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Using probiotics to support digestive health for dogs

By Kelly S. Swanson, PhD, The Kraft Heinz Company Endowed Professor in Human Nutrition, University of Illinois at Urbana-Champaign, USA

Because dogs are considered to be members of the family by most pet owners today, their health and well-being is a top priority. As with humans, nutritional products supporting gastrointestinal health are some of the most popular. Many pets are healthy, but loose stools, constipation, and various gastrointestinal disorders and diseases such as inflammatory bowel disease and irritable bowel syndrome are common. In fact, within the pet food conversation, digestive health improvements have been the most discussed health benefits among social media discussion posts over the past 2 years (see here). Given the high interest in digestive health, it is not surprising that the canine microbiome has been of great interest over the past decade, with many recent reviews reporting on the overall composition of the gastrointestinal microbiota and how it is impacted by diet (Barko et al., 2018; Alessandri et al., 2020; Wernimont et al., 2020). Gastrointestinal microbiome changes contributing to or resulting from digestive diseases have also been documented in dogs (Redfern et al., 2017; Ziese and Suchodolski, 2021). Animals under high levels of stress or undergoing antibiotic therapy are also known to have poor stool quality and an altered gut microbiota (i.e., dysbiosis) (Pilla et al., 2020).

Dietary fibers and prebiotics are commonly used in complete and balanced diets to improve or maintain stool quality, provide laxation, and positively manipulate the microbiota of healthy animals. The use of probiotics is also popular in dogs, but the route of administration, efficacy, and reason for use is usually different than that of fiber and prebiotics. Probiotics are usually provided in the form of supplements (e.g., powders, capsules, pastes) and are most commonly used to treat animals with gastrointestinal disease rather than support the healthy condition. Live microbes are added to many dry extruded foods as ‘probiotics’, but in many cases, maintaining viability and evidence for a health benefit for dogs is lacking for these products. Such microbes would not meet the minimum criteria to be called a ‘probiotic.’ Viability is a challenge because most HACCP plans for producing complete and balanced pet foods include a kill step that inactivates all microorganisms. Therefore, inclusion must be applied post-extrusion on the outside of the kibble. Even if applied in this way, low numbers of viable organisms are common (Weese and Arroyo, 2003). Post-production inclusion is not possible for other diet formats (e.g., cans, pouches, trays). Although spore-forming bacteria that may survive the extrusion process have been of interest lately, evidence of efficacy is lacking thus far.

Picture of Simka (a Samoyed) courtesy of ISAPP board member Dr. Daniel Tancredi

Even though health benefits coming from the inclusion of live microorganisms in dog foods is not supported by the peer-reviewed literature, such evidence exists for many probiotic supplements. The clinical effects of probiotics in the prevention or treatment of gastrointestinal diseases in dogs have been reviewed recently (Schmitz and Suchodolski, 2016; Suchodolski and Jergens, 2016; Jensen and Bjornvad, 2018; Schmitz, 2021). Although some similarities exist, recent research has shown that distinct dysbiosis networks exist in dogs compared to humans (Vazquez-Baeza et al., 2016), justifying unique prevention and/or treatment strategies for dogs.

One population of dogs shown to benefit from probiotics has been those with acute idiopathic diarrhea and gastroenteritis, with a shorter time to resolution and reduced percentage of dogs requiring antibiotic administration being reported (Kelley et al., 2009; Herstad et al., 2010; Nixon et al., 2019). Probiotic administration has also been shown to benefit dogs undergoing antibiotic therapy and those engaged in endurance exercise – two conditions that alter the gastrointestinal microbiota and often lead to loose stools. In those studies, consumption of a probiotic helped to minimize gastrointestinal microbiome shifts and reduced the incidence and/or shortened the length of diarrhea (Gagne et al., 2013; Fenimore et al., 2017). Dogs diagnosed with inflammatory bowel disease have also been shown to benefit from probiotic consumption (Rossi et al., 2014; White et al., 2017). In these chronic conditions, drug therapy is almost always required, but probiotics have been shown to help normalize intestinal dysbiosis, increase tight junction protein expression, and reduce clinical and histological scores.

So what is the bottom line? Well, for dogs with a sensitive stomach and/or digestive health issues, probiotics may certainly help. Rather than relying on live microbes present in the dog’s food or adding a couple spoonfuls of yogurt to the food bowl each day, it is recommended that owners work with their veterinarian to identify a probiotic that has the best chance for success. The probiotic selected should provide an effective dose, be designed for dogs, target the specific condition in mind, and be backed by science. As summarized here, it is important to remember that all probiotics are different so the specific microorganism(s), supplement form, storage conditions, and dosage are all important details to consider.

 

Kelly Swanson joined the ISAPP board of directors in June, 2020, providing valuable expertise in animal gut health and overall health. Swanson also chaired the 2019 ISAPP-led international consensus panel on the definition of synbiotics.

ISAPP board member Prof. Dan Tancredi kindly provided pictures of Simka, pet Samoyed, for the post.

