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Previous data from our research group showed that chitosan-coated poly(isobutylcyanoacrylate) nanoparticles (NPs) (denoted PIBCA/Chito20) exhibited intrinsic anti-Trichomonas vaginalis activity, while PIBCA/pluronic® F68 without chitosan (PIBCA/F68) were inactive. However, the mechanism of anti-T. vaginalis activity of chitosan-coated PIBCA NPs is still unknown. Our hypothesis is that chitosan-coated NPs are internalized by the parasite, contrarily to PIBCA/F68. In this investigation, the impact of NP surface on their internalization by the protozoan was studied using flow cytometry and parasite morphological changes after different incubation times with PIBCA/Chito20 NPs were monitored by electron microscopy. Flow-cytometry revealed that PIBCA/Chito20 NPs were uptaken by T. vaginalis as early as 10-min-incubation. Drastic cell morphological transformations were observed from scanning electron microscopy and transmission electron microscopy after incubation with PIBCA/Chito20 NPs. Numerous pits were seen on cell membrane since 10min. Gradual increase in contact time increased NP endocytosis and induced proportional damages to T. vaginalis membrane. Then, investigation of whether PIBCA/Chito20 NPs can improve MTZ anti-T. vaginalis activity was studied using checkerboard experiment. Calculation of fractional inhibitory concentration index (FICI=3.53) showed an additive effect between NPs and MTZ.