Shotgun sequencing to determine corneal infection.
Cornea
Diagnosis
Infection
Sequencing
Shotgun
Virus
Journal
American journal of ophthalmology case reports
ISSN: 2451-9936
Titre abrégé: Am J Ophthalmol Case Rep
Pays: United States
ID NLM: 101679941
Informations de publication
Date de publication:
Sep 2020
Sep 2020
Historique:
received:
14
08
2019
revised:
30
03
2020
accepted:
05
05
2020
entrez:
22
5
2020
pubmed:
22
5
2020
medline:
22
5
2020
Statut:
epublish
Résumé
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. 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. 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.
Identifiants
pubmed: 32435720
doi: 10.1016/j.ajoc.2020.100737
pii: S2451-9936(20)30089-X
pii: 100737
pmc: PMC7229483
doi:
Types de publication
Case Reports
Langues
eng
Pagination
100737Informations de copyright
© 2020 Published by Elsevier Inc.
Déclaration de conflit d'intérêts
The following authors have no financial disclosures: MP, VR, AF, PL, FB, DB, SBK, DP, SA, SF.
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