Kang Il Jun1, Baek-Lok Oh2, Narae Kim3, Joo Young Shin4, Jangsup Moon5. 1. Division of Infectious Disease, Department of Internal Medicine, Ewha Womans University Seoul Hospital, Seoul, South Korea. 2. Department of Ophthalmology, Seoul National University Hospital, Seoul, South Korea. 3. Department of Neurology, Seoul National University Hospital, Seoul, South Korea. 4. Department of Ophthalmology, Seoul National University College of Medicine, SMG-SNU Boramae Medical Center, Seoul, South Korea. Electronic address: joo0shin@gmail.com. 5. Department of Neurology, Seoul National University Hospital, Seoul, South Korea; Department of Genomic Medicine, Seoul National University Hospital, Seoul, South Korea. Electronic address: jangsup.moon@gmail.com.
Abstract
OBJECTIVES: We investigated whether nanopore amplicon sequencing of aqueous humor was capable of rapid pathogen identification in infectious endophthalmitis. METHODS: 5 cases of culture-positive bacterial endophthalmitis and 3 cases of fungal endophthalmitis (1 culture-positive and 2 presumed) were included. DNA was extracted from the aqueous humor and vitreous specimen, and PCR of bacterial rDNA (16S) and fungal rDNA (ITS1 and D1/2/3) was performed. Then, nanopore amplicon sequencing was performed for 2 h. The results of amplicon sequencing were compared to those of conventional culture studies. RESULTS: In all cases, pathogens were identified by amplicon sequencing of aqueous humor specimens. In 3 cases of bacterial endophthalmitis, the identified microbes were confirmed by culture studies of both aqueous humor and vitreous specimens. In 2 cases of bacterial and 1 case of fungal endophthalmitis, the identified pathogens were confirmed only by culture studies of vitreous specimens. In all cases, amplicon sequencing identified pathogen in a shorter turnaround time than culture studies. In 2 cases with negative culture results, amplicon sequencing of aqueous humor identified fungal pathogens. CONCLUSIONS: Our data demonstrates the potential of amplicon nanopore sequencing using aqueous humor to enable rapid, sensitive and less invasive microbial diagnosis of endophthalmitis.
OBJECTIVES: We investigated whether nanopore amplicon sequencing of aqueous humor was capable of rapid pathogen identification in infectious endophthalmitis. METHODS: 5 cases of culture-positive bacterial endophthalmitis and 3 cases of fungal endophthalmitis (1 culture-positive and 2 presumed) were included. DNA was extracted from the aqueous humor and vitreous specimen, and PCR of bacterial rDNA (16S) and fungal rDNA (ITS1 and D1/2/3) was performed. Then, nanopore amplicon sequencing was performed for 2 h. The results of amplicon sequencing were compared to those of conventional culture studies. RESULTS: In all cases, pathogens were identified by amplicon sequencing of aqueous humor specimens. In 3 cases of bacterial endophthalmitis, the identified microbes were confirmed by culture studies of both aqueous humor and vitreous specimens. In 2 cases of bacterial and 1 case of fungal endophthalmitis, the identified pathogens were confirmed only by culture studies of vitreous specimens. In all cases, amplicon sequencing identified pathogen in a shorter turnaround time than culture studies. In 2 cases with negative culture results, amplicon sequencing of aqueous humor identified fungal pathogens. CONCLUSIONS: Our data demonstrates the potential of amplicon nanopore sequencing using aqueous humor to enable rapid, sensitive and less invasive microbial diagnosis of endophthalmitis.