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Purpose: The purpose of this study was to evaluate the prevalence of SARS-CoV-2 in human postmortem ocular tissues of asymptomatic donors and its implications on our eye banking protocols., Methods: The expression of SARS-CoV-2 RNA was assessed by reverse transcription-polymerase chain reaction in corneal rims and conjunctival tissues from 100 donors who were found suitable for transplantation as per the donor screening guidelines of the Global Alliance of Eye Bank Associations. The donor's clinical history and cause of death were assessed for secondary analysis., Results: Of 200 ocular tissues (100 corneal and 100 conjunctival) from the same 1 eye of 100 surgical-intended donors, between September 2020 and April 2021, the overall positivity rate for SARS-CoV-2 was ∼1% (2/200). Both the ocular samples that tested positive were conjunctival biopsies (2/100, 2%), whereas corneal samples were negative (0/100, 0%) in both donors. The causes of donor death were trauma in 51 donors, suicide in 33, cardiac arrest in 7, electric shock in 5, metabolic cause in 2, malignancy in 1, and snake bite in 1. None of the donors had a medical history suggestive of COVID infection or possible contact. None of the recipients from the donors were reported to have any systemic adverse event after keratoplasty until the follow-up of 6 weeks., Conclusions: The overall prevalence of SARS-CoV-2 was 1% (2% for conjunctival and 0% for corneal samples, P value = 0.5) in the donors who were found suitable for cornea recovery and transplantation. The findings of exceptionally low positive rates in our samples validate the criticality of history-based donor screening and do not support the necessity of postmortem PCR testing as a criterion for procurement and subsequent use for corneal transplantation., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Purpose: To examine corneal tissue for severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) positivity regarding implications for tissue procurement, processing, corneal transplantation, and ocular surgery on healthy patients. We performed quantitative reverse transcription-polymerase chain reaction qRT-PCR-testing for SARS-CoV-2 RNA on corneal stroma and endothelium, bulbar conjunctiva, conjunctival fluid swabs, anterior chamber fluid, and corneal epithelium of coronavirus disease 2019 (COVID-19) postmortem donors., Methods: Included in this study were 10 bulbi of 5 COVID-19 patients who died because of respiratory insufficiency. Informed consent and institutional review board approval was obtained before this study (241/2020BO2). SARS-CoV-2 was detected by using a pharyngeal swab and bronchoalveolar lavage. Tissue procurement and tissue preparation were performed with personal protective equipment (PPE) and the necessary protective measures. qRT-PCR-testing was performed for each of the abovementioned tissues and intraocular fluids., Results: The qRT-PCRs yielded no viral RNA in the following ocular tissues and intraocular fluid: corneal stroma and endothelium, bulbar-limbal conjunctiva, conjunctival fluid swabs, anterior chamber fluid, and corneal epithelium., Conclusions: In this study, no SARS-CoV-2-RNA was detected in conjunctiva, anterior chamber fluid, and corneal tissues (endothelium, stroma, and epithelium) of COVID-19 donors. This implicates that the risk for SARS-CoV-2 infection using corneal or conjunctival tissue is very low. However, further studies on a higher number of COVID-19 patients are necessary to confirm these results. This might be of high importance for donor tissue procurement, processing, and corneal transplantation., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.)

Purpose: To report the clinical outcome and postoperative course of Descemet membrane endothelial keratoplasty (DMEK) in irreversible corneal edema due to herpes simplex virus (HSV) endotheliitis., Methods: This is a retrospective, noncomparative, interventional case series. Nineteen eyes of 19 patients underwent standard DMEK combined with cataract surgery (triple DMEK) between May 2016 and April 2018. All patients received perioperative oral acyclovir (ACV) and prednisolone. Patients were followed up on day 1, on day 7, at 1 month, and then at 3 monthly intervals. Preoperative and postoperative best spectacle-corrected visual acuity (BSCVA), graft clarity, pachymetry, and endothelial cell loss after 1 year were recorded. Postoperative complications and HSV recurrence were noted until the last follow-up visit., Results: All eyes were phakic with variable grades of cataract with a preoperative BSCVA of 1.0 logarithm of the minimum angle of resolution or worse. The mean follow-up period was 19.3 ± 5.4 months. After 1 year, 14 (73.7%) eyes achieved a BSCVA of 0.3 or better. Seventeen (89.5%) patients had a clear graft at the last visit without any rejection episode. One graft failed after 16 months. After 3 months, the mean pachymetry reduced from 667.1 ± 62.1 to 512.8 ± 27.1 μm (P < 0.001). The mean endothelial cell loss after 1 year was 36.7 ± 13.4%. Three (15.8%) eyes had recurrence: one with recurrent endotheliitis and 2 with dendritic keratitis despite oral ACV, which responded to oral valacyclovir and ACV eye ointment. One patient had re-recurrence of endotheliitis after 20 months., Conclusions: DMEK in persistent corneal edema after HSV endotheliitis remains challenging but has encouraging outcomes. The postoperative course may be complicated by HSV recurrence. Prophylactic oral antivirals for 1 year or more and topical antivirals are useful for the prevention of recurrence.

Background: Dengue fever is the most prevalent form of flavivirus infection in humans. We have investigated whether corneoscleral tissue of the donor affected by dengue virus (DENV) harbors the virus., Purpose: To identify the risk for viral transmission through corneal transplants in areas where DENV circulates., Methods: Excised corneoscleral tissue from a cadaver with a history of viral hemorrhagic fever was analyzed using reverse transcriptase-polymerase chain reaction for the presence of DENV and chikungunya virus (CHIV)., Results: DENV was detected in RNA extracted from the donor corneoscleral rim. Further genotyping of the viral isolate from the virus-infected cell harvest revealed DENV type 3 as the causative agent. CHIV was not detected., Conclusions: The data presented in this study recommend the implementation of polymerase chain reaction for detection of DENV and CHIV to analyze excised corneoscleral tissue of a donor with viral hemorrhagic fever.

