The discovery of bacterial DNA in the fetal environment has put an end to the long-term belief that it is sterile. However, a question remains: does the identified DNA come from viable and metabolically active bacteria, originating from the mother? An American team provided an answer by combining studies in humans and mice. First step: Characterizing bacterial populations of mother-child pairs (5 premature babies and 5 born at full term) based on samples extracted post C-section delivery, under optimal sterility conditions. The analysis has made it possible to specify the origin of bacteria found in the child’s mouth and meconium, based on mother’s vaginal, rectal, uterine, placental and amniotic microbiotas. As a result, the existence of a fetal microbiota as soon as 24 weeks of gestation was confirmed. It comes from the uterine environment and is mainly composed of Escherichia and Acinetobacter.

Living bacteria in the fetus...around mid-pregnancy

Second step: in mice, researchers visualized the fetus’ gut flora and observed its viability as well as the frequent changes it undergoes during pregnancy. Their data suggest that in the middle of the gestation period, the fetus is exposed to viable and culturable bacteria of variable maternal origin. On the contrary, at the end of the gestation period, and despite the presence of bacterial DNA mainly of placental and amniotic origin, the samples turned out to be non-culturable. The suggested hypothesis is that the (late) maturation of the immune system leads to the progressive elimination of microorganisms that crossed into the fetal environment.

A viable transmission was confirmed

The team validated its observations on the bacterial transmission during gestation by colonizing the gut of pregnant mice with labeled E. coli, before recovering these viable bacteria in their offspring. All these results make the case for the existence of a fetal microbiota of maternal origin, that changes during pregnancy and is likely to have an impact on the development of the immune system and the constitution of the newborn microbiotas after birth.

The vaginal microbiota is a changing and well characterized microbial ecosystem. It is divided into five large groups based on composition: four are dominated by a single Lactobacillus species (L. crispatus, L. gasseri, L. iners or L. jensenii) and the fifth is heterogeneous with a larger number of anaerobic strains, such as Gardnerella vaginalis, Atopobium vaginae and Prevotella spp. The latter is a sign of bacterial vaginosis and is associated to an increased risk of infertility and sexually transmitted infections.

Considering the impact of the vaginal microbiota of women's intimate health and its consequences on reproduction, an American team focused on the link between bacterial profile and colonization by the Candida yeast. Candida is a commensal member of the vaginal microbiota, but it is responsible for vulvovaginal candidiasis characterized by an aggressive response of the host to the excessive proliferation of the opportunistic fungus. Vaginal swabs were taken from 255 women aged between 14 and 45 years, of Caucasian (53%) or African (47%) descent, and were used to identify dominant lactobacilli and to assess and quantify colonization by Candida.

Ethnic variations of the microbiota...

Test results: 16% of women had candidiasis (90% of C. albicans and about 10% of C. glabrata), with a higher percentage in microbiotas with a predominance of L. iners versus L. crispatus (respectively 39% and 20%). This gap is reflected at ethnic level, since the group with a predominance of L. iners is more frequently associated to women of African descent than to women of Caucasian origin (46.7% vs. 31.9%). This study findings confirm data from the literature.

...and lactic acid regulation

The researchers believe that the correlation between Lactobacillus species and the risk of developing candidiasis is based on the lower or higher ability of each strain to acidify the vaginal environment. In vitro tests have shown that L. crispatus produces larger quantities of lactic acid, leading to a decrease in the local pH (near 4.0) versus a pH of 4.6 with L. iners, which is enough to inhibit C. albicans growth.

Differentiating vaginal bacterial communities could thus allow us to identify predisposition to candidiasis. This is a first step towards tailored preventive and curative strategies based on microbiota modulation.

Bacterial vaginosis is a frequent, difficult-to-treat female disorder caused by an imbalance in the vaginal microbiota and characterized by a decrease in lactobacilli and an increase in potentially harmful bacteria, especially Gardnerella vaginalis. Despite antibiotic treatment, around 60% of affected women will relapse within the following year.

Change of method

Since a link between blood levels of some nutrients and the risk of bacterial vaginosis is suspected, many studies have been conducted but their results have been inconsistent. The authors of a new study published in Reproductive Health believe it is due to the methods used in these studies which are based on vaginal swabs or clinical endpoints and mainly focused on vitamin D. To examine the association between the use of dietary supplements and bacterial vaginosis, they analyzed the composition of the vaginal microbiota of 104 young women, including 25% with bacterial vaginosis. They also analyzed their daily intake on micro- and macronutrients based on their answers to a benchmark questionnaire. The researchers also reviewed the scientific literature on this topic.

