Background: Gastroenteritis has a significant mortality andmorbidity incidence in children globally. Antimicrobial resistance in Enterobacteriaceaeis a serious public health problem, especially in developing countries.Objective: The present study aimed to evaluate the anti-bacterial and anti-biofilmactivities of AgNPs alone and in combination with kanamycin against multidrug-resistant Enterobacteriaceae and P. aeruginosa isolated from diarrheal children.Materials and methods: 90 Enterobacteriaceae and P. aeruginosa isolates fromdiarrheal children were evaluated against 10 antibiotics. Minimum inhibitory doses ofAgNPs and kanamycin were determined using broth microdilution, synergistic wasdetermined using Checkerboard dilution tests, and the Calgary technique was used toanalyze biofilm development.Results: A total of 90 stool cultures were conducted for bacteria associated withdiarrhea among children attending some Baghdad hospitals. The findings revealed thatbacterial diarrhea was most often caused by E. coli 31 (34.5%), followed by S. typhi 19(21.1%), K. pneumoniae 14 (15.5%), P. aeruginosa 11 (12.2%) and S. sonnei 6 (6.7%),and significant variations between the strain's species were discovered using statisticalanalysis (P < 0.05). The present study's findings revealed that bacteria isolated fromchildren with diarrhea were spread significantly in age groups of 37–48 months andsignificantly different between age groups (P < 0.05), with a male/female ratio of0.57/1. Imipenem and amikacin were the most active antibiotics compared to penicillin,which was the least effective antibiotic. The combination of sublethal doses of AgNPswith sub-MIC (½MIC) of kanamycin exhibited substantial synergistic bactericidaleffects against MDR-Enterobacteriaceae. AgNPs inhibited biofilm formation by55%–65% for diarrhea-causing bacteria, while the combination of AgNPs withkanamycin demonstrated the strongest biofilm inhibition of around 80%–90% againstMDR-Enterobacteriaceae with a highly significant variation (P < 0.05).Conclusions: The outcomes of the research shows that the combination of AgNPs withkanamycin has remarkable synergistic bactericidal and anti-biofilm effectivenessagainst MDR-Enterobacteriaceae isolated from diarrheal children.