OBJECTIVE 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. CONCLUSIONS These case reports strongly support the hypothesis that HSV-1 can be the cause of PGF.