Introduction
For decades, research has shown how stress gets “under the skin” by activating neuroendocrine, cardiovascular, and metabolic systems [
1‐
3], ultimately contributing to the development of diseases and conditions such as hypertension, atherosclerosis, diabetes mellitus, nonalcoholic fatty liver disease, Alzheimer’s disease, depression, and cancer. Stress, defined as a state of threatened homeostasis [
4], is becoming more pervasive through psychosocial, environmental, and cultural means in the United States [
5], with discrimination recognized as a stressor likely contributing to ongoing health disparities [
6]. Reported experience of discrimination has been associated with cardiometabolic risk factors including elevated blood pressure and pulse [
7,
8], stress hormones [
9], inflammatory cytokines [
10,
11], obesity [
12,
13], and mental health disorders [
14‐
16], emphasizing its potential role in exacerbating disease risk and prevalence.
More recent inquiry has explored how stress and the social environment can get “into the belly” and affect the diversity, abundance, and function of the gut microbiome [
17,
18], the system microorganisms living within the digestive tract. The gut microbiome is recognized as a vital player in the proper functioning of host physiology through the production of neurotransmitters, bile acids, and short-chain fatty acids [
19,
20]. The “gut-brain axis” is defined as the bidirectional communication between the central and enteric nervous systems, linking emotional and cognitive processes of the brain with peripheral intestinal functioning [
21,
22]. In interventional research with animals and humans (maternal stress, military training, sleep deprivation, and examination stress), induced stress led to changes in stress response hormones, inflammatory cytokine production, intestinal permeability, gastric motility, and behaviors including anxiety and aggression [
22,
23]. Some of these results have also been observed in infants following microbiota dysbiosis of the mother during pregnancy [
24]. Research is emerging linking subjective measures of stress and induced stress models, such as Cohen’s perceived stress score (PSS), school examinations, circadian disruption, and social defeat models in animals, with changes in gut microbiota diversity and abundance and gastrointestinal disorders [
23]. Additionally, research observing associations between gut microbiota and cognitive functioning (emotions, memory, anxiety and depressive feelings) has involved interventions including probiotics, vitamin D, dairy products, and fiber (inulin) [
25]. The experience of discrimination in association with metrics of the gut microbiome has not been reported to date, but is paramount to health disparities research and future interventional strategies [
18,
26].
The association between life stress and gut microbiota may be explained by physiological and behavioral responses. The experience of stress can have an immediate effect on physiology by activating the stress response system, resulting in increased production of cortisol and inflammatory cytokines [
4]. Chronic, on-going stress impairs normal HPA-axis response and immune function, and has been associated with cardiometabolic and gastrointestinal disorders [
27,
28]. Limited research has examined whether chronic cortisol level, an objective measure of stress, mediates associations between subjective stressors and the gut microbiota. Second, diet is recognized as a major modifiable factor in altering microbiota diversity, abundance, and function [
29]. Habitual dietary patterns are associated with microbial clusters, mucosal protection, and anti/pro inflammatory features [
30‐
32]. The intake of micro and macronutrients has also been associated with various gut microbiota taxa, with dietary fiber often being a strong contributor to maintaining bacterial diversity [
33,
34]. Diet quality has also been found to be highly correlated with various types of stressors and disease mortality [
27,
35]. This has led some to observe the mediating and moderating role of diet quality in stress-disease processes [
36,
37], but these mediating and moderating inquiries have not yet included the gut microbiome.
This study aims to explore the gut microbiota profile of generally healthy young women in relation to diet and stress as well as extend upon the interest in uncovering gut microbiota differences by race. We also aim to explore the potential mediating role of diet and cortisol in the association between reported stress, alpha diversity, and specific stress-related gut microbiota. We propose that a diet score developed to assess the dietary inflammatory potential (dietary inflammation score, DIS) may mediate the association between the experience of discrimination, and other stressors and disease-related gut microbiota. We also recognize that individual nutrients may have specific effects on the gut microbiota; thus we are interested in exploring other potential dietary mediators including the ratio of caloric intake over estimated energy expenditure (cal:EER), healthy eating index (HEI), fiber, sugar, and other dietary variables. This research will add to the limited data related to subjective measures of stress, including the experience of discrimination, perceived stress, and depression, and various dietary variables and the gut microbiome of metabolically healthy young adult women.
Discussion
Our results of these analyses reveal interesting findings of the gut microbiota in relation to race, psychosocial stress, and dietary intake. First, ASV count and
Blautia abundance were significantly lower among AA vs EA. Racial differences of the gut microbiota have been reported in previous yet limited research [
26,
52,
53], but our findings regarding differences in
Blautia abundance are new.
