To evaluate the safety and efficacy of netilmicin/dexamethasone combination in the treatment of meibomian gland dysfunction (MGD)-associated posterior blepharitis. In this prospective and controlled study were enrolled 40 patients with MGD and symptoms of dry eye disease. Two groups were established: 20 patients (group 1) received netilmicin 3 mg/ml and dexamethasone 1 mg/ml eye gel, whereas in group 2 (20 patients) received vehicle for 15 days. Patients were evaluated at baseline, 15 and 45 days, including SANDE and VARS questionnaire, non-invasive tear film breakup time (NIBUT), tear meniscus height (TMH), ocular redness and meibography score. Moreover, fluorescein tear-film breakup time (TBUT), fluorescein ocular surface staining, lid margin evaluation including hyperemia, edema and meibum expressibility and quality examinations were carried out. Furthermore, intraocular pressure (IOP) and best-corrected visual acuity (BCVA) were considered as safety parameters. In group 1, at 15 and 45 days there were statistically significant changes in VARS and SANDE score (p < 0.0001) as well as lid margin parameters, TBUT and fluorescein ocular surface staining (p < 0.0001). Comparing the two groups, a significant improvement of SANDE score was observed at 15 days in group 1 as well as lid margin parameters, TBUT and fluorescein ocular surface staining at 15 and 45 days (all p < 0.0001). Netilmicin/dexamethasone combination is effective and safe to treat MGD-associated posterior blepharitis improving both symptoms and ocular surface signs.