Scenes from the many lives of Escherichia coli
A play in three acts
Mark Erickson, Catherine Will
Can bacteria be famous? If they can, then which are the most famous, and why? In this paper we will put our candidate for the world’s most famous bacterium onto the stage, literally, give it a voice – or a few voices – and show how Escherichia coli grew in importance.
We present a play showing E. coli on an odyssey of self-discovery, from a children’s clinic in Bavaria to high-tech university labs, via hospitals, medical schools, wastewater treatment plants and government offices tasked with slowing antimicrobial resistance. Names are important and we’ll see how naming this bacterium changes our relationship to it,1 just as the disciplines and tools of those naming and making these microbes also shift.2
It is not easy to put words into the mouth of a microbe. We confronted the problem of bacteria as plural, and the question of generation. The E. coli made visible (in plural) in laboratories tend to end their lives soon after in the autoclave. Yet we use the singular to gesture to the ontological stakes,3 even as we multiply coliform identities.4 Across the decades, our hero has had a dizzying number of these, though many practices involve familiar actors such as Petri dishes, Falcon® and Durham tubes, agar, Bunsen burners, slides, and microscopes. Nonetheless, we suggest E. coli itself is unsettled – fixed and stained, but often misunderstood and ignored.
Joining a tradition of experimental writing in STS,5 we take inspiration from classical Greek theatre, using a Chorus to represent laboratory assemblages forming and decomposing,6 and its Leader to help navigate. Like the writers of this time we also invoke higher powers, Gods and Muses, to help us tell our tales.
Thalia: Welcome to our performance, a joyous play in three acts. I am Thalia, the muse of blooming. My sister Clio, the muse of history, has helped me with some parts, and where would we be without our darling mama Mnemosyne, whose memory we rely upon?
We are brought together to tell you of a world of microbes; first, as many decades ago we find the wild one. As wild as Enkidu, who is older even than we Muses.8 We tame it through naming it, first, Bacillus coli commune (B. coli), later Escherichia coli. Then bleak-hearted Melpomene scripts our play, as the blame for disease and illness is placed upon E. coli, who turns trickster, hiding and dissembling, only to be revealed as, perhaps, a hero after all. Meanwhile wise Athene guides the molecular biologists’ hands as they grow, break and construct the innards of E. coli, revealing its secrets. But Melpomene drives the humans towards a tragic end as our microbes’ resistance grows. Perhaps it is they who will bloom in the future, while those who have flourished, like all fragrant flowers which abound in the meadow,9 will wither and die.
Act 1, Scene 1
Leader of the Chorus: We begin in the 1880s when the story goes that Theodor Escherich was studying microbes in stool samples. Before this, coliform bacteria were known and seen under the microscope, true, but the quest for understanding this particular companion species was just beginning.
Theodor Escherich MD: It’s good to be in Munich, but here – as in Vienna – we lose too many infants to dysentery. We don’t know what’s responsible, but the microscope might help.
Cultures are grown on plates, then samples placed on slides on a long lab bench, ready to go under the microscope.
Chorus: The party’s assembled; ready for action – lab,
Esteemed doctor, microscope, slide.
Could shit samples reveal the cause of this illness?
Let’s focus and find what our bodies might hide.
Escherich looks and listens carefully.
Theodor Escherich MD: I see distinctive shapes in many of my samples. Tiny organisms, short with rounded ends.
Coliform: I am here.
Theodor Escherich MD: Perhaps they are the source of the dysentery? I will call them Bacteria coli commune and continue my investigations.10
Leader of the Chorus: Meanwhile, in the 21st Century, molecular biologists revive and sequence Escherich’s original bacilli samples, and get a surprise result.
