<p>There have been conflicting results regarding the relationship between mercury (Hg) and semen quality among previous studies, partly because most studies rely on indirect reproductive biomarkers (e.g., whole blood, hair) and no study tested Hg species to understand their specific associations with semen quality. In this study, we collected paired whole semen and seminal plasma samples from 250 male participants aged 26–63 years old in Hong Kong. Total mercury (THg) and methylmercury (MeHg) levels were quantified in both matrices, and inorganic mercury (iHg) level was calculated based on the difference between THg and MeHg. We applied logistic and linear regression to examine associations between Hg levels and semen quality parameters, including normal morphology, total motility, and the number of abnormal semen parameters. To the best of our knowledge, this was the first study reporting Hg species concentrations (THg, MeHg, iHg) in human whole semen and seminal plasma samples. We found that the majority of Hg in whole semen and seminal plasma were iHg (65–70%). Interestingly, increasing MeHg levels in whole semen would accompany with improving semen quality. In contrast, we showed clearly that both iHg and MeHg, when normalized to total sperm count, would be positively associated with sperm morphology and motility impairment. This study clarified a longstanding problem how Hg content is correlated with semen quality, and the findings demonstrated that iHg, not MeHg, is the dominant form while Hg associated with sperm cells is the key determining factor of its toxicity to the male reproductive system.</p>