Higher betaine intake?

They managed to profile women with vaginosis: such women use more frequently vaginal douches, have a high body mass index, and less often use a hormonal method of contraception, compared to women with a balanced vaginal microbiota. Overall, those with lower nutrient intakes have a lower risk of vaginosis. However, betaine is the one exception since a limited intake increases the risk. In vitro, this substance seems to stimulate the survival of lactobacilli and the production of lactic acid, and to prevent colonization by pathogens. According to the authors, it acts directly on the vaginal microbiota by promoting bacterial balance or indirectly through the gut microbiota. This discovery opens up new perspectives to limit the risk of vaginosis, such as increasing betaine intake by changing diet or using dietary supplements.

The study of gut microbiota evolution led many researchers to compare humans and primates. Their works, conducted on a limited number of species, indicated that bacteria found in our gut microbiota descend from bacteria that colonized the gastrointestinal tract of our common ancestors and that they co-evolved. However, although humans are genetically close to large primates (bonobos, chimpanzees), their digestive system is closer to that of Old World monkeys (baboons, macaques) whose environment and dietary habits are quite similar to humans’.

A microbiota closer to that of Old World monkeys

To support the hypothesis stating that these two parameters have a much larger impact on the gut microbiota composition than commonly believed, an American team compared the bacterial flora and its different functions between human populations living in industrialized or non-industrialized countries and 18 wild primate species. While the gut microbiota of populations living in industrialized countries was very different from that of other primates, the gut microbiota of populations living in non-industrialized countries was, on the contrary, very similar to that of primates. There are, however, differences: when compared to the microbiota of other primates, that of humans has unique microbial characteristics (it contains some species, but others are lacking), specific functional pathways and a greater inter-individual variability. The latter could reflect the greater adaptability of humans to new environments. More surprisingly, the gut flora of humans had more similarities with that of baboons than with that of their monkey ancestors.

The influence of environment is underestimated

These results emphasize the influence of environment, diet and physiological adaptations on the gut microbiota composition, especially on their functional capabilities, and belie the idea that it is almost exclusively the result of the co-evolution of bacteria and their host. The authors believe that these discoveries provide a new perspective on the role of the gut microbiota in the evolution of mankind.

Several studies have documented the pivotal role of the gut microbiota in immunity, although clinical evidence remains limited. Better understanding the underlying mechanisms is crucial in terms of public health, especially to develop therapies targeting the microbiota in cases of immune disorders. In this context and based on the observation that vaccine efficacy varies as much as the gut microbiota of individuals on the different parts of the world, researchers focused on the impact of antibiotics on the immune response to vaccination.

Effect of antibiotics on vaccination

To this end, the scientists studied two populations with different levels of anti-flu antibodies at baseline. In the first population, 22 healthy adults were vaccinated against seasonal influenza (vaccine containing 3 strains of attenuated viruses); and 3 days before vaccination, half of them started an antibiotic treatment lasting 5 days. As expected, antibiotics deeply and durably impacted the composition of the gut microbiota. But contrary to the researchers’ hypothesis, the production of antibodies in response to vaccination was not affected by the use of antibiotics. An effect was only observed in the second cohort composed by 11 subjects with no significant pre-existing immunity against influenza (no recent flu or vaccination (sidenote: As proven by low levels of antibodies (1st cohort: serum levels < 320 for at least 2 out of the 3 strains included in the vaccine; 2nd cohort: serum levels ≤ 320 for the 3 strains contained in the vaccine) ) ): antibiotics produced a decrease in the production of IgG1 and IgA antibodies against the H1N1 virus strain. Broad-spectrum antibiotics thus could, in some cases, disrupt the immune response to vaccination.

