Are bacteria important? What do we get by analyzing microbiomes?

By Leila Lemos, PhD candidate, Fisheries and Wildlife Department, OSU

As previously mentioned in one of Florence’s blog posts, the GEMM Lab holds monthly lab meetings, where we share updates about our research and discuss articles and advances in our field, among other activities.

In a past lab meeting we were asked to bring an article to discuss that had inspired us in the past to conduct research in the marine field or in our current position. I brought to the meeting a literature review regarding methodologies to overcome the challenges of studying conservation physiology in large whales [1]. This article discusses different non-invasive or minimally invasive matrices (e.g., feces, blow, skin/blubber) that can be gathered from whales, and what types of analyses could be carried out, as well as their pros and cons.

One of the possible analyses that can be performed with fecal samples that was discussed in the article is the gut microflora (i.e., bacterial gut community) via genetic analysis. Since my PhD project analyzes fecal samples to determine/quantify stress responses in gray whales, we have since discussed the possibility of integrating this extra parameter to our analysis.

But… what is the importance of analyzing the gut microflora of a whale? What is the relationship between microflora and stress responses? Should we really use our limited sample size, time and money to work on this extra analysis? In order to be able to answer all of these questions, I began reading some articles of the field to better understand its importance and what kind of research questions this analysis can answer.

The gut of a mammal comprises a natural habitat for a large and dynamic community of bacteria [2] that is first developed in early life. Colonization of facultative bacteria (i.e., aerobic bacteria) begins at birth [3], and later, anaerobic bacteria also colonizes the gut. In humans, at the age of 1 year old, the microbiome should have a stable adult-like signature (Fig. 1).

Figure 01: Development of the microbiome in early life.
Source: [3]

The gut bacterial community is important for the physiology and pathology of its host and plays an important role in mammal digestion and health [2], responsible for many metabolic activities, including:

  • fermentation of non-digestible dietary residue and endogenous mucus [2];
  • recovery of energy [2];
  • recovery of absorbable nutrients [2];
  • cellulose digestion [4];
  • vitamin K synthesis [4];
  • important trophic effects on intestinal epithelia (cell proliferation and differentiation) [2];
  • angiogenesis promotion [4];
  • enteric nerve function [4];
  • immune structure [2];
  • immune function [2];
  • protection of the colonized host against invasion by alien microbes (barrier effect) [2];

Despite all the benefits, the bacterial community might also be potentially harmful when changes in the community composition (i.e., dysbiosis) occur due to the use of antibiotics, illness, stress, aging, lifestyle, bad dietary habits [4], and prolonged food and water deprivation [5]. Thus, potential pathological disorders might emerge when the microbiome community changes, such as allergy, obesity, diabetes, autism, multisystem organ failure, gastrointestinal and prostate cancers, inflammatory bowel diseases (IBD), and cardiovascular diseases [2, 4].

Changes in gut bacterial composition may also alter the brain-gut axis and the central nervous system (CNS) signaling [3]. More specifically, the core pathway affected is the hypothalamic-pituitary-adrenal (HPA) axis, which is activated by physical/psychological stressors. According to a previous study [6], the microbial community in the gut is critical for the development of an appropriate stress response. In addition, the microbial colonization in early life should occur within a certain time window, otherwise an abnormal development of the HPA axis might happen.

However, the gut microbiome can not only affect the HPA axis, but the opposite can also occur [3]. Signaling molecules released by the axis can alter the gastrointestinal (GIT) environment (i.e., motility, secretion, and permeability) [7]. Stress responses, as well as diseases, may also alter the gut permeability, causing the bacteria to cross the epithelial barrier (reducing the overall numbers of bacteria in the gut), activating immune responses that also alter the composition of the bacterial community in the gut [8, 9].

Figure 02: Communication between the brain, gut and microbiome in a healthily and in a stressed or diseased (mucosal inflammation) mammal.
Source: [3]

Thus, when thinking about whales, monitoring of the gut microflora might allow us to detect changes caused by factors such as aging, illness, prolonged food deprivation, and stressful events [2, 5]. However, since these are two-way factors, it is important to find an association between bacterial composition alterations and stressful events, such as the presence of predators (e.g., killer whales), illness (e.g., bad body condition), prolonged food deprivation (e.g., low prey availability and high competition), noise (e.g., noisy vessel traffic, fisheries opening and seismic surveys), and stressful reproductive status (e.g., pregnancy and lactating period). Examination of possible shifts in the gut microflora may be able to detect and be linked to many of these events, and also forecast possible chronic events within the population. In addition, the bacterial community monitoring study could aid in validating the hormone data (i.e., cortisol) we have been working with.