Molecular biologist: Are you sure you’ve got the right culprit? We’ve brought molecular vision by sequencing genes using SMRT cells and the Pacific Biosciences RS II instrument in these historical samples. They originated in your lab alright, but they don’t look to be pathogenic.11
B. coli: A false accusation, or a case of mistaken identity? Whatever the reason we’re all suspect now.
Leader of the Chorus: Escherich thinks he’s found the culprit but perhaps it’s not so clear. Some of the first to work with his samples focus on hygiene and sanitation. In the Lister Institute of Preventive Medicine, a young Harriette Chick starts what will become an illustrious career in nutrition, with laboratory work on the efficiency of different disinfectants against bacteria.
Harriette Chick (later DBE): They’re dirty and dangerous and I’m going to kill them.12
B. coli: AAAAAAAARGGGGHHH!
Leader of the Chorus: Her colleague Alfred MacConkey – later of agar and broth fame – is more interested in B. coli in water.13 He does a series of experiments to explore ways of identifying it effectively and using it as a signal for faecal contamination.
B. coli: More slander, I assure you!
Leader of the Chorus: But the accusation proves hard to escape. Around the time of the First World War, in a frenzy of nomenclature revisions across scientific communities, the genus Escherichia is named in honour of Theodor as part of the family Enterobacteriaceae.14 Within this genus numerous strains – some pathogenic and some not – are given new names, including Escherichia coli, which will become the most famous of all.
Act 1, Scene 2
Leader of the Chorus: Alfred MacConkey experiments with media to create the perfect conditions for the bacteria to show themselves.
Chorus: With broth and with lactose our friend B. blooms
Let’s join together, say a prayer, make a wish!
Add bile salts and culture. Given time
Coli will appear in clear view on a dish.15
Alfred MacConkey MD: Never mind Hariette’s ‘kill efficiency’. If we’re going to test water samples for B. coli regularly, we need to grow these organisms. Just for a time, here in the lab, we want those bacteria to flourish like the roses in my garden. Fortunately, they do well at body temperature, and they don’t need protection from oxygen, but I’ll offer them lactose, saccharose, dulcit, adonit or inulin. I can show that B. coli likes to ferment sugar and that fact is useful…
Samples of 497 bacilli are meticulously isolated from 76 different substances including human and animal faeces, soil, pond and rain water, oats, beans and cheese, and are put onto plates. Bile salt media is used because of its inhibiting effect on other organisms. Once colonies appear, MacConkey adds lactose and watches for fermentation.
B. coli: Busted! We like the lactose, though the bile salts are not so welcoming. But you’ve certainly found us out: our bacterial cousins can’t grow so well here so we reveal our presence on the plates.
Alfred MacConkey MD: The problem is we keep identifying more types of lactose-fermenting enteric bacilli, and at the moment we’re calling them all B. coli. Still, whatever they are, now we can culture them quickly we can go back to counting… what does our work tell us about the water from which the sample came? Ideally, we’d have some form of numerical interpretation of what we find,16 perhaps by measuring the volume of gas produced in the fermentation.
Chorus: Again, we’re together. Broth, lactose, salts in a tube.
Wait once more. Watch. Hold your breath.
The doughty professor assembles his subjugated workers
But can dodgy B. coli really help confront death?
Alfred MacConkey MD: B. coli might not be a friend, but it could yet be an ally in our war against dirty water if we assume it reflects the presence of other coliforms. Contamination by human faecal matter is our fear but also a practical problem. ‘We all of us always wish to identify organisms as accurately as possible, in as short a time as possible, and with as little trouble as possible’.17
E. coli: Named, tamed and shamed! Established as the villain of the piece, my reputation can only improve as we create new forms of cooperation. But I’m happy to get a job. I’m here to help, believe me…
Leader of the Chorus: MacConkey’s ‘practical’ approach rapidly spreads, appearing in the very first edition of the textbook American Sewerage Practice, published by Leonard Metcalf and Harrison P Eddy in 1914/5, and still a touchstone in the field.
Bacteriologists: Why should we let E. coli speak for all the coliforms? It is only 0.1% of the microbial flora,18 and isn’t even an anaerobe like most of the others.