Blautia was recently found to be inversely associated with visceral fat accumulation [
54] and children with obesity and diabetes. A recent literature review of
Blautia discusses it’s use as a potential beneficial probiotic as it has been found to be involved in flavonoid conversion, free radical scavenging, bacteriocin production thus inhibition of pathogenic bacteria colonization, and maintenance of environmental balance through upregulating T regulatory cells and short-chain fatty acid production [
55].
Several dietary variables were significantly different between AA and EA women. AA women had significantly higher DIS and lower HEI scores, which coincides with higher carbohydrate percentage, fried food servings, and sweet servings, and significantly lower intakes of fiber, beta and alpha carotene, vitamin E, fruit servings, and vegetable servings. Regarding stress variables, AA women also had significantly higher HC levels, and higher reports of lifetime discrimination (LED), and perceived stress (PSS).
Of the top 25 genera of our cohort,
Bacteroides and
Ruminococcus 1,
Ruminococcus 2, and
Ruminococcus torques were among the most abundant genera and
Bifidobacterium,
Lachnospiraceae, and
Lactobacillus were among the least abundant genera, which are similar gut microbiota characteristics of those following a western, animal-based dietary pattern, high-sugar/high-fat diet, and of individuals who have undergone antibiotic treatments [
56,
57].
Blautia and
Faecalibacterium, however, were the second and third most abundant genera of the cohort, genera that are reduced in individuals with cardiometabolic risk and disorders [
54] and increased with plant intake [
58], respectively. Overall, it appears the cohort has a mixed abundance of bacteria important in various nutrient metabolism and host health.
Next, we observed some significant correlations between stress, dietary variables and gut microbiota diversity and genus abundance. Stress variables were not consistently or similarly correlated with gut diversity or genus abundance. PSS was negatively correlated with ASV count and Lachnospiraceae NK4A136 group. Depression did not correlate with any metrics. RED scores were significantly negatively correlated with Blautia, and significantly positively correlated with Alistipes, Ruminococcus UCG 002, and Clostridium sensu. LED scores were significantly positively correlated with Alistipes and Ruminococcus UCG 002. Hair cortisol was significantly positively correlated with Agathobacter, Ruminococcus 1, Bifidobacterium, and Eubacterium coprostanoli. Last, salivary cortisol was significantly positively correlated with Subdoligranulum and Ruminococcus torques.
Regarding diet quality and gut microbiota diversity and abundance, trans fat intake was negatively correlated to ASV, which is similar to recent studies observing dietary fat intake and reduced alpha diversity [
59‐
61]. Adversely, high-glucose and high-fructose diets administered in mice have shown to have this effect on the alpha diversity and increases in proteobacteria phylum, one of the best sources of lipopolysaccharide (LPS), which trigger activation of the innate immune system and inflammatory conditions [
62]. Variables thought to effect gut bacteria diversity were not correlated with ASV including high fiber food groups (fruits, vegetables, fiber), cal:EER ratio (energy balance), and fried food servings. Fruit servings, alpha carotene, vitamin C, and fiber were all positively correlated with the genus
Anaerostipes, a genus that has been found to increase in abundance following the consumption of prebiotic inulin, and improve reports of constipation and stool consistency [
63], possibly through its role in producing butyrate from lactate, contributing to colonic and GI health [
64]. Alpha and beta carotene, vitamin C, and fiber were also positively correlated with
Lachnospiraceae NK4A136, however, this family of bacteria has been found to be controversial in its role in health and disease [
65].
Lachnospiraceae NK4A136 was recently found to be restored after completion of a high-fat diet protocol that induced dysbiosis in mice [
66], was found to be diminished in a small pilot study including individuals with dementia [
67], and was among other short-chain fatty acid producing bacteria that were increased following an inflammation-reducing prebiotic trial in mice [
68,
69]. More recently, a 4-week tannin supplementation trial in humans was found to increase the abundance of healthy gut bacteria, including
Lachnospiraceae NK4A136, and increase short chain fatty acid production [
70].
Variables of fat intake were inversely correlated with
Akkermansia, which is consistent with studies reporting bodyweight and high-fat diet (HFD) in children and pregnant women [
71‐
73].
Akkermansia supplementation in mice with high-fat induced obesity led to beneficial effects on weight, blood glucose control, and memory decay [
71]. In humans,
Akkermansia abundance is understood to be beneficial for cognitive health and may play a role in preventing or delaying neurological disease, such as Parkinson’s disease [
74,
75]. Physiologically,
Akkermansia administration has been shown to improve insulin sensitivity, attenuate adaptive changes related to caloric intake following cold exposure (negative energy balance), increase fat browning, induce anti-inflammatory effects through Treg cell induction in adipose tissue, and provide protective effect against atherosclerosis [
73]. Interestingly, HFD has been shown to decrease
Akkermansia abundance while fish oil consumption has been shown to increase
Akkermansia [
73,
76].
