July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
High Throughput Sequencing reveals predominantly fungal pathogens in the vitreous of patients with presumed infectious, culture negative endophthalmitis in Southern India with implication for diagnostics
Author Affiliations & Notes
  • Joveeta Joseph Ruben
    Jhaveri Microbiology Centre, L V Prasad Eye Institute, Hyderabad, Telangana, India
  • Rajagopalaboopathi Jayasudha
    Jhaveri Microbiology Centre, L V Prasad Eye Institute, Hyderabad, Telangana, India
  • Jaishree Gandhi
    Jhaveri Microbiology Centre, L V Prasad Eye Institute, Hyderabad, Telangana, India
  • Savitri Sharma
    Jhaveri Microbiology Centre, L V Prasad Eye Institute, Hyderabad, Telangana, India
  • Vivek Pravin Dave
    Smt. Kannuri Santhamma Centre for vitreoretinal diseases, L V Prasad Eye Institute, Hyderabad, Telangana, India
  • Footnotes
    Commercial Relationships   Joveeta Joseph Ruben, None; Rajagopalaboopathi Jayasudha, None; Jaishree Gandhi, None; Savitri Sharma, None; Vivek Dave, None
  • Footnotes
    Support  This work was supported by a grant from the DST-SERB to Dr Joveeta Joseph (File Number: EMR/2016/002259) India.
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 829. doi:
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      Joveeta Joseph Ruben, Rajagopalaboopathi Jayasudha, Jaishree Gandhi, Savitri Sharma, Vivek Pravin Dave; High Throughput Sequencing reveals predominantly fungal pathogens in the vitreous of patients with presumed infectious, culture negative endophthalmitis in Southern India with implication for diagnostics. Invest. Ophthalmol. Vis. Sci. 2019;60(9):829.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : To evaluate the clinical utility of High-throughput sequencing (HTS) approach based analysis of bacterial and fungal genome in vitreous samples from patients with presumed infectious endophthalmitis.

Methods : We subjected 75 vitreous samples from clinically presumed infectious endophthalmitis patients to high-throughput sequencing (Illumina HiSeq 2500) after DNA extraction and amplification of the V3–V4 regions of the bacterial 16S rRNA and ITS 2 region of fungus by PCR. As controls, we included 70 vitreous samples from patients undergoing surgery for non-infectious retinal disorders. Paired reads were curated, taxonomically labeled, and filtered to reduce ambiguousness /contaminants in public microbial reference genomes and the environment.

Results : Our genomic database reads revealed in nearly all cases the same microorganism that was determined in cultivation-based analyses (bacterial-14/15, fungal 3/3). HTS additionally diagnosed the presence of microbes in 42/57 (73.7%) patients who were conventionally negative (fungal pathogens in 36/57, bacterial pathogens in 11/57, including five cases that showed presence of both bacteria and fungal organisms). Rarefaction curves indicated that sampling coverage was complete. Aspergillus sp, Fusarium sp, Exserohilum sp. and Candida sp. were the most predominant genera in our cohort of culture negative endophthalmitis cases. Heat map based microbial clustering analysis revealed that these organisms were taxonomically similar to the species identified by conventional culture methods. Interestingly, 4/70 control samples also showed presence of bacteria, including reads of Enterobacteriaceae sp. Streprococcus sp. and Corynebacterium sp., although their clinical significance is uncertain.

Conclusions : HTS is useful in detecting pathogens in endophthalmitis cases that elude conventional attempts at diagnosis and can provide actionable information relevant to management, especially where there is a high index of suspicion of fungal endophthalmitis, particularly in tropical countries. Outcome analyses and clinical trials addressing the success and cost savings of HTS for the diagnosis of endophthalmitis are recommended.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

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