The Probiotic Mood Study: Daily Self-Reports Reveal Gut Bacteria Reduce Negative Feelings After Two Weeks

Based on the Randomized Controlled Trial

Probiotics Reduce Negative Mood Over Time: The Value of Daily Self-Reports in Detecting Effects

Katerina V.-A. Johnson and Laura Steenbergen. npj Mental Health Research. 2025.

Reason Behind the Study

By 2025, the microbiome-gut-brain axis had become one of the most intensely studied areas in mental health research. Animal studies had consistently shown that gut bacteria influence brain development, neurochemistry, and behaviour, with probiotic administration reducing anxiety and depressive-like behaviours in rodents. However, translation to humans had produced frustratingly inconsistent results. Some studies found that probiotics improved symptoms of depression, anxiety, and stress in healthy volunteers, while others reported little or no effect. Meta-analyses suggested overall benefits, but the evidence was considerably stronger for clinically depressed patients than for the general population.

Katerina Johnson from Leiden University and the University of Oxford, together with Laura Steenbergen, recognised a critical methodological flaw underlying this inconsistency. Nearly all previous human studies relied on standard psychological questionnaires administered before and after the intervention. These pre- versus post-intervention assessments might lack the sensitivity to detect subtle shifts in emotional state in healthy individuals who are not experiencing clinically significant symptoms. The researchers hypothesised that a more fine-grained approach, daily self-reports tracking mood in real time over the intervention period, might reveal changes that traditional questionnaires missed. This study, published in npj Mental Health Research in April 2025, directly tested whether daily mood monitoring could detect probiotic-induced emotional changes in healthy young adults.

Goals

The study had a clear primary objective: to investigate the effects of a four-week multispecies probiotic intervention on emotion regulation and mood in healthy volunteers using a comprehensive combination of methods. The key innovation was the inclusion of daily mood self-reports alongside standard pre- and post-intervention questionnaires and computerised tests of emotional processing. A secondary objective was to determine whether probiotics influence how individuals process emotional information, measured through attentional bias and facial expression recognition tasks. A tertiary objective was to identify individual traits, such as baseline risk aversion, that might predict who responds best to probiotic supplementation, enabling future targeting of at-risk individuals.

Key Eye Opening Findings

The study produced three findings that significantly advance the understanding of probiotics and mental health. First, daily self-reports revealed a clear and statistically significant reduction in negative mood in the probiotic group compared to placebo, with the effect emerging after approximately two weeks of daily supplementation. This effect was not detected by any of the standard psychological questionnaires administered before and after the intervention. Second, the probiotic intervention did not affect positive mood ratings, suggesting that the effect is specific to reducing negative feelings such as anxiety, sadness, stress, and fatigue, rather than enhancing positive emotions. Third, exploratory analysis revealed that certain individual traits, particularly a tendency towards risk aversion, were associated with a greater mood-improving response to probiotics, suggesting that some people may benefit more than others.

The discrepancy between the daily reports and the questionnaire findings is the most striking result. Standard questionnaires including the State-Trait Anxiety Inventory (STAI), Penn State Worry Questionnaire (PSWQ), Perceived Stress Scale (PSS), Centre for Epidemiological Studies Depression Scale (CES-D), and Positive and Negative Affect Schedule (PANAS) showed no significant differences between the probiotic and placebo groups after correction for multiple comparisons. Yet daily monitoring captured a clear signal. This finding reconciles the inconsistencies of previous studies and suggests that the field has been using insufficiently sensitive measurement tools.

1. Study in Detail

Design and Participants

The study was a randomised, double-blind, placebo-controlled trial conducted at Leiden University. A total of 88 healthy adult participants were recruited, with 44 allocated to the probiotic group and 44 to the placebo group. The mean age was 22.3 years in both groups, with a standard deviation of 3.1 years. The sample comprised 29 women and 15 men in the probiotic group, and 30 women and 14 men in the placebo group. The mean body mass index was 23.4 in the probiotic group and 23.6 in the placebo group. All participants met strict eligibility criteria: no prior psychiatric diagnoses, no antibiotic or probiotic use in the past three months, BMI between 18 and 30, no significant dietary changes, and no medication use other than hormonal contraceptives. The sample was highly specific: young, healthy, and predominantly female, which limits generalisability to older adults, clinical populations, or males.

Methodology

Each participant underwent a comprehensive pre-intervention assessment, followed by a 28-day intervention period, and then a post-intervention assessment identical to the first.

