Showing posts with label clinical study. Show all posts
Showing posts with label clinical study. Show all posts

Thursday, March 23, 2023

Lactobacillus, Bifidobacterium and other emerging probiotics

In the past few decades, Lactobacillus and Bifidobacterium genera were the main bacteria to be used as probiotics. One of the reasons for such attention was that these bacteria have the ability to thrive aerobically and are simple to package for sale. Cass Nelson-Dooley and Tony Hoffman of Metametrix (acquired by Genova Diagnostics in 2012) talked about these probiotics in MEBO interviews in 2009, emphasizing that high levels of Lactobacillus could be bad in some cases. Diets rich in simple carbs, poor absorption and small intestinal pockets can feed and trap excess Lactobacillus. Adding probiotics Bifidobacteria or Saccharomyces boulardii and prebiotic fiber to the diet could, in their opinion, help the MEBO population. D-lactate was one of the tests they offered that measured the level of this byproduct of bacterial fermentation, indicating an overgrowth of bacteria in the small intestine. According to our early clinical studies, D-lactate, however, was elevated in less than 20% of MEBO and PATM populations. 

Even though Lactobacillus is only a minor member of the human colonic microbiota, the proportions of those bacteria and particular strains are frequently either positively or negatively correlated with human disease and chronic conditions. Although it generally is not considered a pathogen, it can cause disease in compromised hosts, including bacterial endocarditis, pleuropulmonary infections, gastrointestinal abscesses, urinary tract infection, conjunctivitis, dental caries, and endometritis. Lactobacillus bacteremia, presumably secondary to bacterial translocation from the gastrointestinal tract, has been reported in a patient with severe intestinal inflammation caused by ulcerative colitis. A decrease in genus Faecalibacterium and increase in Lactobacillaceae has been previously reported in patients with constipation (Lactiplantibacillus plantarum P9 supplementation, on the other hand, helped to increase frequency of bowel movements). Compared to healthy individuals, pediatric and adult Crohn's disease (CD) patients had an increased relative abundance of Lactobacillus species. In both cases, Lactobacillus enrichment coincided with depletion of F. prausnitzii - that could represent a good candidate as next-generation probiotic.

The percentages of Bifidobacterium and the Lactobacillus group presented a decreasing trend in patients with quiescent ulcerative colitis (UC) compared to active UC, although no significant differences were observed. Excessive amounts of Bifidobacteria can have negative effects on the body, such as in the case of Bifidobacterium breve bacteremia caused by excessive probiotic supplementation in infants with gastrointestinal conditions. On the other hand, Lactobacillus, Bifidobacterium and F. prausnitzii are depleted in IBS patients, resulting in lower SCFA concentrations (short-chain fatty acids, which are organic acids produced by gut bacteria during fermentation of indigestible foods) compared with healthy individuals. Maternal prenatal stress could also cause lower levels of Lactobacillus and Bifidobacterium and keep them low even later in life. With lower levels of Bifidobacterium, celiac patients have an imbalance in the intestinal microbiota, regardless of pH, even while on a gluten-free diet. Hormone treatment affects Bifidobacterium longum's ability to produce acids: progesterone reduces lactic acid and acetic acid production, while estradiol and thyroxine increase levels of both lactic acid and acetic acid (ethinyl estradiol) or lactic acid and butyric acid (thyroxine hormone). 

Data from our microbiome study (NCT03582826: Microbial Basis of Systemic Malodor and "People Allergic To Me" Conditions  - show that while there was a general trend of increasing levels of Bifidobacteria and Lactobacillus in individuals who achieve remission, less than 10% of participants needed to boost levels of both species to improve their condition. In line with previous knowledge, in MEBO/PATM population, Bifidobacteria (one of the first microbes to colonize the human gastrointestinal tract) was more common in younger individuals. About one third of the participants experienced lesser symptoms when increasing levels of their gut Bifidobacteria, another third benefited from more Lactobacillus and the remaining third felt better when increasing levels of Faecalibacterium prausnitzii (a butyrate-producing anaerobe typically associated with good health). Changes in the levels of these bacteria corresponding to recovering are overlayed on Figure 5 from our paper published in JMIR dermatology. It was illustrating changes in microbial diversity vs abundances of selected bacterial species associated with odor (CSS) for 12 female (F1-F12) and 10 male (M1-M19) participants who self-reported both flare-ups (beginning of the arrow) and improvements (end of the arrow). We added text boxes showing whether the levels of F. prausnitzii (F), Bifidobacterium (B) and Lactobacillus (L) are increasing (🡹) , decreasing (🡻) or fluctuating (🡺) as symptoms resolve. It is obvious that individuals with the highest bacterial diversity don't benefit from increasing levels of probiotic diversity, while those with low diversity levels do need more probiotic microbes in their system. The figure demonstrates the complexity of these bacterial perturbations and emphasizes the need to consider more than just three species when predicting which treatment will work best for reducing MEBO and PATM symptoms. 

