Introduction
Ingested food components, preferentially carbohydrates that are not digested and absorbed in the small intestine, reach the colon where they serve as fuel for the inherent microbiota. Hence, the diet is of utmost importance in shaping the composition of the gut microbiota. In turn, the gut microbiota affects the host’s energy metabolism, glucose homeostasis and inflammatory status to a large extent [
1‐
3]. Recent work by Klinder et al. using fluorescence in situ hybridization (FISH) showed that increased intake of fruits and vegetables altered the fecal microbiota in humans, with, e.g., increased levels of
Ruminococcus bromii and increased
Bacteroides/
Prevotella ratio, as well as indicated associations between cardiovascular risk markers and specific bacteria [
4]. Other dietary components, such as walnuts and pomegranate, can also be metabolized by the gut microbiota. These metabolites, urolithins, can be detected in blood and used as markers for cardiometabolic risk in humans [
5]. Other biomarkers more commonly used are c-reactive protein (CRP), cytokines and blood lipids. However, low-grade inflammation observed in individuals with the metabolic syndrome is a complex physiological state that may require analysis of a cluster of biomarkers, as reviewed by Minihane et al. [
6]. New markers for cardiometabolic risk are essential to investigate further, as they are key in developing prevention strategies against cardiovascular disease. A multifunctional diet (MFD) targeting subclinical inflammation was developed as a tool to decrease risk factors for cardiometabolic disease in at-risk individuals, i.e., in mature individuals (> 51 years of age) with overweight or obesity. The diet includes foods and/or meals with anti-inflammatory potential in that they promote low acute glycemic responses, are rich in polyphenols and/or specific dietary fiber with prebiotic action, are rich in omega 3 fatty acids such as oily fish and rapeseed oil, or with anti-oxidative and anti-hypercholesterolaemic effects, e.g., soybeans and almonds [
7].
MFD contains several components that are degraded in the colon by the microbiota, such as dietary fibers from rye, barley, oats and berries. In previous studies, we have observed improved cardiometabolic markers in healthy at-risk individuals after 4–8 week intake of MFD targeting low-grade inflammation [
7,
8]. The diet exerted also extensive impact on the plasma metabolome, particularly the lipidome [
9]. However, whether these improvements can be associated with changes in the gut microbiota composition has not been investigated. In the present study, we analyzed the gut microbiota before and after the 8w dietary intervention with MFD in subjects from [
8] using next-generation sequencing of bacterial 16S rRNA genes. At-risk individuals were given either the MFD or a control diet lacking the functional (“active”) components for 8 weeks in a parallel, randomized design. Fecal samples were collected at baseline and at the end of each study arm for microbiota analysis. We could identify bacterial taxa that were associated with the MFD and, correspondingly, with improved cardiometabolic risk markers.
Discussion
The MFD was effective in improving cardiometabolic risk markers and in the present study, we show that it induced minor shifts in the gut microbiota at species level, with increased abundance of
P. copri. Prevotella has recently been found to be one of the three genera driving enterotypes of the human gut microbiota;
Bacteroides (Enterotype 1),
Prevotella (Enterotype 2)
and Ruminococcus (Enterotype 3) [
20]. A long-term diet enriched in carbohydrates has been linked to the
Prevotella enterotype, while protein and animal fat has been linked to the
Bacteroides enterotype [
21].
Prevotella/
Bacteroides ratio and, particularly,
P. copri has recently been found to be associated with improved glucose tolerance after intake of barley kernels [
22]. In the present study, the MFD resulted in improved fasting glycaemia and insulinaemia, although the effect was not significantly different from that exerted by CD [
8].
We found that a number of bacterial genera were associated with the investigated biomarkers of cardiometabolic risk. Of note,
Treponema correlated positively with blood pressure. This bacterium has been implicated in periodontal disease, a known risk factor for atherosclerosis, and its abundance in the oral cavity has, interestingly, also recently been associated with obesity in humans [
23]. Our study shows that the gut abundance of
Treponema may also be linked to cardiometabolic risk factors such as increased blood pressure. In contrast,
Faecalibacterium showed a negative association with blood pressure, which may be a reflection of its proven anti-inflammatory capacities [
24]. In addition, patients with the metabolic syndrome (MetS) show a reduction in
Faecalibacterium prausnitzii compared to healthy individuals, which was restored upon a dietary intervention with a Mediterranean-type diet [
25]. Three bacterial genera were also found to correlate with blood lipid levels. Certain bacterial species are known to possess bile-salt hydrolase activity, such as some lactobacilli and bifidobacteria species, which may interfere with bile salt activity in the gut and, consequently, bile salt reabsorption and cholesterol synthesis in the liver [
26]. In our study,
Ruminococcus and
CF231 were associated with increased HDL levels, while
Bilophila appeared to be associated with less favorable blood lipid profiles.
Bilophila wadsworthia has been implicated in colitis in mice and it increases after high intake of saturated milk-fat through alterations in bile acid profiles [
27]. Although the MFD did not significantly alter the gut abundance of these bacteria, our correlation analyses of bacterial taxa with cardiovascular risk markers identified members of the gut microbiota that can be targeted in future dietary interventions to improve cardiometabolic risk markers.
The diet is regarded as a major determinant of gut community structure, but in our study the effects of the MFD induced relatively small changes in the gut microbiota composition compared to the control diet, with significant changes only at the species level. Dietary fiber is the main food component reaching the colon undigested, thus acting as substrate for the microbiota. Although the dietary fiber content was higher in the MFD than in the CD (62 versus 24 g/day, respectively), the dietary fiber content in CD almost reaches the level of dietary fiber intake recommendation of 25–35 g/day in adults [
10]. Given that 24 g/day of dietary fiber in CD should be enough to provide substrate for the gut microbiota, appreciable fermentation should take place. The 32% reduction in breath hydrogen, i.e., one of the products from bacterial fermentation, in CD individuals at the endpoint may suggest that some or even most individuals may have included high amounts of dietary fiber in their diets already before the intervention. Further, there was a large variation in the individuals’ microbiota already at the baseline of the study, probably due to variations in their daily dietary fiber sources and intake levels. The period of 8-week intervention may not be enough to induce changes in the gut microbiota towards one direction when the niches are different from the beginning. The large variation of the gut microbiota in these individuals remained at the endpoint of the study, thus possibly reducing the chance of detecting differences due to the diet. Consideration of the daily dietary fiber intake in the inclusion criteria when recruiting subjects for such dietary intervention studies may help to reduce the big gut microbiota variation at baseline. Also, a larger sample size could have helped to clarify further whether the MFD affected also other bacteria than those reported in the present study. In addition, the control diet was a “healthy” diet, formulated according to the Nordic Nutrition Recommendations, while a more extreme, unhealthy control diet may have resulted in larger differences between groups.
In conclusion, the MFD did not alter the gut microbiota at phylum or genus level. At species level, P. copri was identified by the biomarker discovery tool LEfSE as discriminant for the MFD and its role in improvement of risk markers should be investigated further. Furthermore, gut abundance of several bacteria correlated with blood pressure, including Faecalibacterium that correlated negatively with blood pressure and Treponema that correlated positively, while Bilophila appeared to associate with an unfavorable blood lipid profile. Thus, the results can be used to further optimize health effects of the MFD, by addressing bacteria associating with cardiometabolic risk markers. Taken together, results from the present study may be used in the further development of effective dietary concepts capable of reducing cardiometabolic risk markers in humans through a targeted modulation of the gut microbial community.