The inclusion characteristics and microstructure in the low carbon microalloyed steel with addition ZrO2 nanoparticles were investigated by high temperature experiment and metallurgical analysis. The results showed that after ZrO2 nanoparticles were added, a large number of Zr–Al–Si–O+MnS inclusion, ZrO2+Al–Si–O+MnS inclusion, and ZrO2+MnS inclusion appeared in the steel, and these Zr-containing inclusions were effective to induce acicular ferrite (AF) formation. With the amount of added ZrO2 nanoparticles increased from 0.013% to 0.054%, the inclusions types had no significant effect, but the inclusions size distribution, number density and average diameter were affected. When the addition amount of ZrO2 nanoparticles was 0.027%, the proportion of large inclusions (larger than 5.0 µm), the inclusions number density and average diameter all were reached the extremum values, respectively 10.81%, 111/mm2, 2.62 µm. Moreover, as ZrO2 nanoparticles adding into steel, the majority solidification microstructure changed from bainitic ferrite (BF) to AF. With the adding amount of nanoparticles increase, the proportion of AF first increased and then decreased, also reached the maximum of 67.65% as the addition amount was 0.027%. Finally, Mn-depletion zone (MDZ) in the vicinity of Zr-containing inclusion was observed, and the MDZ was believed to be one of the possible mechanisms of Zr-containing inclusions inducing AF formation.