Au nanowire/Pichia pastoris cell composites were prepared using a microorganism-mediated method, where closely packed Au nanowires (AuNWs) are synthesized at high yields in the presence of hexadecyltrimethylammonium bromide (CTAB) without auxiliary seeds. The initial concentrations of ascorbic acid (AA), CTAB, and tetrachloroaurate trihydrate, as well as the amount of Pichia pastoris cells (PPCs), were optimized for the formation of AuNWs. For the formation of AuNWs, Au(III) ions were initially bound by the PPCs and then reduced by AA to produce Au nanoparticles (AuNPs) via preferential nucleation. The linear fusion of these AuNPs influenced by CTAB resulted in the formation of growing AuNWs, whereas the secondary nucleation beyond the PPCs produced small AuNPs that were subsequently consumed through Ostwald ripening during the aging of AuNWs. These AuNW/PPCs composites could be effectively used as surface-enhanced Raman scattering substrates for the detection of rhodamine 6G.