Probiotic Intervention

Participants in the probiotic group received daily sachets containing 2 grams of freeze-dried powder of the multispecies probiotic mixture Ecologic Barrier (Winclove Probiotics B.V.). Each sachet contained 2.5 × 10^9 colony forming units (CFU) per gram of nine bacterial strains: Bifidobacterium bifidum W23, B. lactis W51 and W52, Lactobacillus acidophilus W37, Levilactobacillus brevis W63 (formerly classified as Lactobacillus brevis), Lacticaseibacillus casei W56 (formerly Lactobacillus casei), Ligilactobacillus salivarius W24 (formerly Lactobacillus salivarius), and Lactococcus lactis W19 and W58. The placebo group received sachets containing the carrier powder of maize starch and maltodextrins, indistinguishable in colour, taste, and smell. Participants dissolved the powder in lukewarm water daily for four weeks. Compliance was assessed by counting returned sachets.

Daily Mood Monitoring

The critical innovation was daily electronic self-reports. Each day, participants received a reminder and rated their mood on a sliding scale from 0 to 100 for two statements: "How much positive feeling do you have today?" and "How much negative feeling do you have today?". This daily tracking provided temporal resolution that pre- and post-intervention questionnaires could not offer. Participants also reported stool consistency using the Bristol Stool Scale to monitor any gastrointestinal effects.

Questionnaire Measures

Both before and after the four-week intervention, participants completed a comprehensive battery of standardised psychological questionnaires: the State-Trait Anxiety Inventory (STAI), Penn State Worry Questionnaire (PSWQ), Perceived Stress Scale (PSS), Leiden Index of Depression Sensitivity Revised (LEIDS-R), Centre for Epidemiological Studies Depression Scale (CES-D), Positive and Negative Affect Schedule (PANAS), Emotion Reactivity Scale (ERS), Multidimensional Assessment of Interoceptive Awareness (MAIA), Bermond-Vorst Alexithymia Questionnaire (BVAQ), and Buss-Perry Aggression Questionnaire (BPAQ). These represented the standard tools used in the field.

Tests of Emotional Processing

Participants completed two computerised tasks to assess how they processed emotional information. The emotional dot-probe task measured attentional bias towards or away from emotional facial expressions. The facial expression recognition task measured accuracy in identifying emotions from facial stimuli. These tasks were included because they have been shown to detect early effects of pharmaceutical drugs on emotional processing in depression and are validated in healthy individuals.

Statistical Analysis

For questionnaire scores and dot-probe reaction times, independent two-sample t-tests compared the change following intervention between groups. For the facial expression recognition task, a linear mixed-effects model was conducted with emotion, intensity, group, session, and the group-session interaction as independent variables. For daily mood measures, linear mixed-effects models were constructed with group, time, and their interaction as independent variables. Exploratory analysis used Kendall Tau-b correlations to determine whether any pre-intervention questionnaire scores predicted which individuals responded best to the probiotic treatment.

1. Key Findings

Daily Self-Reports Detect a Clear Reduction in Negative Mood

The primary finding was unambiguous when daily self-reports were analysed. The probiotic group showed a statistically significant reduction in daily self-reported negative mood compared to the placebo group. The effect began to emerge after approximately two weeks of supplementation and persisted across the four-week intervention period. The time series graphs showed a clear divergence between the two groups after the 14-day mark.

This finding is remarkable not only because it demonstrates a probiotic effect on mood, but because the effect was undetectable by any other measure. The researchers concluded that daily self-reports provided the necessary sensitivity to detect probiotic-induced changes in healthy subjects' emotional state, whereas standard questionnaires administered at only two time points were insufficient.

Standard Questionnaires Show No Significant Effects

In contrast to the daily reports, the questionnaire data showed almost no differences between the probiotic and placebo groups. The most notable difference was on the Penn State Worry Questionnaire, where both groups decreased in worry scores following the intervention, but only the placebo group demonstrated a significant reduction (P = 0.018). The probiotic group scored lower on the not-distracting subscale of the MAIA, indicating a greater tendency to ignore or distract themselves from sensations of pain or discomfort following the intervention compared to the placebo group (P = 0.017). However, given the number of questionnaires administered, these differences would not survive correction for multiple comparisons.

The PANAS, which includes both positive and negative affect subscales, showed no significant group differences. This aligns with the finding that the probiotic effect was specific to reducing negative feelings rather than enhancing positive ones.

