I Contain Multitudes: The Microbes Within Us and a Grander View of Life

by Ed Yong

the world of symbiosis is one in which our allies can disappoint us and our enemies can rally to our side. It’s a world where mutualisms shatter for the matter of a few millimetres.

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Why do microbes so easily slide between pathogen and mutualist? For a start, these roles are not as contradictory as you might imagine. Think about what a ‘friendly’ gut microbe needs to set up a stable relationship with its host. It must survive in the gut, anchor itself so it doesn’t get swept away, and interact with its host’s cells. These are all things that pathogens must do, too. So both characters – mutualists and pathogens, heroes and villains – often use the same molecules for the same purposes.

Some of these molecules get saddled with negative names, like ‘virulence factors’, because they were first discovered in the context of disease, but they are inherently neutral. They are just tools, like computers, pens, and knives: they can be used to do wonderful things and terrible things.

Almost every major partnership in the natural world is like this. Cheats are always a problem. Betrayal lurks perpetually on the horizon. Couples might work well together, but if one partner can get the same benefits without spending as much energy or effort, it will do so unless punished or policed. H. G. Wells wrote about this in 1930: ‘Every symbiosis is, in its degree, underlain with hostility, and only by proper regulation and often elaborate adjustment can the state of mutual benefit be maintained. Even in human affairs, the partnerships for mutual benefit are not so easily kept up, in spite of me being endowed with intelligence and so being able to grasp the meaning of such a relation. But in lower organisms, there is no such comprehension to help keep the relationship going. Mutual partnerships are adaptations as blindly entered into and as unconsciously brought about as any others.’

We like our black-and-white narratives, with clear heroes and villains. In the last few years, I’ve seen the viewpoint that ‘all bacteria must be killed’ slowly give ground to ‘bacteria are our friends and want to help us’, even though the latter is just as wrong as the former. We cannot simply assume that a particular microbe is ‘good’ just because it lives inside us. Even scientists forget this.

Take oxpeckers. These brown birds can be found in Africa, clinging to the flanks of giraffes and antelope. They’re classically viewed as cleaners that pick ticks and blood-sucking parasites off their hosts. But they also peck at open wounds – a less helpful habit that stymies the healing process and increases the risk of infection. These birds crave blood, and they satisfy that craving in ways that either profit their hosts, or punish them.

A similar dynamic goes on in coral reefs, where a small fish called the cleaner wrasse runs a health spa. Big fish arrive, and the wrasse picks parasites from their jaws, gills, and other hard-to-reach places. The cleaners get meals, and the clients get healthcare. But the cleaners sometimes cheat by nipping bits of mucus and healthy tissue. The clients punish them by taking their business elsewhere, and the cleaners themselves will castigate any colleagues that annoy potential customers.

Meanwhile, in South America, acacia trees rely on ants to defend them from weeds, pests, and grazers. In return, they give their bodyguards sugary snacks to eat and hollow thorns to live in. It looks like an equitable relationship, until you realise that the tree laces its food with an enzyme that stops the ants from digesting other sources of sugar. The ants are indentured servants. All of these are iconic examples of cooperation, found in textbooks and wildlife documentaries. And each of them is tinged with conflict, manipulation, and deceit.

wow. Acadia, sugar and ants.

How, in other words, do I contain my multitudes? Containing our multitudes is not unlike a bit of agriculture. We use fences and barriers to mark the boundaries of our gardens. We use fertiliser to feed the plants. We uproot and poison incipient weeds. And we set the garden in a place with the right temperature, soil, and levels of sunlight to nourish whatever we want to grow. Animals use equivalents of all of these measures to set the terms and conditions for their microbial partnerships.17 We will meet each one in turn.

To begin with, every body part on every species has its own zoological terroir – its unique combination of temperature, acidity, oxygen levels, and other factors that dictate what kinds of microbe can grow there. The human gut might seem like nirvana for microbes, with its regular baths of food and fluid. But it is a challenging environment, too. That food supply comes in a fast-flowing torrent, so microbes need to grow quickly or carry molecular anchors to maintain a foothold. The gut is a dark world, so microbes that depend on sunlight to make their food cannot thrive. It lacks oxygen, which explains why the overwhelming majority of gut microbes are anaerobes – organisms that ferment their food, and grow without this supposedly essential gas. Some of them are so reliant on the absence of oxygen that they die in its presence.

molecular anchors to maintain foothold. anaerobes (organisms that ferment their food)

The skin is different: it varies from cool, dry deserts like the forearm to warm, humid jungles like the groin or armpits. Sunlight is abundant, but is also a problem because of the ultraviolet radiation it contains. Oxygen matters here too, and since most of the skin is exposed to fresh air, aerobes thrive.

concealed niches, like sweat glands, can support the growth of oxygen-hating anaerobes like Propionibacterium acnes, the microbe that causes acne. All over our bodies, the laws of physics and chemistry sculpt bundles of biology.