 

Bacterial vesicles: Emerging potential postbiotics

By Dr. Gabriel Vinderola, PhD,  Associate Professor of Microbiology at the Faculty of Chemical Engineering from the National University of Litoral and Principal Researcher from CONICET at Dairy Products Institute (CONICET-UNL), Santa Fe, Argentina

The recently published ISAPP consensus paper defines a postbiotic as “a preparation of inanimate microorganisms and/or their components that confers a health benefit on the host“. Such a definition quickly brings to mind that a postbiotic is not equivalent to microbial metabolites. A postbiotic should also contain inanimate microbial cells or cell fragments. Metabolites or fermentation products may be present, but they are not required.

Because microbes are complex entities, we must be open to innovative understandings of what a postbiotic might entail. Indeed, although not explicitly mentioned in the ISAPP consensus paper, extracellular membrane vesicles may comprise an innovative conceptualization of a postbiotic, falling within the ‘cell component’ part of the postbiotic definition.

Bacterial vesicles

Extracellular membrane vesicles (EMV) are universal carriers of biological information produced in all domains of life. Bacterial EMV are small, spheroidal, membrane-derived proteoliposomal nanostructures, typically ranging from 25 – 250 nm in diameter, containing proteins, lipids, nucleic acids, metabolites, numerous surface molecules and many other biomolecules derived from their progenitor bacteria (Figure 1). Bacterial vesicles have been known for more than 50 years as structures able to carry cellular material (Ñahui Palomino et al. 2021).  However, studies on membrane vesicles derived from Gram-positive bacteria are more recent as it was for a time believed they were incapable of producing vesicles due to their thick and complex cell walls, and the lack of an outer membrane. Today, EMVs have been isolated from Gram-positive probiotic bacteria, including those belonging to the Lactobacillaceae family (under which Lactobacillus was recently split into many new genera) and the Bifidobacterium genus. In probiotic bacteria, vesicles may mediate quorum sensing and material exchange. Perhaps even more important, they can act as mediators of bacteria-to-cell and bacteria-to-bacteria interactions. As bacterial EMV are inanimate structures that cannot replicate, they fit the postbiotic definiton as cell components as long as other criteria stipulated by the definition are met.

Figure 1. Membrane vesicles budding on the surface of L. reuteri DSM 17938 and released into the surrounding medium. These vesicles were described in by Grande et al. 2017. Photo used with permission of BioGaia.

Functions of bacterial vesicles related to potential health benefits

Underlying mechanisms and corresponding molecules driving health effects of bacterial vesicles are not well understood, in part due to reliance on in vitro models. Bacterial EMV derived from Lactobacillaceae spp., Bifidobacterium spp., and Akkermansia spp. have been reported to alleviate metabolic syndrome and allergy symptoms, promote T-cell activation and IgA production, strengthen gut barrier function, and exhibit anti-viral and immunomodulatory properties (Kim et al. 2016; Tan et al. 2018; Ashrafian et al. 2019; Molina-Tijeras et al. 2019; Palomino et al. 2019; Shehata et al. 2019; Bäuerl et al. 2020). Interestingly, vesicles from Limosilactobacillus reuteri DSM 17938 (West et al. 2020) and Lacticaseibacillus casei BL23 (Domínguez Rubio et al. 2017) may accomplish some of the the effects of these probiotic bacteria. In fact it is not unreasonable to think that EMVs may be already present and active in probiotic products.

Challenges for bacterial vesicle production

To develop a postbiotic from microbial EMVs, many challenges need to be overcome.  Defining optimal cultivation conditions, and methods for vesicle release, isolation and scaling up are some of the challenges of bacterial vesicle production. There are several studies showing that altering the cultivation parameters can impact vesicle production. Examples of treatments shown to increase vesicle release include exposure to UV radiation and antibiotic pressure (Gamalier et al. 2017; Gill et al. 2019). Exposure to glycine has also been shown to increase vesicle production (Hirayama & Nakao 2020). Interventions during culture, for example by introducing agitation and varying pH, can possibly be ways to potentiate vesicle release and increase their bioactivity (Müller et al. 2021). A recent report also revealed that B. longum NCC2705 released a myriad of vesicles when cultured in human fecal fermentation broth, but not in basal GAM anaerobic medium (Figure 1). Moreover, the B. longum vesicle production pattern differed among individual fecal samples suggesting that metabolites derived from symbiotic microbiota stimulate the active production of vesicles in a different manner (Nishiyama et al. 2020). Whether any of these treatments and culture conditions are general or strain specific remains to be elucidated. Large differences in the number of vesicles that may be obtained by different extraction methods can occur (Tian et al. 2020). Tangential flow filtration or the use of antibodies targeting specific epitopes of the vesicles are some of the options proposed for the large scale isolation of EMV (Klimentová & Stulík 2015).

Figure 2. Left: Bifidobacterium longum NCC2705 grown on GAM broth. Right: secretion of membrane vesicles by Bifidobacterium longum NCC2705: the strain was cultured in bacterial-free human fecal fermentation broth and secreted a myriad of membrane vesicles. Reported and adapted from Nishiyama et al. 2020.

