Introduction: There are few studies on sex difference in patients with infectious mononucleosis caused by Epstein-Barr virus (EBV-IM). We performed a retrospective study to evaluate the sex difference in clinical presentation of patients with EBV-IM. Methods: We performed a single-center retrospective study evaluating >14-year-old patients with serologically confirmed EBV-IM during 2006e2017. We compared the patients' age, symptoms, physical findings, and laboratory data between male and female patients. To adjust for confounding factors, we performed a logistic regression analysis based on the results of univariate comparisons. Result: Of the 122 eligible patients (56 male and 66 female, ratio: 1:1.2), the median ages were 26 years old (interquartile range [IR], 22e31.5 years old]) and 22 years old (IR, 20e25 years old) for males and females, respectively (p < 0.001). Headache was significantly more prevalent in males (25.0% vs. 10.6%, p ¼ 0.036). Leukocyte count was also significantly higher in males (11,400/mm 3 [IR, 7,600e14,100/mm 3 ] vs. 9,400/mm 3 [IR, 6,600e11,600/mm 3 ], p ¼ 0.021). The prevalence of periorbital edema (male: 3.6% vs. female: 18.1%, p ¼ 0.012) and severity of transaminase elevation were significantly higher in females. The regression analysis evaluating clinical characteristics of male patients showed that age >30 years old, headache, and leukocyte >11,000/mm 3 had high odds ratios. Conclusion: Our single-center retrospective study suggests that older age of onset, headache, and leukocytosis are more likely to be characteristics of male patients with EBV-IM. Our study also underscores the importance of periorbital edema as a clue for early diagnosis of EBV-IM, especially in female patients.