Propionibacterium acnes

Bacteriocytes have repeatedly evolved in different lineages. Some insects slot them between other cells; others bundle them together into organs called bacteriomes, which branch off from the gut like clusters of grapes. Whatever their origin, their functions are the same: contain and control bacterial symbionts; stop them from spreading into other tissues; and hide them from the immune system. Bacteriocytes are not luxury accommodation. A single one can contain tens of thousands of bacteria, packed so tightly that they make sardine cans look roomy. They are cells in more ways than one.

Bacteriocytes

Despite the old and mutually dependent relationships that many insects have with their symbionts, there is still plenty of room for conflict. If that sounds strange, think about the millions of people diagnosed with cancer every year. Cancer is a disease of cellular rebellion, where a cell strikes out against the regulations of its own body. It grows and divides uncontrollably, producing tumours that can jeopardise the life of its host. If human cells can do this when they are actually part of the same animal, it is easy to imagine that a bacterium like Blochmannia, which is still a separate organism from its ant host, might do the same. It could turn into a kind of symbiotic cancer that replicates unchecked, soaks up energy that the ant needs for itself, and invades cells it shouldn’t.

Containment is tougher for backboned animals like ourselves. We have to control a far larger consortium of microbes than any insect, and we have to do it without bacteriocytes. Most of our microbes live around our cells, not inside them. Just think about your gut. It’s a long and heavily folded tube that, if spread out fully, would cover the surface of a football field. Swarming within that tube are trillions of bacteria. There’s just one layer of epithelial cells – the ones that line our organs – stopping them from penetrating the walls of the gut and reaching the blood vessels that could carry them to other parts of the body. The gut epithelium is our main point of contact with our fellow microbes, but also our greatest point of vulnerability.

epithelium

Simple aquatic animals like corals and sponges have it even worse. Their entire bodies are little more than layers of epithelium immersed in a bath of microbes. And yet they too can control their symbionts. How? For a start, they use mucus, the same slimy goo that clogs your nose when you have a cold. ‘You can’t go wrong with mucus, because mucus is cool,’ says Forest Rohwer.22 He should know – he has been collecting samples of the stuff from across the animal kingdom for years. Nearly all animals use mucus to cover tissues that are exposed to the outside world.

Mucus is made from giant molecules called mucins, each consisting of a central protein backbone with thousands of sugar molecules branching off it. These sugars allow individual mucins to become entangled, forming a dense, nearly impenetrable thicket – a Great Wall of Mucus that stops wayward microbes from penetrating deeper into the body. And if that wasn’t deterrent enough, the wall is manned by viruses. When you think of viruses, you probably think of Ebola, HIV, or influenza: well-known villains that make us sick. But most viruses infect and kill microbes instead. These are called bacteriophages – literally, ‘eaters of bacteria’ – or phages, for short.

phages as a wall to protect foreign microbes

In a typical environment, there will be 10 phages for every bacterial cell.23 In mucus, there will be 40. The same fourfold spike in phage concentrations exists in human gums, mouse guts, fish skins, marine worms, sea anemones, and corals.

Phages are 15 times more likely to find a victim if they stick to mucus. And since mucus is universal in animals, and phages are universal in mucus, this partnership probably started at the dawn of the animal kingdom. In fact, Rohwer suspects that phages were the original immune system – the means through which the simplest animals controlled the microbes at their door.24 These viruses were already plentiful in the environment. It was a simple matter of concentrating them by giving them a layer of mucus in which to anchor themselves.

The mucus that covers it comes in two layers: a dense inner one that sits directly on top of the epithelial cells, and a loose outer one beyond that. The outer layer is full of phages, but it’s also a place where microbes can anchor themselves and build thriving communities. They abound here. By comparison, very few of them exist in the dense inner layer. That’s because the epithelial cells liberally spray this zone with antimicrobial peptides (AMPs) – small molecular bullets that take out any encroaching microbes.

If any microbes successfully weave their way through the mucus, run the gauntlet of phages and AMPs, and sneak through the epithelium, there’s a battalion of immune cells on the other side to swallow and destroy them. These cells aren’t just sitting around, waiting for the worst to happen. They are surprisingly proactive. Some reach through the epithelium to check for microbes on the other side, as if feeling around through the slats of a fence. If they find bacteria in the demilitarised zone, they capture them and bring them back across. By taking these prisoners, the immune system gets regular intel about the species that dominate the mucus, and can prepare antibodies and other appropriate countermeasures.

third layer, immune cells, anti-bodies

These measures – the mucus, the AMPs, and the antibodies – also determine the species that get to stay in the gut.27 We know this because scientists have bred many lines of mutant mice that lack one or more of these components. They all end up with irregular collections of microbes, and usually some kind of inflammatory disease.

inflammatory disease

many bacterial molecules stimulate gut cells to produce more mucus; the more bacteria there are, the more heavily fortified the gut becomes.

