Effects of a live vs heat-inactivated probiotic Bifidobacterium spp in preterm infants (the ProPara study)- a randomized clinical trial

Background: Heat-inactivated probiotics may provide an effective alternative to live probiotics by avoiding the risk of probiotic sepsis, altered immune responses and antimicrobial resistance while retaining probiotic benefits. Objective We assessed safety and efficacy of a heat-inactivated probiotic in very preterm (VP: gestation < 32 weeks) infants. Methods VP infants were recruited including a pre-planned subgroup of extremely preterm (EP: gestation < 28 weeks). Mixture of heat-inactivated (HP) or live probiotic (P) strains B. breve M-16V, B. longum subsp. infantis M-63, B. longum subsp. longum BB536 (Total 3 x10 9 CFU/day) assuring blinding. Primary outcomes included fecal calprotectin (FCP) levels and safety. Secondary outcomes included fecal microbiota assessed by 16S ribosomal RNA and shotgun sequencing and short chain fatty acid (SCFA) levels in samples collected after the 1st (T1) and 3rd (T2) week of supplementation. Results 86 VP (P:43; HP:43) infants were randomized. Median (range) FCP was lower at T2 vs T1 in both HP [75 (8-563) vs 109 (5.1–725) µg/g; p = 0.22] and P [80 (21–277) vs 105 (11–842) µg/g; p = 0.4] groups. Total FCP and SCFA were comparable between HP vs P groups at T1 and T2 (p > 0.05). Propionate was significantly raised in both groups, whilst butyrate was significantly raised in HP group (all p < 0.01). At T2, alpha diversity increased but was comparable and beta diversity showed significantly different community structures in both groups (all p < 0.01). Actinobacteria significantly increased and Bacteroidetes decreased at T2 vs T1 for both groups (p < 0.05). Bifidobacteriacae increased (p < 0.001) whilst Staphylococcaceae decreased (p < 0.05) for both groups. Bifidobacteriacae, B. longum subsp. infantis and B. longum subsp. longum levels were comparable. Clinical outcomes were comparable and there were no adverse events. Conclusions Heat-inactivated probiotic was safe and well tolerated, with an intestinal anti-inflammatory effect comparable to live probiotic. Adequately powered randomised trials are needed to assess its clinically significant effects in preterm infants. Trial Registration ID and URL Australian and New Zealand Clinical Trials Registry (ACTRN12618000489291); ANZCTR - Registration