Objectives: We aimed to comprehensively explore the etiology of granulomatous lobular mastitis (GLM) to optimize treatment programs.

Methods: We collected 30 fresh mastitis samples for metagenomic next-generation sequencing, morphological observation, and analysis of the clinical information.

Results: Of the 30 samples, 25 were GLM; pathogens were detected in 17, these were: Corynebacterium kroppenstedtii (10 of 25, 40%); C. kroppenstedtii and Pseudomonas oleovorans (3 of 25, 12%); C. kroppenstedtii and human gammaherpesvirus 4 (1 of 25, 4%); Acinetobacter baumannii and C. kroppenstedtii (1 of 25, 4%); P. oleovorans (1 of 25, 4%); and Tepidiphilus thermophilus (1 of 25, 4%). Abnormal sex hormone levels (mainly prolactin) and/or autoimmune function were found in 12 of the 25 samples. Lipophilic antibiotics (rifampicin) were found to work effectively in patients with slow-healing wounds after surgery.

Conclusions: The main pathogenic factor of GLM is C. kroppenstedtii infection, but other unusual pathogens (P. oleovorans, human gammaherpesvirus 4, A. baumannii, T. thermophilus) are likely to be closely related to GLM, particularly human gammaherpesvirus 4 (Epstein-Barr virus)-associated mastitis, which may be a new entity of mastitis. Abnormal levels of sex hormones and autoimmune function are also common causes. Therefore, lipophilic antibiotics (rifampicin) and prolactin inhibitors may be an effective treatment.

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