Radhika Tandon, MD, DNB, FRCS, FRCOphth,

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COMPARISON OF TEAR COLLECTION TECHNIQUE FOR
DETECTION OF HSV DNA BY PCR

• Radhika Tandon, MD, DNB, FRCS, FRCOphth,
• Manoj Sharma, MD Gita Satpathy, MD
• Department of Cornea, Cataract Refractive
  Surgery and Ocular Microbiology
  Dr Rajendra Prasad Centre For Ophthalmic
  Sciences, AIIMS

The authors have no financial interest in the
subject matter of this poster.
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Review of Literature- Shedding of Herpes simplex
virus type 1 (HSV-1) DNA in active stromal
keratitis and / or endotheliitis, as detected by
polymerase chain reaction (PCR) analysis of tear
samples obtained by different methods
Authors (year) PCR type Subjects, proportion () Subjects, proportion () Sample Method Samples Collected per Subject
Authors (year) PCR type Active stromal keratitis Endotheliitis Sample Method Samples Collected per Subject
Yamamoto S et al (1994) Conventional PCR 2/6 (33.33) - Schirmer Strips 1
Kudo E et al (1996) Nested PCR 5/15 (33.33) - Schirmer Strips 3.3
Pramod NP et al (2000) (Conventional PCR 12/40 (30) - Schirmer Strips 2
Fukuda M et al (2003) Real-time PCR 8/14 (57) 0/3 Eye Rinse 1
Fukuda M et al (2008) Real-time PCR 13/22 (59.1) 0/3 Eye Rinse 1
Total no. of subjects with shedding of HSV-1
DNA/total no. of subjects assessed (percentage
of patients with shedding of HSV-1 DNA)
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AIM

• To compare PCR results in tear samples obtained
  by two different methods
• Schirmer's strip fire-polished micro capillary
  tube
• in patients with clinically diagnosed
• viral stromal keratitis and endotheliitis

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Patients

• Inclusion Criteria
• Clinically diagnosed cases of active stromal
  keratitis and endotheliitis
• Exclusion criteria
• Pure epithelial keratitis with no stromal
  involvement
• History of oral acyclovir use within 1 month

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Study Design

• Study group 66 eyes( 59 patients)
• 52 unilateral 7 bilateral
• Control group 130 eyes of 90 patients
• Contralateral eye of 50 unilateral affected
  patients
• Both eyes of 40 normal volunteers
• Of 52 unilaterally affected, 2 patients had
  contralateral phthisis bulbi, sample was not
  taken from the phthisical eye

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Tear Collection Methods

• Tear samples of each patient were collected
• using Schirmers filter paper strip and fire
  polished micro- capillary tubes in a randomized
  sequence
• from the anaesthetized inferior fornix
• and were subjected to PCR for detection of HSV
  DNA

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RESULTS
Fig. 1 Distribution of tear samples from various
clinical categories PUK Peripheral ulcerative
keratitis A Active Q Quiescent P with
perforation V Viral
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Fig 2 PCR result in different clinical
categories with capillary Schirmers method
PUK Peripheral ulcerative keratitis A
Active Q Quiescent P with perforation V
Viral
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Figure 3 PCR result using Capillary and
Schirmer's tear sampling method

• PCR positivity in tear samples collected by
  either method were not significantly different
  (p0.23).

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Conclusion

• 20 cases of active Herpetic stromal keratits
  and endotheliitis had positive tear PCR test
  result
• Tear samples collected by Schirmers strip and
  fire- polished micro capillary tube are equally
  effective for HSV DNA detection

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• Address for Correspondence
• Dr Radhika Tandon
• Professor of Ophthalmology
• radhika_tan_at_yahoo.com
• Dr. RP Centre for Ophthalmic Sciences,
• All India Institute of Medical Sciences
• (AIIMS)
• New Delhi, India