Herpes simplex virus type 1 (HSV-1) establishes a latent infection in sensory neurons that can sometimes be reactivated. HSV-1 keratitis often recurs and can be vision threatening. Reactivation of the latent virus can be stimulated by stress, immunosuppression, trauma, adrenergic iontophoresis, and UV radiation. Healthy and asymptomatic individuals are known to shed HSV-1, and this is a major factor in the spread of the virus. We investigated the frequency of shedding of HSV-1 DNA in tears of dry eye patients and individuals with conjunctivitis. Subjects were divided into 3 groups: normal (12 eyes), dry eye (11 eyes), and conjunctivitis (15 eyes). Quantitative real-time polymerase chain reaction was used for HSV DNA detection. The incidences of HSV positivity in the normal, dry eye, and conjunctivitis groups were 1 of 12 (8.3%), 3 of 11 (27.3%), and 4 of 15 (26.7%), respectively. We have previously shown that bromfenac sodium eye drops, intramuscular adenosine monophosphate, and geldanamycin effectively lower HSV-1 recurrence rates in a mouse model. Recently, we also found that nuclear factor κ-B, an IκB kinase-β inhibitor, could be a candidate for reducing HSV-1 reactivation. We sampled recipients' corneal buttons during keratoplasty and performed polymerase chain reaction. Cytomegalovirus (CMV) DNA was detected in corneas obtained from some patients, and the copy number of the detected CMV DNA was quantified. CMV DNA-positive samples were taken from 2 of the 3 patients with ocular pemphigoid; thus, in future work, the relationship between CMV in the cornea and the incidence/onset of ocular diseases of the anterior segment needs to be evaluated.

Purpose: To present a review supporting and refuting evidence from mouse, rabbit, nonhuman primate, and human studies of herpes simplex virus type 1 (HSV-1) concerning corneal latency., Methods: More than 50 research articles on HSV-1 published in peer-reviewed journals were examined., Results: Infectious HSV-1 has been found in mouse denervated tissues and in tissues with negative cultures from the corresponding ganglion. However, the different mouse strains have shown varied responses to different strains of HSV, making it difficult to relate such findings to humans. Rabbit studies provide excellent evidence for HSV-1 corneal latency including data on HSV-1 migration from the cornea into the corneoscleral rim and on the distribution of HSV-1 DNA in the cornea. However, the available methods for the detection of infectious HSV-1 may not be sensitive enough to detect low-level infection. Infectious HSV-1 has been successfully isolated from the tears of nonhuman primates in the absence of detectable corneal lesions. The recurrence of corneal ulcers in nonhuman primates before the appearance of infectious HSV-1 in tears suggests that the origin of the HSV-1 is the cornea, rather than the trigeminal ganglion. Human studies presented evidence of both ganglion and corneal latency., Conclusions: Understanding HSV-1 disease progression and the possibility of corneal latency could lead to more effective treatments for herpetic keratitis. However, it is unlikely that operational latency in the cornea will be definitively proven unless a new method with higher sensitivity for the detection of infectious virus is developed.

Purpose: : The purpose of this study was to determine the association of human herpes virus 6 (HHV-6) and/or other human herpesviruses in corneal inflammation using polymerase chain reaction (PCR)., Methods: : We collected tear films, conjunctival smears, and a corneal button of inflamed cornea, and the presence of HHV-6 and other herpesviruses in these samples were assessed by a nested PCR., Results: : In tear films collected from 3 of 9 patients with dendritic keratitis, HHV-6 DNA was positive twice, together with herpes simplex virus (HSV) or varicella zoster virus DNA most often, during the acute phase of the disease. Two other patients in this group were either positive for HSV-1 and varicella zoster virus or for HSV-1 and Epstein-Barr virus DNA but negative for HHV-6. When another 12 patients' smear samples from corneal ulcer or keratouveitis were examined, 9 were positive for HHV-6 DNA. Of these, 4 were positive for HSV-1 simultaneously, whereas the remaining 5 patients were negative for HSV-1. One patient's smear was positive for HSV-1 but not for HHV-6. In the corneal button, both HSV and HHV-6 DNAs were positive by nested PCR. HHV-6 was also positive by nested PCR in the conjunctival swab obtained from the contralateral inflamed eye of the patient., Conclusions: : In 22 patients with corneal inflammation, HHV-6 was positive in 14 of 22 patients and HSV-1 was found in 9 of those patients. These data indicated that the association of HHV-6 with disease was more frequent than with other herpesviruses and that HHV-6 may be another sole causative agent for corneal inflammation.

Purpose: We detected herpes simplex virus types 1 (HSV-1) and 2 (HSV-2), varicella zoster virus (VZV), and cytomegalovirus (CMV) DNAs in recipient corneal buttons taken at the time of penetrating keratoplasty., Methods: Twenty-seven corneal buttons were obtained from 27 patients (10 men and 17 women), 7 of whom had a history of HSV keratitis. Excised corneal buttons were immediately frozen in liquid nitrogen in the operating theater and then stored at -80°C until DNA extraction. The detection of HSV-1, HSV-2, VZV, and CMV DNAs was carried out by a nested polymerase chain reaction (PCR) method. The genome copy numbers for the nested PCR-positive samples were subsequently quantified by real-time PCR., Results: HSV-1, HSV-2, VZV, and CMV DNAs were detected in 10, 1, 9, and 2 of the 27 recipient corneal buttons, respectively. HSV-1 or HSV-2 DNAs were also detected in 5 of 7 patients with a history of HSV keratitis. Both CMV-positive patients (patients 2 and 3) had ocular pemphigoid. Among the nested PCR-positive samples, 2 HSV-1, 1 HSV-2, 1 VZV, and 1 CMV sample could be quantified by real-time PCR. Copy numbers ranged from 19 to 928 copies., Conclusions: All 4 herpesviruses, including CMV, were detected in the corneal buttons. The relationship between CMV in the cornea and ocular diseases of the anterior segment should be further evaluated.

Aim: The aim of this study was to attempt to visualize herpes simplex keratitis in an ex vivo model using currently available ophthalmological equipment and anti-herpes simplex virus 1 (HSV-1) fluorescein isothiocynate-labeled antibody., Methods: Sixteen donor human corneas were included in this study. Eight corneas were infected with HSV-1, whereas 8 remained uninfected. Abrasions were made on 2 infected and 2 uninfected corneas to assess a possible nonspecific absorption of antibodies in the sites of corneal epithelial defects. Corneas were examined before and after antibody application using a slit lamp, the fluorescein enhancing filter settings of fundus camera, and Confoscan 3. All corneas were further imaged using multiphoton laser confocal microscopy., Results: Before anti-HSV-1 antibody application, no fluorescence was detected in donor corneas with the blue light of the slit lamp and fundus camera at fluorescein enhancing filter settings. Examination with the fundus camera after antibody application detected increased background fluorescence in all the corneas with more highlighted areas of epithelial defects in abraded infected and uninfected corneas. Confoscan 3 did not show a significant difference between the appearances of HSV-1-infected and control corneas with and without application of the antibody. However, specific staining was confirmed by multiphoton laser confocal microscopy in all infected corneas., Conclusion: Further refinement of currently available ophthalmological tools is required to aid in vivo visualization of herpes simplex keratitis using fluorescein isothiocynate-labeled antibodies.

