Novel nanofibrous materials composed of polyelectrolyte complexes (PECs) between N,N,N-trimethylchitosan iodide (TMCh) and poly(acrylic acid) (PAA) or poly(2-acrylamido-2-methylpropanesulfonic acid) (PAMPS) were prepared. This was achieved by facile one-pot electrospinning of solutions of the oppositely charged polyelectrolyte partners. It was rendered possible by using a solvent system containing formic acid and/or by adding a strongly ionized low-molecular-weight salt (CaCl2). Use of formic acid enabled TMCh/PAA nanofibers containing in situ synthesized silver nanoparticles (AgNPs) to be electrospun. The AgNPs had an average diameter of 3.0 +/- 0.8 nm and were uniformly distributed in the nanofibers as evidenced by the performed transmission electron microscopic (TEM) analyses. The prepared nanofibers preserved their morphology and did not dissolve in phosphate-buffered saline (PBS). Hybrid AgNPs-containing PEC nanofibrous materials showed good antibacterial activity against Gram-positive bacteria Staphylococcus aureus and Gram-negative bacteria Escherichia coli and possessed higher efficacy than that of the nanofibers of the same composition without AgNPs and TMCh.