Infant colic is a condition characterised by excessive crying of unknown cause. Despite more than sixty years of research the aetiology of infant colic remains unclear and there is no widely accepted treatment. Recently, a pathophysiological relationship due to an imbalance in microbiota and/or inflammation has been proposed as a cause of infant colic.  This has led to the Baby Biotics trial, which investigated whether administration of the probiotic bacterium Lactobacillus reuteri was an effective treatment for colic, and examined the effects on intestinal microbiota and intestinal inflammation.  In the current study, infants with colic were given daily drops of a L. reuteri solution or placebo for 28 days.  We examined faecal samples collected from 65 infants and assessed E. coli colonisation, intestinal microbial diversity, intestinal inflammation (calprotectin levels) and infant crying time at day 28 of the trial. We found that L. reuteri supplementation and/or colonisation did not significantly modulate intestinal microbial diversity or intestinal inflammation and did not significantly reduce crying time in infants with colic. In contrast, we observed a non-significant, positive trend for a positive association between L. reuteri and E. coli and a significant positive correlation between L. reuteri colonisation levels and crying time.  These findings suggest that as L. reuteri colonisation did not modulate E. coli colonisation, diversity or inflammation, the positive association with infant crying time may be due to other, unknown causes. These results highlight the need for further studies to facilitate the development of evidence-based treatment strategies for infant colic.