Ocular Microbiology and Immunology Group
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2017 OMIG Abstract 12

A step forward in the diagnosis of infectious endophthalmitis:
Comparison of next generation sequencing results in culture negative
and culture positive vitreous samples 
Savitri Sharma, Moumita Chakrabarti, Dhanshree Deshmukh, Joveeta Joseph, Rajagopalaboopathi Jayasudha, Kalyana C Sama, Bhavani Sontam, Mudit Tyagi, Raja Narayanan, S Shivaji
Brien Holden Eye Research Centre, L. V. Prasad Eye Institute, Hyderabad, India

Purpose: To investigate the presence of microbial DNA in culture-positive and -negative clinically diagnosed infectious endophthalmitis using next generation sequencing (NGS) of vitreous samples.

Methods: Vitreous biopsy samples from 34 patients with clinical diagnosis of infectious endophthalmitis, and ten vitreous samples from patients undergoing surgery for non-infectious retinal disorders were included in the study. One half of the sample was used for routine microbiological work up by direct microscopy (calcofluor white, Gram and Giemsa stains and culture for bacteria and fungus) and the other half was processed for NGS. Total DNA was extracted using the Qiagen mini kit and the V3–V4 regions of the bacterial 16S rRNA gene and ITS 3 and ITS 4 regions of fungal genome were amplified by PCR and deep sequenced on an Illumina HiSeq 2500 Machine. Paired reads were curated, taxonomically labeled, and filtered.

Results: Of the 34 patients with clinical infectious endophthalmitis 15 (44%) were culture positive (Bacterial-14, fungal- 1) Comparatively NGS diagnosed the presence of microbes in 30/34 patients, which included bacteria in 26/30, fungi in 2/30 and mixed infections in 2/30 cases amounting to an increase by 44% (overall positivity in 30/34, 88%). Lack of adequate DNA in 4 out of 19 culture-negative vitreous samples precluded application of NGS. All 10 vitreous samples from control eyes undergoing routine vitreous surgery were negative for bacteria or fungus by NGS. There was good agreement between culture and NGS for culture-positive cases. Additionally, NGS showed presence of polymicrobial infectionsin almost all culture-negative samples and many culture positive samples.

Conclusions: Culture, and NGS provide complementary information in presumed infectious endophthalmitis, thus making this approach a promising diagnostic platform for the diagnosis of infectious endophthalmitis. Clinical validity and impact on treatment are being evaluated.

Disclosure: N

Financial Support: Hyderabad Eye Research Foundation and Department of Science and Technology, Government of India

2017 Agenda and Abstracts | < Previous Next >