Progress has been made on the production of bacterial vesicles in recent years, yet several issues remain to be clarified including how vesicles are generated from the progenitor microbe, how the composition of vesicles changes according to the culture conditions, how to target specific bacterial vesicle purification from a pool of vesicles derived from other organisms (for example, bacterial vesicles produced in milky media can be accompanied by vesicles from eukaryotic cells present in the milk), safety aspects, quantification methods and the regulation of their use by the corresponding authority.

Their future as potential postbiotics

Membrane vesicles are an exciting opportunity for the development of postbiotics. A potential benefit of vesicles is that their small size compared to whole cells may enable them to more readily migrate to host tissues that could not be otherwise reached by a whole cell (Kulp & Kuehn 2010). Their nanostructure enables them to penetrate through the gut barrier and to be delivered to previously unreachable sites through the bloodstream or lymphatic vessels, and to interact with different cell types (Jones et al. 2020). For example, bacterial rRNA and rDNA found in the bloodstream and the brain of Alzheimer’s patients were postulated to have originated from bacteria vesicles (Park et al. 2017). Safety of EMVs must be carefully considered and assessed, even if they are derived from microbes generally recognized as safe, as their small size may increase penetration capacity with potential and yet unknown systemic effects. Novel postbiotics derived from microbial membrane vesicles is an intriguing area for future research to better understand production parameters, safety and functionality.

Thanks to Cheng Chung Yong, postdoctoral researcher at Morinaga Milk Industry Co., LTD (Japan) and Ludwig Lundqvist, industrial PhD student at BioGaia AB (Sweden) for their contributions to this blog, and Mary Ellen Sanders and Sarah Lebeer from ISAPP for fruitful discussions.

References

Ashrafian, F., Shahriary, A., Behrouzi, A., Moradi, H.R., Keshavarz Azizi Raftar, S., Lari, A., Hadifar, S., Yaghoubfar, R., Ahmadi Badi, S., Khatami, S. and Vaziri, F., 2019. Akkermansia muciniphila-derived extracellular vesicles as a mucosal delivery vector for amelioration of obesity in mice. Frontiers in microbiology10, p.2155.

Bäuerl, C., Coll-Marqués, J.M., Tarazona-González, C. and Pérez-Martínez, G., 2020. Lactobacillus casei extracellular vesicles stimulate EGFR pathway likely due to the presence of proteins P40 and P75 bound to their surface. Scientific reports10(1), pp.1-12.

Domínguez Rubio, A.P., Martínez, J.H., Martínez Casillas, D.C., Coluccio Leskow, F., Piuri, M. and Pérez, O.E., 2017. Lactobacillus casei BL23 produces microvesicles carrying proteins that have been associated with its probiotic effect. Frontiers in microbiology8, p.1783.

Gamalier, J.P., Silva, T.P., Zarantonello, V., Dias, F.F. and Melo, R.C., 2017. Increased production of outer membrane vesicles by cultured freshwater bacteria in response to ultraviolet radiation. Microbiological research194, pp.38-46.

Grande, R., Celia, C., Mincione, G., Stringaro, A., Di Marzio, L., Colone, M., Di Marcantonio, M.C., Savino, L., Puca, V., Santoliquido, R. and Locatelli, M., 2017. Detection and physicochemical characterization of membrane vesicles (MVs) of Lactobacillus reuteri DSM 17938. Frontiers in microbiology8, p.1040.

Gill, S., Catchpole, R. & Forterre, P., 2019. Extracellular membrane vesicles in the three domains of life and beyond. FEMS microbiology reviews, 43(3), pp.273–303.

Hirayama, S. & Nakao, R., 2020. Glycine significantly enhances bacterial membrane vesicle production: a powerful approach for isolation of LPS-reduced membrane vesicles of probiotic Escherichia coli. Microbial biotechnology, 13(4), pp.1162–1178.

Jones, E.J., Booth, C., Fonseca, S., Parker, A., Cross, K., Miquel-Clopés, A., Hautefort, I., Mayer, U., Wileman, T., Stentz, R. and Carding, S.R., 2020. The uptake, trafficking, and biodistribution of Bacteroides thetaiotaomicron generated outer membrane vesicles. Frontiers in microbiology11, p.57.

Kim, J.H., Jeun, E.J., Hong, C.P., Kim, S.H., Jang, M.S., Lee, E.J., Moon, S.J., Yun, C.H., Im, S.H., Jeong, S.G. and Park, B.Y., 2016. Extracellular vesicle–derived protein from Bifidobacterium longum alleviates food allergy through mast cell suppression. Journal of Allergy and Clinical Immunology137(2), pp.507-516.

Kulp, A. & Kuehn, M.J., 2010. Biological functions and biogenesis of secreted bacterial outer membrane vesicles. Annual review of microbiology, 64, pp.163–184.

Molina-Tijeras, J.A., Gálvez, J. & Rodríguez-Cabezas, M.E., 2019. The immunomodulatory properties of extracellular vesicles derived from probiotics: a novel approach for the management of gastrointestinal diseases. Nutrients, 11(5), p.1038.

Müller, L., Kuhn, T., Koch, M. and Fuhrmann, G., 2021. Stimulation of probiotic bacteria induces release of membrane vesicles with augmented anti-inflammatory activity. ACS Applied Bio Materials4(5), pp.3739-3748.

