Lactoferrin plays an important role in the defense against infections, including herpes simplex virus (HSV) keratitis. We studied the impact of three single nucleotide polymorphisms in the human lactoferrin gene on the susceptibility to HSV infections of the eye and the severity of such infections. Lactoferrin gene polymorphisms were determined by PCR combined with restriction fragment length analysis in 105 HSV keratitis patients and 145 control subjects. Bilateral tear samples were harvested from 50 patients and 40 healthy controls and tear lactoferrin concentrations were determined by ELISA. Patients' records were used to acquire information about the severity of the HSV keratitis. The frequencies of the Glu561Asp polymorphism, but not those of the Ala11Thr and Lys29Arg polymorphisms, differed significantly between patients and control subjects with an under-representation of the Asp561 allele in the patient group. Furthermore, the values for best corrected visual acuity, frequency of recurrences since onset, and average duration of clinical episodes did not differ among patients with various lactoferrin genotypes. In addition, tear lactoferrin concentrations were the same in patients with HSV keratitis and healthy controls and also did not differ among patients with various lactoferrin genotypes. Lactoferrin Glu561Asp polymorphism is associated with the susceptibility to HSV keratitis with a protective role for lactoferrin variants comprising Asp561. However, no beneficial effects of this lactoferrin variant on the clinical outcome of ocular HSV keratitis were noted.