You could view this as the immune system calibrating the microbiome: the more microbes there are, the more strongly the immune system pushes back against them. Alternatively, you could say that the microbes are calibrating the immune system, triggering responses that create a suitable niche for themselves while pushing out their competitors. This latter view makes sense when you consider that many of our most common gut microbes have adaptations for coexisting with the immune system.

The rangers only get time off at the very start of our lives, when in microbiological terms we are blank slates. To allow our first microbes to colonise our newborn bodies, a special class of immune cells suppresses the rest of the body’s defensive ensemble, which is why babies are vulnerable to infections for their first six months of life.

not because their immune system is immature, as is commonly believed: it’s because it is deliberately stifled to give microbes a free-for-all window during which they can establish themselves. But without the immune system’s full selective powers, how can a mammalian baby ensure that it gets the right communities? Its mother helps. Mother’s milk is full of antibodies which control the microbial populations of adults – and babies take up these antibodies during breastfeeding.

mother's milk and antibodies

Every mammal mother, whether platypus or pangolin, human or hippo, feeds her baby by literally dissolving her own body to make a white fluid that she secretes through her nipples. The ingredients of that fluid have been tweaked and perfected through 200 million years of evolution to provide all the nutrition that infants need.

Those ingredients include complex sugars called oligosaccharides. Every mammal makes them but human mothers, for some reason, churn out an exceptional variety – scientists have identified over 200 human milk oligosaccharides, or HMOs, so far.32 They are the third-biggest part of human milk, after lactose and fats, and they should be a rich source of energy for growing babies.

But babies cannot digest them. When German first learned about HMOs, he was gobsmacked. Why would a mother spend so much energy manufacturing these complicated chemicals if they were indigestible and therefore useless to her child? Why hasn’t natural selection put its foot down on such a wasteful practice? Here’s a clue: these sugars pass through the stomach and the small intestine unharmed, and land in the large intestine where most of our bacteria live. So, what if they aren’t food for babies at all? What if they are food for microbes?

Wow

No other Bif has this genetic cluster; it is unique to B. infantis. Human milk has evolved to nourish this microbe and it, in turn, has evolved into a consummate HMOvore. Unsurprisingly, it is often the dominant microbe in the guts of breast-fed infants.

It earns its keep. As it digests HMOs, B. infantis releases short-chain fatty acids (SCFAs) that feed an infant’s gut cells – so while mothers nourish this microbe, the microbe in turn nourishes the baby. Through direct contact, B. infantis also encourages gut cells to make adhesive proteins that seal the gaps between them, and anti-inflammatory molecules that calibrate the immune system. These changes only happen when B. infantis grows on HMOs; if it gets lactose instead, it survives but doesn’t engage in any repartee with the baby’s cells. It unlocks its full beneficial potential only when it feeds on breast milk. Likewise, for a child to reap the full benefits that milk can provide, B. infantis must be present.

Human breast milk stands out among that of other mammals: it has five times as many types of HMO as cow’s milk, and several hundred times the quantity. Even chimp milk is impoverished compared to ours. No one knows why this difference exists, but Mills offers a couple of good guesses. One involves our brains, which are famously large for a primate of our size, and which grow incredibly quickly in our first year of life. This fast growth partly depends on a nutrient called sialic acid, which also happens to be one of the chemicals that B. infantis releases while it eats HMOs.

This might explain why, among monkeys and apes, social species have more milk oligosaccharides than solitary ones, and a greater range of them to boot. Larger groups mean more social ties to remember, more friendships to manage, and more rivals to manipulate. Many scientists believe that these demands drove the evolution of primate intelligence; perhaps they also fuelled the diversity of HMOs.

Diversity of HMOs

HMOs bear a striking resemblance to these intestinal glycans, so pathogens sometimes stick to them instead. They act as decoys to draw fire away from a baby’s own cells. They can block a roll call of gut villains including Salmonella; Listeria; Vibrio cholerae, the culprit behind cholera; Campylobacter jejuni, the most common cause of bacterial diarrhoea; Entamoeba histolytica, a voracious amoeba that causes dysentery and kills 100,000 people every year; and many virulent strains of E. coli. They may even be able to obstruct HIV, which might explain why most infants who suckle from infected mothers don’t get infected despite drinking virus-loaded milk for months.

This helps to explain both why breast-fed babies have fewer gut infections than bottle-fed ones and why there are so many HMOs. ‘It makes sense that they would need to be diverse enough to handle a range of pathogens, from viruses to bacteria,’ says Mills. ‘I think it’s the amazing diversity that provides a constellation of protections.’