Ñahui Palomino, R.A., Vanpouille, C., Costantini, P.E. and Margolis, L., 2021. Microbiota–host communications: Bacterial extracellular vesicles as a common language. PLoS Pathogens17(5), p.e1009508.

Nishiyama, K., Takaki, T., Sugiyama, M., Fukuda, I., Aiso, M., Mukai, T., Odamaki, T., Xiao, J. Z., Osawa, R., & Okada, N. 2020. Extracellular vesicles produced by Bifidobacterium longum export mucin-binding proteins. Applied and Environmental Microbiology, 86(19), e01464-20.

Palomino, R.A.Ñ., Vanpouille, C., Laghi, L., Parolin, C., Melikov, K., Backlund, P., Vitali, B. and Margolis, L., 2019. Extracellular vesicles from symbiotic vaginal lactobacilli inhibit HIV-1 infection of human tissues. Nature communications10(1), pp.1-14.

Park, J.Y., Choi, J., Lee, Y., Lee, J.E., Lee, E.H., Kwon, H.J., Yang, J., Jeong, B.R., Kim, Y.K. and Han, P.L., 2017. Metagenome analysis of bodily microbiota in a mouse model of Alzheimer disease using bacteria-derived membrane vesicles in blood. Experimental neurobiology26(6), p.369.

Shehata, M.M., Mostafa, A., Teubner, L., Mahmoud, S.H., Kandeil, A., Elshesheny, R., Boubak, T.A., Frantz, R., Pietra, L.L., Pleschka, S. and Osman, A., 2019. Bacterial outer membrane vesicles (omvs)-based dual vaccine for influenza a h1n1 virus and mers-cov. Vaccines7(2), p.46.

Tan, K., Li, R., Huang, X. and Liu, Q., 2018. Outer membrane vesicles: current status and future direction of these novel vaccine adjuvants. Frontiers in microbiology9, p.783.

Tian, Y., Gong, M., Hu, Y., Liu, H., Zhang, W., Zhang, M., Hu, X., Aubert, D., Zhu, S., Wu, L. and Yan, X., 2020. Quality and efficiency assessment of six extracellular vesicle isolation methods by nano-flow cytometry. Journal of extracellular vesicles9(1), p.1697028.

West, C.L., Stanisz, A.M., Mao, Y.K., Champagne-Jorgensen, K., Bienenstock, J. and Kunze, W.A., 2020. Microvesicles from Lactobacillus reuteri (DSM-17938) completely reproduce modulation of gut motility by bacteria in mice. PloS one15(1

New ISAPP-led paper calls for investigation of evidence for links between live dietary microbes and health

The past two decades have brought a massive increase in knowledge about the human gut microbiota and its links to human health through diet. And although many people perceive that regular consumption of safe, live microbes will benefit their health, the scientific evidence to date has not been sufficiently developed to justify adding a daily recommended intake of live microbes to food guides for different populations.

Recently, a group of seven scientists, including six ISAPP board members, published their perspective about the value of establishing the link between live dietary microbes and health. They conclude that although the scientific community has a long way to go to build the evidence base, efforts to do this are worthwhile.

The collaboration on this review was rooted in an ISAPP expert discussion group held at the 2019 annual meeting in Antwerp, Belgium. During the discussion, various experts presented evidence from their fields—addressing the potential health benefits of live microbes in general, rather than the narrow group of microbial strains that qualify as probiotics.

Below, the authors of this new review answer questions about their efforts to quantify the relationship between greater consumption of live microbes and human health.

Why is it interesting to look at the potential importance of live microbes in nutrition?

Prof. Joanne Slavin, PhD, RD, University of Minnesota

Current recommendations for fiber intake are based on protection against cardiovascular disease—so can we do something similar for live microbes? We know that intake of live microbes is thought to be health promoting, but actual recommended intakes for live microbes are missing.  Bringing together a talented group of microbiologists, epidemiologists, nutritionists, and food policy experts moves this agenda forward.

Humans need proper nutrition to survive, and a lack of certain nutrients creates a ‘deficiency state’. Is this the case for live microbes?

Dr. Mary Ellen Sanders, PhD, ISAPP Executive Science Officer

I don’t think we’ll find that live microbes are essential in the same way that vitamins and minerals lead to deficiency diseases. After all, gnotobiotic animal colonies are viable. But I believe there is enough evidence to suggest that consumption of live microbes will promote health. Exactly how and to what extent remains to be established.

Why think about intake of ‘live microbes’ in general, rather than intake of probiotic & fermented foods specifically?

Prof. Maria Marco, PhD, University of California Davis

We are constantly exposed to microorganisms in our foods and beverages, in the air, and on the things we touch. While much of our attention has been on the microbes that can cause harm, most of our microbial exposures may not affect us at all or, quite the opposite, be beneficial for maintaining and improving health. Research on probiotic intake as a whole supports this possibility. However, probiotic-containing foods and dietary supplements are only a part of our dietary connection with live microbes. Non-pasteurized fermented foods (such as kimchi and yogurts) can contain large numbers of non-harmful bacteria (>10^7 cells/g). Fruits and vegetables are also sources of living microbes when eaten raw.  Although those raw foods they may contain lower numbers of microbes, they may be more frequently eaten and consumed in larger quantities. Therefore, our proposal is that we take a holistic view of our diets when weighing the potential significance of live microbe intake on health and well-being.