No Effect on Positive Mood

The daily reports also revealed that the probiotic intervention did not significantly affect positive mood ratings. Participants did not report feeling happier or more positive; they simply reported less negative feelings such as anxiety, sadness, stress, and fatigue. This specificity is notable because some antidepressants can blunt both negative and positive emotions. Probiotics appear to selectively reduce the negative component of emotional experience.

Emotional Processing: Mixed Findings

The emotional dot-probe task showed no evidence of a difference between the probiotic and placebo groups in terms of any change in attentional bias towards the five emotions tested (sadness, fear, anger, happiness, surprise, and neutral). However, the facial expression recognition task revealed a marginally significant group-by-session interaction (P < 0.05), such that the probiotic group showed improved accuracy in recognising facial expressions following the intervention. This suggests that probiotics may subtly influence how individuals process social and emotional cues, although the effect was not strong enough to be considered robust.

Baseline Risk Aversion Predicts Response

Exploratory analysis revealed that individuals with a greater tendency towards risk aversion, measured by the LEIDS-R subscale, showed a greater improvement in negative mood in response to probiotics. No similar predictive relationship was found in the placebo group. This finding suggests that probiotics may be particularly beneficial for individuals who are prone to anxiety or worry, traits that are transdiagnostic risk factors for mental disorders.

No Effect on Bowel Symptoms

The study found no evidence that the probiotic intervention affected bowel complaints, frequency, or stool consistency as measured by the Bristol Stool Scale. This is important because it suggests that the mood effect was not simply a consequence of resolving gastrointestinal discomfort. The psychological benefits were independent of gastrointestinal changes, supporting a direct microbiome-gut-brain axis mechanism rather than an indirect effect mediated by digestive symptom relief.

1. Lessons Learned

Measurement Matters: Daily Self-Reports Are More Sensitive Than Questionnaires

The most important lesson from this study is that the choice of measurement tool fundamentally determines whether an effect is detected. The standard approach in the field, using pre- and post-intervention questionnaires, failed to detect an effect that daily tracking revealed clearly. This suggests that many previous null findings may have been false negatives, with the effect existing but remaining undetected due to insensitive measurement. The researchers concluded that daily self-reports should be incorporated more widely in future studies of psychological interventions.

Probiotics Affect Negative Mood Specifically, Not Positive Mood

The finding that probiotics reduce negative feelings without enhancing positive ones is clinically relevant. Negative mood is a core feature of depression and anxiety disorders, and reducing it is a primary therapeutic goal. The fact that probiotics do not artificially elevate positive mood may be an advantage, as some pharmacological interventions can cause emotional blunting. However, the researchers emphasised that probiotics are not a replacement for antidepressants but may serve as an early intervention or adjunctive strategy.

Individual Differences Determine Response

Not everyone responds equally to probiotics. The finding that risk aversion predicted greater response suggests that probiotics may be most beneficial for individuals with a tendency towards anxiety or worry. This opens the possibility of targeted interventions, where probiotics are prescribed based on individual psychological profiles rather than used universally. The researchers concluded that identifying traits of individuals who derive greatest benefit will allow future targeting of at-risk individuals.

The Microbiome-Gut-Brain Axis Mechanisms Are Plausible but Not Proven

The study did not measure microbiome composition directly, so it cannot establish the mechanism through which probiotics affected mood. However, the researchers discussed plausible pathways based on existing evidence: neural signalling via the vagus nerve, immune modulation through reduction of pro-inflammatory cytokines, and endocrine effects through reduction of cortisol levels. Animal studies have shown that probiotics can reduce anxiety and depressive-like behaviours only when the vagus nerve is intact, and human studies have found evidence that probiotics and prebiotics can lower cortisol levels. The present study adds to this mechanistic foundation by demonstrating that a psychological effect exists in healthy humans.

The Two-Week Latency Is Clinically Relevant

The finding that the mood improvement began after approximately two weeks is notable because it parallels the latency period of many antidepressants, which typically take two to four weeks to show clinical effects. This suggests that probiotics may act through similar mechanisms involving gradual neurochemical or neuroendocrine changes rather than immediate psychotropic effects. It also provides a practical guideline for individuals trying probiotics for mood: at least two weeks of consistent use may be required before any benefit is noticed.