Purpose: Detailed description of the morphology of rubeola keratitis lesions is missing from textbooks and published reports. We performed a detailed analysis of rubeola keratitis lesions by using a photographic slit lamp and ophthalmic dyes., Methods: Thirty-four eyes of 17 male young adult patients with rubeola keratitis were examined. Patients were examined at 3.6 days after the development of rash. Ocular symptoms were recorded, and patients underwent complete ocular examination including photography of corneal lesions, with and without instillation of ophthalmic dyes (rose Bengal and fluorescein)., Results: Patients complained of foreign-body sensation (88%), photophobia (65%), tearing (65%), and burning sensation (47%). Visual acuity was unaffected (26%) or mildly affected (71%). Conjunctivitis was observed in 74% and keratitis in 100% of eyes. Corneal photographs were studied, and 4 types of corneal lesions were identified: small, punctate epithelial lesions staining only with rose Bengal (100%); small, round, or larger and irregular when they coalesced epithelial defects (100%); large or tiny filaments (39%); and target lesions (100%). Target lesions appeared when both dyes were instilled. The outline of target lesions stained with rose Bengal, and they consisted of a pattern of alternating concentric zones of staining with rose Bengal and fluorescein. Keratitis was strictly confined to the epithelium and resolved gradually in all patients without the appearance of complications., Conclusions: Rubeola keratitis in healthy young adults runs a benign course. Recognition of the specific to rubeola keratitis corneal target lesions could aid in the differential diagnosis and prevent the transmission of the disease.

Purpose: To detect herpes simplex virus (HSV) genome in the cornea, we sampled the limbal corneas and scleras of the imported eye bank eyes and recipient's corneal buttons and quantitated HSV genome in them by real-time polymerase chain reaction (PCR)., Methods: Forty-four recipient corneas including 7 corneas with and 37 corneas without a history of herpetic keratitis, 70 eye bank donor limbal corneas, and 35 eye bank donor scleras were obtained. Primers for real-time PCR were synthesized using the HSV-1 and -2 common regions of the viral DNA polymerase. Primers for conventional PCR were designed to detect HSV-1 and -2 and varicella zoster virus (VZV)., Results: Significantly higher copy number of HSV DNA was detected in corneas with a history of herpetic keratitis 85.7% (6/7), with an average of 1.6 x 10(4) copies/mg tissue weight than in corneas without a history of herpetic keratitis 10.8% (4/37), with an average of 8.7 copies/mg tissue weight (P < 0.05, Mann-Whitney U test). HSV DNA was detected in 5.7% (4/70) of the eye bank donor corneas, with an average of 4.9 x 10(2) copies/mg tissue weight, and in 8.6% (3/35) of the donor scleras, with an average of 10.6 copies/mg tissue weight. HSV-2 and VZV-DNA were not detected in these samples., Conclusions: Real-time PCR quantitated HSV genome in the cornea even at a quiescent phase of infection. HSV genome was detected in the corneas and scleras without a past history of herpetic keratitis by this method.

Purpose: To examine the efficacy of valaciclovir (VACV) oral formulation as an alternative to topical treatments in a case of herpetic keratitis., Methods: The patient was a 61-year-old man who presented with dendritic keratitis in his left eye. After recognizing his difficulty in using eye ointment, we prescribed oral VACV 500 mg tablets twice daily for 7 days. We also describe our experiments with orally administered VACV in a mouse model of this disease. In this study, 143 Balb/c mice were inoculated with herpes simplex virus type 1 (HSV)-1 in each eye and treated with oral VACV 50 or 100 mg/kg twice daily, oral acyclovir (ACV) 50 mg/kg 5 times/d, 3% ACV eye ointment (ACV-O) 5 times/d, 3% ACV eye drops 5 times/d, or control for 5 days., Results: After 7 days, the patient's lesion was observed healed. Corrected left visual acuity was also improved, and HSV DNA was below detectable level. In the mouse study, slit-lamp examination on days 3, 4, 5, and 7 revealed that all 5 ACV and VACV treatment groups were significantly more effective in improving symptoms of herpetic epithelial keratitis versus control (P < 0.05). Moreover, VACV 100 mg/kg was superior to other treatments. Viral titers in mouse eyeball and trigeminal ganglia were lowest in the VACV 100 mg/kg group versus other treatment groups., Conclusion: Our case example suggests that when frequent application, blurred vision, and foreign body sensation after ACV-O application cause difficulty for patients to follow treatment, oral VACV might be an effective and safe option for patients with herpetic keratitis.

Purpose: To report 2 cases of herpes simplex keratitis misdiagnosed as rheumatoid arthritis (RA)-related peripheral ulcerative keratitis (PUK), where isolation of the herpes simplex virus (HSV) led to a complete modification in management., Methods: This is a case report., Results: Two patients with RA presented with painful red right eyes. Ocular examination in both revealed an ulcer involving the peripheral cornea. The adjacent conjunctiva was infected, and the underlying sclera appeared inflamed. A diagnosis of corneal PUK secondary to RA was therefore made. The first patient had corneal scrapes taken for routine microbiological examination, which included polymerase chain reaction (PCR) for HSV. In the second patient, despite systemic immunosuppressive therapy, the ulcer progressed to involve deeper stroma and more central cornea. The conjunctiva adjacent to the ulcer was resected, and healthy conjunctival tissue was mobilized to cover the peripheral corneal ulcer. Resected conjunctival and corneal tissue was histopathologically assessed. In the first patient, PCR for HSV yielded a positive result. This prompted treatment with immediate systemic and topical acyclovir. The ulcer responded well to treatment. In the second patient, histopathological assessment and electron microscopy identified HSV. Treatment with topical trifluorothymidine and steroids was started, and a good recovery was made., Conclusions: Treatment of PUK is with systemic immunosuppressive therapy, and such therapies have serious side effects. PUK may have an occult cause in RA, and a search for a secondary agent may be beneficial. In particular, occult HSV infection must be ruled out before commencing immunosuppressive therapy.