What are dietary sources of live microbes? And do we get microbes in foods besides fermented & probiotic foods?

Prof. Bob Hutkins, PhD, University of Nebraska Lincoln

For tens of thousands of years, humans consumed large amounts of microbes nearly every time they ate food or drank liquids. Milk, for example, would have been unheated and held at ambient temperature with minimal sanitation and exposed to all sorts of microbial environments.  Thus, a cup of this milk could easily have contained millions of bacteria. Other foods like fruits and vegetables that were also exposed to natural conditions could have also contained similar levels of microbes. Even water would have contributed high numbers of live microbes.

Thanks to advances in food processing, hygiene, and sanitation, the contemporary western diet generally contains low levels of microbes. Consider how many foods we eat that are canned, pasteurized, or cooked – those foods will contain few, in any live microbes. Fresh produce can serve as a source of live microbes, but washing, and certainly cooking, will reduce those levels.

For sure, the most reliable sources of dietary microbes are fermented foods and beverages. Even if a fresh lettuce salad were to contribute a million bacteria, a single teaspoon of yogurt could contain 100 times more live bacteria. Other popular fermented foods like kefir, kimchi, kombucha, and miso, can contain a large and relatively diverse assortment of live microbes. Other fermented foods, such as cheese and sausage, are also potential sources, but the levels will depend on manufacturing and aging conditions. Many fermented, as well as non-fermented foods are also supplemented with probiotics, often at very high levels.

What’s the evidence that a greater intake of live microbes may lead to health benefits?

Prof. Dan Merenstein, MD, Georgetown University

Studies have shown that fermented foods are linked to a reduced risk of cardiovascular disease, reduced risk of weight gain, reduced risk of type 2 diabetes, healthier metabolic profiles (blood lipids, blood glucose, blood pressure and insulin resistance), and altered immune responses. This link is generally from associative studies on certain fermented foods. Many randomized controlled trials on specific live microbes (probiotics and probiotic fermented foods) showing health benefits have been conducted, but randomized controlled trials on traditional fermented foods (such as kimchi, sauerkraut, kombucha) are rare. Further, no studies have aimed to assess the specific contribution of safe, live microbes in diets as a whole on health outcomes.

Why is it difficult to interpret past data on people’s intake of live microbes and their health?

Prof. Colin Hill, PhD, University College Cork

It would be wonderful if there were a simple equation linking the past intake of microbes in the diet and the health status of an individual (# MICROBES x FOOD TYPE = HEALTH). In reality, this is a very complex challenge. Microbes are the most diverse biological entities on earth, our consumption of microbes has not been deliberately recorded and can only be estimated, and even the concept of health has defied precise definitions for centuries. To further confuse the situation microbes meet the host in the gastrointestinal tract, the site of our enormously complex mucosal immune system and equally complex microbiome.  But the complexity of the problem should not prevent us from looking for prima facie evidence as to whether or not such a relationship is likely to exist.

Databases of dietary information have data on people’s intake of live microbes, but what are the limitations of our available datasets?

Prof. Dan Tancredi, PhD, University of California Davis

Surveys often rely on food frequency questionnaires or diaries to determine consumption of specific foods. These are notoriously prone to recall error and/or other types of measurement error. So, even just measuring consumption of foods is difficult. For researchers seeking to quantify survey respondents’ consumption of live microbes, these challenges become further aggravated because the respondents would not typically know the microbial content in the foods they consumed. Instead, we would have to have them tell us the types and amounts of the foods they ate, and then we would need to translate that into approximate microbial counts—but even within a particular food, the microbial content can vary, depending on how it was processed, stored, and/or prepared prior to consumption.

See ISAPP’s press release on this paper here.

Current status of research on probiotic and prebiotic mechanisms of action

By Mary Ellen Sanders, PhD, ISAPP Executive Science Officer

Human intervention studies in the fields of probiotics and prebiotics assess the health effects of these ingredients, whether it’s improving specific symptoms or preventing the occurrence of a health condition. Yet scientists in the field recognize the importance of learning the ‘chain of events’ by which probiotics and prebiotics are able to confer health benefits. Such mechanistic insights allow better probiotic selection and development of therapeutic approaches, as well as more precise dosing.

Mechanisms of action for probiotics and prebiotics are complex and often difficult to pinpoint, especially since any given health benefit may derive from multiple co-functioning mechanisms. However, scientists have made incremental gains in understanding these mechanisms. This scientific progress was covered in a recent webinar co-presented by ISAPP and ILSI-Europe, titled Understanding Prebiotic and Probiotic Mechanisms that Drive Health Benefits. Speakers for the webinar were:

  • Sarah Lebeer, University of Antwerp, Belgium
  • Colin Hill, University College Cork, Ireland
  • Karen Scott, University of Aberdeen, UK
  • Koen Venema, Maastricht University – campus Venlo, The Netherlands

The webinar was held live on September 17, 2020. Of the 499 webinar registrants, 357 attended the webinar live from 57 countries, from Australia to the US. ISAPP and ILSI-Europe hope the webinar will serve as a resource for people who want a rapid overview about mechanisms of action.

