Background: Acute diarrhea continues to be one of the main causes of morbidity in children, however the etiologica diagnosis presents limitations given the low sensitivity of traditional methods. Aim: To describe the microorganisms identified in children who attended the emergency department (ED) in Santiago, Chile, due to acute diarrhea and to whom a gastrointestinal panel was requested as part of their study. Material and Methods: Clinical records and results of the gastrointestinal panel carried out between June 2017 and March 2020 were reviewed. Results: 198 patients were included, the average age was 54.5 months and 60.6% (120/198) were males. Positivity was 78.8% (156/198) with 35.3% (55/156) of the samples being polymicrobial. 229 microorganisms were identified, of which 72.9% (167/229) co-rresponded to bacteria, 25.8% (59/229) to viruses, and 1.3% (3/229) to parasites. Campylobacter spp. and enteropathogenic Escherichia coli (EPEC) were the most frequently identified bacteria. Patients with detection of Campylobacter spp. presented a higher frequency of fever (p = 0.00). EPEC was isolated in 82.5% of the cases in polymicrobial samples. Discussion: The results emphasize the potential of molecular studies to improve the etiological diagnosis of diarrhea and at the same time lead to question the pathogenic role of some microorganisms.