Objective: To report recurrent herpetic keratouveitis following YAG laser peripheral iridotomy., Methods: Case report., Results: A 64-year-old woman with a history of dendritic keratitis underwent Nd:YAG laser peripheral iridotomy and developed geographic epithelial keratitis with stromal keratouveitis. Culture of a corneal swab grew herpes simplex virus type I. Keratouveitis subsided with acyclovir and corticosteroid therapy., Conclusions: Recurrent herpetic keratouveitis may be induced by YAG laser iridotomy.

Objective: To report three cases of herpetic infection in recipients of organ-cultured donor corneas among 586 consecutive corneal transplantation procedures., Methods: Three patients with no history of symptomatic herpes infection underwent corneal transplantation for keratoconus (2 patients) and Fuchs dystrophy (1 patient). Two patients developed keratouveitis and primary graft failure. The third patient developed dendritic keratitis in the graft. Culture of corneal scrapings and the patient's bandage contact lens were positive for herpes simplex virus type 1 (HSV-1). Donor and recipient sera were tested for HSV serology by EIA. Recipient corneal buttons were studied by means of transmission electron microscopy and immunohistochemistry. The three HSV-1 strains were genotyped by sequencing part of a variable antigenic domain of glycoprotein B (gB)., Results: None of the donor corneas showed endothelial cell necrosis after organ culture. All keratoplasties performed with the three mate donor corneas had an uncomplicated course. All three donor sera were positive for HSV. Preoperative recipient sera were positive for HSV. Analysis of the recipient corneal buttons showed no evidence of herpetic infection. Sequence analysis revealed three different gB genotypes., Conclusion: Ascertaining that a postoperative herpetic infection in a corneal transplant originates from the donor tissue is still difficult. Although some features of the reported cases suggest donor-to-host transmission of herpes simplex virus, the recipients could have been the source of the virus.

Purpose: To review our previous studies regarding herpes simplex virus type 1 (HSV-1) corneal latency in the rabbit lamellar keratoplasty (LK) and penetrating keratoplasty (PKP) models., Methods: Rabbits latently infected with HSV- I received allografts from naive rabbits, and naive rabbits received grafts from rabbits latently infected with HSV-1. In rabbits undergoing LK, viral shedding in tear film and the occurrence of herpetic lesions were investigated for 7 days after operation. In rabbits undergoing PKP, latency-associated transcript (LAT)-positive and -negative HSV- I mutants were used to establish latency. Ninety days after PKP, reactivation of HSV-1 was induced by transcorneal iontophoresis of epinephrine. Viral shedding was then assessed by tear-film swabbing. Donor corneal buttons, recipient corneal rims, and corresponding trigeminal ganglia were analyzed for HSV DNA concentration and viral transcription., Results: In rabbits undergoing LK, the occurrence of positive tear-film cultures and number of days on which corneal epithelial lesions were observed were significantly higher in the operated eyes of latently infected rabbits as compared with controls. In rabbits undergoing PKP, HSV- I could transmit between host and donor tissues both in anterograde and retrograde fashion. LAT-positive virus had a significantly greater ability to transmit. Higher concentrations of HSV DNA detected in cornea and trigeminal ganglia correlated with active viral transcription and higher percentage of viral shedding., Conclusion: Corneas from latently infected rabbits contain HSV-1 DNA that can replicate and transmit after induced reactivation. Our studies provide further evidence for corneal latency of this virus.

Purpose: To determine the effect of topical therapy with several corticosteroids with limited potency on viral clearance in the adenovirus type 5 (Ad5) rabbit ocular model., Methods: Sixty rabbits were inoculated in both eyes with Ad5. On the first day, the rabbits were equally divided into four topical treatment groups: 0.12% prednisolone acetate (PA), 0.1% fluorometholone (FM), 1% rimexolone (RMX), and control. Treatment was administered four times daily, in both eyes, for 3 days. All eyes were cultured for virus on days 0, 1, 3, 4, 5, 7, 9, 11, 14, 16, 18, and 21., Results: Compared with the control group, treatment with PA, FM, and RMX significantly increased the number of Ad5 positive eye cultures from days 7-21. Fluorometholone and RMX prolonged the duration of Ad5 shedding, and FM increased the mean combined Ad5 titer from days 1-5 and 7-21., Conclusions: Treatment of an experimental ocular adenovirus infection with PA, FM, and RMX for 3 days significantly enhanced adenovirus replication compared with the control group. Short-term treatment of EKC with several commercially available topical corticosteroids with limited potency may offer symptomatic relief, but may also delay viral clearance and promote office and community epidemics.

Purpose: Herpes simplex virus (HSV) can cause corneal infections in humans and lead to permanent scarring, loss of vision, and blindness. Current treatment of epithelial HSV keratitis consists of using antiviral DNA analogs. In this study, we used in vitro and in vivo models to evaluate the efficacy of six polyclonal antibodies to HSV recombinant surface glycoprotein D in treating ocular epithelial HSV., Methods: Confluent cultures of African Green monkey kidney fibroblasts (Vero cells) and normal 3-to 5-lb female New Zealand White rabbits were infected with HSV type 1, strain RE. In vitro virucidal and antiviral assays were performed, and the best of the compounds was chosen for the in vivo stage. Animals were carefully monitored until day 5 after HSV-1 inoculation, then arbitrarily divided into groups receiving, for 14 days, varying doses of: polyclonal antibodies four times a day, polyclonal antibodies three times a day, trifluorothymidine (current treatment of choice and the positive control) nine times a day, or 0.9% physiologic saline nine times a day. The animals were followed up in a masked fashion and carefully monitored for severity and resolution of the HSV infection by biomicroscopy (slit lamp) examination and viral cultures using standardized plaque assays., Results: All six of the compounds tested were effective in vitro, but one compound in particular, SP-510-50, was superior. It was used for the in vivo testing and showed antiviral efficacy in a dose-dependent manner, and at dosing four times a day, it was of comparable efficacy to trifluorothymidine (nine times a day)., Conclusions: We conclude that polyclonal antibodies to glycoprotein D appear to be effective antiviral agents in vitro and in vivo in a rabbit model of HSV-1 keratitis and show promise as a new antiviral treatment for ophthalmic use.