Watch the full webinar here, and read further for a summary of key points from these experts.

Prebiotic benefits and mechanisms of action

Prebiotics are defined as substrates that are “selectively utilized by host microorganisms conferring a health benefit”. ‘Utilization’ in the gut may involve crossfeeding, which means products produced by the first microbes degrading the prebiotic can then be used by different members of the host microbiota – so it may take a series of complex steps to get to a final health outcome. However, selective utilization and health benefit are always required for a substance to meet the definition of a prebiotic.

The health benefit of a prebiotic can be local (in the gut) or systemic. Locally, prebiotics can act via fecal bulking, as they are typically types of fiber. In addition, they can produce short-chain fatty acids (SCFAs), which reduce gut pH and thereby can discourage pathogenic and toxigenic activity of gut microbes, increase calcium ion absorption and provide energy for gut epithelial cells.

Systemic functions of prebiotic metabolism include them being used as substrates for microbes that produce or interact with host cells to produce molecules with neurochemical, metabolic or immune activity. Further, SCFAs can end up in the blood and can reach the liver, muscles and the brain. The SCFAs interact with specific host receptors and can lead to the release of satiety hormones or interact with receptors in the liver, adipose tissue and muscle tissue, leading to reduced inflammation. Prebiotics can also interact directly with immune cells.

Probiotic health effects and mechanisms of action

Health and disease are the end results of complex interactions on a molecular scale within a human or animal host.  Host molecules also interact with microbial molecules, including those molecules introduced with or produced by probiotics. Designing studies to discover probiotic mechanisms in human research is extremely challenging because both host and probiotic are very complex systems that most probably engage with one another on multiple levels. Probiotic molecules can have direct effects and downstream effects, and we are aware of only a few cases where a health effect can be tied to one specific probiotic molecule.

Probiotics can interact directly with the host, but also can act indirectly by influencing the microbiome. There may be many different mechanisms by which a given probiotic interacts with the host.

It is interesting to note that probiotics use some of the same types of mechanisms (pili, small molecule production, etc.) that are used by pathogens, microbes that have a detrimental effect on the host.  But these shared mechanisms are usually connected to surviving or colonising strategies, not those that cause damage to the host.

L. rhamnosus GG is a well-researched model probiotic, for which many mechanisms have been identified, including pili, immune modulators and lactic acid production, some mechanisms shared with other probiotic strains and species. Other studies have identified mechanisms for novel types of probiotics. For example, in mice and humans taking a strain of Akkermansia, heat killed cells had the same or even better effect on markers of metabolic health, which implies that the molecules (perhaps proteins in the bacteria, unaffected by heat treatment) are mediating the effect in this case.

See here to watch the webinar in full.

 

 

ISAPP partners with British Nutrition Foundation for fermented foods webinar

Did you miss the live webinar? Access the archived version here. Read the speaker Q&A here.

From sourdough starter tips to kombucha flavor combinations – if you’ve checked a social media feed lately, you’ll know how many people are sharing an interest in fermented foods as they self-isolate during the pandemic. And with this rise in popularity comes a host of questions about the practice and the science of fermented foods.

To meet the need for science-based information about fermented foods, ISAPP has partnered with the British Nutrition Foundation (BNF) on a free webinar titled ‘Fermented Food – Separating Hype from Facts.’ The BNF is a UK-based registered charity that brings evidence-based information on food and nutrition to all sectors, from academia to medicine.

The webinar, designed for practicing dietitians and nutrition-savvy members of the public, featured three leading scientific experts who explained the microbiology of fermented foods, the evidence for their health effects, and who might benefit from making these foods a regular part of the diet. Viewers will come away with a clear understanding of what fermented foods are and what evidence exists for their health benefits.

The webinar was held live on Wednesday, July 1, 2020 from 1pm-2pm (BST).

Webinar speakers & topics

 Understanding fermented foods: Dr. Robert Hutkins, University of Nebraska, USA

Exploring the evidence for effects of fermented foods on gastrointestinal health – how strong is it? Dr. Eirini Dimidi, Kings College London

What role can fermented foods have in our diet? A public health perspective, Anne de la Hunty, British Nutrition Foundation

For a quick primer on fermented foods, see the short ISAPP video here or the ISAPP infographic here.

Is probiotic colonization essential?

By Prof. Maria Marco, PhD, Department of Food Science & Technology, University of California, Davis

It is increasingly appreciated by consumers, physicians, and researchers alike that the human digestive tract is colonized by trillions of bacteria and many of those bacterial colonists have important roles in promoting human health. Because of this association between the gut microbiota and health, it seems appropriate to suggest that probiotics consumed in foods, beverages, or dietary supplements should also colonize the human digestive tract. But do probiotics really colonize? What is meant by the term “colonization” in the first place? If probiotics don’t colonize, does that mean that they are ineffective? In that case, should we be searching for new probiotic strains that have colonization potential?