E. coli: Because the standard guide to Wastewater Engineering tells you ‘The presence of E. coli in drinking water is an indication that there is a greater risk that disease causing pathogens could also be present’.19 In this field you respect traditions, and these stretch back decades. I can represent the bigger group, and with some amplification I do that pretty well. Look how safe the drinking water is, if you don’t believe me.
Act 2, Scene 1
Leader of the Chorus: We’ve given you one history, here’s another. From their earliest identification B. coli have been blamed for more than diarrhoea. And in the early 20th-century hospital the laboratory is growing in importance, meaning they play more of a role…20
Chorus: Big hospital, small lab. With samples coming
All the time it’s getting a bit tight.
Clinicians and technicians, they all join in the hunt
Examining slides late into the night.
B. coli: Why is it always my fault?
Leader of the Chorus: Urine has been examined for centuries, but from the 1880s bedside pronouncements were enhanced by chemical testing using test papers and now ‘piss pots’ are travelling down to the lab for some bacteriology.21
Chorus: Another glass jar viewed with suspicion.
Our microscopes are ready though so we’ll
Plate piss – and culture – leave them to grow.
Tomorrow we’re back for the great big reveal.
Leonard S. Dudgeon, MRCP (Lond): But it is hard to interpret the results of these cultures, even working long hours and following careful procedures. What’s the meaning of our frequent discovery of B. coli? ‘Some pathologists go so far as to say that in the female this condition is of no importance; certainly there may be no associated symptoms of disease unless constipation is considered as such. At times, however, there will be headache, slackness, and general malaise’.22
B. coli: Slackness and general malaise sound a bit vague. What exactly is the accusation here?
Leader of the Chorus: You’re in the wrong place in short. But attention is focused on the female urinary tract, not sewers. Are you causing symptoms or are you somehow incidental?
Act 2, Scene 2
Leader of the Chorus: By 1957 doctors have antibiotics that can act effectively against E. coli. But the cultures are confusing, and they are still debating which results to act on.
Edward H Kass MD: Here in Boston, we test pretty much everyone who comes into hospital. But we keep finding bacteria. There’s a considerable number of cases of what I call ‘asymptomatic bacteriuria’. I propose we distinguish between severe and mild infection by counting the colonies on an agar dish. If you’re comparing in this way you have to follow careful steps to do the culture, but numbers can be multiplied to give an estimate of bacteria per ml.23 In my view, more than 1,000,000 per ml should be taken as evidence of infection. Less than 10,000 per ml should be taken as a sign of contamination or as clinically irrelevant.
Chorus: For sure we’re doing what we can to help here
More knowledge, more medics – the brightest and best –
The mid-century height of clinical hopes
With more counting we could have a new type of test.
Leader of the Chorus: With this approach, E. coli could be blamed for silent infections as well as those with symptoms.
Act 2, Scene 3
Leader of the Chorus: In family medicine too, examining urine samples remains a staple of practice.24 By the 1950s, general practitioners have the ‘dipstick’ – combining test papers on a single card for glucose, white blood cells and nitrites or proteins without the fuss of sending samples off for culture.25
Chorus: With antibiotics E. coli’s distressed, but bacteriologists are still in a mess.
We’ll need to gather all our wits and make some diagnostic kits.
Bacteria, samples, doctors, women,
Journals, dipsticks and – ampicillin.
A patient awkwardly hands over a small pot with yellow liquid, still warm. The doctor examines it visually, then puts in a card indicator for 1-2 seconds. He pulls it out and examines the coloured squares. Moving to purple and pink in the first indicates the presence of leukocytes, the body’s response to infection, and nitrites which are reduced by E. coli. He pulls a prescription pad towards him and writes the name Penbritin.26
E. coli: Now I’m seriously worried I’m a target. Your efforts to make and use antibiotics are pretty concerning. Just because you can see me does not mean I’m causing any trouble. You’re just picking me out because it’s easy. But should you really try to kill me off?
Leader of the Chorus: Short answer is no. In the 1980s, doctors are still arguing. They can see colonisation without symptoms or white blood cells in some patients, but they also see symptoms that look or feel like urinary tract infections when their tests can’t find bacteria.27 Through these investigations – mainly from a clinical perspective – E. coli emerges as a tricky character.
