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Utility of broad-range 16S rRNA PCR assay versus conventional methods for laboratory diagnosis of bacterial endophthalmitis in a tertiary care hospital
  1. Deepanshi Mishra1,
  2. Gita Satpathy1,2,
  3. Rohan Chawla3,
  4. Pradeep Venkatesh3,
  5. Nishat Hussain Ahmed1,
  6. Subrat Kumar Panda4
  1. 1 Ocular Microbiology, Dr R P Centre for Ophthalmic Sciences, All India Institute of Medical Sciences, New Delhi, India
  2. 2 Ocular Microbiology, Dr R P Centre for Ophthalmic Sciences, Department of Microbiology, All India Institute of Medical Sciences, New Delhi, India
  3. 3 Vitreo Retinal Services, Dr R P Centre for Ophthalmic Sciences, All India Institute of Medical Sciences, New Delhi, India
  4. 4 Department of Pathology, All India Institute of Medical Sciences, New Delhi, India
  1. Correspondence to Dr Gita Satpathy, Rajendra Prasad Centre for Ophthalmic Science, All India Institute of Medical Sciences, New Delhi 11002, India; gita.satpathy{at}gmail.com
  • Present affiliation The present affiliation of Gita Satpathy is: Ocular Microbiology, Dr R P Centre for Ophthalmic Sciences and Deptt. of Microbiology, AIIMS, All India Institute of Medical Sciences , New Delhi, India

Abstract

Background Endophthalmitis, a sight-threatening intraocular infection, can be of postsurgical, post-traumatic or endogenous origin. Laboratory diagnosis-based appropriate therapy can be vision-saving. Conventional culture-based laboratory diagnosis takes time and lacks sensitivity. In this study a broad-range PCR assay was assessed against conventional and automated culture methods in vitreous specimens for accurate microbiological diagnosis.

Aims To use broad-range PCR assay targeting 16S ribosomal RNA (rRNA) region of bacteria and to assess its performance vis-à-vis conventional and automated culture methods in the laboratory diagnosis of endophthalmitis.

Methods Vitreous specimens from 195 patients with clinically diagnosed endophthalmitis were processed for classical and automated culture methods, antimicrobial sensitivity and broad-range PCR assay targeting 762 bp region of 16S rRNA followed by nucleotide sequencing by Sanger’s method. Causative agents were identified from the nucleotide sequences analysed against the GenBank database, and organisms were identified using the Clinical and Laboratory Standards Institute (CLSI) MM18A guidelines.

Results Bacteria could be detected from 127 (65.13%) of the 195 vitreous specimens by broad-range PCR assay; bacterial isolation was possible from 17 (8.7%) and 60 (30.76%) of these specimens by conventional and automated culture methods, respectively (p<0.0001). PCR assay could detect two uncultured bacterium, and in five cases the bacterial identity could not be determined from NCBI database matching.

Conclusion Broad-range PCR assay could provide definitive microbial diagnosis within 24 hours in significantly more patients (p<0.0001). Some rare organisms could be detected, useful in treatment modalities. Automated culture was significantly more sensitive than conventional culture.

  • infection
  • vitreous
  • microbiology
  • diagnostic tests/investigation

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Footnotes

  • Contributors DM did the experimental work. GS planned the work, arranged the funds, and reviewed the data and the manuscript. RC and PV did the clinical work and collected patient specimens. NHA did the classical culture technique. SKP planned the work, arranged the funds and reviewed the data.

  • Funding Internal funding from All India Institute of Medical Sciences.

  • Competing interests None declared.

  • Patient consent Obtained.

  • Ethics approval Ethics Committee of All India Institute of Medical Sciences, New Delhi.

  • Provenance and peer review Not commissioned; externally peer reviewed.

  • Data sharing statement There are no additional unpublished data.

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