My answer to the first question is no – probiotics generally do not colonize the digestive tract or other sites on the human body. Before leaping to conclusions on what this means for probiotic efficacy, “colonization” as defined here means the permanent, or at least long-term (weeks, months, or years) establishment at a specific body site. Colonization can also result in engraftment with consequential changes to the gut microbiota composition and function. For colonization to occur, the probiotic should multiply and form a stably replicating population. This outcome is distinct from a more transient, short-term (a few days to a week or so) persistence of a probiotic. For transient probiotics, it has been shown in numerous ways that they are metabolically active in the intestine and might even grow and divide. However, they are not expected to replicate to high numbers or displace members of the native gut microbiota.

Although some studies have shown that digestive tracts of infants can be colonized by probiotics (weeks to months), the intestinal persistence times of probiotic strains in children and adults is generally much shorter, lasting only few days. This difference is likely due to the resident gut microbiota that develops during infancy and tends to remain relatively stable throughout adulthood. Even with perturbations caused by antibiotics or foodborne illness, the gut microbiome tends to be resilient to the long-term establishment of exogenous bacterial strains. In instances where probiotic colonization or long-term persistence was found, colonization potential has been attributed more permissive gut microbiomes specific to certain individuals. In either case, for colonization to occur, any introduced probiotic has to overcome the significant ecological constraints inherent to existing, stable ecosystems.

Photo by http://benvandenbroecke.be/ Copyright, ISAPP 2019.

This leads to the next question: Can probiotics confer health benefits even if they do not colonize? My answer is definitely yes! Human studies on probiotics with positive outcomes have not relied on intestinal colonization by those microbes to cause an effect. Instead of colonizing, probiotics can alter the digestive tract in other ways such as by producing metabolites that modulate the activity of the gut microbiota or stimulate the intestinal epithelium directly. These effects could happen even on short-time scales, ranging from minutes to hours.

Should we be searching for new probiotic strains that have greater colonization potential? By extension of what we know about the resident human gut microbiota, it is increasingly attractive to identify bacteria that colonize the human digestive tract in the same way. In some situations, colonization might be preferred or even essential to impacting health, such as by engrafting a microbe that performs critical metabolic functions in the gut (e.g. break down complex carbohydrates). However, colonization also comes with risks of unintended consequences and the loss of ability to control the dose, frequency, and duration of exposure to that particular microbe.

Just as most pharmaceutical drugs have a transient impact on the human body, why should we expect more from probiotics? Many medications need to be taken life-long in order manage chronic conditions. Single or even repeated doses of any medication are similarly not expected to cure disease. Therefore, we should not assume a priori that any observed variations in probiotic efficacy are due to a lack of colonization. To the contrary, the consumption of probiotics could be sufficient for a ripple effect in the intestine, subtly altering the responses of the gut microbiome and intestinal epithelium in ways that are amplified throughout the body. Instead of aiming for engraftment directly or hand-wringing due to a lack of colonization, understanding the precise molecular interactions and cause/effect consequences of probiotic introduction will lead to a path that ultimately determines whether colonization is needed or just a distraction.

blog post resilience figure 1

Resilience as a measure of health: implications for health claims for foods

January 16, 2018. By Mary Ellen Sanders PhD, Sylvie Binda PhD, Seppo Salminen PhD, Karen Scott PhD

Demonstrating health benefits for healthy people is a challenge faced by those attempting to communicate claims on a health promoting food. Foods, in many global regulatory frameworks, are intended for the general population. Therefore, any benefits ascribed to them, the logic goes, must be demonstrated in the generally healthy population.

An old concept has new-found notoriety in the context of offering an approach for establishing health benefits for healthy people. It is the concept of resilience. In an ecological sense, resilience refers to the ability of an ecosystem to withstand perturbation and continue normal function, i.e. maintain homeostasis. In the context of human physiology, resilience enables a host to remain healthy even when exposed to a stress, or to recover from a stress faster. A variety of external challenges such as drugs, pathogens, emotional stress, poor diet among others, may perturb normal physiological function or disrupt the gut ecosystem. Individuals more able to maintain stability of physiological functions when exposed to such challenges would be healthier than those who cannot maintain stability.  Thus, a food would be considered to have a beneficial effect if it could increase the resilience of the consumer to a challenge.

This concept was described in an EFSA guidance document on biological relevance of data in scientific assessments:

“When subject to a disturbance, a biological system enters in a transient state: a process variable has been changed and the system has not yet reached steady state. Some systems, including humans, have the capacity to regulate their internal environment and to maintain a stable, relatively constant condition of properties; it is called ‘homeostatic capacity’. Resilience represents the amount of disturbance that can be absorbed by a system before the system changes or loses its normal function, or the time taken to return to a stable state, within the normal operation range following the disturbance…” [Reducing] “homeostatic capacity … might be detrimental, whereas increasing the capacity could be beneficial.”

This concept aligns with the definition of ‘health’, which includes the ability to adapt to the environment.