Purpose: Testing for the p24 antigen of the human immunodeficiency virus (HIV) may detect early HIV infection in the seronegative window; however, falsely reactive results may occur in cadaver specimens. Although neither the Food and Drug Administration (FDA) nor the Eye Bank Association of America requires p24 testing of cornea donors, many tissue banks using other organs from cornea donors do perform this assay, and the FDA requires that eye banks reject corneal tissue if a reactive p24 assay is reported. We investigated the impact of p24 testing on eye banking and corneal transplantation., Methods: Two clinical cases and records from the Lions Eye Bank of Delaware Valley (LEBDV) were reviewed retrospectively., Results: Two corneas from the LEBDV were transplanted before the reporting of p24 reactivity by other tissue banks. In one case, because of the young age of the recipient, the surgeon elected to replace the cornea with new tissue hours after the original transplant, and later polymerase chain reaction (PCR) testing was negative. In the other case, there was not enough specimen to perform Western blot or PCR confirmatory testing. The patient was followed with periodic serologic testing for HIV and has remained seronegative. To avoid such problems in the future, the LEBDV initiated testing of all donors with p24 and other nonrequired screening tests. Over a 2-month period, 22 corneas (from 11 donors) were discarded because of these tests: 4 donors had reactive p24 tests, 6 were reactive for antibody to hepatitis B core antigen, and 1 had a reactive syphilis test., Conclusions: Results from p24 assays by other tissue banks may cause difficult clinical situations when the results are received after transplantation of the tissue, but the use of the p24 assay in the screening of cornea donors may result in excessive waste of donor tissue. Further guidance is needed regarding the management of positive results from this and other nonrequired screening tests.

Purpose: To present a cluster of four patients with primary graft failure (PGF) who consecutively underwent a penetrating keratoplasty (PKP) during a period of 17 days in one institution. PKP was performed for reasons unrelated to herpes simplex infection. Herpes simplex virus type 1 (HSV-1) is presented as the possible cause of these PGFs., Methods: Viral culture of conjunctival swabs and of a bandage contact lens was performed on VERO, MRC-5, and Hep-2 cells. The four patients underwent subsequent regrafting. Polymerase chain reaction (PCR) for HSV-1 was carried out on aqueous humor and on a sample of iris and cornea with primers. Aqueous humor specimens were pretreated by boiling, and a qiagen extraction was performed according to the instructions of the manufacturer on biopsies of iris and cornea. Immunohistopathology was performed with polyclonal antibodies directed against HSV-1 and -2., Results: Culture of a conjunctival swab in three patients and culture of a bandage contact lens in the fourth patient were positive for HSV-1. In three of the four patients, PCR was positive for HSV-1 on aqueous humor and corneal graft tissue. PCR on iris tissue was positive in all patients. In three patients, culture for HSV-1 of aqueous humor and of iris tissue could not be carried out because of insufficient sample. Viral culture of the iris tissue in one patient and of the corneal graft in the four patients were negative. Immunohistopathologic examination was positive for HSV-1 in three cases., Conclusion: These case reports strongly support the hypothesis that HSV-1 can be the cause of PGF.

Purpose: There have been no reported cases of hepatitis C virus (HCV) transmission by corneal transplantation. Previous studies have also shown no correlation between HCV seropositivity and the presence of HCV RNA in the corneal tissues. This study aims to investigate such correlation and to provide further evidence to the possible transmissibility of HCV via corneal grafts., Methods: Of the 1,619 potential corneal donors to the Eye Bank of Canada over a 1-year period, 15 tested HCV-positive by the second-generation Abbott HCV enzyme immunoassay (EIA) 2.0 assay. Their sera were further tested with second-generation radio-immunoblot assay (RIBA-II), and their corneas (29 altogether) were processed for identification of HCV RNA using polymerase chain reaction (PCR)., Results: Of the 29 corneas from seropositive donors, HCV RNA was detected in 7 (24.1%)., Conclusion: This is the first study in the literature that demonstrates a significant correlation between HCV seropositivity and the presence of HCV in the corneas. Routine HCV serologic testing for all potential corneal donors and rejection of corneal tissues based on HCV seropositivity is certainly justifiable.

Purpose: To examine and report the results of penetrating keratoplasty performed in patients with varicella-zoster virus keratopathy., Methods: The authors retrospectively reviewed the records of 15 patients who had penetrating keratoplasty for varicella-zoster virus keratopathy from January 1989 through December 1998 on the Cornea Service at Wills Eye Hospital., Results: Twelve patients had a preoperative diagnosis of herpes zoster ophthalmicus, and three, of varicella. Four eyes had lateral tarsorrhaphies performed in conjunction with penetrating keratoplasty. Three eyes had endothelial rejection episodes that responded well to treatment with topical steroids. One eye had a regraft 1 month after primary failure, and this second graft also failed because of recurrent neurotrophic keratopathy. Three eyes that had repeated penetrating keratoplasty for graft failure had clear grafts at the last examination. At an average follow-up time of 50 months, 13 (86.7%) grafts remained clear, and the best corrected visual acuity was 20/100 or better in eight (53.3%) eyes. Five patients had decreased visual acuity because of retinal diseases., Conclusion: Although varicella-zoster virus keratopathy is an uncommon indication for penetrating keratoplasty, effective visual rehabilitation can be achieved in these patients. Careful postoperative management, frequent lubrication, and lateral tarsorrhaphies to protect the corneal surface are major factors in the successful outcome of these cases.

Purpose: To demonstrate the presence of an active herpetic dendrite in an eye-bank cornea., Case Report: One eye-bank cornea was studied. Viral cultures and polymerase chain reaction (PCR) testing were performed 4 days after tissue preservation. The presence or absence of herpes simplex virus (HSV) type 1 was investigated., Results: The presence of an active HSV dendrite in an eye-bank cornea was verified. HSV type 1 was confirmed using PCR amplification and restriction endonuclease DNA fragment analysis., Conclusion: This case suggests that HSV may remain viable in stored corneal tissue at 4 degrees C.

Purpose: We propose a nomenclature for classification of herpes simplex virus (HSV) keratitis. We hope that a more consistent classification system will lead to a better understanding of the disease processes, thus resulting in improved diagnosis, treatment, and patient outcomes., Methods: A review of the literature was performed to evaluate current HSV classification systems. These systems were evaluated in the context of both current clinical and basic science studies and our own clinical observations., Results: The proposed classification system is based on the anatomy and pathophysiology of the specific presentations of HSV keratitis. Anatomically, the primary level of corneal involvement, whether epithelium, stroma, or endothelium, must be elucidated. Pathophysiologically, the cause of the inflammation. whether immunologic, infectious, or neurotrophic, must be determined. There are four major categories of HSV keratitis. (1) Infectious epithelial keratitis, which is made up of cornea vesicles, dendritic ulcer, geographic ulcer, and marginal ulcer. (2) Neurotrophic keratopathy, which includes punctate epithelial erosions and neurotrophic ulcer. (3) Stromal keratitis, which is subdivided into necrotizing stromal keratitis and immune stromal keratitis. (4) Endotheliitis, which has three clinical presentations: disciform, diffuse, and linear., Conclusion: We believe that by categorizing cases of HSV keratitis by their primary anatomic and pathophysiologic etiologic characteristics, clinicians can better understand and therefore treat all types of HSV keratitis. The four main categories of HSV keratitis are infectious epithelial keratitis, neurotrophic keratopathy, stromal keratitis, and endotheliitis. Each of these is subdivided to more specific clinical presentations.

