Background: The prevalence of human papillomavirus (HPV) infection in infertile men is significantly higher than that in the general population. Nevertheless, it is unclear whether the presence of HPV in semen samples is related to reduced semen quality. Objectives: The present article aims to elucidate the possible association between HPV infection and semen parameters in Iranian infertile men. Methods: This cross-sectional study included semen samples from 140 infertile Iranian male partners of couples seeking fertility assessment. Standard semen analysis was performed. Sperm DNA fragmentation index (DFI), sperm chromatin maturation, anti-sperm antibodies, leukocyte count, fructose concentration, and zinc concentration were also assessed. Nested polymerase chain reaction (PCR) and INNO-LiPA were used to identify HPV DNA sequences in semen samples and to determine HPV genotype, respectively. Independent samples t-tests were used to compare sperm parameters between HPV-positive and HPV-negative groups. Results: Human papillomavirus DNA was detected in 18 (12.85%) out of 140 semen samples (10% high-risk genotypes, 2.14% low-risk genotypes, and one sample was excluded). Our findings revealed that HPV18 was the most prevalent type (8.57%). According to our results, the presence of HPV DNA in semen samples was associated with higher semen anti-sperm antibodies (P = 0.001), higher white blood cell (WBC) counts (P = 0.02), and increased DFI values (P = 0.03). Conclusions: Human papillomavirus -positive patients had higher semen anti-sperm antibodies, semen WBC counts, and sperm DFI values. These results highlight the importance of careful screening for the presence of HPV in semen samples from infertile men. It should also be noted that more studies are required.