Background: Mushrooms are a good source of many nutrients which are potentially beneficial for chronic diseases. We speculated that due to its abundant nutrients edible mushrooms might have a beneficial effect on the prevention of subclinical thyroid dysfunction (SCTD). Therefore, we designed a large-scale cohort study to examine whether mushrooms consumption is a protective factor for SCTD in adults. Methods: This prospective cohort study investigated 6 631 participants (mean age: (45.0 ± 10.2) years; 55.1% men). Edible mushrooms consumption was measured at baseline using a validated food frequency questionnaire. SCTD was defined as abnormal serum thyroid-stimulating hormone levels and normal free thyroxine. Cox proportional hazards regression models were used to examine the association of edible mushrooms consumption with incident SCTD. Results: During follow-up period, a total of 262 new cases of SCTD were identified, the incidence rate of subclinical hypothyroidism was 8.9/1 000 person-years and subclinical hyperthyroidism was 7.2/1 000 person-years. After adjusting potential confounding factors, the multivariable hazard ratios (95% confidence intervals) for subclinical hypothyroidism were 1.00 (reference) for almost never, 0.53 (0.29, 0.97) for 1–3 times/week and 0.30 (0.10, 0.87) for ≥ 4 times/week (P for trend = 0.02). It also showed edible mushrooms consumption was inversely associated with subclinical hypothyroidism in obese individuals but not non-obese individuals, the final hazard ratios (95% confidence intervals) were 0.14 (0.03, 0.73) (P for trend < 0.01). Conclusions: This population-based prospective cohort study has firstly demonstrated that higher edible mushrooms consumption was significantly associated with lower incidence of subclinical hypothyroidism among general adult population, especially in obese individuals.