Quaternized chitosan (HTCC)-organic rectorite (OREC) intercalated composites based electrospun nanofibrous mats were fabricated from HTCC-OREC/polyvinyl alcohol (PVA) solutions for the first time. The morphology, intercalated structure, and antibacterial activity of the as-spun mats were investigated. The transmission electron microscopy images taken from HTCC-OREC composites showed that HTCC chains were inside the OREC interlayer. Scanning electron microscopy results verified that more typical fibrous structure would be generated by adding OREC. Fourier transform infrared spectra and energy-dispersive X-ray spectroscopy results indicated that OREC existed in the HTCC-OREC/PVA nanofibrous mats. The intercalation structure in nanofibrous mats was confirmed by X-ray diffraction results, which confirmed that HTCC and PVA chains could intercalate into the interlayer of OREC. The antibacterial activity of the electrospun mats was enhanced when the amount of the OREC increased. Therefore, the novel ternary nanofibrous mats could be used in the field of food packaging and biomedical applications.