Purpose: To review the classification of herpes simplex virus (HSV) keratitis and anterior uveitis using the available clinical, pathologic, and laboratory models of disease., Methods: The literature was reviewed to establish prior classifications of herpes simplex keratitis and uveitis. The author introduces a classification attempting to encompass all patients with this disease process, taking into account the newer proposed mechanisms of disease., Results: A classification of HSV keratitis and uveitis is introduced based on the literature and the experience of the author in treating patients with this condition., Conclusions: There are multiple mechanisms of disease after ocular HSV infection, including damage from live virus, from immune and inflammatory mechanisms, and from structural damage in the aftermath of HSV infection (metaherpetic disease). Clinical, pathologic, and etiologic descriptions of these disease manifestations are offered. This may allow better communication between clinicians and authors of research protocols by defining the different aspects of this complex disease process.

Purpose: To evaluate the efficacy of an antiherpetic vaccine in recurrent herpetic ocular infections., Methods: Twenty patients with herpes simplex virus 1-related recurrent keratitis/keratouveitis were prospectively enrolled and randomly assigned to receive either a specific vaccination with heat shock-inactivated herpes simplex virus type 1 (10 patients) or to be observed as controls (10 patients). The number, duration, and anatomic localization of relapses were recorded in all the patients for 12 months before inclusion in the study and for a similar period after the assignment of each subject to vaccine or control group., Results: In the vaccine group, we observed a reduction both in the number (p = 0.016) and average duration (p = 0.050) of recurrences, whereas in the control group, no significant change was found comparing a 12-month period before and after inclusion in the study. The comparison between the two groups highlighted a significant reduction in the number (p = 0.013) and average duration (p = 0.051) of relapses in treated subjects, who did not show any significant vaccine-induced side effects., Conclusion: The use of a vaccination with heat shock-inactivated herpes simplex virus 1 seems to be able to reduce the number and duration of relapses in herpes simplex virus 1-related keratitis/keratouveitis.

Purpose: A new class of antiviral agent, cobalt chelates (the CTC series), was evaluated for treating epithelial herpetic keratitis, consequent stromal disease being the major infectious cause of blindness in industrial nations., Methods: Effects of CTC complexes were monitored in cell cultures and in a rabbit eye model, either infected with herpes simplex virus type 1 (HSV-1) or uninfected. Several antiviral concentrations of CTC complexes nontoxic to Vero cells were administered to rabbit eyes with HSV-1-induced keratitis. Corneal surface virus titers were measured, and corneal lesions of epithelial keratitis were monitored by slit-lamp microscopy and scored. Recovery rates and incidence were compared in eyes treated with CTC complexes, placebo, or clinically formulated trifluorothymidine (Viroptic), using nonparametric statistics., Results: All CTC complexes inhibited HSV-1 replication in vitro, CTC-96 being best. CTC-96, CTC-23, and CTC-67 eliminated (<1 plaque-forming unit[pfu]) corneal surface HSV-1 (otherwise >10(5) pfu) in order of descending potency, but CTC-82 was ineffective. CTC-96 (either 5 microg/ml six times daily or 10 microg/ml five times daily) accelerated herpetic dendritic keratitis recovery better than or the same as trifluorothymidine (10 mg/ml nine times daily). CTC complexes were nontoxic to Vero cells continuously exposed to < or =25 microg/ml; 50 microg/ml of CTC 96 nine times daily did not irritate uninfected rabbit eyes., Conclusion: Topical CTC-96 applications were at least as effective as Viroptic in diminishing disease signs and corneal surface virus at concentrations less than one-thousandth that of Viroptic.

Purpose: To determine the ability of norepinephrine to modulate proliferation, adhesion, and migration of SV-40 transformed human corneal epithelial cells., Methods: Assays were performed using SV-40 transformed human corneal epithelial cells. For proliferation assays, cells were plated in 96-well plates coated with fibronectin and collagen (FNC). A dose-response curve was generated for norepinephrine in concentrations of 100 nM-100 microM. The cell number in each well was evaluated using the fluorochrome Calcein AM (an intracellular esterase cleavage substrate), and fluorescence was determined using an automated fluorescent plate reader. For cell adhesion, 25 x 10(-3) cells were plated onto FNC-coated 96-well plates, incubated in 10 nM-100 microM norepinephrine for 90 min, gently irrigated, and the remaining adherent cells quantitated. Cell migration was measured using blind-well migration chambers with a 10-microm pore size and FNC-coated filters. Cells (250 x 10(3)) were added to the upper chamber, incubated for 18 h in the presence of factors, after which time the cells that had migrated through the filter were quantitated. The toxicity of norepinephrine was evaluated using a standard Live/Dead assay employing the combined fluorochromes of ethidium homodimer (to indicate dead cells) and Calcein AM (to indicate viable cells). Varying concentrations of norepinephrine were added, and the cells incubated for 3 h and the fluorometric assay performed., Results: Norepinephrine stimulated corneal epithelial cell proliferation and migration over a wide range of concentrations. It did not modulate cell adhesion and demonstrated cell toxicity only at the highest (supraphysiologic) concentration tested., Conclusions: Norepinephrine is normally found in the cornea and may be important in the maintenance of normal corneal homeostasis and in wound-healing processes.

Purpose: To determine the prevalence of hepatitis B (HBV), hepatitis C (HCV), and human immunodeficiency virus (HIV) in the serum of cornea donors who had been previously screened by social and medical history by the Eye Bank of Canada, Ontario Division., Method: A retrospective examination of the donor records of the Eye Bank of Canada, Ontario Division, was conducted. A total of 3,228 records were examined covering the period from May 17, 1993 to May 31, 1996., Results: The prevalence of HBV was determined to be 0.25%, HCV, 0.93%; and HIV, 0.031%., Conclusion: The data revealed that despite the use of medical and social history to prescreen, a small percentage of prescreened donors test serologically positive. A comparison of the prevalence of HBV, HCV, and HIV in the prescreened cornea donors to a similarly screened cohort of blood donors over a similar time period reveals a higher prevalence in all three diseases in the cohort of cornea donors. An examination of the demographic characteristics of the population as a whole versus the three groups with confirmed serology failed to show a significant difference between the seropositive and seronegative groups. This study confirms the value and necessity of serologic prescreening of cornea donors as is currently the standard of practice.

