I haven’t, surprisingly, posted about probiotics recently, but there are some new findings in the literature that warrant discussion.
Granger C, et al. Necrotising enterocolitis, late-onset sepsis and mortality after routine probiotic introduction in the UK. Arch Dis Child Fetal Neonatal Ed. 2021. The majority of pre- post- studies of introduction of probiotics that have been published have shown a significant reduction in NEC. Of course, there could well be a huge publication bias in such publications. I am pleased therefore, that this study was published, even though it showed little impact on NEC, overall. Especially as it is from a centre with a huge interest in intestinal microbiome development in the preterm, and in NEC, that is Newcastle upon Tyne, UK. They showed a small reduction from 10.6% of babies <32 weeks to 9.2% after routine introduction of probiotics, intially Infloran (Lactobacillus acidophilus and Bifidobacterium Bifidum) then Labinic (same species plus B longum ssp infantis)in the last 30% of the treated cohort. Overall there was no major impact on their outcomes, but subgroup analysis suggested a reduction in NEC in the more mature babies >28 weeks, and a reduction in late-onset sepsis in the less mature <28 weeks. Overall mortality, and mortality associated with those complications, were not affected.
Tobias J, et al. Bifidobacterium infantis EVC001 Administration Is Associated With a Significant Reduction In Incidence of Necrotizing Enterocolitis In Very Low Birth Weight Infants. J Pediatr. 2022. In this centre, in contrast, there was a dramatic reduction in NEC when they started routine supplementation with B. infantis. The strain they used, noted in the title, is a commercially produced strain produced by a company which sponsored this study, and of which one of the authors is an unpaid consultant. In this centre in Oregon, they showed a reduction in NEC from 11% to 2.6% of VLBW infants after introduction of the probiotic. Mortality related to NEC was also reduced, from 2.7% to 0. The impact applied to the ELBW subgroup also, 19.2% NEC incidence reduced to 5.3%. The relative risk of NEC after probiotic introduction was 0.27, compatibility intervals (0.09-0.61).
Another study using the same strain is Nguyen M, et al. Impact of Probiotic B. infantis EVC001 Feeding in Premature Infants on the Gut Microbiome, Nosocomially Acquired Antibiotic Resistance, and Enteric Inflammation. Front Pediatr. 2021;9:618009. Of note, many of the authors were employees of the company which makes the probiotic preparation, and they also funded the study. In this project VLBW babies <32 wks were given probiotics, bigger preterm babies were not. They also differed slightly in diet, as they all predominantly received human milk, but formula was only given to the larger babies that needed a supplement, rather than donor milk given to the smaller ones. There are several very pretty figures in the publication, some parts of which I actually understand! The following for example, graphic B, I think, shows the overall percentage of all the bugs they found in all the stools, that were in each of the 3 groups noted by the colours, I couldn’t tell you exactly what they are percentages of, as all the other OTUs are missing from the graph, but it is designed to convince you that there are a lot more Bifidobacteria around.
As for panels C and D, they are from the 2 GA groups, C from the more immature babies who received the probiotic, and D from the bigger babies. They also looked at whether the stool samples from the babies had bacteria capable of metabolising HMOs, and I will have to take their word for it, but it sure looks like the babies fed the probiotic had more ability to metabolise HMOs.
The 2 centres involved had a different pattern of antibiotic resistance genes, which the babies acquired throughout their stays in the NICU. As a non-specialist, I am not sure how well known this is, it certainly was something I was not aware of, but it is not a surprise, that different NICUs have different antibiotic resistance gene populations. Different NICUs tend to have differing bacteria causing their sepsis, and intestinal colonisation during NICU stays. They also showed that babies transferred from one hospital to the quaternary centre acquired the receiving centre’s antibiotic resistance genes after they arrived. The B infantis reduced the abundance of those genes compared to more mature, untreated controls. There was also, finally, a reduction in pro-inflammatory biomarkers among the immature, probiotic receiving babies. They state that their data show that gut microbiome composition can be altered by feeding probiotics. But the data don’t show that. As there are no randomized controls, all they can say is that the 2 groups are different, and that there is an association with probiotic administration. Which is my major criticism of all these EVC001 trials, why haven’t they performed an RCT powered to detect a reduction in NEC? There are still many centres in the USA who don’t use probiotics, partly because of the lack of an FDA licensed preparation containing, probably, the most important species of probiotic organism for the newborn, which is extremely well standardised, with the quality control of a medical product, like this one.
They might be able to convince the FDA, for that latter requirement, to give them a license for preterm babies, for the prevention of NEC, if their trial proved the value of Bifidobacterium longum susp infantis, strain EVC001 for the prevention of this devastating condition in preterm infants.
As one of the corresponding authors of the Nguyen et al paper, I would be happy to explain the figures to you and the underlying methodologies as you don’t seem to grasp them, based on your characterization of our work in this post.
Please explain where I erred in my interpretations. I would be happy to correct, my post, there are certainly many aspects of the methods that I don’t understand in any detail.
Happy to offer some clarifying thoughts here:
“The following for example, graphic B, I think, shows the overall percentage of all the bugs they found in all the stools, that were in each of the 3 groups noted by the colours, I couldn’t tell you exactly what they are percentages of, as all the other OTUs are missing from the graph, but it is designed to convince you that there are a lot more Bifidobacteria around.”
