Development of a DNA microarray for simultaneous detection and genotyping of lyssaviruses

Gurrala, R.; Dastjerdi, Akbar; Johnson, Nicholas; Nuñéz-García, Javier; Grierson, Sylvia S.; Steinbach, Falko; Banks, M.

doi:10.1016/j.virusres.2009.04.028

Cited by 22 publications

(12 citation statements)

References 35 publications

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“…As underlined by Fooks et al [2], the development of new molecular tools for the detection of rabies virus, such as microarrays for lyssavirus detection [37], [38] or antibody titrations using lentiviral pseudotypes [39] are aimed at improving rabies surveillance and control activities. However, their applicability in laboratories in most rabies-endemic countries is debatable.…”

Section: Resultsmentioning

confidence: 99%

More Accurate Insight into the Incidence of Human Rabies in Developing Countries through Validated Laboratory Techniques

Dacheux

Wacharapluesadee²,

Hemachudha³

et al. 2010

PLoS Negl Trop Dis

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Section: Resultsmentioning

confidence: 99%

More Accurate Insight into the Incidence of Human Rabies in Developing Countries through Validated Laboratory Techniques

Dacheux

Wacharapluesadee²,

Hemachudha³

et al. 2010

PLoS Negl Trop Dis

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“…Recently, microarray protocols for the detection of lyssaviruses have been able to demonstrate a high species-level concordance with standard reference assays (7,18). Although potentially suitable for a diagnostic purpose, the method is still not used on a routine basis.…”

mentioning

confidence: 99%

Lyssavirus Detection and Typing Using Pyrosequencing

et al. 2011

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Rabies is a fatal zoonosis caused by a nonsegmented negative-strand RNA virus, namely, rabies virus (RABV). Apart from RABV, at least 10 additional species are known as rabies-related lyssaviruses (RRVs), and some of them are responsible for occasional spillovers into humans. More lyssaviruses have also been detected recently in different bat ecosystems, thanks to the application of molecular diagnostic methods. Due to the variety of the members of the genus Lyssavirus, there is the necessity to develop a reliable molecular assay for rabies diagnosis able to detect and differentiate among the existing rabies and rabies-related viruses. In the present study, a pyrosequencing protocol targeting the 3 terminus of the nucleoprotein (N) gene was applied for the rapid characterization of lyssaviruses. Correct identification of species was achieved for each sample tested. Results from the pyrosequencing assay were also confirmed by those obtained using the Sanger sequencing method. A pan-lyssavirus one-step reverse transcription (RT)-PCR was developed within the framework of the pyrosequencing procedure. The sensitivity (Se) of the one-step RT-PCR assay was determined by using in vitro-transcribed RNA and serial dilutions of titrated viruses. The assay demonstrated high analytical and relative specificity (Sp) (98.94%) and sensitivity (99.71%). To date, this is the first case in which pyrosequencing has been applied for lyssavirus identification using a cheaper diagnostic approach than the one for all the other protocols for rapid typing that we are acquainted with. Results from this study indicate that this procedure is suitable for lyssavirus detection in samples of both human and animal origin.

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“…23 Other groups have reported using a similar method to randomly amplify virus for microarray detection from other sources such as nasal lavages, 12 endotracheal aspirates, 28 and homogenized mouse brains. 11 Using RT-rPCR amplicons, we were able to detect virus derived from cell culture but not from infected mosquitoes (Figure 1). Perhaps the ratio of viral RNA compared with non-viral RNA in samples from previous studies was greater than that found in infected arthropods, making the amplification of viral RNA less efficient in the presence of excessive amounts of arthropod RNA and DNA.…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, the platform was shown to be a useful tool for Influenza A virus genotyping, providing evidence that the microarray can be used for the detection and genetic characterization of pathogens. 10,11 Even though some microarrays have been developed to target mosquito-borne viruses, particularly flaviviruses pathogenic to humans, none were evaluated with infected mosquitoes. [12][13][14][15][16][17] Recently, a microarray was adapted to identify Culicoides species belonging to the Obsoletus group, potential bluetongue virus vectors, however it was not tested in the field.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of a Field-Portable DNA Microarray Platform and Nucleic Acid Amplification Strategies for the Detection of Arboviruses, Arthropods, and Bloodmeals

Grubaugh¹,

Petz²,

Melanson³

et al. 2013

The American Society of Tropical Medicine and Hygiene

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Abstract. Highly multiplexed assays, such as microarrays, can benefit arbovirus surveillance by allowing researchers to screen for hundreds of targets at once. We evaluated amplification strategies and the practicality of a portable DNA microarray platform to analyze virus-infected mosquitoes. The prototype microarray design used here targeted the nonstructural protein 5, ribosomal RNA, and cytochrome b genes for the detection of flaviviruses, mosquitoes, and bloodmeals, respectively. We identified 13 of 14 flaviviruses from virus inoculated mosquitoes and cultured cells. Additionally, we differentiated between four mosquito genera and eight whole blood samples. The microarray platform was field evaluated in Thailand and successfully identified flaviviruses (Culex flavivirus, dengue-3, and Japanese encephalitis viruses), differentiated between mosquito genera (Aedes, Armigeres, Culex, and Mansonia), and detected mammalian bloodmeals (human and dog). We showed that the microarray platform and amplification strategies described here can be used to discern specific information on a wide variety of viruses and their vectors.

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Development of a DNA microarray for simultaneous detection and genotyping of lyssaviruses

Cited by 22 publications

References 35 publications

More Accurate Insight into the Incidence of Human Rabies in Developing Countries through Validated Laboratory Techniques

More Accurate Insight into the Incidence of Human Rabies in Developing Countries through Validated Laboratory Techniques

Lyssavirus Detection and Typing Using Pyrosequencing

Evaluation of a Field-Portable DNA Microarray Platform and Nucleic Acid Amplification Strategies for the Detection of Arboviruses, Arthropods, and Bloodmeals

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