Three new naphthoquinone-based meroterpenoids (1–3), including spironaphthomeroterpenoid (1), a novel naphthoquinone-based meroterpenoid with a rare [5, 6] spiro-ring skeleton, and naphthablins D and E (2–3), along with three known analogues (4–6) were isolated from Streptomyces sp. HBERC-16614. The structures of these compounds, including their absolute configurations, were established by extensive spectroscopic analysis and electronic circular dichroism (ECD) spectrum. The antibacterial activities of compounds (1–6) were evaluated against Staphylococcus aureus, Streptococcus suis, and Escherichia coli. Among them, compound 1 demonstrated potent antibacterial activity against S. aureus, with a minimum inhibitory concentration (MIC) value of 3.13 µg/mL.