Translational vision science & technology · 2025
A Tear-Based Approach for Rapid Identification of Bacterial Pathogens in Corneal Ulcers Using Nanopore Sequencing.
СтатьяНаблюдательное исследованиеНазвание и аннотация: Перевод в очереди
Обработка статьи · этапы, ошибки и управление
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Dibbs Mark, Matesva Mitchelle, Theotoka Despoina, Jayaraj Christina, Metiku Beruk, Demkowicz Patrick, Heng Jacob S, Wang Yvonne, Bakhoum Christine Y, Chow Jessica, Bakhoum Mathieu F
Аннотация
PURPOSE: This prospective observational study assesses the efficacy of using portable next-generation sequencing directly on tear samples to identify bacterial pathogens in corneal ulcers.
METHODS: Tear samples were collected from ulcerated and contralateral eyes using Schirmer strips. Corneal scrapings and cultures were performed as medically indicated. The 16S rRNA gene was amplified from tear samples using polymerase chain reaction (PCR), and Nanopore sequencing was used for bacterial species identification and taxonomic classification.
RESULTS: Bacterial DNA was identified in 8 of 10 samples using the tear-based sequencing method. Nanopore sequencing accurately identified the causative bacteria in all four samples that exhibited bacterial growth on culture and detected bacterial pathogens in two of the four ulcers that did not show bacterial growth on culture. In two cases where cultures could not be obtained due to the ulcer's small size, tear sequencing successfully identified bacterial species. Among the nine contralateral tear samples collected, Nanopore sequencing identified commensal bacteria in four samples.
CONCLUSIONS: PCR amplification of 16S rRNA directly from tears followed by Nanopore sequencing is an effective, noninvasive method to identify bacterial pathogens in corneal ulcers, offering noninferior results to traditional culture methods.
TRANSLATIONAL RELEVANCE: By eliminating the need for corneal scrapings and nucleic acid extraction, this tear-based method improves the timing and accuracy of bacterial pathogen diagnosis in corneal ulcers, allowing for prompt detection of causative organisms and enabling earlier targeted antimicrobial therapy, thereby improving patient outcomes.
METHODS: Tear samples were collected from ulcerated and contralateral eyes using Schirmer strips. Corneal scrapings and cultures were performed as medically indicated. The 16S rRNA gene was amplified from tear samples using polymerase chain reaction (PCR), and Nanopore sequencing was used for bacterial species identification and taxonomic classification.
RESULTS: Bacterial DNA was identified in 8 of 10 samples using the tear-based sequencing method. Nanopore sequencing accurately identified the causative bacteria in all four samples that exhibited bacterial growth on culture and detected bacterial pathogens in two of the four ulcers that did not show bacterial growth on culture. In two cases where cultures could not be obtained due to the ulcer's small size, tear sequencing successfully identified bacterial species. Among the nine contralateral tear samples collected, Nanopore sequencing identified commensal bacteria in four samples.
CONCLUSIONS: PCR amplification of 16S rRNA directly from tears followed by Nanopore sequencing is an effective, noninvasive method to identify bacterial pathogens in corneal ulcers, offering noninferior results to traditional culture methods.
TRANSLATIONAL RELEVANCE: By eliminating the need for corneal scrapings and nucleic acid extraction, this tear-based method improves the timing and accuracy of bacterial pathogen diagnosis in corneal ulcers, allowing for prompt detection of causative organisms and enabling earlier targeted antimicrobial therapy, thereby improving patient outcomes.