Purpose: The differential diagnosis of Acanthamoeba keratitis frequently includes herpes simplex viral keratitis. Previous in vitro studies with chlorhexidine, a drug with antiacanthamoebic action, have suggested concomitant antiviral activity against herpes simplex virus. We tested another related antiacanthamoebic compound, polyhexamethylene biguanide (PHMB), to determine its activity against herpes simplex virus (HSV) in vitro and herpes simplex viral keratitis in vivo., Methods: Equal aliquots of HSV-1 (McKrae) strain were incubated in a medium with no PHMB or with PHMB at 0.01, 0.02, or 0.05 for 5 min at 35 degrees C and the inoculum was then titered on a monolayer of E-2 cells (human corneal fibroblasts). Monolayers were examined on consecutive days and the percentage of plaque reduction was calculated. Eighteen rabbits (36 eyes) were inoculated with HSV-1 McKrae strain (10(5) pfu [plaque-forming units]/per eye). Rabbits were divided into three groups and treatment was initiated on day 3 postinfection. Group I received trifluorothymidine, group II received PHMB, and group III received artificial tears, each given five times daily in both eyes until day 10. Daily corneal swabbing to detect viral shedding and slit-lamp examination every 3 days were performed during this period., Results: In vitro studies showed 62.5, 100, and 100% plaque reduction with 0.01, 0.02, and 0.05% PHMB, respectively. Slit-lamp examination of the rabbit corneas revealed faster resolution of dendrites in animals in group I treated with trifluorothymidine. Virus was not recoverable from corneal swabs in nine of 10 rabbits in group I by day 5, but all animals in groups II and III were still shedding HSV through day 8., Conclusion: Although PHMB has potent in vitro activity against HSV, it was not an effective treatment in the in vivo rabbit model of primary HSV keratitis at the concentration commonly used for treatment of Acanthamoeba infection. This suggests that 0.02% PHMB will not provide adequate antiherpetic coverage with treatment of keratitis of undetermined etiology in which the clinical differential diagnosis includes both herpes simplex and Acanthamoeba.

Purpose: Ganciclovir is a broad-spectrum virustatic agent. Its efficacy and safety after ocular application have been demonstrated in studies of herpetic keratitis in rabbits. Two strengths of ganciclovir gel (0.05 and 0.15%) were compared with 3% acyclovir ointment in the treatment of superficial herpes simplex keratitis in humans., Methods: Two multicenter randomized clinical trials were carried out in Africa (Trial 1) and Europe (Trial 2). Sixty-seven patients (Trial 1) and 37 patients (Trial 2) from herpetic ulceration were recruited., Results: The results showed no statistically significant difference between the treatment groups, although the healing rates tended to be better in the group receiving 0.15% ganciclovir gel, with healing rates of 85% (Trial 1) and 83% (Trial 2) as compared with 72% (Trial 1) and 71% (Trial 2) in the group receiving acyclovir ointment. Local tolerance was found to be superior with the gel formulation of ganciclovir with fewer complaints of discomfort (stinging, burning) or blurred vision after application of the drug. Systemic absorption of the drug was low. No hematologic changes were detected., Conclusions: These findings support the efficacy of ganciclovir gel in the treatment of ulcerative herpes simplex keratitis and demonstrate its superior local tolerance when compared with acyclovir ointment.

Purpose: We examined the efficiency and kinetics of recombinant adenovirus vector-mediated gene transfer to rat and rabbit cornea in culture ex vivo., Methods: A recombinant replication-defective adenovirus was used to transfer a lacZ marker gene to whole rat and rabbit corneas in culture. Histochemistry was used to localise transgene expression and a colorimetric assay to quantify recombinant protein expression., Results: After infection with recombinant virus and culture for 3 days, high-efficiency gene transfer was found, with expression in most endothelial cells of both species. Minimal expression was found in other corneal cell types. On histochemistry, longer duration of expression was found in rat than in rabbit endothelium. In both rat and rabbit cornea, highest levels of recombinant protein were found at days 3-7 after incubation with virus, decreasing to low or undetectable levels at 21 days., Conclusion: Adenovirus vectors allow high-efficiency transgene expression in cornea, largely restricted to the endothelial cells of ex vivo cultured cornea. Kinetics of expression differ according to the species of cornea studied, a factor that must be considered if this vector is used in further studies.

Purpose: Two cases of hepatitis B virus (HBV) infection after penetrating keratoplasty are presented., Methods: An extensive clinical and serologic investigation of these two transplant recipients was performed. In addition, the medical histories, autopsy reports, and specimens of blood from the two deceased corneal tissue donors were retrieved and studied., Results: Serum from both donors was positive for hepatitis B surface antigen; the clinical history and serologic testing of both recipients strongly suggest that the HBV infection in each case was acquired from donor corneal tissue., Conclusion: To our knowledge, these are the first documented cases of HBV infection after corneal transplantation. Eye banks should continue to screen donors for HBV.

Herpes simplex virus (HSV) infection is one of the leading causes of corneal blindness. This study compared the clinical, virologic, and immunopathologic features of primary and recurrent murine models of herpes simplex keratitis (HSK) in the National Institutes of Health (NIH) inbred mouse strain. Primary infection resulted in multiple epithelial dendrites, followed by diffuse stromal opacification, symptoms that do not mimic what is seen in human HSK. In contrast, recurrent infection presented clinical features that included microdendrites, focal stromal opacities, disciform endotheliitis, and corneal neovascularization, which were similar to those observed in human disease. Immunohistochemical characterizations indicated that the number and duration of T cells and macrophages in recurrent HSK resemble those observed in primary disease. Results also indicated that the amount of infectious virus detected in the cornea during primary and recurrent disease was similar. However, when corneas were stained for HSV-1 antigens, mice with primary HSK displayed diffuse HSV antigen expression throughout the cornea, whereas HSV antigens were more focally distributed in recurrent disease. These data suggest that the clinical differences between the recurrent and primary herpetic keratitis may, in part, reflect the different distribution of HSV-1 antigens within the cornea.