As we mentioned in the previous blog post, Lactobacillus is the main genera responsible for odors of the uncooked food waste. Some strains of Lactobacillus produce lactic acid, which can contribute to a more acidic environment in the gut. This can be beneficial for some people, but for others it can exacerbate odor issues. Bifidobacteria could also produce organic acids such as acetic acid and lactic acid in addition to short-chain fatty acids (SCFAs) like butyrate, which can have anti-inflammatory effects and promote gut health, but some people may find it unpleasantly smelling. F. prausnitzii produces a range of metabolites including one of the worst smelling compounds Putrescine. F. prausnitzii  plays important role in balancing immunity. Its butyrate production has been correlated to the capacity to induce IL-10, an anti-inflammatory cytokine, in peripheral blood mononuclear cells (PBMC) but not to the ability to block IL-8 secretion in TNF-α-stimulated HT-29 cells. In COVID-19, Faecalibacterium prausnitzii has been inversely correlated with disease severity. 

Research has shown that the gut microbiome is far more complex than previously thought, with hundreds of bacterial species interacting in a delicate balance.  Therefore, focusing solely on Lactobacillus and Bifidobacterium, or any other bacterium, may not be enough to promote a healthy gut microbiome for everyone. Besides, different strains of bacteria within the same species can have different effects on the body.

After more than a century of research, the following species are identified as the primary probiotic species of Lactobacillus species: L.acidophilus, L.casei, L.fermentum, L.gasseri, L.johnsonii, L.paracasei, L. plantarum, L. rhamnosus, and L. salivarius. For Bifidobacteria these are: B.adolescentis, B.animalis, B.bifidum, B.breve, and B.longum. We are learning more about strains of these species. Novel probiotic strain Bifidobacterium bifidum CECT 7366 is active against the pathogenic bacterium Helicobacter pylori - and so are variations of Saccharomyces Boulardii and Lactobacilli johnsonii. Another strain L. plantarum 299v can directly interfere with E. coli colonization and improve the immunological status of the intestinal mucosa although these results have not yet been confirmed in humans. Lactobacillus GR-1 and B-54 or RC-14 strains in the vagina has been shown to reduce the risk of urinary tract infections. Mileti et al. found that Lactobacillus paracasei displayed a delay in the development of colitis and a decreased severity of disease but that L. plantarum and L. rhamnosus GG exacerbated the development of dextran sodium sulfate (DSS)-induced colitis. Bifidobacterium supplemented with various fiber additives can prevent the growth of Clostridium difficile, while Lactobacillus salivarius protects the broiler chicks from Salmonella infections. Several strains of other species promising as probiotics are from Roseburia spp., Akkermansia spp., and Faecalibacterium spp. Probiotic potential risk score (PPRS) allowed to classify 84 Faecalibacterium prausnitzii strains  into low-, medium-, and high-risk groups. 15 strains identified as low-risk strains are prioritized for clinical application. 

Unfortunately, the extreme oxygen sensitivity of F. prausnitzii imposes practical challenges to the production, transportation, storage, and manufacturing of probiotic products to be evaluated in a clinical setting. Instead, we have to turn to prebiotics - a type of dietary fiber - in our food . F. prausnitzii's growth is promoted by inulin, inositol (vitamin B8) and fructo-oligosaccharides, chicory roots, wheat, onion, banana, garlic, and leek, wheat, rye, rice, barley, oat, and sorghum, and gold kiwifruit

Gut microbiota can be also influenced by phytochemicals. In one study, black raspberry (BR) reduced the serum levels of trimethylamine-N-oxide and cholesterol in rats fed excessive choline with a high-fat diet (HFC). The authors hypothesized that since gut microbiota plays a crucial role in the production of trimethylamine and microbial metabolites, BR could influence gut microbial composition. Their study examined microbiomes and metabolomes of rats and showed that the BR supplementation enriched Bifidobacterium and reduced Clostridium cluster XIVa. 