Act 2, Scene 4
Leader of the Chorus: There is still debate about the right approach to treatment today, with fears of growing resistance in antibiotics.
Sally Davies, Chief Medical Officer of Health UK: Over-treatment with antibiotics represents a waste of antibiotics’ declining efficacy. If E. coli is exposed to too many antibiotics it can learn to live with them and the drugs won’t work! Antimicrobial resistance (we call it AMR) is a major threat to modern medicine.
Leader of the Chorus: Automation makes it possible to review larger numbers of samples more quickly and more cheaply. It might be more reliable, but it might not.
Adam, director of a hospital lab: There’s still lots of uncertainty. Numerous factors affect the results we get from urine samples coming into our lab after a positive dipstick result. Still we put them all through our system. Now when the counts look high, we do sensitivity tests with the MAST-URI system against different antibiotics. We notice up to 15% error in antibiotic quantity in the prepared discs we use, though the machines have got the depth of agar fairly consistent now. Is it MacConkey agar? No, I’m sure it’s not, but I couldn’t tell you what it is these days. Everything is proprietary in this set up.
Chorus: It’s all automated here. Once again progress is in the air.
Look on us E. coli and despair!
But we’re not sure it’s the end of the argument…
A clamour of voices from journal science: There’s so much more to the human microbiome than we think. ‘The urinary tract is not sterile’!28
The urine is unlikely to contain all pathogenic agents. ‘Bacteria may invade the epithelial lining cells finding sanctuary from immune surveillance and urinary clearance mechanisms’.29
There might even be such a thing as ‘polymicrobial UTI’.30
Leader of the Chorus: Given this, the industry built around culturing urine samples might need rethinking. If E. coli is even more ubiquitous than previously thought, the search is on to understand its role in illness and how it relates to other organisms.
E. coli: Oh yes, we’re also good at hiding, and we have friends who are even better at concealment. For us this is more than just a visit. We’ve settled in to stay. If we stick together we can avoid being flushed away by the urine flow. The epithelial lining is a wonderful thing. It’s not home exactly but it’s a lovely spot.
Act 2, Scene 5
Leader of the Chorus: By the start of the 21st century, all this leads to considerable caution about the use of laboratory cultures for UTI. They are still done, especially for pregnant women, but not on apparently straightforward urinary tract infections in primary care. The principle becomes treating people quickly if they have the expected symptoms and culturing the tricky cases.
Public Health England: Our new guidance is for GPs to not send samples for testing, just prescribe the recommended antibiotics anyway for uncomplicated UTIs. We know what’s happening and who is responsible.
Leader of the Chorus: In the hospital lab however there’s a whole suite of technology designed for direct susceptibility testing.
Microbiologists: We see more and more nasty infections in the blood stream: if E. coli makes it in there, we have to look for the right antibiotic for each case.
Chorus: More helpers required! Resistance tests aren’t so simple.
Pour broth libations. Stack up 8x12 racks.
So much work to show whether drugs kill the bugs.
Spotlight falls on the machines designed to give a measure of resistance, processing 95 samples at a time to examine susceptibility to different concentrations of antibiotics.
E. coli (wild type): I get it, you’re scared. Well maybe you should be! I’m feeling pretty good, getting wise to your tricks, and I’ve got some of my own.31 I’m doing so well, I’m travelling all over.32 You can throw ampicillin at me and half the time I bounce right back.33 Maybe trimethoprim still knocks me out mostly, but don’t take it for granted. Ciprofloxacin and amoxicillin are not a problem I assure you. I’ve hung out with them so often they’re like mates. See how resistant I can be!
European Committee on Antimicrobial Susceptibility Testing (EUCAST): Whatever you say. We now know exactly how to define susceptibility and resistance right across Europe. We’re onto this.34
Leader of Chorus: Meanwhile, routine samples checking for UTIs in pregnant women are seen as valuable for tracking the problem of antimicrobial resistance.
