Intravaginal sponges (IS) impregnated with progestagens are widely used for oestrous synchronization in ewes. As progestogens depress the immuno response, the first aim was to determine whether medroxiprogesterone acetate (MAP) content affects the vaginal bacteria number (VBN) in IS-treated anoestrous ewes. The second aim was to compare the effectiveness of different antibiotic treatments to control the VBN increase caused by IS. In both experiments, IS were inserted during 14 days in anoestrous ewes. In the first, 11 ewes received commercial sponges (50 mg MAP), and 10 ewes received placebo sponges. For the second experiment, IS were inserted in three groups (n = 12/group), containing oxytetracycline im (20 mg/kg); injected into the sponge (0.02 mg), or control (no antibiotic). At sponge withdrawal, all ewes received 300 UI eCG. Mucous samples were collected from the vagina before sponge insertion, at sponge withdrawal, 24, 48 and 72 h later, and the VBN (colony-forming units per ml; CFU/ml) was counted after 48-h incubation. Medroxiprogesterone content did not affect VBN (log CFU/ml: 4.3 ± 0.2 vs 4.4 ± 0.2 with and without MAP, respectively). Bacterial number increased from 3.5 ± 0.2 at sponge insertion to 6.9 ± 0.1 at sponge withdrawal (p < 0.0001) and decreased the following day to 4.3 ± 0.2 (p < 0.0001). In the second experiment, VBN increased at sponge withdrawal (p < 0.0001) in all groups and decreased the following day (p < 0.0001). The CFU/ml at sponge withdrawal was lower in ewes treated with antibiotics (p < 0.0001), being even lower when local rather than systemic antibiotic was administered (log CFU/ml: 3.3 ± 1.8 vs 7.2 ± 1.8). The day of oestrous VBN was similar for all treatments and similar to that observed before sponge insertion. We concluded that MAP does not influence the increase in VBN, as the main effect is provoked by the sponge device itself, and local antibiotic treatment resulted in a lower bacterial growth than systemic treatments.