoes, environmental samples, and clinical specimens. The methods covered encompass a spectrum of approaches, including conventional PCR, isothermal amplification, real-time RT-PCR, Sanger sequencing, and whole-genome sequencing, providing a holistic overview of the available techniques. These methods play pivotal roles in clinical diagnosis, outbreak analysis, vector surveillance, and vaccine development. Furthermore, the article underscores the importance of integrating these techniques into existing healthcare systems, emphasizing their significance in ensuring precise dengue diagnostics to enhance the efficiency of disease management. These molecular methods are indispensable tools that contribute to accurate diagnosis, enable effective outbreak investigation, facilitate vector surveillance, and support vaccine preparation, thereby enhancing the overall management and control of dengue, ultimately working toward mitigating its impact on public health. Bangladesh J Microbiol, Volume 40, Number 1, June 2023, pp 41-49

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Section: Rna Extraction From Clinical Samplesmentioning

confidence: 99%

Molecular Detection and Genotyping of Dengue Viruses: Current Techniques and Future Prospect

Lee,

Morshed

2024

Bangla. J. Microbiol.

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“…However, they have limited real-world use because sample preparation, which plays a vital role in POC-Dx, is still a significant obstacle for obtaining nucleic acids with enough quality for accurate and specific results ( Ali et al., 2017 ). Although efforts have been made ( Ali et al., 2020 ; Hernandez et al., 2022 ), a simple and effective sample preparation method or portable instrument is still a major challenge for the widespread use of POC-Dx, especially considering the economic conditions of the countries that would most need such tests ( Land et al., 2019 ). However, new materials and reagents, as well as new instruments and technologies, are being developed to overcome such hurdles ( Ferreira et al., 2009 ; Escobedo et al., 2013 ; Perez-Guaita et al., 2018 ; Rampazzo et al., 2019 ; Cereda et al., 2022 ).…”

Section: Introductionmentioning

confidence: 99%

Editorial: New strategies and technologies enabling point of care diagnosis of neglected or tropical diseases

Costa

Santos

2022

Front. Cell. Infect. Microbiol.

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SARS-CoV-2 genotyping and sequencing following a simple and economical RNA extraction and storage protocol

Hernandez

Nguyen²,

Taz³

et al. 2023

PLoS ONE

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Since the beginning of the SARS-CoV-2 pandemic, supply chain shortages have caused major disruptions in sourcing the materials needed for laboratory-based molecular assays. With increasing demand for molecular testing, these disruptions have limited testing capacity and hindered efforts to mitigate spread of the virus and new variants. Here we evaluate an economical and reliable protocol for the extraction and short-term ambient temperature storage of SARS-CoV-2 RNA. Additional objectives of the study were to evaluate RNA from this protocol for 1) detection of single nucleotide polymorphisms (SNPs) in the spike gene and 2) whole genome sequencing of SARS-CoV-2. The RNAES protocol was evaluated with residual nasopharyngeal (NP) samples collected from Emory Healthcare and Emory Student Health services. All RNAES extractions were performed in duplicate and once with a commercial extraction robot for comparison. Following extraction, eluates were immediately tested by rRT-PCR. SARS-CoV-2 RNA was successfully detected in 56/60 (93.3%) RNAES replicates, and Ct values corresponded with comparator results. Upon testing in spike SNP assays, three genotypes were identified, and all variant calls were consistent with those previously obtained after commercial extraction. Additionally, the SARS-RNAES protocol yield eluate pure enough for downstream whole genome sequencing, and results were consistent with SARS-CoV-2 whole genome sequencing of eluates matched for Ct value. With reproducible results across a range of virus concentrations, the SARS-RNAES protocol could help increase SARS-CoV-2 diagnostic testing and monitoring for emerging variants in resource-constrained communities.

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Simple and Economical Extraction of Viral RNA and Storage at Ambient Temperature

Abstract: RNA extraction methods and ultra-cold storage requirements limit molecular testing for common viruses. We developed a simple, flexible, and economical method that simultaneously addresses these limitations.

Cited by 4 publications

References 42 publications

Molecular Detection and Genotyping of Dengue Viruses: Current Techniques and Future Prospect

Molecular Detection and Genotyping of Dengue Viruses: Current Techniques and Future Prospect

Editorial: New strategies and technologies enabling point of care diagnosis of neglected or tropical diseases

SARS-CoV-2 genotyping and sequencing following a simple and economical RNA extraction and storage protocol

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