Polymerase chain reaction in the diagnosis of herpetic keratitis: experience in a developing country.
BACKGROUND
Herpetic ocular disease is a major cause of blindness. Rapid and accurate diagnosis is essential for prompt, proper treatment. We evaluated the usefulness of detection of herpes simplex virus (HSV) DNA by polymerase chain reaction (PCR) in the laboratory diagnosis of herpetic keratitis.


METHODS
A retrospective study was conducted involving 234 patients who attended the cornea clinic at the Regional Ophthalmic Institute, Chennai, India, between March 1995 and September 1997. Inclusion in the study was based on clinical diagnosis of herpetic keratitis. Oligonucleotide primers directed against the HSV-I thymidine kinase gene were used, yielding a 110 base pair amplicon. The utility of PCR analysis was assessed against other diagnostic markers: HSV isolation on cell culture, HSV antigen detection by indirect immunofluorescence, detection of anti-HSV IgG by enzyme-linked immunosorbent assay (ELISA) and detection of HSV-specific tear secretory IgA (sIgA) by ELISA. These tests showed overall sensitivity values of 22.4%, 39.8%, 30.4% and 20.3% respectively.


RESULTS
In epithelial keratitis all 35 specimens from which virus was cultured were positive by PCR. PCR gave a positive result in 23 (82.1%) of the 28 specimens in which HSV antigen was detected and in 4 (57.1%) of the 7 specimens that showed HSV-specific IgG. In addition, PCR detected HSV DNA in 5 of the 30 cases in which these three tests gave a negative result. PCR of two pooled tear samples (collected 1 week apart from the same patient) from 40 patients with stromal keratitis gave a positive result in 12 cases (30%). In stromal keratitis the sensitivity of PCR in detecting HSV DNA in tear samples was 85.7% with culture, indirect immunofluorescence and detection of anti-HSV IgG as the gold standard, and 80% with detection of sIgA as the gold standard.


CONCLUSIONS
The results confirm the good correlation with the clinical picture that can be obtained with PCR analysis. They also highlight the diagnostic utility of PCR in detecting HSV DNA in tear samples. This is particularly important in herpetic stromal keratitis, in which collection of corneal scrapings is not advised and, hence, conventional techniques such as virus isolation and antigen detection become difficult.
Pramod
N P
NP
Department of Microbiology, University of Madras, Taramani, Chennai, India.
Thyagarajan
S P
SP
Mohan
K V
KV
Anandakannan
K
K
D000914:Antibodies, Viral / Q000032:analysis; D000956:Antigens, Viral / Q000032:analysis; D003319:Corneal Stroma / Q000276:immunology / Q000821:virology; D003430:Cross-Sectional Studies; D017931:DNA Primers / Q000737:chemistry; D004279:DNA, Viral / Q000032:analysis; D003906:Developing Countries; D004797:Enzyme-Linked Immunosorbent Assay; D019573:Epithelium, Corneal / Q000276:immunology / Q000821:virology; D019084:Fluorescent Antibody Technique, Indirect; D018259:Herpesvirus 1, Human / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D006801:Humans; D007071:Immunoglobulin A, Secretory / Q000032:analysis; D007074:Immunoglobulin G / Q000032:analysis; D007194:India; D016849:Keratitis, Herpetic / Q000175:diagnosis* / Q000821:virology; D016133:Polymerase Chain Reaction / Q000379:methods*; D011237:Predictive Value of Tests; D015203:Reproducibility of Results; D012189:Retrospective Studies; D012680:Sensitivity and Specificity; D013666:Tears / Q000276:immunology / Q000821:virology
D000914:Antibodies, Viral; D000956:Antigens, Viral; D017931:DNA Primers; D004279:DNA, Viral; D007071:Immunoglobulin A, Secretory; D007074:Immunoglobulin G