The purpose of this study was to compare the effects of adding Lacticaseibacillus (previously Lactobacillus) rhamnosus GG (LGG) to a partially hydrolyzed protein formula on the behavioral state, stool microbiota composition, and calprotectin levels in infants with infantile colic. Infants diagnosed with colic (using modified Wessel’s criteria: cried and/or fussed ≥3 h/day for ≥3 days/week, in a 1-week period) were randomly assigned to receive either a commercially available partially hydrolyzed cow’s milk-based infant formula (PHF, n=35) or a similar formula with added LGG (PHF-LGG, n=36) over a 3-week feeding period. Infant behavior was reported by parents at 3 different points (Study Days 2–4, 10–12, and 18–20). The primary outcome was the mean duration of weeping or fussing throughout a 3-day period (in hours). At the beginning of the study (Days 1-2) and at the end of the study (Days 19-21), feces samples were taken to analyze for LGG colonization (using quantitative polymerase chain reaction) and microbial abundance (16S rRNA gene sequencing) and calprotectin (μg/g). Over the course of the trial, there was no statistically significant difference between the groups in terms of the mean SE duration of crying/fussing or awake/content behavior. By the end of the research, there were no significant differences between the groups in the percentage of infants who experienced colic. For both groups, the incidence of colic decreased by the end of the study compared to the beginning. Also, there was no significant difference in the calprotectin change in the feces across the groups. The PHF-LGG group had higher LGG abundance at the end of the study compared to the baseline (P<0.001), whereas the PHF group had higher alpha diversity (P=0.022). At the study’s End, there was a statistically significant difference in beta diversity between PHF and PHF-LGG. When comparing the PHF group and the Baseline group, the PHF-LGG had a greater relative abundance of L. rhamnosus at the end of the study. Both study formulae were well tolerated in this small pilot trial of infants with colic. Throughout the course of the trial, both groups showed a decrease in crying/fussing and an increase in awake, content behavior. In other words, the probiotic was successfully integrated into the microbiome, as shown by the results of the study. The addition of LGG to the partially hydrolyzed protein mix was linked to substantial alterations in the gut microbiome.