Abstract
Purpose: To compare the clinical and molecular diagnoses of Herpes Simplex Keratitis (HSK). Materials and Methods: Conjunctival swabs (after fluorescein and anaesthetic wash out) and detailed questionnaire data were obtained from 146 participants. Corneal rims and conjunctival epithelial cells were infected with Herpes Simplex Virus (HSV) type 1 or HSV2 and supernatant collected. HSV1; HSV2; Varicella Zoster Virus (VZV) and Adenovirus (ADV) DNA was assessed using two real time Polymerase Chain Reaction (PCR) methods. Results: Of the 146 participants recruited, 54 were clinically diagnosed with typical epithelial lesions and 38 with atypical epithelial lesions, 17 with old inactive HSK and 37 healthy volunteers. HSV1 DNA was detected in 28 (30%) of the 92 participants with clinically suspect HSK. Patients who presented with typical epithelial lesions had a higher positive rate (46%) than those who presented with atypical type lesions (8%), when using primers against the Glycoprotein (Gp) G region of the virus. When the same samples were retested with primers against the GpB region, the positive rate for the typical and atypical cases increased to 52% and 11% respectively. Antiviral use at the time of sampling reduced the rate of PCR positivity by 20% (p in vitro control investigations of HSV1 and 2 infected corneal rims and conjunctival epithelial cells were 100% positive for infected and 100% negative for uninfected samples when assessed using both PCR methods. Conclusions: Clinical diagnosis of typical HSK is not always confirmed by PCR. Concomitant use of an antiviral reduces levels of PCR positivity. Given this and the findings that other ocular surface pathogens may mimic HSK pathology, and that choice of gene amplification region can also affect accurate detection of HSV1 by PCR, we propose the use of a multiplex assay. This would perform PCR using primers spanning a number of different regions within one gene and would also target a number of different viral genes to ensure potentially different HSV1 viral strains or other viruses do not affect the test and lead to disagreements between the clinical and molecular diagnosis of HSK. From these findings, this paper proposes a clinical supportive algorithmic guide to manage such disagreements.
Highlights
The predominant cause of infectious corneal blindness in the developed world is Herpes Simplex Virus type 1 (HSV1) infection of the eye [1]
Concomitant use of an antiviral reduces levels of Polymerase Chain Reaction (PCR) positivity. Given this and the findings that other ocular surface pathogens may mimic Herpes Simplex Keratitis (HSK) pathology, and that choice of gene amplification region can affect accurate detection of HSV1 by PCR, we propose the use of a multiplex assay
The type of lesion had no significant affect on ADV PCR incidence. These results demonstrate an agreement level of only 46% (25 patients PCR positive for HSV1 DNA) between the clinical and molecular diagnosis of patients presenting with typical HSK epithelial lesions
Summary
The predominant cause of infectious corneal blindness in the developed world is Herpes Simplex Virus type 1 (HSV1) infection of the eye [1]. HSV1 is responsible for the development of Herpes Simplex Keratitis (HSK) and has a global incidence of up to 1.5 million people per year including 40,000 cases with severe visual impairment or blindness due to this disease [2]. The diagnosis of HSK is primarily dependent on the clinical opinion of the ophthalmologist. Marangon et al [5] reported that in their study, approximately 5% of the clinically diagnosed cases of HSK were caused by adenovirus (ADV) and 2.7% by enterovirus rather than HSV1. They found that 3.2% of the suspected herpes conjunctivitis cases were due to cytomegalovirus (CMV). Confirmatory or additional molecular tests are important to avoid potential lifelong inappropriate diagnosis of HSK and to enable appropriate treatment
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