While we have made significant progress in understanding the role of probiotics and prebiotics in supporting our gut health, it's important to recognize that there is still a great deal that we don't know. It's becoming increasingly clear that a personalized approach to prebiotic and probiotic use may be necessary to achieve optimal health outcomes. This approach involves paying attention to how different foods make you feel and adjusting your diet accordingly, while eating a diverse and balanced diet that incorporates a variety of whole foods and phytochemicals, staying hydrated, getting enough sleep and exercise, and managing stress levels. Focusing on overall health and well-being will help to support the body's natural ability to maintain a healthy microbiome and avoid dysbiosis-related health issues.


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Bai Z, Zhang N, Jin Y, Chen L, Mao Y, Sun L, Fang F, Liu Y, Han M, Li G. Comprehensive analysis of 84 Faecalibacterium prausnitzii strains uncovers their genetic diversity, functional characteristics, and potential risks. Frontiers in Cellular and Infection Microbiology. 2023 Jan 4;12:1924.

Gabashvili I.S. Cutaneous Bacteria in the Gut Microbiome as Biomarkers of Systemic Malodor and People Are Allergic to Me (PATM) Conditions: Insights From a Virtually Conducted Clinical Trial. JMIR Dermatol 2020;3(1):e10508 URL: DOI: 10.2196/10508

Wednesday, December 1, 2021

FMO3 and COVID-19

Flavin-containing monooxygenase 3 (FMO3) enzyme is a seemingly insignificant enzyme that normally converts fishy-smelling trimethylamine (TMA) into a neutral trimethylamine-N-oxide (TMAO). The amounts of this highly-specialized detoxifying enzyme are highly variable. It depends on the age, sex hormones, infections (estradiol and testosterone, hepatitis virus have been found to reduce FMO3 capacity), obesity traits and diseases such as diabetes. The difference can be up to 20-fold between individuals. Mutations in the FMO3 gene cause low metabolic capacity associated with the disorder trimethylaminuria (TMAU) that attracts little biomedical interest.  This condition, however, might matter more than it seems.

Could there be a link between FMO3 and SARS-CoV-2 infection and vaccination? 

Individuals differ in their susceptibility to viral infections and genes contribute to the risk score. Less than 10% of humans infected with Mycobacterium tuberculosis develop TB, partially because of polymorphism in Tyrosine kinase (TYK2, P1104A) also responsible for severe COVID-19. Early in the pandemic, it was discovered that SARS-CoV-2 infection is dependent on the ACE2 receptor for cell entry and the serine protease TMPRSS2 for spike protein priming. ACE2 expression, indeed, influences COVID-19 risk and a rare variant located close to this gene was found to confer protection against COVID-19, possibly by decreasing ACE2 expression. Interestingly, FMO3 is one of the few genes with expression correlated to ACE2 [Sungnak et al, 2020] along with genes associated with immune functions. 

Coronavirus disease is associated with increased risk of thrombotic events. According to recent research, low levels of FMO3 protect against thrombosis [Shih et al, 2019] while some FMO3 mutations confer higher risk [Oliveira-Filho et al, 2021]. FMO3 rs1736557 might increase the anti‐platelet efficacy of clopidogrel [Zhu et al, 2021]. Genetic risk can be mediated by gut microbiota [Gabashvili, 2020]. There are also associations with other diseases such as diabetes, renal and cardiovascular conditions increasing risk of severe COVID-19. 

Studying trimethylaminuria-like conditions might help in developing strategies for prevention and therapy of other diseases, including COVID-19.

Our COVID-19 disease and vaccines study [NCT04832932, Gabashvili, 2021] compares side-effects of vaccines and clinical course of infections (including vaccine breakthroughs) in several cohorts including MEBO and TMAU. You can help by enrolling and participating in this online survey in English or Spanish.


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Gabashvili IS. Cutaneous Bacteria in the Gut Microbiome as Biomarkers of Systemic Malodor and People Are Allergic to Me (PATM) Conditions: Insights from a Virtually Conducted Clinical Trial. JMIR Dermatology. 2020 Nov 4;3(1):e10508.  

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Liu W, Wang C, Xia Y, Xia W, Liu G, Ren C, Gu Y, Li X, Lu P. Elevated plasma trimethylamine-N-oxide levels are associated with diabetic retinopathy. Acta Diabetologica. 2021 Feb;58(2):221-9.

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Oliveira-Filho AF, Medeiros PF, Velloso RN, Lima EC, Aquino IM, Nunes AB. Trimethylaminuria and Vascular Complications. Journal of the Endocrine Society. 2021 Apr;5(Supplement_1):A313-4. 

