To investigate the surgical outcomes of vesiculoscopy on refractory hematospermia and ejaculatory duct obstruction (EDO), the clinical data (including pelvic magnetic resonance imaging (MRI) examinations and the long-term effects of endoscopic treatment) from 305 patients were analyzed. Four main etiologic groups were found on MRI. We found that 62.0% (189/305) of patients showed abnormal signal intensity in MRI investigations in the seminal vesicle (SV) area. Cystic lesions were observed in 36.7% (112/305) of the patients. The third sign was dilatation or enlargement of unilateral or bilateral SV, which were observed in 32.1% (98/305) of the patients. The fourth sign was stone formation in SV or in an adjacent cyst, which was present in 8.5% (26/305) of the patients. The transurethral endoscopy or seminal vesiculoscopy and the related procedures, including fenestration in prostatic utricle (PU), irrigation, lithotripsy, stone removal, biopsy, electroexcision, fulguration, or transurethral resection/incision of the ejaculatory duct (TURED/TUIED), chosen according to the different situations of individual patients were successfully performed in 296 patients. Fenestrations in PU+ seminal vesiculoscopy were performed in 66.6% (197/296) of cases. Seminal vesiculoscopy via the pathological opening in PU was performed in 10.8% (32/296) of cases. TURED/TUIED + seminal vesiculoscopy was performed in 12.8% (38/296) of cases, and seminal vesiculoscopy by the natural orifices of the ejaculatory duct (ED) was performed in 2.4% (7/296) of cases. Electroexcision and fulguration to the abnormal blood vessels or cavernous hemangioma at posterior urethra were performed in 7.4% (22/296) of cases. Two hundred and seventy-one patients were followed up for 6-72 months. The hematospermia of all the patients disappeared within 2-6 weeks, and 93.0% of the patients showed no further hematospermia during follow-up. No obvious postoperative complications were observed. The transurethral seminal vesiculoscopy technique and related procedures are safe and effective approaches for refractory hematospermia and EDO.