Microbiologist: In many parts of the world, we test pregnant women because we know urinary tract infections are dangerous for the health of their child. We can use those samples to picture the spatial (or even social) distributions of the more resistant E. coli.35
Chorus: No, the work never ends, and nor do the grants.
They’re calling committees and funding reports.
It’s dizzying how many people turn up.
Multiplying disciplines, all with their thoughts.
Data scientist: Yes, the figures are big enough for us to play now. Away from the smell of stale pee in the laboratories, we crunch numbers in bright white offices. But beware. The resistance percentages are acutely sensitive to the full set of samples being tested by each lab.36 More testing of routine samples and the ‘resistance rate’ might look artificially low. If testing of routine samples is reduced – say because prescribing is increasingly allowed to follow clinical symptoms – then it may look artificially high. ‘Instead of trying to sum up from laboratory results at a regional or national level perhaps we should instead create a surveillance system. We have more to learn about how far prescribing for one condition increases the chance of suffering resistance in another and about the mechanisms behind resistance within the host or patient’.37
Chorus: A system, let us have a system. That’s always the solution in our experience.
Immunologist: Please don’t forget us and the contribution of lab research. In this funding bonanza you’ll want interdisciplinary teams, and now we know there’s plasmid transfer of resistance we should study the ‘bacterial gene pool’ too. ‘Even a transient effect of antibiotic use on the carriage of resistant organisms by an individual could have a major impact on the endemic level of resistance in the population’.38
Act 3, Scene 1
A molecular biology laboratory at a university in the West Midlands
Chorus: Here we all are again
And what a proliferation!
Microbiologists, sociologists, biochemists
Careers, papers and prep-kits
Microbes, vectors, equipment shiny
Will make our esoteric thought community39
Working away on our usual routine
Looking for a β-barrel protein
Molecular biologist 1: The laboratory is a secure and hazardous environment, so we need to don our PPE; lab coats, goggles and nitrile gloves. We’ve got a big job: to work out the structure and function of a piece of E. coli’s cell wall, its BamA β-barrel protein. This could give us a new antibiotic if we’re lucky.
Leader of the Chorus: The molecular biologists’ task is large. They will need to enlist the help of a large number of actors to bring this about, but the star here will be their old ‘workhorse’ E. coli, here in the K-12 strain which is an established model organism in the field.40
The team of molecular microbiologists in the lab are surrounded by machines, Bunsen burners, assorted glassware, reagents, family photographs, discarded PPE, pens and papers, fridges and microwave ovens. Is this a mess?
Leader of the Chorus: Eventually, the molecular biologists will triumph in identifying the structure and function of E. coli’s BamA β-barrel protein,41 but (spoiler alert) on the way they experience some doubt.
E. coli K-12: Take me apart, look inside me, tell me my secrets. I am at your disposal, a willing helper in the quest for complete knowledge.
Molecular biologist 1: Thank you E. coli K-12 but be aware that this will be a long, complicated process. And we should say, we have nagging doubts about your identity.
E. coli K-12: Why? You’ve worked with me before many, many times and know me almost personally. I’m here to help – I’m as accommodating as possible.
Molecular biologist 2: That might be a problem E. coli. Maybe you’re too accommodating, too refined, cossetted, too removed from your wild cousins? You’re right, we know you personally and have even given you your own name: E. coli K-12 RLG221 to be precise,42 which indicates that you are a very specialised and refined bacterial strain, bred in captivity as a clone with special characteristics, not least of which is our ability to work with you easily. Your cell wall, which is what we are interested in, is much easier to penetrate than that of your wild cousins.
E. coli K-12 RLG221: Is that what you are going to do to me?
Molecular biologist 1: Yes – first through electroporation, to insert those plasmids and vectors, then through sonication to break you apart and take out your new DNA. Then we’ll look at what the ‘new you’ is capable of doing before finally taking you apart again and inferring the structure of that BamA β-barrel protein.
