Should the Faecal Microbiota Composition Be Determined to Certify a Faecal Donor?

Faecal microbiota transplantation (FMT) is considered a safe and effective therapy for recurrent Clostridioides difficile infection. It is the only current clinical indication for this technique, although numerous clinical research studies and trials propose its potential usefulness for treating other pathologies. Donor selection is a very rigorous process, based on a personal lifestyle interview and the absence of known pathogens in faeces and serum, leading to only a few volunteers finally achieving the corresponding certification. However, despite the high amount of data generated from the ongoing research studies relating microbiota and health, there is not yet a consensus defining what is a “healthy” microbiota. To date, knowledge of the composition of the microbiota is not a requirement to be a faecal donor. The aim of this work was to evaluate whether the analysis of the composition of the microbiota by massive sequencing of 16S rDNA could be useful in the selection of the faecal donors.

The results suggest that donor certification does not need to rely exclusively on their microbiota composition, as it is unique to each donor. While one donor showed greater microbial diversity and richness, clear criteria for microbial normality and health have yet to be established. Therefore, donor certification should focus more on clinical and lifestyle aspects. FMT is a safe and effective treatment for rCDI, the aim of FMT is to restore gut microbiota alterations, which is in contrast to antibiotic treatment that acts as a disruptor. However, the concrete stool component that restores microbiome competence has not been identified.
Finding adequate faecal donors is quite difficult; in fact, different faecal banks have reported that only 2–5% of the candidates complete the selection process.⁽¹⁾

A systematic review in 2013 ⁽²⁾ evaluated the efficacy of FMT in 273 patients diagnosed with rCDI across 11 different studies, reporting a remarkable 90% success rate. Most importantly, it found no significant difference in treatment efficacy between related and anonymous donors.
In our case, the donors studied have been used in real rCDI treatment situations, with no differences observed between them. These results suggest that donor selection should prioritize other criteria rather than their relationship to the recipient.

To date, it is still unknown if using multiple donors could be more effective than the use of just one for each patient, and this is certainly a key point to be addressed by the manufacturers. In the present study, 16S rDNA massive sequencing was performed on consecutive faecal samples from certified donors at Mikrobiomik Company to evaluate whether knowledge of microbiota composition is necessary to improve donor selection criteria. Donor selection based on a healthy microbiota has always been the main goal of FMT; however, it has been limited by the inability to define what constitutes a healthy microbiota. Nevertheless, it is generally accepted that a stable and diverse microbiota, characterized by a high number of different species (alpha diversity) and richness in the abundance of taxa (beta diversity), is correlated with a healthy gut state.

The results obtained in this study showed that most of the faecal donors had similar alpha diversity values measured by the Chao 1 and Shannon indices. However, donor 23 had a low Shannon index, suggesting that the microbiota of donor 23 may lack some of the variety found in the other samples. In contrast, donors 17 and 19 had a richer and more diverse gut microbiota than the others. Furthermore, the comparation of microbiota diversity across donors (beta diversity) using the Bray–Curtis index indicated that each donor has a distinct microbial ecosystem but with some similarities, except donors 23 and 29 whose microbial composition differed from the other donors. Additionally, some variability was observed among the stool samples collected from the same donor, indicating fluctuations in their microbiota composition over time. These fluctuations can be consequences of different factors, such as diet, environment and overall health.⁽³⁾

In our study, we examined the taxonomy of the gut microbiota of the donors using 16S rDNA sequencing, paying special attention to the phyla and genera composition. We found that the phylum Bacillota (64.5%) was the most abundant in all donors, followed by Bacteroidota (16.7%), Actinomycota (6.7%), Verrucomycota (6.7%) and Pseudomonadota (2.9%), with some inter-donor and intra-donor variations. We also observed a notable variability in the phyla distribution among samples from the same donor taken at different time points.
In conclusion, the core microbiome, which includes the genus Agathobacter, Eubacterium, Bacteroides, Clostridia UCG-014 and Akkermansia, likely reflects a microbial community that contributes to intestinal health in our 10 donors, with uncertainty as to whether these results can be extrapolated to other donors.

To complete this investigation, we represented the genera composition of the ten certified donors in a heatmap to illustrate and compare the characteristic microbiota patterns of each donor. These results further emphasize that each donor has a unique gut composition with intra-individual variations.
The concept that a healthy microbiome can be defined by specific populations of certain bacteria is too limited ⁽⁴⁾. It is essential to include metabolomic studies to determine whether these differences in microbial composition are translated into variations in ecosystem functionality.

Our conclusion is that the extensive knowledge of the microbiota composition is not essential for stool donor certification, since, like all healthy individuals, each one has its own pattern, without interfering with the result of the FMT. Since different microbiota patterns can have the same metabolic functionality, studying molecules such as short-chain fatty acids could be useful in the future; however, this aspect has yet to be explored in healthy donors. Currently, the inability to incorporate microbial and metabolic analyses of donors still limits their certification to lifestyle factors and the absence of known pathogens and diseases.

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