1. How This Research Can Help Humanity

Improving Study Design in Mental Health Research

The study provides a methodological lesson that extends far beyond probiotics. Daily self-reports of mood, which are simple and inexpensive to implement, may be more sensitive than standard questionnaires for detecting subtle psychological changes. The researchers hope that their findings will encourage other investigators to incorporate simple daily measures of mood into their studies. This could improve the detection of effects across a wide range of interventions, including pharmaceuticals, behavioural therapies, and nutritional supplements.

Informing Personalised Probiotic Recommendations

The finding that risk aversion predicts response to probiotics suggests that probiotics could be recommended more selectively. Rather than advising everyone to take probiotics for mental health, clinicians could assess individual traits and recommend probiotics primarily to those who are prone to anxiety, worry, or stress. This personalised approach could improve the cost-effectiveness of probiotic interventions and reduce the number of people who take probiotics without benefit.

Providing an Evidence Base for Probiotic Use in the General Population

Previous meta-analyses had found stronger evidence for probiotics in clinically depressed populations than in healthy individuals, leaving uncertainty about whether healthy people benefit. This study provides clear evidence that healthy young adults can experience meaningful reductions in negative mood from probiotic supplementation. While the effect size was not quantified in the published abstract, the statistical significance and the two-week timing of emergence suggest a clinically relevant effect.

Supporting the Microbiome-Gut-Brain Axis as a Therapeutic Target

The study adds to the growing evidence that the gut microbiome influences mental health in humans, not just in animals. This supports the broader goal of developing microbiome-focused interventions for mental health, including probiotics, prebiotics, and dietary modifications. The researchers noted that psychological conditions are often comorbid with gastrointestinal problems, suggesting that gut-brain interactions may be relevant to a wide range of mental health conditions.

Offering a Low-Risk Intervention for At-Risk Individuals

Probiotics are generally safe and well tolerated, with few side effects compared to antidepressants and other psychotropic medications. The study suggests that probiotics could serve as a low-risk early intervention for individuals who are at risk of developing clinical depression or anxiety, such as those with high trait anxiety or risk aversion. By reducing negative mood before it escalates to clinical levels, probiotics might help prevent the onset of mental disorders.

1. Final Summary

Most Important Takeaways

1. Daily self-reports reveal that probiotics reduce negative mood in healthy adults. The Johnson and Steenbergen study found a clear and statistically significant reduction in daily self-reported negative mood in the probiotic group compared to placebo, an effect that emerged after approximately two weeks of supplementation.

2. Standard psychological questionnaires failed to detect the effect. Despite a comprehensive battery of well-validated questionnaires including the STAI, PSWQ, PSS, CES-D, and PANAS, none showed significant differences between the probiotic and placebo groups after correction for multiple comparisons. This reconciles the inconsistencies of previous studies and reveals that common pre- versus post-intervention assessments are not sensitive enough to detect probiotic-induced changes in healthy subjects.

3. The effect is specific to negative mood, not positive mood. Probiotics reduced negative feelings such as anxiety, sadness, stress, and fatigue without significantly affecting positive mood ratings. This selectivity may be an advantage over some antidepressants that blunt all emotions.

4. Individual traits predict response. Exploratory analysis revealed that baseline risk aversion, measured by the LEIDS-R subscale, significantly predicted improvement in negative mood in the probiotic group. This suggests that some people, particularly those prone to anxiety and worry, may benefit more than others.

5. The two-week latency is clinically relevant. The effect began to emerge after approximately two weeks, which parallels the latency of many antidepressants. Consistent daily supplementation for at least two weeks may be required before any benefit is noticed.

6. The study was conducted in young, healthy adults. The sample comprised 88 predominantly female participants with a mean age of 22.3 years. The findings may not generalise to older adults, clinical populations, or males.

Action Points

For Individuals Considering Probiotics for Mood

If you are a healthy adult experiencing low-level negative mood, anxiety, or stress, a multispecies probiotic containing Bifidobacterium and Lactobacillus strains may help. The effect begins after approximately two weeks of daily use, so consistent daily supplementation is essential. Do not expect immediate results. If you are prone to worry or risk aversion, you may be more likely to benefit. Probiotics are not a replacement for antidepressants or therapy, but they may serve as a complementary or early intervention strategy.

For Clinicians

Consider asking patients about their use of probiotics and discussing the evidence for mood effects. The Johnson and Steenbergen study provides evidence that probiotics can benefit mental health in the general population, not just in clinically depressed patients. However, the evidence is limited to healthy young adults, and probiotics should not be prescribed as a substitute for evidence-based treatments for clinical depression. For patients who are at risk of developing depression, such as those with high trait anxiety or rumination, probiotics may be a low-risk adjunctive intervention.

