Transcript Detection of Herpes Simplex Virus Genome in Tear Fluid of Patients

Detection of Herpes Simplex Virus Genome in Tear Fluid
of Patients with Herpetic Keratouveitis and Endotheliitis
Radhika Tandon, MD, DNB, FRCS, FRCOphth
Dr. Manoj Sharma, MD,
Prof.Jeewan.S.Titiyal, MD
Dr Tushar Agarwal, MD
Dr Namrata Sharma, MD
Prof. 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
Aim

To evaluate the role of the polymerase chain
reaction (PCR) for detecting Herpes virus DNA
in tear samples of patients with herpetic
keratouveitis and endotheliitis.
Patients

Inclusion criteria



Clinically diagnosed
Keratouveitis and/or endotheliitis
Exclusion criteria

Oral acyclovir use within one month
Study Design

Study group: 20 eyes (17 patients)




12 unilateral
3 bilaterally active
2 bilaterally affected with fellow eye quiescent*
Control group: 94 eyes of 54 patients


Contra lateral eye of 14 unilateral active disease
Both eyes of 40 normal volunteers
*59% (10/17) had definite history of recurrent disease
Laboratory Diagnosis

Before starting treatment, tear samples were collected
with fire polished microcapillary tubes and subjected to
PCR for HSV DNA detection

PCR Protocol
1. DNA extraction: commercial QI Amp DNA blood kit
2. Polymerase chain reaction
Primer-111 bp region of HSV 1 thymidine kinase gene
(Hofgartner W T et al Clinical chemistry, 1999) Amplificationthermal cycler (Gene Amp PCR system 9700, applied
biosystem, USA)
3. Electrophoreses- in 2% agarose gel
Treatment


Tab acyclovir 400 mg (5 times/day) × 7 days followed by
Tab acyclovir 400 mg (BD) × 6 months
Topical steroid (1% prednisolone acetate)
Adjunct therapy was given as required

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Topical antibiotic
Topical mydriatic (2% homatropine)
Topical lubricant (preservative free)
Analgesics (if required)
Result
Clinical type
Viral keratouveitis



PCR
Positive
Negative
2
9
Total
11
Endotheliitis
2
7
9
Control
0
80
80
Fellow unaffected eyes
0
12
12
Fellow quiescent eye
0
2
2
Four of twenty tear samples (20%) from patients yielded PCR positive for
Herpes virus DNA, of whom 2 had acute endotheliitis and 2 had keratouveitis.
All four PCR positive cases had recurrent disease.
No HSV DNA was detected in tear samples from both eyes of 40 normal
volunteers and fellow eye of unilateral active cases
Figure : PCR result In different clinical categories
2 cases of Herpetic Keratouveitis who tested PCR
positive in tear samples from the affected eye
At presentation
(1a, 2a)
1a
1b
1c
2a
2b
2c
1 week
(1b, 2b)
1 month
(1c, 2c)
A patient with bilateral Herpetic Endotheliitis who tested
PCR positive in tear samples from both eyes
1a
1b
2a
At presentation
(1a, 2a)
1c
2c
2b
1 week
(1b, 2b)
3 weeks
(1c, 2c)
Conclusion

20% cases of active keratouveitis and
endotheliitis had positive tear sample PCR
test result for HSV DNA

Herpes Virus DNA is detectable in tear
samples of such patients and could prove
useful in supporting the clinical diagnosis of
these cases.
Address for Correspondence
Dr Radhika Tandon
Professor of Ophthalmology
[email protected]
Dr. RP Centre for Ophthalmic Sciences
All India Institute of Medical Sciences ( AIIMS)
New Delhi -110029
India