A Starved Bacterium is the Mirror of Civilisation

It's amazing what bacteria will do to keep themselves, or others of their kind, alive.

The invention of agriculture, which enabled food security, was a critical turning point in the history of mankind. It resulted in the formation of large societies, cities and civilisation as we know it today. From then to this day, malnutrition has also been among the greatest evils of civilised societies. Over-utilisation of resources, including for example the ability of the soil to provide for agriculture, has often resulted in the abandonment and dissipation of cities.

Nutrition is essential not only to humans and animals but for every form of life that lives on this planet, including bacteria – the most numerous of free-living life forms on earth. Bacteria are single-celled organisms. They eat food and, like larger animals, different types of bacteria are able to eat and digest different types of food. They grow in size and reproduce. Reproduction in bacteria involves a bunch of complicated molecular processes but the bottomline is that each bacterium duplicates its DNA and splits down the middle into two.

When a human cell duplicates into two, it leaves a mark on its DNA. In other words, the DNA of a cell carries a mark of its age. This however is not the case for bacteria – the reason for the difference being hidden in the way DNA replication occurs and the fact that human DNA is linear, whereas most bacterial DNA is circular. With each bacterium carrying pretty much as many newly-produced molecules as any other bacterium, and in the absence of genetic signatures of ageing, bacteria do not age much as long as they’re eating, dividing and multiplying.

To reiterate, for a bacterial cell to divide and multiply, it has to make new molecules: make new DNA, make new proteins and make new oils and sugars that make up the cell surface. And to do all this, a bacterial cell needs food – it needs a lot of carbon, nitrogen, sulphur, phosphorus, calcium, iron, magnesium and many other things. So, the rate at which a population of bacteria doubles in size depends on how much food it has. And a bacterium that is starved of food becomes unable to reproduce and starts to age as well, as the molecules that it contains somehow become ‘older’ with time. Ageing in any organism is poorly understood and our understanding of bacterial ageing is especially skeletal.

A start with a fight for food

It turns out that unlike many affluent human beings, many bacteria may often be starved of food. Take the case of Escherichia coli, which many of us know as the inhabitants of our guts and as agents of food poisoning, and which scientists know and love as laboratory workhorses for helping us understand life.

In the lab, where we pamper E. coli with some seriously rich food, one bacterium becomes two every 20-30 minutes. In 1983, Michael Savageau estimated that in nature, an average E. coli might reproduce once every 40 hours. Between the cushy lab and the harsh natural world, there is nearly a 50-100-fold difference in reproduction times of these bacteria. Savageau’s number is an estimate and might even have been an exaggeration; moreover, the number is also an average over the many different circumstances – favourable or unfavourable to growth – that E. coli faces in life. These notwithstanding, we can be sure that the average E. coli in nature needs a lot more than 20-30 minutes to reproduce. As a result, its nutritional status is often much less than what it sees as optimal.

Bacteria have numerous ways to make sure that they get nutrients even out of unfavourable circumstances. For example, some members of a bacterial population might commit altruistic suicide, releasing their contents out for the rest of the population to eat and survive.

Finding food ‘out of nowhere’ is very important for a bacterium during pathogenesis, the process of causing disease. Let’s imagine a bacterium capable of causing disease after entering the human body. The human is rich in benign bacteria with whom the invader has to compete for food. And that’s not easy at all. However, pathogens such as Helicobacter pylori, an inhabitant of many human tummies and in some of its genetic forms a cause of stomach ulcer and even gastric cancer, manipulate host immune response to obtain nutrients.

Lactoferrins v. siderophores

The inflammatory response of the human immune system to Helicobacter and other bacterial pathogens damages our tissue layers, which can result in the leakage of nutrients that the pathogen can use. If the pathogen has mechanisms by which it can uniquely resist the killing agents that the process of inflammation releases, then this can give it a selective advantage over the innumerable human-friendly bacteria that it competes with. Tissue damage, resulting in breaches to various physical barriers, can also provide the pathogen access to new sites in the human body where it can freely do its job in the absence of competitors. Access to such sites can even make otherwise benign bacteria the agents of disease!

For a pathogenic bacterium, it’s not just competition with other bacteria that’s inhibitory to nutrient acquisition. The human system itself does everything it can to deny pathogenic bacteria essential nutrients. One such nutrient is the metal iron – and iron wars are critical determinants of the outcome of tussles between a pathogenic bacterium and its large potential victim. The presence of iron is essential for the functioning of many proteins, including those involved in producing energy from food. Iron being very plentiful on the planet, it’s not surprising that life evolved making use of this element; without iron, there’d be no life as we know it today.

