In this study, an industrially scalable method is reported for the fabrication of polylactic acid (PLA)/silver nanoparticle (AgNP) nanocomposite filaments by an in-situ reduction reactive melt mixing method. The PLA/AgNP nanocomposite filaments have been produced initially reducing silver ions (Ag+) arising from silver nitrate (AgNO3) precursor mixed in the polymer melt to elemental silver (Ag0) nanoparticles, utilizing polyethylene glycol (PEG) or polyvinyl pyrrolidone (PVP), respectively, as macromolecular blend compound reducing agents. PEG and PVP were added at various concentrations, to the PLA matrix. The PLA/AgNP filaments have been used to manufacture 3D printed antimicrobial (AM) parts by Fused Filament Fabrication (FFF). The 3D printed PLA/AgNP parts exhibited significant AM properties examined by the reduction in Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) bacteria viability (%) experiments at 30, 60, and 120 min duration of contact (p < 0.05; p-value (p): probability). It could be envisaged that the 3D printed parts manufactured and tested herein mimic nature’s mechanism against bacteria and in terms of antimicrobial properties, contact angle for their anti-adhesive behavior and mechanical properties could create new avenues for the next generation of low-cost and on-demand additive manufacturing produced personal protective equipment (PPE) as well as healthcare and nosocomial antimicrobial equipment.