Healthcare workers in the hospital environment are at risk of infection and body fluids such as saliva, bacterial contamination, oral bacteria, etc. directly or indirectly exacerbate this issue. These bio-contaminants, when adhered to hospital linens and clothing, grow substantially, as conventional textile products provide a favorable medium for bacterial and viral growth, adding to the risk of transmitting infectious diseases in the hospital environment. Textiles with durable antimicrobial properties prevent microbial colonization on their surfaces and help contain the spread of pathogens. This longitudinal study aimed to investigate the antimicrobial performance of PHMB-treated healthcare uniforms during prolonged usage and repetitive laundry cycles in a hospital environment. The PHMB-treated healthcare uniforms displayed non-specific antimicrobial properties and remained efficient (>99% against S. aureus and K. pneumoniae) after use for 5 months. With the fact that no antimicrobial resistance was reported towards PHMB, the presented PHMB-treated uniform may reduce infection in hospital settings by minimizing the acquisition, retention, and transmission of infectious diseases on textile products.