They have already found something weird: phages are great at sticking to mucus, but they do so ten times more efficiently if there’s breast milk around. Something in the milk helps them anchor in place. The culprits seem to be little spheres of fat, encased in proteins that resemble those in mucus. If you let a glass of milk sit in the open, the layer of fat that forms on the surface is full of these globules. They provide nutrition to a baby, but they might also give baby’s first viruses a foothold in the gut.

Sphere of fat and mucus

When animals get sick, we frequently lose our appetite – a sensible tactic that diverts energy from foraging and towards getting better. It also means that our gut microbes experience a temporary famine. Sick mice deal with this problem by releasing emergency rations: a simple sugar called fucose. Gut microbes can snip off this sugar and feed on it, staying alive while they wait for their hosts to resume normal service.

The Bacteroides group, which excels at eating these glycans, soon become the most common microbes in the gut. But crucially, glycans are so diverse that no single species of bacterium has the right tools for eating all of them. This means that by swallowing or making a wide range of glycans we can support an abundance of different bacteria. Some are unfussy generalists like pigeons or raccoons; others are choosy specialists like pandas or anteaters. They form food webs where some microbes break down the biggest and hardiest molecules and release smaller fragments that others mop up. They make pacts, where two species feed each other, each digesting a different food while creating waste chemicals that its partner can use. They form truces by adjusting their metabolic antics to avoid competing with their neighbours.

This is the price of symbiosis. Even when microbes aren’t as crucial to their hosts as a cicada’s symbionts are, they still exert a powerful influence on our lives and our health. When they go rogue, the consequences can be disastrous. That’s why humans and other animals have evolved so many ways of stabilising their multitudes.

Stablising the multitude

We restrict them by relying on the chemistry of our bodies. We corral them with physical barriers. We can go for the carrot, by nourishing them with dedicated foods. We can beat them with the stick, by using phages, antibodies, and other parts of our immune system. We have many solutions to the ever-present conflicts that exist with our microbes, and many ways of enforcing our contracts with them.

All the stressors that weaken the corals – the warming seas, acidic waters, and nutrient overloads – disrupt their partnerships with their microbes, leaving them with distorted and impoverished communities that are vulnerable to disease, or that might even cause disease.

If the algae rise, the corals fall, and vice versa. In most reefs, fleshy algae are kept in check by grazers like surgeonfish and parrotfish, which nibble them down to well-trimmed lawns. But humans kill the grazers with spears, hooks, and nets. We also kill top predators like sharks, leading to population explosions of medium-sized predators, which then take out the grazers. Either way, we give the algae an advantage.

That something turned out to be dissolved organic carbon (DOC); essentially, sugars and carbohydrates in the water. When algae get too numerous on a reef they make huge amounts of DOC and create a banquet for coral microbes. These algal sugars would normally flow up the food chain to be locked away in the bodies of grazers and, ultimately, sharks; a single shark represents the stored energy of several tons of algae. But if all the sharks die, those sugars remain at the bottom of the food web where, instead of fuelling the flesh of fish, they build the cells of microbes. Nourished by this feast, the microbes bloom so explosively that they consume all the surrounding oxygen, choking the corals.

dissolved organic carbon (DOC)

The algae then trigger Rohwer’s cycle: more DOC, more microbes, more pathogens, more disease, more dead corals.

Rohwer’s work with corals hints at a different type of microbial disease, one without a single obvious culprit.4 These illnesses are caused by communities of microbes, which have shifted into configurations that harm their hosts. None is a pathogen in its own right; instead, the entire community has shifted to a pathogenic state. There’s a word for such a state: dysbiosis.5 It is a term that evokes imbalance and discord in place of harmony and cooperation. It is the dark reflection of symbiosis, the antithesis of all the themes we have seen so far.

dysbiosis

This altered community still communicates with its host, but the tenor of their conversation changes. Sometimes it becomes, quite literally, inflammatory, as microbes over-stimulate the immune system or wheedle their way into tissues where they don’t belong. In other cases, microbes might start to opportunistically infect their hosts.

They exemplify the concept of ‘gnotobiosis’, from the Greek for ‘known life’. We know exactly what lives in these animals – which is nothing.

gnotobiosis

Normally, germ-free rodents can eat as much as they like without putting on weight, but this enviable ability disappeared once their guts were colonised. They didn’t start eating any more food – if anything, they ate slightly less – but they converted more of that food into fat and so piled on the pounds.

interesting

the team showed that obese people (and mice) have different communities of microbes in their guts.10 The most obvious difference lay in the ratio of the two major groups of gut bacteria: obese people had more Firmicutes and fewer Bacteroidetes than their leaner counterparts.

That’s where Peter Turnbaugh came in. Then a graduate student in the lab, he harvested microbes from fat and lean mice, and then fed them to germ-free rodents. Those that got microbes from lean donors put on 27 per cent more fat, while those with obese donors packed on 47 per cent