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

Grubaugh, Nathan D.; Petz, Lawrence N.; Melanson, Vanessa R.; McMenamy, Scott S.; Turell, Michael J.; Long, Lewis S.; Pisarcik, S.; Kengluecha, Ampornpan; Jaichapor, Boonsong; O’Guinn, Monica L.; Lee, John S.

doi:10.4269/ajtmh.2012.12-0048

Cited by 17 publications

(21 citation statements)

References 34 publications

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“…Subsequent detection is done by either agarose gel electrophoresis (followed by ethidium bromide staining) or hybridization with molecular probes (Houng et al, 2001; Scaramozzino et al, 2001; de Morais Bronzoni et al, 2005; Chao et al, 2007; Naze et al, 2009; Yeh et al, 2010; Grubaugh et al, 2013a; Grubaugh et al, 2013b). …”

Section: Introductionmentioning

confidence: 99%

High-throughput multiplexed xMAP Luminex array panel for detection of twenty two medically important mosquito-borne arboviruses based on innovations in synthetic biology

Glushakova

Bradley

et al. 2015

Journal of Virological Methods

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Mosquito-borne arboviruses are emerging world-wide as important human and animal pathogens. This makes assays for their accurate and rapid identification essential for public health, epidemiological, ecological studies. Over the past decade, many mono- and multiplexed assays targeting arboviruses nucleic acids have been reported. None has become established for the routine identification of multiple viruses in a “single tube” setting. With increasing multiplexing, the detection of viral RNAs is complicated by noise, false positives and negatives. In this study, an assay was developed that avoids these problems by combining two new kinds of nucleic acids emerging from the field of synthetic biology. The first is a “self-avoiding molecular recognition system” (SAMRS), which enables high levels of multiplexing. The second is an “artificially expanded genetic information system” (AEGIS), which enables clean PCR amplification in nested PCR formats. A conversion technology was used to place AEGIS component into amplicon, improving their efficiency of hybridization on Luminex beads. When Luminex “liquid microarrays” are exploited for downstream detection, this combination supports single-tube PCR amplification assays that can identify 22 mosquito-borne RNA viruses from the genera Flavivirus, Alphavirus, Orthobunyavirus. The assay differentiates between closely-related viruses, as dengue, West Nile, Japanese encephalitis, and the California serological group. The performance and the sensitivity of the assay were evaluated with dengue viruses and infected mosquitoes; as few as 6–10 dengue virions can be detected in a single mosquito.

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Section: Introductionmentioning

confidence: 99%

High-throughput multiplexed xMAP Luminex array panel for detection of twenty two medically important mosquito-borne arboviruses based on innovations in synthetic biology

Glushakova

Bradley

et al. 2015

Journal of Virological Methods

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“…Probes were designed to target unique viral sequences, 30 to 45 nucleotides in length, between the GSPs using methods previously described [30]. A total of 2,097 oligonucleotide probes were selected for inclusion on the ArboChip5.1.…”

Section: Methodsmentioning

confidence: 99%

“…Most of the assays included an unbiased, random amplification method for microarray detection. We have previously reported that such techniques are efficient for the amplification viral nucleic acids isolated from cultured cells, but not from a complex matrix of RNA from mosquito homogenates [30]. Additionally, most of the microarray designs are not wide-ranging and only cover a few of the most concerning viral pathogens.…”

Section: Introductionmentioning

confidence: 99%

“…For each genus, at least three sets of consensus gene-specific primers (GSPs) were designed for efficient PCR amplification of the viral nucleic acids isolated from infected mosquitoes and virus-specific microarray capture probes were designed to differentiate between the PCR amplicons. We previously demonstrated that a portable microarray platform, the ElectraSense 4×2K (CustomArray, Inc., Bothell, WA), is practical for field diagnostics [30]. The platform included a small and rugged microarray reader that was able to analyze four samples simultaneously against 2,240 oligonucleotide DNA probes using electrochemical detection (ECD) [31], [32], [33], [34].…”