Bile acids: inflammation messengers

Researchers also observed that changes induced by vaccination at the blood metabolites level were different between the group who received antibiotics and the control group, especially for substances derived from the metabolism of bile acids. It should be reminded that the microbiota transforms primary bile acids secreted by the gallbladder and supplied to the intestines, into secondary bile acids. They are then partly reabsorbed. But in subjects who received an antibiotic therapy, an increase in blood levels of primary bile acids and a major decrease in secondary bile acids were observed. Antibiotic therapy could even reduce by a factor of 1,000 the most important of them, i.e. lithocholic acid. This decrease in secondary bile acids was strongly correlated to the increase of pro-inflammatory molecules. This suggests that a change in the ratio of bile acids induced by antibiotic therapy could be one of the mechanisms regulating the inflammatory response.

Two different response pathways

Is there a link between the pathway modulating the production of antibodies and that involving bile acids and inflammation? According to the researchers there is no such link: they mapped all the different signaling pathways, from the microbiota disruption following the administration of antibiotics, to the changes in blood metabolites; and they concluded that the two pathways are independent. In conclusion, the profound disruption of the microbiota induced by the use of antibiotics could modulate the immune function through two distinct mechanisms: by directly interacting with immune cells, or in a systemic way, by modulating the production of some key metabolites.

Fecal microbiota transplant: a miracle cure?

The frenzy surrounding fecal microbiota transplant (FMT) is real and it needs to be slowed down a bit: some patients have unrealistic expectations regarding the benefits of FMT in their particular case. Every week I receive dozens of letters about anything and everything. However, FMT is a not a magical cure! For now, it is indicated to treat a single disease: recurrent C. difficile infection. For all other indications, it is just a potential therapeutic avenue which cannot replace current treatments. Moreover, the future most likely lies in treatments combining stool transplant (or other therapies that target the microbiota) and more standard treatments that target the immune system, for instance.

Why does C. difficile respond that well to fecal microbiota transplant?

This infection is almost exclusively related to a disruption in the gut microbiota, while in other diseases, the role of the gut microbiota–although presumed– is only one of several contributing factors and its importance probably varies significantly from one disease to another. For instance, in the case of ulcerative colitis, for which we have the strongest data: clinical trials indicate a 20 to 30% rate of remission within a 8-12 week period; which is good, but very far from the results obtained in C. difficile infections (near 90%). This clearly shows that other factors (immune, genetic...) also play a role.

Are there obstacles to the development of clinical research focused on FMT?

Research on FMT is still at a very early stage since it started less than 10 years ago; which is why we must take the time needed to assess it properly. In France, the handling of feces is subjected to major regulations and the selection of donors is strictly regulated. As a result, clinical trials are expensive and require complex logistics. In addition, since hospitals do not automatically assign a budget to FMT, the mobilization of health professionals varies from one facility to another, thus depriving researchers from a specific structure to rely on. It is time for public authorities to better understand this issue and to invest in order to provide hospitals with the means to develop this line of research. In Assistance Publique-Hôpitaux de Paris (Paris public hospital system), we hope to quickly see the emergence of a structured approach to FMT in the healthcare system.

In animals, the physical trait “obese” or “thin” can be transferred through fecal microbiota transplant with a clearly established causal link. In humans, the situation is a little more complex but the presence of dysbiosis associated to metabolic disorders in obese or hypertensive subjects led scientists to conclude that FMT could be a promising avenue. Works are ongoing to assess the impact of the change in gut microbiota experienced by patients with metabolic syndrome^17,14.

Mixed benefits

Several clinical studies were conducted on obese patients with a metabolic syndrome. The first one was carried out on a small group of individuals and showed that stool transplant from thin donors improved the metabolic profile of recipients. The second included a larger number of patients and produced more mixed results. Only a few participants had an improved metabolic profile after the FMT, namely those who initially had a gut microbiota that was not very diversified. The response to transplant thus seems to be dependent on the patient’s initial gut microbiota. However, the benefits did not withstand the test of time...nor did the transformation of the gut microbiota, which quickly returned to its initial composition.

Complex relationships

Overall, these results highlight the complexity of the link between the gut microbiota and metabolic functions. According to some scientists¹³, metabolic and microbial responses to FMT could be based on interactions between the donor’s microbiota and the recipient’s. Several trials are ongoing to assess this technique’s ability to reduce metabolic disorders as well as several obesity-related parameters. These eagerly awaited results should open the way to new strategic approaches in the treatment of metabolic syndrome.

