The revolutionary development of green technology, such as ultrasound-assisted extraction, has had a significant impact on the extraction methods of natural compounds with pharmacological potential. These methods are considered green because they are environmentally friendly, minimizing the use of toxic solvents, energy consumption, and CO2 generation. Antimicrobial resistance is a global public health problem. Pathogens of dermatological and nosocomial relevance are more frequent because they are resistant to antimicrobials. Agave nuusaviorum leaves are used in traditional medicine to treat onychomycosis and inflammatory processes. Here, the antimicrobial activity of hydroethanolic extract of A. nuusaviorum leaves obtained by ultrasound-assisted extraction was evaluated. The antimicrobial activity was tested against 14 pathogens with dermatological and nosocomial relevance by the agar diffusion technique. The extract exhibited an antimicrobial effect on all microorganisms tested, showing inhibition zones of 9–16 mm for Staphylococcus aureus, Candida albicans, Candida parapsilosis, and eleven isolates of Trichophyton. The extract’s minimum inhibitory concentration ranged between 0.5 mg/mL for Staphylococcus aureus, 0.25–1 mg/mL for yeast, and 0.4–1.25 mg/mL for molds. The phytochemical screening revealed the presence of terpenes, phenolic acids, tannins, carbohydrates, and steroids in the extract. The results showed that A. nuusaviorum leaves have broad-spectrum antimicrobial potential.