E. coli K-12 RLG221: OK. You’re right, it does sound complicated. Painful, too.
Molecular biologist 2: And we were only giving you a tiny part of the story! But first, can we resolve the question of your identity K-12 RLG221?
E. coli K-12 RLG221: Some more experiments, perhaps?
Act 3, Scene 2
Two other molecular biology laboratories at universities in the Midlands
Leader of the Chorus: So, the molecular biologists design and run their experiments.
Molecular biologist 1 [to E. coli K-12]: One thing we know about you, K-12, is that you can’t express the O antigen, but your wild cousin can. I wonder what difference that makes.45
E. coli K12 RLG221: OK, I admit I can’t express the O antigen, but maybe I can be just like the true wild type, maybe I too can infect your other favourite model organism, the little worm Caenorhabditis elegans.46 Perhaps that would be a good test?
Molecular biologist 3: Nice idea K-12 RLG221; we’ve used C. elegans to test the pathogenicity of other strains of E. coli for years. We’ll reverse-engineer you and make you express O antigen. [Time passes…] Now you have a new identity: E. coli DFB 1655 L9.47 Like the name?
E. coli DFB 1655 L9: Neat! But that was a lot of hard work too. So – gonna test me? Bring on those worms!
Chorus: We’re adding to our assembly again,
This time a worm and the O antigen
The worms are in a different laboratory
So, our new microbe needs to make a journey.
Molecular biologist 1: Well, putting that O antigen back into K-12 made DFB 1655 L9 kill those little worms with a vengeance! E. coli K-12 RLG221 without the ability to express the O antigen are non-pathogenic for C. elegans, but then we knew that already. But all the true wild type E. coli do have the ability to synthesise the O antigen. So – have we got the right model for our lab studies? These results certainly call things into question.48 These guys are quite different from one another!
E. coli K-12 RLG221: I am here to help you. I thought I was helping you really well.
Molecular biologist 3: And we do thank you for it – but things are different outside the lab.
E. coli (true wild type): You better believe it – those wimpy lab strains wouldn’t last a moment in the hostile environments I hang out in! You think you’re tough ‘cos you like MacConkey Bouillon and TBX agar – you should try living in some of the places I do – frozen seagull poo,49 sewage systems,50 air conditioning units…51
Act 3, Scene 3
The original molecular biology laboratory at a university in the Midlands
Molecular biologist 1: Interesting results regarding K-12! Do you think we should change how we run the BamA β-barrel protein experiments?
Molecular biologist 2: Not right now – stick to the protocol!
E. coli K-12 RLG221: Yay! Happy to be working with you guys again. Let’s go.
Chorus: E. coli K-12, K-12 RLG221, K-12 DFB1655 L9 all tamed like Enkidu
But which is the wild type like that in our poo?
We’ve done a lot of experiments, but what have we found?
Are we nearer to showing our knowledge is sound?
E. coli: From your perspective I suppose this looks like a mess. From mine, a great success. Whatever you threw at me, I learned to cope and shared my skills. I still want to assist, but you’re going to have to treat me with a bit more respect. Stop trying to get rid of me all the time and recognise I can help in more ways than you knew. Our long collaboration may be entering a new phase but please, respect my creativity and complexity rather than just what you call ‘virulence’ or ‘resistance’. I know you’re a bit uneasy about how my lab self – K-12 – may vary from free-living E. coli but try to reconcile this. Your skills in ensuring my cooperation in the laboratory may yet give us a way to thrive together. But you need to lose your anthropocentric view of bacteria!52
Back on Mount Olympus, the Muses look down.
Clio: I wonder what the mortals have learned from our tour? They have tools and motivation now, whatever the twists of the story, they just keep going!
Thalia: How joyous, how beautiful our actors working together are. From bacteriology to molecular and cell biology, and biotech, E. coli is a constant partner to humans!
Melpomene: Have you learnt nothing? Look at the storm clouds on the horizon. Those mortals have sowed their doom with their reliance on this organism.
Clio: Only I can know the future, sisters, but let us leave it to our audience to decide which voice they will hear, Thalia or Melpomene.