E. coli is a microbe used in a number of school practicals but recently there have been concerns about its safety. Microbiology Today Editor Gavin Thomas clears up these issues.

Escherichia coli K-12 is a great organism to use in the school lab for a variety of different experiments. This is because it has actually been developed as a tool in research labs to study basic principles of biology. In this article I will introduce the species and general properties of E. coli and compare the strains that are used in the lab to the pathogenic ones that cause disease in an attempt to convince the reader that they are certainly not the same beast.

What are E. coli?
Escherichia coli are Gram-negative bacteria that belong to the g-proteobacteria. As they primarily live in the mammalian gut they have been grouped with other related bacteria as 'enteric' bacteria. They are straight rod shaped cells of about 2 µm long and 0.5 µm wide, which can grow and divide rapidly by binary fission.

There are many different types of E. coli and the chief way they are distinguished is immunologically using serotyping. The current typing system is based mainly on three types of antigen: the somatic (O) antigen which corresponds to terminal sugars on the cell surface lipopolysaccharide (LPS), the capsular (K) antigens and the flagellar (H) antigen. There are over 170 O antigens, over 100 K antigens and over 50 H antigens. Hence, when we refer to pathogenic strain O157:H7, it means that this E. coli has O antigen 157 and H antigen 7. Many other strains cause disease as well, like O26:H11.

Why do we use E. coli K-12?
While there is a great diversity of strains in the environment, only a few are used in the lab. The majority are a derivative of a commensal strain called K-12. One of the main reasons why this microbe is a key research tool is that it is safe to handle; you could drink a culture of the stuff and not notice any effect (not to be recommended, however!). As well as being safe to use, K-12 is ridiculously easy to grow. It is usually cultured in the lab on a rich nutrient broth or agar, which supplies plenty of goodies for rapid growth. Whilst it is often said to be able to divide every 20 minutes, that is really only under absolutely optimal conditions. However, it still grows very quickly compared to other microbes. This is a big advantage in school as a culture can be set up one evening and by the following day nice clear and distinct colonies are visible on an agar plate.

Growing E. coli in nutrient broth is a quick and simple way of propagating this microbe, but does not exploit one of E. coli's most important properties. Unlike humans and many other microbes, it doesn't need lots of complex chemicals, like vitamins, to grow. Just provide a solution of some sugar (glucose is best), ammonium sulphate, salt and phosphates and grow it aerobically at the 37^oC used in research laboratories and it's perfectly happy. Such incubation temperatures are not allowed in schools, but even at the permitted maximum of 25^oC, K-12 still grows well. Basically, it can synthesise everything it needs to make a completely new cell from these few simple molecules, which is a seriously impressive feat.

All E. coli are not the same.
While K-12 and B strains are safe microbes, we know that there are other E. coli out there like O157:H7 that can kill people. However, these are quite different from K-12 even though they have the same species name. This is illustrated very clearly when the DNA sequences (genomes) that make up K-12 and O157:H7 are compared. They are 25% different from each other! As humans share about 99% of their DNA with chimps, this gives an indication of how much evolution and movement of genes have occurred in the environment since these 2 strains of E. coli last had a common ancestor.

Scientists now know why K-12 is not harmful. Many of the known properties of the bacteria that allow them to cause disease, called virulence factors, are seen in pathogenic strains but not in K-12. In fact, the K-12 strain used in the laboratory is even less dangerous than a commensal strain living in your own gut that you might isolate from your stools. K-12 has been grown in the lab for many generations and so has adapted to live there rather than the intestine. It wouldn't stand a chance in the hugely competitive environment that is your gut where bacteria are constantly evolving to keep their 'cutting edge' and not be pushed out by other microbes. Getting K-12 to establish itself in the gut would be like trying to qualify for a Formula 1 race with a car from 1922 (which is when K-12 was taken from the somebody's gut)! It was competitive at the time, but is now way off the pace.

E. coli K-12 is a friendly bacterium
Some studies that suggest E. coli could be used as a probiotic, but when you browse the web for information about commensal E. coli you will find a statement something like 'E. coli is a friendly bacterium as it can produce vitamins that we require, especially vitamin K'. Not trusting the internet as a particularly reliable source, I searched for experimental data that supports this assertion.

Vitamin K is essential in humans and most animals as we cannot synthesise the compound ourselves. In humans, vitamin K is used by the liver to synthesise prothrombin, which in turn is processed to form the enzyme thrombin; a key enzyme involved in the blood clotting process and there is increasing evidence that vitamin K has additional roles in maintaining in bone health. There are two forms of vitamin K, vitamin K1 and vitamin K2. Vitamin K1 is called phylloquinone and comes from our diet. It is found in some oils, especially soybean oil, and in dark-green vegetables such as spinach and broccoli. Vitamin K2 is menaquinone, which can come from the bacteria in the gut and indeed E. coli can synthesise menaquinone because it uses it during respiration. As E. coli lives and dies in the gut, the dead cells release vitamin K2, which can then, theoretically, be absorbed and utilised by the body.

Evidence suggesting vitamin K derived from E. coli can improve diet

A number of studies have given support to the idea that vitamin K2 produced by the gut flora, and specifically by E. coli, has an important function in keeping us healthy. One study looked at rats that were born and raised in a sterile environment in the absence of any bacteria (gnotobiotic). They infected different rats with different bacteria, including E. coli, and found that the ones that were known to make menaquinone in the lab also made it when they were growing in the rats. They also showed that the concentration of the menaquinone in the liver (a site where vitamin K functions) was increased in the rats that had been infected with bacteria that made menaquinones. Hence, in rats at least, there is evidence that menaquinone made by E. coli can be taken up by the host organism and concentrated in the liver (Kindberg et al., 1987).

A second more recent study looked at the reverse process. This study was in humans that had a normal fully formed gut flora (of which E. coli only makes a tiny proportion). Like the study above, they measure d the concentration of menaquinone in the livers of patients who had just died. They compared the amount of menaquinone in individuals who had been taking broad spectrum antibiotics before they died (in whom most of the gut flora would have been absent) to individuals who had not (who should have had a normal gut flora). They found that individuals who had been treated with antibiotics had a much reduced menaquinone content in their livers. The authors suggest that a reduction in the gut flora responsible for menaquinone production (which includes E. coli) leads to reduced stores of this form of the vitamin in the liver (Conly and Stein, 1994).

While both these studies demonstrate that menaquinone produced by E. coli can be utilised by humans neither demonstrate that this is i) giving a benefit to health and ii) that E. coli is really contributing to this in the mixed population of the gut. Also, it is clear that the majority of the vitamin K that we obtain is as vitamin K1 from our diet and so the often quoted benefit of E. coli in our guts does still seem a possibility but has no real experimental support.

Dr Gavin Thomas is a lecturer at the University of York. He has set up a website on E. coli . If you have any questions about E. coli, email Gavin at: ght2@york.ac.uk

References
Conly J, and Stein K. (1994) Reduction of vitamin K2 concentrations in human liver associated with the use of broad spectrum antimicrobials. Clin Invest Med 17(6):531-9.

Kindberg C, Suttie JW, Uchida K, Hirauchi K and Nakao H. (1987) Menaquinone production and utilization in germ-free rats after inoculation with specific organisms J Nutr 117(6):1032-5.