Therefore, the main research questions that arise in this context that can aid in elucidating the stress physiology in gray whales are:

  1. What is the microflora community content in guts of gray whales along the Oregon coast?
  2. Is it possible to detect shifts in the gut microflora from our gray fecal samples over time?
  3. How do gut microflora and cortisol levels correlate?
  4. Am I able to correlate shifts in gut microflora with any of the stressful events listed above?

We can answer so many other questions by analyzing the microbiome of baleen whales. Microbiomes are mainly correlated with host diet [10], so the composition of a microbiome can be associated with specific diets and functional gut capacity, and consequently, be linked to other animal populations, which helps to decode evolutionary questions. Results of a previous study on baleen whale microbiomes [10] point out that whales harbor unique gut microbiomes that are actually similar to those of terrestrial herbivores. Baleen whales and terrestrial herbivores have a shared physical structure of the GIT tract itself (i.e., multichambered foregut) and a shared hole for fermentative metabolisms. The multichambered foregut of baleen whales fosters the maintenance of the gut microbiome that is capable of extracting relatively unavailable nutrients from zooplankton (i.e., chitin, “sea cellulose”).

Figure 03: The similarities between whale and other terrestrial herbivore gut microbiomes: sea and land ruminants.
Source: [11]

Thus, the importance of studying the gut microbiome of a baleen whale is clear. Monitoring of the bacterial community and possible shifts can help us elucidate many questions regarding diet, overall health, stress physiology and evolution. Thinking about my PhD project, it may also help in validating our cortisol level results. I am confident that a microbiome analysis would significantly enhance my studies on the health and ecology of gray whales.

 

References

  1. Hunt, K.E., et al., Overcoming the challenges of studying conservation physiology in large whales: a review of available methods.Conservation Physiology, 2013. 1: p. 1-24.
  2. Guarner, F. and J.-R. Malagelada, Gut flora in health and disease.The Lancet, 2003. 360: p. 512–519.
  3. Grenham, S., et al., Brain–gut–microbe communication in health and disease.Frontiers in Physiology, 2011. 2: p. 1-15.
  4. Zhang, Y., et al., Impacts of Gut Bacteria on Human Health and Diseases.International Journal of Molecular Sciences, 2015. 16: p. 7493-7519.
  5. Bailey, M.T., et al., Stressor exposure disrupts commensal microbial populations in the intestines and leads to increased colonization by Citrobacter rodentium.Infection and Immunity, 2010. 78: p. 1509–1519.
  6. Sudo, N., et al., Postnatal microbial colonization programs the hypothalamic-pituitary-adrenal system for stress response in mice.The Journal of Physiology, 2004. 558: p. 263–275.
  7. Rhee, S.H., C. Pothoulakis, and E.A. Mayer, Principles and clinical implications of the brain–gut–enteric microbiota axis Nature Reviews Gastroenterology & Hepatology, 2009. 6: p. 306–314.
  8. Kiliaan, A.J., et al., Stress stimulates transepithelial macromolecular uptake in rat jejunum.American Journal of Physiology, 1998. 275: p. G1037–G1044.
  9. Dinan, T.G. and J.F. Cryan, Regulation of the stress response by the gut microbiota: Implications for psychoneuroendocrinology.Psychoneuroendocrinology 2012. 37: p. 1369—1378.
  10. Sanders, J.G., et al., Baleen whales host a unique gut microbiome with similarities to both carnivores and herbivores.Nature Communications, 2015. 6(8285): p. 1-8.
  11. El Gamal, A. Of whales and cows: the baleen whale microbiome revealed. Oceanbites 2016[cited 2018 07/31/2018]; Available from: https://oceanbites.org/of-whales-and-cows-the-baleen-whale-microbiome-revealed/.

 

“Evolution”: a board game review

By Florence Sullivan MSc student, Department of Fish and Wildlife.

Another grad student once told me that in order to survive grad school, I would need three things:

(1) an exercise routine, (2) a pet, and (3) a hobby. My Pilates class on Wednesdays is a great mid-week reminder to stretch. I don’t have a pet, so that advice gets fulfilled vicariously through friends. As for my hobby, I think you’ll find that even when scientists take a break from work, we really don’t get that far away from the subject matter…..

Board games have evolved significantly since the early ‘90s when I grew up on such family staples as Monopoly, Risk, Sorry!, Candyland, and Chutes and Ladders, etc. Now, table-top games tend to fall into three loose categories – “Euro-games” that focus on strategy and economic themes as well as keeping all players in the game until the end, “American-style” that tend toward luck and direct player contact so that not everyone plays until the end, and “Party” that are easy to learn and are often played in large groups as social icebreakers or to provide entertainment.

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A few of my favorite games.

