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Laboratory science
New test for the diagnosis of bacterial endophthalmitis
  1. P Goldschmidt1,
  2. S Degorge1,
  3. D Benallaoua1,
  4. E Basli2,
  5. L Batellier1,
  6. S Boutboul2,
  7. C Allouch2,
  8. V Borderie2,
  9. L Laroche2,
  10. C Chaumeil1
  1. 1
    Laboratoire du Centre National d’Ophtalmologie des Quinze-Vingts, 28 rue de Charenton, Paris, France
  2. 2
    Service 5 du Centre National d’Ophtalmologie des Quinze-Vingts, 28 rue de Charenton, Paris, France
  1. Correspondence to Dr P Goldschmidt, Laboratoire du Centre National d’Ophtalmologie des Quinze-Vingts, 28 rue de Charenton, 75012 Paris France; pablogol{at}aol.com; goldschmidt{at}quinze-vingts.fr

Abstract

Background: Diagnosis of bacterial endophthalmitis (BE) often fails due to: (1) insufficient volumes of vitreous fluid (VF) and aqueous humour (AH); (2) lack of sensitivity of culture; (3) antibiotic treatments; (4) polymerase chain reaction (PCR) cross-contamination; and (5) limitations on the interpretation of the real-time PCR melting curve. We developed a fast real-time (f-real-t) PCR to improve the performance of the laboratory diagnosis of BE.

Methods: The following samples were processed after adding an internal control: phosphate buffered saline (PBS); VF, AH and cell suspensions spiked with Bacteria (Bac); VF and AH from patients with endophthalmitis; and VF and AH from non-infective patients. DNA was extracted (MagNA Pure®) and added to four tubes containing selected primers and probes for the identification and quantification of all Bac and eight genera by f-real-t PCR. Diagnostic performances based on direct microscopic examination, culture and f-real-t PCR were compared.

Results: The f-real-t PCR detected at least 0.01 colony-forming units (CFU) of Bac/μl with no cross-reactivity with fungi. Correlation with culture-positive results was 100%. Sixty per cent of BE samples tested culture-positive, but f-real-t PCR tested positive for 90%. Samples from non-infective cases were negative.

Conclusion: The f-real-t PCR detected and quantified Bac, Staphylococci, Streptococci, Haemophilus, Pseudomonas, Enterobacteria, Acinetobacter, Propionibacteriacae and Corynebacteria in one run. Cultures required several hours to days (with a non-negligible number of false-negative results) and the f-real-t PCR was completed in 90 min. The f-real-t PCR is presented as a new tool for the diagnosis of BE: its usefulness requires validation with larger series of samples.

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Footnotes

  • Competing interests None

  • Ethics approval Investigations were performed in accordance with the Declaration of Helsinki (http://www.wma.net/e/policy/).

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