Section: Introductionmentioning

confidence: 99%

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Multi-Gene Detection and Identification of Mosquito-Borne RNA Viruses Using an Oligonucleotide Microarray

et al. 2013

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BackgroundArthropod-borne viruses are important emerging pathogens world-wide. Viruses transmitted by mosquitoes, such as dengue, yellow fever, and Japanese encephalitis viruses, infect hundreds of millions of people and animals each year. Global surveillance of these viruses in mosquito vectors using molecular based assays is critical for prevention and control of the associated diseases. Here, we report an oligonucleotide DNA microarray design, termed ArboChip5.1, for multi-gene detection and identification of mosquito-borne RNA viruses from the genera Flavivirus (family Flaviviridae), Alphavirus (Togaviridae), Orthobunyavirus (Bunyaviridae), and Phlebovirus (Bunyaviridae).Methodology/Principal FindingsThe assay utilizes targeted PCR amplification of three genes from each virus genus for electrochemical detection on a portable, field-tested microarray platform. Fifty-two viruses propagated in cell-culture were used to evaluate the specificity of the PCR primer sets and the ArboChip5.1 microarray capture probes. The microarray detected all of the tested viruses and differentiated between many closely related viruses such as members of the dengue, Japanese encephalitis, and Semliki Forest virus clades. Laboratory infected mosquitoes were used to simulate field samples and to determine the limits of detection. Additionally, we identified dengue virus type 3, Japanese encephalitis virus, Tembusu virus, Culex flavivirus, and a Quang Binh-like virus from mosquitoes collected in Thailand in 2011 and 2012.Conclusions/SignificanceWe demonstrated that the described assay can be utilized in a comprehensive field surveillance program by the broad-range amplification and specific identification of arboviruses from infected mosquitoes. Furthermore, the microarray platform can be deployed in the field and viral RNA extraction to data analysis can occur in as little as 12 h. The information derived from the ArboChip5.1 microarray can help to establish public health priorities, detect disease outbreaks, and evaluate control programs.

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“…There are a large number of recently launched portable devices that allow testing of forensic samples outside the laboratory. Many of these systems are based on familiar laboratory platforms, either utilising capillary electrophoresis (CE) based detection [22], microarray [23] or utilise melt curve detection of PCR amplicons [24,25].…”

Section: Introductionmentioning

confidence: 99%

Species detection using HyBeacon ® probe technology: Working towards rapid onsite testing in non-human forensic and food authentication applications

Dawnay

Hughes

Court

et al. 2016

Forensic Science International: Genetics

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26Identifying individual species or determining species' composition in an unknown sample is important 27 for a variety of forensic applications. Food authentication, monitoring illegal trade in endangered 28 species, forensic entomology, sexual assault case work and counter terrorism are just some of the fields 29 that can require the detection of the biological species present. Traditional laboratory based approaches 30 employ a wide variety of tools and technologies and exploit a number of different species specific traits 31 including morphology, molecular differences and immuno-chemical analyses. A large number of these 32 approaches require laboratory based apparatus and results can take a number of days to be returned to 33 investigating authorities. Having a presumptive test for rapid identification could lead to savings in terms 34 of cost and time and allow sample prioritisation if confirmatory testing in a laboratory is required later. 35This model study describes the development of an assay using a single HyBeacon ® probe and melt curve 36 analyses allowing rapid screening and authentication of food products labelled as Atlantic cod (Gadus 37 morhua). Exploiting melt curve detection of species specific SNP sites on the COI gene the test allows 38 detection of a target species (Atlantic cod) and closely related species which may be used as substitutes. 39The assay has been designed for use with the Field Portable ParaDNA system, a molecular detection 40 platform for non-expert users. The entire process from sampling to result takes approximately 75 41 minutes. Validation studies were performed on both single source genomic DNA, mixed genomic DNA 42 and commercial samples. Data suggests the assay has a lower limit of detection of 31 pg DNA. The 43 specificity of the assay to Atlantic cod was measured by testing highly processed food samples including 44 frozen, defrosted and cooked fish fillets as well as fish fingers, battered fish fillet and fish pie Ninety-45 six (92.7 %) of all Atlantic cod food products tested provided a correct single species result with the 46 remaining samples erroneously identified as containing non-target species. The data shows that the assay 47 was quick to design and characterise and is also capable of yielding results that would be beneficial in 48 a variety of fields, not least the authentication of food. 49 50 3

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

Cited by 17 publications

References 34 publications

High-throughput multiplexed xMAP Luminex array panel for detection of twenty two medically important mosquito-borne arboviruses based on innovations in synthetic biology

High-throughput multiplexed xMAP Luminex array panel for detection of twenty two medically important mosquito-borne arboviruses based on innovations in synthetic biology

Multi-Gene Detection and Identification of Mosquito-Borne RNA Viruses Using an Oligonucleotide Microarray

Species detection using HyBeacon ® probe technology: Working towards rapid onsite testing in non-human forensic and food authentication applications

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