As board games proliferate, we see the use of many themes and often, there are valuable educational lessons included in the game design!  There are militaristic or survival games (Betrayal at the House on the Hill, Dead of Winter), economic and engineering (Settlers of Catan, Istanbul, Ticket to ride, Carcassonne), fantasy and art (Small World, Dixit), cooperative vs competitive (Hanabi, Forbidden Desert vs. 7 Wonders), and some of my favorites – the sciences (Compounded, Bioviva, Pandemic).

Today, let’s talk about my current favorite – Evolution. It is immediately obvious that the game designers responsible are either giant nerds (I use this in the most loving way possible) or have spent some quality time with ecologists.  Not only is the art work beautiful, and the game play smooth, but the underlying mechanics allow serious ecological theories such as ‘predator and prey mediated population cycles’, ‘co-evolution’ and ‘evolutionary arms-races’ to be acted out and easily understood.

Players set up their species around the watering hole, and contemplate their next moves.
Players set up their species (1 green/yellow tile = 1 species) around the watering hole, and contemplate their next moves.

In game play, as in life, the point of the game is to eat – victory is achieved by the player who has managed to ‘digest’ the most food tokens. All players begin with a single species, and with each turn, can either add traits (ie. fat tissue, scavenger, etc.) to the species, increase the body size of a species, gain a population level, or gain additional species.  Next, players take food from a limited, random supply until there is no food left. Species that have not been fed to their full capacity (population levels) will starve, and can even become extinct – much like the reality of environmental cycles.  Finally, all food that has been ‘eaten’ is digested, and the next round begins.

Since a player can never be sure how much food will appear on the watering hole each turn, it is a good strategy to capitalize on traits like foraging which allows a species to take twice as much food every time it feeds.  If your species cooperates with another, that means that it gets to eat every time you feed the first species. A player who combines foraging traits with multiple cooperating species in a “cooperation chain” can quickly empty the watering hole before any other players get a chance.  Much like a species perfectly adapted to its niche in the real world will out compete more generalist species.

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The pack-hunting carnivore on the left can easily take down the fertile defensive herding species in the upper right. The efficient foraging species in the middle is protected by its horns, and cooperates with the next species to the right. The burrowing species is protected from carnivores only as long as it is full (and presumably no longer needs to venture out of its burrow).

One way to avoid the competition for food at the watering hole is to play the carnivore trait.  This species must now consume other species in order to feed itself.  A few caveats; a carnivore must be larger in body size than anything it tries to eat, and can no longer eat plant food as it is an obligate carnivore. As soon as a carnivore appears on the board, the evolutionary arms-race begins in earnest!  Traits such as burrowing, climbing, hard shells, horns, defensive herding and warning calls become vital to survival.  But carnivores can be clever, and apply ambush to species with warning call, or pack-hunting to a species with defensive herding.  In everything, there is a certain balance, and quickly, players will find themselves acting out a classic ‘boom and bust population growth cycle’ scenario, where herbivores go extinct due to low food supply at the watering hole and/or high predation pressure, and carnivores soon follow when there are no un-protected species for them to feed upon.

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A flying creature must first pay the ‘upkeep cost’ of its body size in food, before it can feed its population. Good thing it has the extra cliff-side food source that is only accessible to other species with wings!

An expansion has been released for the game – it is called Flight – and introduces traits such as flight, camouflage, good eyesight, and others.  From an ecologist’s perspective, it fits the original game well both scientifically and thematically.  To achieve flight, a higher price must be paid (in terms of cards discarded) to gain the trait card, and unlike other species, an ‘upkeep cost’ must be gathered in food tokens before the species actually eats any food tokens during the round.  However, flight also gives access to a cliff-side watering hole that is not accessible to earthbound species. This neatly mirrors the real world where flight is an energetically costly activity that also opens new niches.

The next expansion is just arriving in stores, and I can’t wait to play it! It’s called Climate, and adds traits such as nocturnal, claws, and insectivore. Perhaps more exciting though, are the ‘event cards’ which will trigger things like desertification, cold snaps, heatwaves, volcanic eruptions and meteor strikes. A climate tracker will keep track of whether the planet is in an ice age or a warming period, and certain traits will make your species more or less likely to survive – can you guess which ones might be useful in either scenario? I think it will be enormously fun to play through different climate scenarios and see how traits stack and species interactions evolve.  Perhaps this new addition to the game will even cause a new game review in Nature – check out their initial assessment here: http://www.nature.com/nature/journal/v528/n7581/full/528192a.html

Games like evolution are useful thought exercises for students and researchers because they promote discussion of adaptive traits, predator-prey cycles, climate, and ecosystem dynamics as related to our own projects. Watching a story unfold in front of you is a great way to truly understand some of the core principles of ecology (and other subjects). This is especially relevant in the GEMM lab where we continuously ask ourselves why our study species act the way they do? How do they find prey, and how are/will they adapt(ing) to our changing climate?