I respectably disagree with your conclusion of this figure. If one notes the text in the paper and the figure legend, this is not a representation of OTU abundance or ‘the overall percentage of all the bugs in the stools’. This figure describes the proportion of all infant fecal samples that were either fed the probiotic or not that were clustered into each of the discriminative sample ‘cluster types.’ The three colors represent the three ‘cluster types’ observed among these samples. For a more exhaustive discussion of the methodology we employed, see “Taxonomic Cluster Grouping Assignment” in the methods. Thus, while one might assume that this means there is a lot more bifidobacteria around, it would be an extrapolation we do not directly infer in panel B. I would direct your attention to Panel A in Figure 3, as this figure does directly illustrate the abundance of Bifidobacteriaceae, in case readers would like to view the abundance for each sample collected.
The Figure legend here is somewhat instructive in describing the graph in Panel B, and I’d direct readers to it for a concise explanation. We wrote, “B) Fecal sample composition was clustered by species-level signature taxa (e.g., B longum species, Escherichia coli and unclassified Escherichia, and S. epidermidis), as indicated by color. Infants in the EVC001-fed feeding protocol more frequently developed a gut microbiome composed of primarily B. longum species (blue), while the infants not assigned to receive the probiotic frequently had compositions typified by high levels of Enterobacteriaceae (e.g., Escherichia species) and/or Staphylococcus (red and orange, respectively).”
“As for panels C and D, they are from the 2 GA groups, C from the more immature babies who received the probiotic, and D from the bigger babies.”
Panels C and D show the relationship between Bifidobacteriaceae abundance over time (blue), in relation to Enterobacteriaceae (red) among infants that have been fed the probiotic (Panel C) or not (Panel D). Panel D is quite illustrative in showing that absent introduction of the probiotic, we do not see later recovery in the microbiome of these infants prior to discharge.
As we stated in the limitations and throughout the paper, one of the confounding issues with our cohort is the difference in gestational ages at birth, and by extension the age in days during sample collection. In Panels C and D, we show what the populations of two key taxa are, in aggregate, across these populations (e.g., those who received the probiotic and those that did not).
“There was also, finally, a reduction in pro-inflammatory biomarkers among the immature, probiotic receiving babies. They state that their data show that gut microbiome composition can be altered by feeding probiotics. But the data don’t show that.“
As we showed in Figure 2, after correcting for all other metadata variables (as noted in the figure and in the text), we still find probiotic supplementation is the only variable that explains the abundance of Bifidobacterium, the only organism in the probiotic. Second, Figure 6 shows the same effect on inflammatory cytokines and no clinical metadata explains the difference as strongly, nor does it come close in terms of the magnitude of the effect.
It would be exceptionally difficult to assert the abundance of the B. infantis observed among these infants is the result of another factor and (I would argue) remarkably illogical, given virtually all data we have on well adapted strains of infant-associated Bifidobacterium species and their ability to colonize the infant gut and the evidence provided by metagenomic sequencing of these samples. Indeed, as we show in Figure 3 (the panels noted above), the infants that were not fed the probiotic do not develop a high mean abundance of Bifidobacterium. If another feature of the infant’s clinical care or a difference in their gestational age at birth or any other metadata variable is responsible for the differences observed, what possible explanation can be identified that would account for the differences in abundance of an exceptionally rare organism that is genetically identical to the probiotic administered?
While randomized clinical trials clearly hold significant power in shaping clinical practice and provide a strong rationale for changes in clinical care, compelling data showing the impact and benefit of a probiotic organism cannot simply be ignored and dismissed because it does not have the study design of a pharmaceutical intervention. As I have not been involved with the company for some time, I can’t speak to the rationale for not pursuing a clinical trial in premature infants, but I can assert that our data do indeed show that feeding this organism (the product is categorized as a food for special dietary use (FSDU), rather than a dietary supplement) that has a known mechanism and is ecologically adapted to the breastfed infant gut microbiome. Moreover, I would encourage skeptical researchers to analyze our findings (as the data are publicly deposited).
Thank you for those clarifications. My comment about being unable to state that the “gut microbiome can be altered” was a very narrow criticism, based on the lack of randomized controls, as differences between the groups are not just in the probiotic administration, but in other characteristics also. However, I think it is highly unlikely that the differences in intestinal microbiome development that are demonstrated are just because the intervention group was less mature; to my mind, by far the most likely explanation for the preponderance of Bifidobacteria, is the receipt of B infantis by the intervention group, it would take some real mental gymnastics to come up with another explanation. That leaves some uncertainties, however; how much of the “secondary” differences, such as the associated reduction in Enterobacteriaceae, are due to the B infantis administration? I am someone who has promoted the use of probiotics for years, based on results of randomized trials, supported by mechanistic studies, of which this is one of the best. I think the evidence suggests that B infantis is the most important probiotic for NEC prevention, and I would love to see studies that show that the changes in microbiome development that this study showed, which are mostly probably due to the B infantis administration, lead to significant reductions in NEC.
10%-11% NEC is an excessive amount to begin with. I am interested to know if probiotics would be useful in a NICU that doesn’t have NEC.
I’ve never seen an NICU with significant numbers of extremely preterm babies that doesn’t have NEC, but if it is so, then probiotics also seem to improve feeding tolerance, and decrease systemic sepsis. The effect on systemic sepsis are not seen in all studies, so I would check which strains seem to be most effective. If you don’t have late onset sepsis either , then Bravo!