Zhu KX, Song PY, Li MP, Du YX, Ma QL, Peng LM, Chen XP. Association of FMO3 rs1736557 polymorphism with clopidogrel response in Chinese patients with coronary artery disease. European Journal of Clinical Pharmacology. 2021 Mar;77(3):359-68.

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Wednesday, December 4, 2019

Saturday, November 30, 2019

Concluding the MEBO-uBiome study

We are very close to concluding our clinical trial NCT03582826 ​according to the pre-specified protocol. 

Recruitment for our microbiome study commenced in June 2018 and completed in December 2018. The last batch of raw FASTQ sequences and raw taxonomy-level abundance summaries was received in October 2019. We had to spend more time on collecting and processing annotation data (medical histories, dietary intakes, medications, etc), ensuring quality, completeness, consistency and validity, and defining best approaches of dealing with "missing values".

We have now completed final data collection for the primary and secondary outcome measures.

...  Read more ... 

Thursday, August 29, 2019

Reducing the stigma of odor disorders

Some conditions - such as obesity, depression and functional odor disorders - come with a social stigma. Understanding the etiology of these conditions helps to avoid stereotypes and find remedies. 

Metabolomics analysis of morning urine samples helped to evaluate the heterogeneity of MEBO population as good as challenge tests, procedures used to induce symptoms and assess resilience to perturbations caused by sugars. 

...  Read more ... 

Sunday, July 21, 2019

How do you feed your microbiome?

As we are gearing up for final stage of our clinical study NCT03582826, we are looking at different subgroups of our participants, to find more precise, personalized and powerful solutions for everyone. 

We know, that genes and environment always combine to make us who we are. We already talked about some of those seemingly less important genes - such as ABO blood group. The environmental factors include diet, exercise, social environments (such as social support), varying conditions and situations.   

According to descriptions of diets and lifestyle submitted by our participants, about 25% of them are taking commercial probiotic products, about the same number as those who had chicken as the main staple of their kitchen (see word cloud depiction of diet keywords on the right). About 2% take commercial prebiotics, but many more eat prebiotic foods (such as oats/oatmeal, bananas, apples and nuts shown in green). 4% eats onions and about the same percentage actively avoids it.  Milk, Rice, Bread and Pasta were among the most popular foods, after chicken. 

The world is eating less meat overall, and a few years ago, among the meat eaters, the scale tipped from greater consumption of beef to greater consumption of chicken.
This is even more pronounced in MEBO population. 

Half of MEBO population takes vitamins and  supplements.  The most popular among them is Zinc closely followed by Magnesium and Enzymes. Also popular are multivitamins, Calcium,  Resveratol, Vitamin B12, Charcoal, Chlorophyll, Ashwagandha, Biotin, Primrose oil, Omega 3 and Livercare. Blood pressure drugs are among the most used medications.

This information will be useful in understanding differences in microbiomes of subjects with similar symptoms. We might already be observing interesting trends. Certain supplements, for example, seem to benefit some blood group more than others. Same about foods naturally rich in certain minerals and vitamins.

We'll talk about this next time.

Friday, November 30, 2018

On what's in your head, the downside of diversity and deodorizing bacteria: preliminary results of NCT03582826

Mirrored from Aurametrix, preliminary results of NCT03582826:

By now, 105 uBiome kits were registered in our Quality of Life form (out of 300+ sets sent). 16 volunteers have submitted all 3 samples and 23 registered one or two kits. Over half of our participants have not yet started the study. We hope that this post will answer some of their questions on where to start.

The goal of MEBO Gut Microbiome study is to identify microbial communities associated with flare ups and remission of systematic malodor or PATM conditions.  Participants are asked to answer MEBO Quality of Life (QoL) questionnaire about the last 24 hrs before every sample is taken. The Questionnaire is embedded into English and Spanish posts about the study.  The answers are used to calculate total QoL score that increases with less odor and more wellbeing. The questionnaire evaluates physical health, psychological health, social support and environment such as diet and exercise. A separate score is calculated for MEBO (malodor/PATM) symptoms.

This post is about preliminary results of the first 41 uBiome kits representing samples from 24 participants. Quality of Life scores varied from 55 to 149 (best, "ideal" quality of life would be 150), while MEBO scores ranged from 3 (experienced MEBO/PATM symptoms "all the time" and odor was lingering for hours at 10 feet and farther) to 25 (100% odor free).

