Background: In a pulpectomy, the eradication of microbes from the primary root canal is accomplished through biomechanical preparation, which could be carried out with either manual or rotary instruments. Aims: The objective of this clinical trial was to evaluate the efficiency of manual K-files, H-files, and Kedo-S Square rotary files in reducing microbial flora after canal preparation in primary molars. Materials and Methods: This randomized clinical trial consisted of 45 primary molars requiring pulpectomy. The teeth were randomly allocated to one of the three groups: Group I: Manual K-files, Group II: Manual H-files, and Group III: Kedo-S Square rotary files, based on the type of instrumentation. Pre-and Post-instrumentation sampling was performed using clean absorbent paper points and kept in a clean Eppendorf tube having thioglycolate broth as the transport medium. Culturing was performed on agar media from which both aerobic and anaerobic microbial counts were estimated. Collected data were statistically analyzed using one-way analysis of variance (ANOVA) and Wilcoxon signed-rank test. Following root canal preparation, 87-89% reduction of the aerobic and anaerobic microbial load was noted in group I, whereas it was an 89-92% reduction in group II and a 93-95% reduction in group III. Results: Biomechanical preparation with Kedo-S Square rotary file showed higher efficacy in microbial reduction compared to manual instrumentation. Conclusion: Manual and rotary files were equally effective in removing root canal microbes. Biomechanical preparation with a KedoS Square rotary file resulted in greater microbial efficacy. Hence In children, effective root canal cleaning in a short period of time is a major consideration.