Background Healthcare-associated infections (HCAI) are a worldwide threat in intensive care units particularly in the pediatric intensive care units with a major cause of morbidity and mortality among this age group. The aim of the study is to determine the prevalence and risk factors predisposing to multidrug-resistant organisms (MDROs) infections among pediatric intensive care unit (PICU) patients at Ain Shams Pediatric University Hospitals as well as determining mortality and morbidity rates along with the length of stay at PICU. Results Culture results revealed that of the 282 patients evaluated, only 26 (9.2%) were MDROs (half of the affected patients had Acinetobacter species (50%) and the rest of them were free, 256 (90.8%). Our study revealed that the majority of MDROs were isolated from sputum in more than half of the patients 19/32 (59.3%) followed by whole blood in 10/32 (31.2%) and urine in 3/32 (9.4%). Pulmonary system was the most common affected site and was mainly colonized by MDR Acinetobacter (71.4%) followed by MDR Klebsiella (41.6%). Regarding MDR risk factors in our PICU, multivariate logistic regression analyses showed significant relationship between MDROs and age under 1 year (odds ratio [OR] 2.4554; 95% confidence interval [95% CI] (1.072–5.625); p = 0.043) and underlying pulmonary disease (OR 2.417; 95% CI (1.014–5.761); p = 0.592). A statistically significant higher mortality was detected in patients colonized with MDROs 9/26 (34.6%) versus MDROs non-colonized patients 32/256 (12.5%) [P=0.002]. Moreover, MDROs infection has negative significant risk with discharged patients ([OR] 0.269; [95% CI] (0.111–0.656); p = 0.002). Additionally, patients infected with MDROs did have significantly greater PICU stay than those non-infected [median (IQR), 16.5 (10.7–22), 5 (4–8), P=0.00] and have longer ventilation [median (IQR), 15.5 (10–18), 3 (2–10), P=0.00]. Conclusion Prevalence of MDROs (9.2%) was low among PICU cases at Ain Shams University Hospitals. Most common MDROs were Acinetobacter and Klebsiella followed by pseudomonas species. The frequency of gram-negative organisms is much more common than gram-positive organisms. An increasing rate of antimicrobial resistance with increasing mortality and morbidity among PICU patients is observed worldwide; even for new categories, so, strict infection control programs should be implemented.