A novel dengue virus detection method that couples DNAzyme and gold nanoparticle approaches

Virology Journal

Malcolm J Fraser¹  Tresa S Fraser¹  Cheryl A Kucharski¹  Velmurugan Balaraman¹  James R Carter¹ 

[1] Department of Biological Sciences, Eck Institute of Global Health, University of Notre Dame, Notre Dame, IN 46556, USA

关键词: Detection;    Arbovirus;    Gold;    Nanoparticles;    DNAzyme;    Flavivirus;    Dengue;   

Others  :  1149365
DOI  :  10.1186/1743-422X-10-201

 received in 2012-11-12, accepted in 2013-04-08,  发布年份 2013

【 摘 要 】

Background

Recent epidemics of dengue viruses (DENV) coupled with new outbreaks on the horizon have renewed the demand for novel detection methods that have the ability to identify this viral pathogen prior to the manifestation of symptoms. The ability to detect DENV in a timely manner is essential for rapid recovery from disease symptoms. A modified lab-derived 10-23 DNAzyme tethered to gold nanoparticles provides a powerful tool for the detection of viruses, such as DENV.

Results

We examined the effectiveness of coupling DNAzyme (DDZ) activation to the salt-induced aggregation of gold nanoparticles (AuNP) to detect dengue virus (DENV) progeny in mosquito cells. A DNAzyme was designed to recognize the 5’ cyclization sequence (5’ CS) that is conserved among all DENV, and conjugated to AuNPs. DDZ-AuNP has demonstrated the ability to detect the genomic RNA of our model dengue strain, DENV-2 NGC, isolated from infected Aedes albopictus C6/36 cells. These targeting events lead to the rapid aggregation of AuNPs, resulting in a red to clear color transition of the reaction mixes, and thus positive detection of the DENV RNA genome. The inclusion of SDS in the reaction mixture permitted the detection of DENV directly from cell culture supernatants without additional sample processing. Specificity assays demonstrated detection is DENV-specific, while sensitivity assays confirm detection at levels of 1 × 10¹ TCID50 units. These results demonstrate DDZ-AuNP effectively detects DENV genomes in a sequence specific manner and at concentrations that are practical for field use.

Conclusions

We have developed an effective detection assay using DNAzyme catalysis coupled with AuNP aggregation for the detection of DENV genomes in a sequence specific manner. Full development of our novel DDZ-AuNP detection method will provide a practical, rapid, and low cost alternative for the detection of DENV in mosquito cells and tissues, and possibly infected patient serum, in a matter of minutes with little to no specialized training required.

【 授权许可】

   
2013 Carter et al.; licensee BioMed Central Ltd.

【 图 表 】

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