The human body produces many proteins such as lactoferrins, which bind to iron with high affinity and keep the metal inaccessible to invading bacteria. But bacteria throw out molecules called siderophores, which hug iron more tightly than the lactoferrins do, and so tear the metal out and attach it to themselves. These siderophores then find their way back into bacterial cells, where chemical reactions release iron from these molecules so that the metal ions can be used in metabolism.

In these examples, the bacteria do what needs to be done to find existing, but limiting, nutrients. What if there isn’t any food to find? Many bacteria will probably die – but many also survive, and survive in interesting ways. A mechanism by which some bacteria respond to extreme starvation is by forming what are called spores. A severe lack of food is sensed by the bacterium and a series of molecular events results in the organism forming a powerful, robust coat around the cell. Within this coat, the bacterium sits dormant or nearly dormant. And in dormancy, the bacterium needs little food to survive and survives like a bear in the winter. The spore also protects the bacterium against exposure to lethal UV radiation, desiccation and temperature variations. Needless to say, spore formation is reversible and, under favourable conditions, the spore can unravel to release a happy-to-eat-and-reproduce bacterium.

Spores are a serious medical problem when it comes to an opportunistic pathogen called Clostridium difficile. In most healthy people, this bacterium causes no harm. But in individuals with a weakened immune system, and in people hospitalised and being treated with antibiotics, this bug becomes a major problem. It forms hardy spores that are impervious to chemical disinfectants – including many that are commonly used by doctors for sanitation – and antibiotics. In the absence of competing bacteria wiped out by antibiotics, Clostridium comes into its own and causes diarrhoea which can be fatal. It has already become a major problem in the West, resulting in several hospital-associated outbreaks across which hundreds of patients hospitalised for other things have died in the last ten years.

No keeping down a bacterium

Spore formation can also be cool in other contexts, such as in the soil bacterium Myxococcus xanthus. When starved for nutrients, many Myxococcus individuals start ‘talking’ to each other, come together and form large structures called fruiting bodies, made of many individual cells. Starvation resistant spores form on these fruiting bodies. Dramatic stills of the process of fruiting body formation were first published more than 30 years ago by J.M. Kuner and Dale Kaiser, and this has been a talismanic example of multicellular behaviour shown by a collection of unicellular organisms.

Bacteria such as E. coli do not form spores but can still survive extended periods of starvation. In the lab, E. coli grown in the pamper-media keeps doubling its population size several times over a few hours before exhausting nutrients and entering a period of little or no population growth, called the stationary phase. This is characterised by the slowing down of many metabolic processes, including DNA replication and protein synthesis. A few days into it, 99% of cells die but the remaining 1% keep going – and they can keep going for years! Though the population size doesn’t increase during this phase, it doesn’t mean that the phase is uneventful.

In a classic paper published in the late 1990s, Roberto Kolter and his colleagues at Harvard University showed that during the stationary phase, an E. coli population finds novel mutations that enable it to outcompete compatriots not carrying such mutations.

It’s believed that, during this period of stasis, there are multiple waves of growth-and-death of distinct small subpopulations of mutants, the former uniquely capable of growth under these conditions and the latter not; each wave resulting in the utilisation and release of distinct sets of molecules. Let’s assume that at the beginning of the stasis phase, there are two subpopulations of E. coli – A and B – in the growth pot, which contains the nutrient N. If E. coli A can eat N and release molecule M, but E. coli B is unable to use N, then A reproduces while B dies out. After a while, N is exhausted but M is available. E. coli A cannot use M but a new mutant C can, and in turn release molecule P. Then the population of E. coli C grows, while A dies out. And this can go on indefinitely. The growth and death of different subpopulations might average out to a net growth around zero.

There is also evidence that cellular processes that generate increased genetic diversity, such as by introducing high error rates while replicating the DNA, are essential for long-term survival. And increasing genetic diversity also means generation of novel variants that may be able to survive a range of stresses, including antibiotic onslaughts. And this keeps going on forever. So if you’re worried about infectious diseases, do fear the starved bug! And if you find bacterial life fascinating, do love the starved bug!

Aswin Sai Narain Seshasayee runs a laboratory researching bacterial biology at the National Centre for Biological Sciences, Bengaluru. Beyond science, his interests are in classical art music and history.