Talaromyces marneffei (T. marneffei) is one of the most important opportunistic human pathogens endemic in Southeast Asia. Talaromycosis, which was once regarded as an opportunistic infectious disease in patients with acquired immunodeficiency syndrome, is being increasingly reported in HIV-negative populations. Since T. marneffei infection can be localized or disseminated, patients may present with a variety of symptoms. However, mediastinal infection attributed to T. marneffei is extremely rare. We report the case of a 32-year-old female who manifested a large mediastinal mass and was eventually diagnosed as acute T. marneffei mediastinitis. The patient was HIV-negative and had no direct contact with intermediate hosts. We successfully managed to treat the patient with inhaled amphotericin B deoxycholate and observed lesion absorption in subsequent CT examinations. To our knowledge, this is the first published case of T. marneffei mediastinitis and first use of inhaled antifungal monotherapy on patients with T. marneffei infection.