Some of our correlational findings between dietary variables and gut microbiota genera were unexpected. For example, fiber was inversely correlated with
Eubacterium coprostanoli, a genus that has been found aid in the conversion of cholesterol to coprostanol, which is important in cholesterol excretion [
77]. Next,
Lactobacillus was not correlated any dietary variables. Because strains of
Lactobacillus are often one of the main sources of bacteria in probiotics, inquiry into the associations and effects of different strains on human health is ongoing and somewhat controversial as obese individuals were found to have less
L. caseae and
L. plantarum, and greater abundance of
L. reuteri [
78]. Vegetable servings were inversely correlated with
Alistipes, another controversial genus [
74] that has been shown to increase in individuals following a calorie restricted high-fat diet that induced weight loss.
Last, the inflammatory score of one’s dietary pattern, captured by DIS, did not correlate with specific gut genera or diversity, unlike findings from a similar study by Zheng et al [
79] who found differentially abundant species between dietary inflammatory index (DII) tertiles, and research observing adherence to Mediterranean diet and beneficial changes in gut microbiota abundance [
80]. This may be due to potential significant compositional differences at both ends of the DIS spectrum (animal vs. plant-dominant patterns), and thus the most diverse microbiota may be characterized with a more neutral or balanced DIS.
Finally, variables we hypothesized to mediate the association between subjective stress and gut microbiota parameters did not hold true. In the current study, AA women reported higher perceived stress and lifetime discrimination, greater intakes of sweet and fried food servings, lower diet quality (increased DIS and reduced HEI scores), increased hair cortisol, and reduced ASV count. We hypothesized that a worse dietary intake or elevated cortisol in response to stress may be a way that AA have reduced alpha diversity and reduced
Blautia abundance. Although diet did not mediate the association between PSSUM and
ASV count, the simple regressions between PSSUM and dietary variables (fiber, DIS, HEI) were significant (Table
3), and is important to note as young adults, especially those between the ages of 18 and 25, were shown to have experienced the greatest increase in symptoms consistent with major depression, suicidal thoughts, and serious psychological distress over the last decade (2008–2017). Additionally, RED and LED were independently significantly associated with
Alistipes and
Blautia abundance. These separate associations are significant and reveal the independent associations between stress, diet quality, and specific gut bacteria abundance. The prevalence of mental illness symptoms is cause for concern as individuals are less likely to engage in health-promoting behaviors. To date, it appears that greater microbial diversity is indicative of resilience and good health [
83], and reduced diversity is associated with autoimmune disorders and cardiometabolic disease [
20]. Choosing low-nutrient, high-calorie, sugary and fatty foods as a response to stress may alter the health and diversity of a young individuals gut microbiome, potentially leading to changes in metabolic health.
Previous research has similarly explored associations between added sugar intake, the abundance of various pathogenic gut bacteria, and cognitive function. Added sugar intake was found to increase inflammatory-related bacteria such as
Proteobacteria, reduce beneficial bacteria,
Bacteroidetes [
81] and
Lachnobacterium [
82], and increase species of
Parabacteroides, which were found to impair memory performance [
83]. The alterations in these bacteria abundances can lead to lipopolysaccharide-induced inflammation and impaired gut integrity through modified tight junctions and increased intestinal permeability. Further experimental research is needed to test if induced stress and changes in sugar intake, independently, lead to alterations of beneficial bacteria that are integral in managing lipopolysaccharides. Still, subjective stress questionnaires may not be the best predictor of dietary, physiologic, or gut microbiota metrics because of the nature of the variable accounting for past and cumulative experience, not current or acute response. Additionally, RED, LED, and other discrimination questionnaires do not capture one’s resilience or emotional response to these experiences. Induced stress interventions, such as the Trier social stress test [
84], and observation of immediate food choice, stress response, and gut microbiota changes will better capture these relationships and mediations. Further research in this area is important to explore effective protocols in reducing the burden of stress.
This research is not without limitations. First, the cross-sectional, observational design of this study cannot describe causal relationships. Additionally, the sample size and metabolically healthy state of our cohort may limit the strength of our findings. Prospective studies observing the gut microbiota across the young adult period would be beneficial in learning about the resilience of one’s gut microbiome and any significant changes that may be associated with declining or stable health, chronic experiences of stress, and diet quality. Future research should continue to observe the relationships among stress, diet, and the gut microbiome in early adolescence and young adulthood and conduct interventions involving mental health treatment and lifestyle modification, with hopes to halt or reverse concerning trends in obesity, prehypertension and hypertension, and pre-diabetes and diabetes rates of this population [
14,
85‐
87].