For Researchers

Incorporate daily self-reports of mood into studies of psychological interventions. The Johnson and Steenbergen study demonstrates that pre- and post-intervention questionnaires may miss effects that daily tracking reveals. This finding is likely to extend beyond probiotics to other nutritional, behavioural, and pharmaceutical interventions. As the researchers concluded, in our attempt to unravel the complexity of the human brain and emotions, we must not forget to ask the obvious questions. Sometimes the simplest questions yield the most meaningful answers.

For Public Health Authorities

Consider the potential for probiotics as a population-level mental health intervention. With mental health disorders representing a growing burden, low-risk and low-cost interventions such as probiotics deserve attention. However, the evidence is still emerging, and public health recommendations should be cautious. The researchers noted that probiotics are widely available over the counter in many countries, and their findings suggest that these products may provide mental health benefits beyond their well-known gastrointestinal effects.

For the Supplement and Food Industry

The study provides evidence to support the marketing of probiotics for mental health benefits, but claims must be carefully worded to avoid misleading consumers. The effect size and clinical significance remain to be fully characterised. Industry investment in further research, particularly larger and longer-term trials with daily monitoring, would strengthen the evidence base and potentially expand the market for mood-targeting probiotics.

Recommended Follow Up Study

The High-Risk Population Probiotic Prevention Trial

The Johnson and Steenbergen study found that risk aversion predicted response to probiotics and that the effect emerged after two weeks in healthy young adults. The logical next step is a randomised controlled trial in individuals at high risk of developing depression, such as those with subclinical depressive symptoms, high trait anxiety, or a family history of depression. A larger sample, at least 200 participants, would provide adequate power to detect effects. The trial would compare a four-month probiotic intervention to placebo, with daily mood monitoring as the primary outcome and clinically validated depression scales as secondary outcomes. The follow-up period would extend to 12 months to assess whether probiotics prevent the onset of clinically significant depression. If positive, this trial would provide the evidence base for probiotics as a scalable, low-cost depression prevention strategy.

List of Other Related and Connected Studies

The Cryan and Dinan Microbiome-Gut-Brain Axis Foundational Review (2012, Nature Reviews Neuroscience)

This landmark review established the conceptual framework for the microbiome-gut-brain axis, synthesising evidence from animal studies that gut bacteria influence behaviour, stress responses, and neural development. The Johnson and Steenbergen study builds on this foundation by demonstrating a psychological effect in humans.

The Messaoudi Probiotic Anxiety and Depression Study (2011, British Journal of Nutrition)

This randomised controlled trial found that a four-week probiotic intervention reduced psychological distress and improved mood in healthy volunteers, but the effect was measured with questionnaires. The Johnson and Steenbergen study extends this by showing that daily reports are more sensitive than questionnaires.

The Steenbergen Probiotic and Cognitive Reactivity Study (2015, Brain, Behavior, and Immunity)

This study, by co-author Laura Steenbergen, found that the same Ecologic Barrier probiotic used in the Johnson study reduced cognitive reactivity to sad mood in healthy volunteers. The Johnson study replicates and extends this finding by using daily mood reports.

The Schmidt Prebiotic Stress Study (2015, Psychopharmacology)

This study found that prebiotic supplementation reduced cortisol awakening response in healthy volunteers, suggesting that gut-focused interventions can affect the stress hormone system. The Johnson study did not measure cortisol but provides evidence of subjective mood effects that may be mediated by similar pathways.

The Bastiaanssen Microbiome and Depression Review (2020, Biological Psychiatry)

This review synthesised evidence that gut microbiome composition differs between depressed patients and healthy controls and that microbiome-targeted interventions may be effective in depression. The Johnson study contributes to this literature by showing effects in healthy individuals.

The ADEPT Probiotic and Depression Trial (2022, JAMA Psychiatry)

This large randomised controlled trial found no benefit of probiotics for major depression, highlighting the difficulty of translating microbiome interventions to clinical populations. The Johnson study suggests that daily monitoring might have detected effects that were missed by standard assessments in the ADEPT trial.

The Kortman Gut-Brain Axis and Microbiome Review (2024, Annual Review of Clinical Psychology)

This comprehensive review concluded that the microbiome-gut-brain axis holds promise for mental health but that methodological inconsistencies have hampered progress. The Johnson study directly addresses one such inconsistency by demonstrating the importance of measurement sensitivity.