Systemic antibiotics are extensively used to control moderate to severe acne. Hence, it is crucial to understand their impact on the skin microbiota, which is supposedly perturbed. The purpose of this study was to compare the makeup and diversity of the skin microbiota in acne patients before and after taking oral antibiotics. A longitudinal cohort study was performed on 20 participants with moderate to severe facial acne with no recent use of oral and topical antibiotics/retinoids. Patients were prescribed oral doxycycline, 100 mg, twice daily for six weeks. Skin areas on the cheek were sampled for 16S ribosomal RNA gene sequencing at baseline, and after six weeks of doxycycline treatment. Ten males and 10 females aged 11 to 44 years with a median Investigator’s Global Assessment score of 3 (moderate) were enrolled. At baseline, Cutibacterium acnes (formerly Propionibacterium acnes) was the most dominant species followed by Staphylococcus epidermidis. Acne severity showed a positive correlation with the abundance of Cutibacterium acnes. Across all subjects, antibiotic treatment reduced clinical acne grades and was associated with a 1.96-fold reduction in the relative abundance of Cutibacterium acnes (p = 0.01, 95% CI −22% to −3%). Marked changes were also identified in other bacterial species, such as Cutibacterium granulosum (formerly Propionibacterium granulosum), which increased by 4.46-fold (p = 0.02, 95% CI 0.004% to 0.9%) in the treated samples. In general, antibiotics administration was associated with an increase in bacterial diversity (alpha diversity). Principal coordinates analysis showed mild clustering of samples by patient (analysis of similarity, R = 0.135, p = 0.04) whereas there was scant clustering with treatment (ANOSIM, R = 0.005; p = 0.29). In conclusion, we found individuals with acne to have a unique microbial signature. Acne treatment with systemic antibiotics was associated with changes in the composition and diversity of skin microbiota, especially Cutibacterium acnes, which correlates with acne severity. Our study provides insight into the skin microbiota in acne and how it is modulated by systemic antibiotics.