We report the development of cytomegalovirus (CMV) keratitis in the penetrating keratoplasty of a 59-year-old human immunodeficiency virus-negative woman after uncomplicated corneal transplantation. Immunosuppression with topical cyclosporine A 2% in corn oil and topical prednisolone acetate 1% suspension was used postoperatively. The 15-month postoperative course was complicated by multiple episodes of endothelial rejection, medically controlled elevated intraocular pressure, polymicrobial bacterial (coagulase-negative staphlococcus and alpha-hemolytic streptococcus) keratitis, and endothelial plaque formation with associated hypopyon and epithelial defect. The graft failed and penetrating keratoplasty was repeated. Cytomegalovirus infection of superficial keratocytes in a region of scarring was identified in histological sections stained with hematoxylin and eosin and confirmed using mouse monoclonal anti-cytomegalovirus antibodies. Excision of the diseased corneal button with no additional treatment appears to have been curative. Low-grade keratitis was the only manifestation of the CMV infection, and it has not recurred 6 months postoperatively.

The treatment of herpes simplex keratitis has ranged from simple debridement to the use of prototypic antiviral agents specific to the herpes family, such as acyclovir. The various treatment modalities for dendritic keratitis, for the most part, have been effective. However, in recent years numerous reports of antiviral-resistant strains of herpes simplex have appeared, particularly in immunocompromised patients receiving chronic acyclovir. Interferon has been proposed as a possible treatment adjunct in these cases. We report the case of a 46-year-old man on chronic immunosuppressive and acyclovir therapy after a renal transplant who developed ulcerative keratitis due to a strain of herpes simplex virus that was resistant to multiple antiviral agents both in vivo and in vitro. After 7 weeks without improvement on multiple treatment regimens, including trifluridine, vidarabine, debridement, and topical and systemic acyclovir, interferon alpha-2a was added to topical acyclovir therapy, and the keratitis quickly resolved. Topical interferon alpha-2a appears to be an effective treatment adjunct for refractory herpes simplex keratitis in patients with cell-mediated immune dysfunction and subsequent lack of endogenous interferon. Its value in nonimmunocompromised patients remains to be determined.

Experimental animal virus replication models make it possible to study the role of viral gene products in adenovirus (Ad)-induced ocular disease. This study tested the hypothesis that the early region 3 (E3) of the human Ad genome plays an important role in the pathogenesis of Ad-induced ocular disease. Both Ad5 wt300, a genetically defined E3+ parent, and Ad5 dl327, a deletion mutant without E3 (E3-), replicated in ocular-derived cell cultures. Ad5 E3+ and Ad8 replicated more efficiently than did Ad5 E3- in cornea and conjunctival cell cultures. Lacrimal gland-derived cell cultures supported human Ad8 replication significantly more efficiently than did either Ad5 E3+ or Ad5 E3-. After intrastromal and topical inoculation of rabbits with either Ad wt300 or Ad dl327, a specific immune response was elicited that coincided with the appearance of subepithelial opacities that mimicked human disease both clinically and histologically. The clinical features (i.e., conjunctivitis, iritis, and corneal edema) were not significantly different for Ad5 E3(+)- and Ad5 E3(-)-induced ocular infection. Ad5 E3(+)- and Ad5 E3(-)-inoculated eyes shed virus for up to 7 and 5 days, respectively, and occasionally established persistent and/or latent infections in corneal, conjunctival, and, infrequently, lacrimal gland cells. Both Ad5 E3+ and Ad5 E3- spread from virus-inoculated animals to the cornea and conjunctiva of normal animals. Under current experimental conditions, expression of the E3 gene does not appear to affect the degree of virulence in ocular disease induced by Ad5 in the rabbit eye model. Deletion of the E3 gene from Ad5 does not make the model more like human disease.

Corneal subepithelial infiltrates are a known complication of adenoviral conjunctivitis. In this article, we present a case of a 32-year-old man who developed these classic infiltrates after a well-documented adenoviral conjunctivitis. What is unique about this case is that the infiltrates recurred 8 months later after an upper respiratory infection without any clinical or laboratory evidence of a viral conjunctivitis. The possible confounding variable of corticosteroid withdrawal being responsible for the recurrence is highly unlikely because he had not used topical corticosteroids for almost 4 months.

The authors sought to determine whether adenovirus could infect human corneal epithelium in vivo. They reviewed the medical records of six patients with adenovirus-positive viral corneal cultures who were examined at the Bascom Palmer Eye Institute between March 21, 1986, and December 31, 1992. The six patients with adenovirus-positive viral corneal cultures included one patient with dendritic epithelial keratitis, one with geographic epithelial ulceration, three with both geographic ulceration and contiguous dendrites, and one with heaped-up corneal epithelium but no ulceration. Four patients had rose bengal solution applied to their ocular surface, and in all four patients rose bengal uptake was seen at the epithelial edges of the dendrite or geographic ulceration in a manner characteristic of herpes simplex viral keratitis. Serotype determination of the isolates showed adenovirus type 3 (one patient), type 8 (three patients), type 19 (one patient), and indeterminate (one patient). Results of monoclonal antibody staining of cultures against herpes simplex virus (types 1 and 2) antigens was negative for all six cases. Adenovirus epithelial keratitis may result from infection of human corneal epithelium by the virus and rarely may mimic infection of corneal epithelium by herpes simplex virus.

Three corneoscleral discs (from two donors) underwent subtotal endothelial loss during routine "long-term" organ culture storage. Laboratory studies of these corneas revealed evidence of herpes simplex virus (HSV) infection. The fellow cornea from one of the donors had been issued for transplant to a patient with keratoconus. Deterioration of the graft was noted 5 days after surgery; the disc was removed at 2 months and was shown to be infected with HSV. In an experiment designed to simulate initial "cleansing" of donor globes, 0.1% polyvinylpyrolidone-iodine protected cells from infection with HSV. It was concluded that the detection of HSV in these corneas could not be explained by external contamination of the ocular surface. Furthermore, culture of conjunctival and pharangeal swabs taken from 47 consecutive donors confirmed that HSV is rarely isolated at or around the time of death. Five pairs of donor corneas destined for use in transplantation were selected at random and investigated for the presence of HSV. HSV DNA was detected by polymerase chain reaction (PCR) in tissue from two of the corneal donors. Sequential stepwise sectioning suggested that HSV DNA when present was distributed in discrete foci within the cornea. These observations suggest that HSV infection may be a cause of severe endothelial loss during corneal organ culture and possibly provide an explanation for some "failures" of corneal grafting.