Background: Early host-microbe interaction provides important maturational stimuli for the developing immune system. The role of prenatal microbial contact remains elusive. Objectives: Our aim was to investigate whether microbes in placenta or amniotic fluid affect fetal innate immune gene expression during late pregnancy and whether innate immune gene expression profiles in the placenta and the fetal gut may be modulated by dietary supplementation with specific probiotics. Methods: Altogether 43 pregnant women were randomized to receive (1) Bifidobacterium lactis, (2) B. lactis in combination with Lactobacillus rhamnosus GG (LGG) or (3) placebo for 14 days before elective cesarian section at full term in a double-blind clinical trial. Bacteria in amniotic fluid and placenta were detected by quantitative (q)PCR. The expression of Toll-like receptor (TLR)-related genes in the placenta and meconium samples was assessed by qPCR. Gene expression patterns in meconium were interpreted to reflect immune physiology in the fetal gut. Results: The study was completed by 29 mother-infant pairs. Bacterial DNA was detected in all placenta samples. Microbial DNA in amniotic fluid and placenta was associated with changes in TLR-related gene expression in the fetal intestine. Maternal probiotic supplementation significantly modulated the expression of TLR-related genes both in the placenta and in the fetal gut. Conclusions: Microbial contact in utero is associated with changes in fetal intestinal innate immune gene expression profile. Fetal and placental immune physiology may be modulated by maternal dietary intervention using specific probiotics.