Objective: To evaluate the association between the vaginal microenvironment and fecundability among women. Design: Register‐based nationwide cohort study. Setting: Chinese National Free Pre‐conception Check‐up Project from 2015 to 2018. Population: Our study included a total of 3 388 554 eligible women who were attempting to become pregnant. Method: We assessed the vaginal microenvironment at baseline by considering four indices: vaginal pH, clue cell examination, whiff test and vaginal cleanliness grading. If any of these indicators was abnormal, the vaginal microenvironment was defined as poor. Propensity score matching was used to control for potential confounders and reduce bias. Logistic models were used to estimate the fecundability odds ratios (FORs) after adjustment for covariates. Main outcome measures: Achievement of a pregnancy within 1 year. Results: Of the total study population, 379 718 women (11.2%) had a poor vaginal microenvironment and their pregnancy rate after 1 year was significantly lower than the group with a normal microenvironment (71.8% versus 76.1%, P < 0.001). After adjusting for potential confounders, the women with a poor vaginal microenvironment were associated with a 9% reduction in fecundability compared with the normal microenvironment group (FOR 0.91, 95% CI 0.90–0.92). The adverse effects of a poor vaginal microenvironment were stronger among multipara (FOR 0.89, 95% CI 0.87–0.90) or women with irregular menstruation (FOR 0.86, 95% CI 0.84–0.89). Conclusion: There was a negative association between a poor vaginal microenvironment and the fecundability of women. These findings highlight the significance of assessing the vaginal microenvironment during pre‐pregnancy health examinations. Women with a poor vaginal microenvironment were associated with a reduction in fecundability. Women with a poor vaginal microenvironment were associated with a reduction in fecundability. [ABSTRACT FROM AUTHOR]