Resilience of gut microbiota

This concept of resilience can be applied to the human microbiota as an ecosystem. Once established in early childhood, our colonizing microbiota reaches a relatively stable state. Although brief fluctuations occur, especially in relation to daily diet and medicines used, the microbial ecosystem of a healthy adult provides relatively stable functionality.  Disruption of the microbiota by repeated stressors can be associated with poorer health. There seems to be a solid rationale that the ability of the colonizing microbiota to resist, or recover quickly from, perturbations reflects a person’s ability to remain healthy. The microbiota stability may be indicated in either populations of bacteria or their metabolic output.

Homeostasis and health: a statistical approach

“A statistical approach to measuring improved health was proposed by Dr. Dan Tancredi at the 2010 ISAPP meeting. It is reprinted here from: Sanders, et al. 2011. Health claims substantiation for probiotic and prebiotic products. Gut Microbes 2:3, 1-7.

An approach to measuring improved health may be to measure homeostasis, as suggested by D. Tancredi. From a statistical point of view, if an intervention were able to minimize the variation around the mean for a specific measure (even in the absence of changing the mean; Fig. 1), it could be a reflection of improved health, assuming a biological rationale exists that tighter control of the parameter is physiologically advantageous. In other words, lessening the fluctuation around an individual’s biomarker could be interpreted as contributing to improving health. This novel idea emphasizes the importance of homeostasis as a focus of studies on health, and provides a rationale based in solid statistical theory as a way to measure this.

One challenge to demonstrating the value of this approach is to identify appropriate biomarkers that could be studied. The following properties would be important to a relevant biomarker for homeostasis:

blog resilience figure one

  • maintaining moderate levels of the biomarker is associated with good health;
  • high or low values are associated with ill health;
  • biomarker levels in the same person can fluctuate over time; and
  • reducing the magnitude or duration of such fluctuations in healthy people is considered desirable (Fig. 2).

Such a biomarker could be an individual endpoint or be formed as a ratio of two other biomarkers, when maintaining the same relative amounts of the two component biomarkers would be desirable.

Assuming a biomarker with the above properties is available, it could be used as the outcome measure in a randomized controlled trial to provide evidence that the experimental food is able to improve the maintenance of health in humans. Statistically, the trial would be set up to address the hypothesis that the experimental substance is associated with lower variation in biomarker levels, compared to the control arm, in subjects who were healthy at baseline. Such a trial would be able to use information on within-person variations in biomarker levels, even those who did not become ill. Partly as a result of the more efficient use of study data, such a trial would require far fewer subjects than an intervention that instead addressed the hypothesis that treatment is associated with fewer healthy persons becoming ill.

A mounting understanding of the value of stability of the colonizing microbial communities makes this endpoint an attractive one to consider. Perturbation of gut microbiota is associated with intestinal dysfunction, as illustrated during antibiotic treatment. Specific probiotics have been shown to promote a quicker rebound from antibiotic-induced microbiota disruption, including a study on Lactobacillus rhamnosus GG (LGG) (Cox et al. 2000). This paper concludes ‘…that a key mechanism for the protective effect of LGG supplementation on the subsequent development of allergic disease is through the promotion of a stable, even and functionally redundant infant gastrointestinal community.’

However, it would be useful to define additional biomarkers that would be appropriate targets for this type of investigation.

In pediatric nutrition, the measurement of metabolic homeostasis has become a standard approach when developing infant formulas (Heird, 2005).  The concept of homeostasis as a model to distinguish between foods (including food supplements) and medicinal products was explored by the Council of Europe (2011), and is an interesting correlate to the above hypothesis.”

Conclusions

The recent recognition by EFSA that maintenance of homeostasis is a valid measure of health provides an opportunity to apply this concept to validate health benefits of specific foods and food ingredients. Stability of microbial populations, microbial metabolism or host physiological readouts could be measured to reflect the concept of resilience. Since there is no definitive composition of a ‘healthy human microbiota’, a more reasonable target for measuring positive impacts of a probiotic on the microbiota would be reflected not in absolute levels of specific microbes but in the ability of a specific probiotic or prebiotic to bolster the resilience of the microbiota.

 

References:

Council of Europe. Homeostasis, a model to distinguish between foods (including nutritional supplements) and medicinal products 2008; (Accessed February 24, 2011, at http://www.coe.int/t/e/social_cohesion/soc-sp/homeostasis%20%282%29.pdf ).

Cox MJ, Huang YJ, Fujimura KE, Liu JT, McKean M, Boushey HA, et al. Lactobacillus casei abundance is associated with profound shifts in theGunderson LH, 2000. Ecological resilience: in theory and application. Annual Review of Ecology and Systematics, 31, 425–439.

EFSA guidance document:  Guidance on the assessment of the biological relevance of data in scientific assessments; July 12, 2017; EFSA Journal 2017;15(8):4970

Heird WC. Biochemical homeostasis and body growth are reliable end points in clinical nutrition trials. Proceedings of the Nutrition Society 2005; 64:297-303.

Huber M, Knottnerus JA, Green L, van der Horst H, Jadad AR, Kromhout D, Leonard B, Lorig K, Loureiro MI, van der Meer JW, Schnabel P, Smith R, van Weel C, Smid H (2011). “How should we define health?” BMJ. 343:d4163.

Sanders, et al. 2011. Health claims substantiation for probiotic and prebiotic products. Gut Microbes 2:3, 1-7; May/June 2011