The presence of a gut dysbiosis–characterized by reduced levels of beneficial SCFAs-producing bacteria and increased levels of harmful bacteria associated to cognitive disorders–was observed in patients with hepatic encephalopathy. These observations led researchers to consider fecal microbiota transplant as a therapeutic alternative to antibiotics¹⁶.

A unique donor

An initial study focused on this indication was conducted on a small cohort of only twenty cirrhotic men, who were administered either the standard treatment, or FMT combined with an antibiotic pretreatment aiming at preparing the recipient’s digestive tube. The fecal microbiota came from one single donor, selected through an AI software based on the abundance in his/her microbial flora of bacteria that are precisely lacking in these patients.

A promising approach

None of the transplant recipients had an additional episode of encephalopathy, while 5 out of the 10 control patients relapsed. A slight increase in lactobacilli and bifidobacteria was also observed in the former, while no change was observed in the latter. Finally, only FMT produced an improvement in cognitive functions which led researchers to advocate for further investigation.

The microbiota of patients with irritable bowel syndrome is less diversified and shows greater abundance of enterobacteria, but with a lower content in bifidobacteria and lactobacilli. Functional gastrointestinal disorders are characterized by a decreased production of butyrate and an increased production of acetic and propionic acids, which are three substances associated with bloating. Abdominal pain, diarrhea and constipation are other symptoms of these disorders. In the US, an estimated 20% of the population is affected¹⁴.

Controversial effects

Aside from ulcerative colitis, irritable bowel syndrome is the only other gastrointestinal condition for which a treatment with fecal microbiota transplant has been studied in clinical trials¹⁴. One of them showed a decrease in intestinal discomfort, abdominal pain and flatulence in transplant recipients. However, the results varied depending on the initial nature of the gut microbiota: the best responders to fecal microbiota transplant had an initial high content of Streptococci and they had a greater increase in biodiversity. Other works confirm the increased diversity and abundance of the microbiota after the administration of fecal matter capsules; however, patients reported a better improvement in symptoms when they received... placebo! Although these results do not undermine the efficacy of FMT in people with functional gastrointestinal disorders, detailed microbiota analyses before and after the transplant are necessary, according to the researchers.

What about constipation?

Constipation is potentially associated to gut dysbiosis and has been the focus of several works aiming at assessing the usefulness of FMT in this transit disorder¹³. In a study conducted in about sixty adults suffering from slow transit that compared the standard treatment to 6 courses of FMT, the latter was shown to cause a significant improvement of symptoms and transit, and more generally, in the quality of life¹⁵. These encouraging results still need to be confirmed: studies are also ongoing with specific strains, lactobacilli and bifidobacteria¹³.

CIBDs progress as inflammatory flareups of variable duration and frequency depending on the patient, alternating with periods of remission. They cause acute abdominal pain, severe diarrhea (between 5 and 10 bowel movements per day), associated with blood and pus in ulcerative colitis (UC); and in severe forms, other symptoms and complications can occur such as fever, tachycardia, nausea and vomiting, weight loss and dehydration. CIBDs also trigger extra-intestinal symptoms, especially articular pain, skin and mucosa lesions (skin ulcerations, mouth ulcers, glossitis, i.e. tongue inflammation...) as well as hepatic and ocular disorders¹³.

Unbalanced microbiota

The gut microbiota analysis of patients with UC revealed they had a decreased diversity of microbial species¹⁴, especially a lower content of Firmicutes and Bacteroidetes. But it is mainly the low content of Faecalibacterium prausnitzii and the excessive content of Proteobacteria and Actinobacteria that are associated with these CIBDs. This imbalance causes a decrease in the production of short-chain fatty acids, beneficial substances which are food to colon cells and play an important role in the regulation of the immune system. That is why fecal microbiota transplant (FMT) has been considered to treat this disorder.

Moderate benefits in UC

Three of the four published clinical trials on UC concluded that this approach was beneficial. Overall, the beneficial effects were much more moderate than they were in the treatment of C. difficile colitis and depend on the donor–which is why donor selection is so important. And several results raise new questions: are only some microorganisms efficient? And if so, which ones? Should the patient receive an enema or an antibiotic therapy beforehand? Which administration route is preferable? Does the restoration of the gut microbiota work long-term or must transplants need to be repeated? These questions need to be answered before fecal microbiota transplant can be seriously considered as an alternative for the treatment of ulcerative colitis.