Parekh, M;
Romano, V;
Franch, A;
Leon, P;
Birattari, F;
Borroni, D;
Kaye, SB;
... Ferrari, S; + view all
(2020)
Shotgun sequencing to determine corneal infection.
American Journal of Ophthalmology Case Reports
, 19
, Article 100737. 10.1016/j.ajoc.2020.100737.
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Abstract
Purpose: To investigate if shotgun-sequencing method could be useful in detailed diagnosis of herpes simplex virus (HSV) infection and compare it with the conventional diagnostic method. Observations: Using a sterile scraper, the infectious part of the ocular surface was scraped gently and placed on a glass slide for conventional diagnosis using PCR and histology and in RNA stabilizing reagent for shotgun sequencing respectively. Concentration of the DNA was determined using a sensitive fluorescence dye-based Qubit dsDNA HS Assay Kit. Shotgun-sequencing libraries were generated using the NEBNext DNA ultra II protocol. The samples were sequenced on the Illumina NextSeq 500 in high output mode with 2X150 bp paired-end sequencing. Taxonomic and functional profiles were generated. Conventional diagnostic method suspected herpetic keratitis. The results indicated presence of an amplified product of 92 bp positive HSV-DNA. Conventional diagnostic method detected the presence of Herpes Simplex Virus DNA (type 1). Shotgun sequencing confirmed the diagnosis of HSV along with the taxonomical profiling of the virus. These results were achieved using 1.9 ng/μL of DNA concentration (114 ng in 60 μL) of the total sample volume. Conclusions and importance: Shotgun sequencing is a hypothesis-free approach that identifies full taxonomic and functional profile of an organism. This technology is advantageous as it requires smaller sample size compared to conventional diagnostic methods.
| Type: | Article |
|---|---|
| Title: | Shotgun sequencing to determine corneal infection |
| Open access status: | An open access version is available from UCL Discovery |
| DOI: | 10.1016/j.ajoc.2020.100737 |
| Publisher version: | https://doi.org/10.1016/j.ajoc.2020.100737 |
| Language: | English |
| Additional information: | This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/ |
| Keywords: | Sequencing, Shotgun, Diagnosis, Cornea, Infection, Virus |
| UCL classification: | UCL UCL > Provost and Vice Provost Offices UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Brain Sciences > Institute of Ophthalmology |
| URI: | https://discovery.ucl.ac.uk/id/eprint/10098323 |
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