title: Prospective Study of the Diagnostic Accuracy of the In Vivo Laser Scanning Confocal Microscope for Severe Microbial Keratitis.
abstract: To determine the diagnostic accuracy of in vivo confocal microscopy (IVCM) for moderate to severe microbial keratitis (MK).
  
  
  
  Double-masked prospective cohort study.
  
  
  
  Consecutive patients presenting to Aravind Eye Hospital, Madurai, India, between February 2012 and February 2013 with MK (diameter ≥3 mm, excluding descemetocele, perforation, or herpetic keratitis).
  
  
  
  Following examination, the corneal ulcer was scanned by IVCM (HRT3/RCM, Heidelberg Engineering, Heidelberg, Germany). Images were graded for the presence or absence of fungal hyphae or Acanthamoeba cysts by the confocal microscopist who performed the scan (masked to microbial diagnosis) and 4 other experienced confocal graders (masked to clinical features and microbiology). The regrading of the shuffled image set was performed by 3 graders, 3 weeks later. Corneal-scrape samples were collected for microscopy and culture.
  
  
  
  The main outcome measures were sensitivity, specificity, and positive and negative predictive values of IVCM compared with those of a reference standard of positive culture or light microscopy. Sensitivities and specificities for multiple graders were pooled and 95% confidence intervals calculated using a bivariate random-effects regression model.
  
  
  
  The study enrolled 239 patients with MK. Fungal infection was detected in 176 (74%) and Acanthamoeba in 17 (7%) by microbiological methods. IVCM had an overall pooled (5 graders) sensitivity of 85.7% (95% confidence interval [CI]: 82.2%-88.6%) and pooled specificity of 81.4% (95% CI: 76.0%-85.9%) for fungal filament detection. For Acanthamoeba, the pooled sensitivity was 88.2% (95% CI: 76.2%-94.6%) and pooled specificity was 98.2% (95% CI: 94.9%-99.3%). Intergrader agreement was good: κ was 0.88 for definite fungus; κ was 0.72 for definite Acanthamoeba. Intragrader repeatability was high for both definite fungus (κ: 0.88-0.95) and definite Acanthamoeba classification (κ: 0.63-0.90). IVCM images from 11 patients were considered by all 5 graders to have a specific organism present (10 fungus, 1 Acanthamoeba) but had negative results via culture and light microscopy.
  
  
  
  Laser scanning IVCM performed with experienced confocal graders has high sensitivity, specificity, and test reproducibility for detecting fungal filaments and Acanthamoeba cysts in moderate to large corneal ulcers in India. This imaging modality was particularly useful for detecting organisms in deep ulcers in which culture and light microscopy results were negative.
authors.affiliation: London School of Hygiene and Tropical Medicine, London, United Kingdom. Electronic address: Jaya.Chidambaram@Lshtm.ac.uk.
authors.forename: Jaya D
authors.initials: JD
authors.lastname: Chidambaram
authors.affiliation: Aravind Eye Hospital, Madurai, Tamil Nadu, India; Aravind Medical Research Foundation, Madurai, Tamil Nadu, India.
authors.forename: Namperumalsamy V
authors.initials: NV
authors.lastname: Prajna
authors.affiliation: London School of Hygiene and Tropical Medicine, London, United Kingdom.
authors.forename: Natasha L
authors.initials: NL
authors.lastname: Larke
authors.affiliation: Aravind Eye Hospital, Madurai, Tamil Nadu, India.
authors.forename: Srikanthi
authors.initials: S
authors.lastname: Palepu
authors.affiliation: Aravind Eye Hospital, Madurai, Tamil Nadu, India.
authors.forename: Shruti
authors.initials: S
authors.lastname: Lanjewar
authors.affiliation: Aravind Medical Research Foundation, Madurai, Tamil Nadu, India.
authors.forename: Manisha
authors.initials: M
authors.lastname: Shah
authors.affiliation: Aravind Medical Research Foundation, Madurai, Tamil Nadu, India.
authors.forename: Shanmugam
authors.initials: S
authors.lastname: Elakkiya
authors.affiliation: Aravind Eye Hospital, Madurai, Tamil Nadu, India; Aravind Medical Research Foundation, Madurai, Tamil Nadu, India.
authors.forename: Prajna
authors.initials: P
authors.lastname: Lalitha
authors.affiliation: Moorfields Eye Hospital, London, United Kingdom.
authors.forename: Nicole
authors.initials: N
authors.lastname: Carnt
authors.affiliation: Moorfields Eye Hospital, London, United Kingdom.
authors.forename: Minna H
authors.initials: MH
authors.lastname: Vesaluoma
authors.affiliation: Moorfields Eye Hospital, London, United Kingdom.
authors.forename: Melanie
authors.initials: M
authors.lastname: Mason
authors.affiliation: Moorfields Eye Hospital, London, United Kingdom.
authors.forename: Scott
authors.initials: S
authors.lastname: Hau
authors.affiliation: London School of Hygiene and Tropical Medicine, London, United Kingdom; Moorfields Eye Hospital, London, United Kingdom.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Burton
mesh_terms: D015823:Acanthamoeba Keratitis / Q000175:diagnosis* / Q000469:parasitology; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D003320:Corneal Ulcer / Q000175:diagnosis* / Q000469:parasitology; D003937:Diagnosis, Differential; D004311:Double-Blind Method; D015822:Eye Infections, Parasitic / Q000175:diagnosis* / Q000469:parasitology; D005260:Female; D005500:Follow-Up Studies; D006801:Humans; D008297:Male; D018613:Microscopy, Confocal / Q000379:methods*; D008875:Middle Aged; D011446:Prospective Studies; D015203:Reproducibility of Results; D012720:Severity of Illness Index; D055815:Young Adult