The pie chart on the right shows social environment for MEBO volunteers: almost half of study participants had active social lives interacting with over 8 people per day. Almost 20% of social interactions were entirely positive, while 10% were entirely negative. Social support did not correlate with MEBO symptoms and it was not predictor nor protective of malodor.

Graphs of depressive feelings vs odors is another prove that even with stress as a trigger, body odors are not all in your head. Obviously, those free of malodors tend to be happier, but it's possible to control depression even with worst MEBO symptoms and happy mood, alone, does not help to control the symptoms.

uBiome provides a multitude of measurements characterizing gut microbiomes. One of the most widely used metrics is gut diversity metrics.

Microbial diversity is cornerstone of a good health and decreased diversity is likely to be associated with poor wellbeing including malodors. Our data, however, show that most MEBO volunteers have healthy diversity scores that do not correlate with incidence of odors. Correlation is observed only for  cases of very low, unhealthy diversity, especially for total quality of life scores as opposed to just malodor symptoms. Even more so, MEBO volunteers in remission, completely free of odors, had healthy,  but not the very best diversity scores.

In extremely diverse communities, people may be less inclined to trust each other, vote and volunteer for activities. In people not suffering from uncontrollable odors, microbes and hosts have a balanced dialogue, and form symbiotic relationships beneficial for both parties.

Could it be that MEBO sufferers have more "smelly" bacteria and less bacteria neutralizing the smell?  Levels of traditional probiotic strains in the study participants do not seem to correlate with their odor levels. As more data is accumulated, interesting patterns are emerging for less common bacteria. An example is Anaerovorax, bacterium that processes putrid-smelling compounds produced by clostridia, pseudomonads, lactic acid and other bacteria from amino acids in food. Anaerovorax is observed in only 3% of uBiome samples, but it is present in a quarter of MEBO participants. Plotting its levels in all samples with Anaerovorax present against the scores representing improvement of MEBO symptoms (from worst odor to no odor), shows a clear trendline - the more Anaerovorax, the less odor. We note that this seem to be the case for ONLY a subset of study participants. For example, participants #901298 and #901240 who contributed 2 samples each into the first 41-sample-set, had 0 Anaerovorax on their worse day and over 0.03% Anaerovorax on their better days. # 901240 submitted 2 "bad day" samples (which is perfectly fine - as it still yielded much needed data!) had a little more Anaerovorax on a slightly better day. #90346 had experienced exactly the same MEBO symptoms on both days of sampling, but the result with 0 Anaerovorax corresponded to a worse Quality of Life score. #90365 experienced exactly the same symptoms for days with and without Anaerovorax, and had the same Quality of Life Score, but there was a noticeable difference in diet. This person had more Anaerovorax when consumed more odor triggers in food (onions and vinegar).

There is not much known about the role of Anaerovorax in human gut. It is proposed to increase when adding meat to a usually vegetarian diet and with increased fasting insulin levels. It is also increased in healthy preadolescent children (7 to 12 years old) and by minerals such as Zinc and Manganese.

Levels of TMA-producing bacteria such as Desulfibrio and Bacteroides seem to lower when odor levels are decreasing in most cases, although signal-to-noise ratio is weak and the dependence is very individual (see Figure). The trend is similar for #901214, #90265, #900841 and #901273. #901253 is one of the outliers: MEBO symptoms in this case were improving with higher levels of Bacteroides and lover diversity (and a much stricter diet).

Bacteroides maintain a complex and generally beneficial relationship with the host. The genus Bacteroides makes up to 30% of the bacteria in the human gut. Levels of Bacteroides increase with rapid weight loss. Specific species of Bacteroides, have a role in preventing infection with Clostridium difficile. Level of Bacteroides, however, increase (or significantly decrease) in inflammatory bowel disease and chronic fatigue syndrome.
Lactobacillus and bifidobacteria probiotics, garlic, walnuts and barley could all decrease levels of Bacteroides. Polydextrose (dietary fiber found in traditional bakery items, beverages, dairy products and sauces) not only decreases Bacteroides but also increases probiotic strains of Lactobacillus and Bifidobacterium.

These are only preliminary results. We'll be looking at more complex patterns in bacterial community structures associated with different subgroups of study participants. We'll be posting new findings as we mine more data and get additional results.

If you have not started the study yet, we hope that this post encourages you to start participating. As you see, every data point counts, even if the results of your experimentation and odor reduction approaches are not as good as we wished.


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