MadSci Network: Microbiology

Re: Why haven't harmful heat resistant bacteria evolved?

Date: Thu Dec 22 21:13:19 2005
Posted By: Steve Mack, Post-doc/Fellow, Molecular and Cell Biology
Area of science: Microbiology
ID: 1135282176.Mi

Hi Mike,

Thanks for submitting your question to the MadSci network. You are asking two questions here; (1) why have pathogenic antibiotic-resistant bacteria evolved in response to the use of antibiotics so rapidly (i.e., in under 100 years); and (2) why haven't pathogenic heat-resistant bacteria evolved in response to cooking.

With respect to question (1), you seem to be assuming that bacterial antibotic resistance arose de novo in bacteria as a result of human antibiotic use. However, it is not true that antibiotics have only been in use for 80 years. They have actually been used for hundreds of millions of years, by fungi. Remember that the original antibiotic, penicillin, was isolated from colonies of Penicillium chrysogenum. Over the hundreds of millions of years that have passed since these antibiotic compounds were first recruited, some bacterial species have been able to develop means of protecting themselves by reducing the effectiveness of the compound, or completely neutralizing the compound's activity.

For example, many antibiotics interfere with bacterial protein synthesis, either stopping protein synthesis or reducing the fidelity of the process to the point where the organism can't make sufficient functional product. If the bacteria can't make proteins, then they can't replicate, carry out metabolisms, or repair damage to themselves, and they eventually die. Many of the mutations that confer resistance to specific antibiotics are single nucleotide rRNA mutations (e.g., Streptomycin resistance, a single base mutation in the 16S rRNA, and Chloramphenicol resistance, a single base mutation in the 23S rRNA), which protect the bacterial cell from the method of action of the antibiotic (often at the expense of a sub-optimal rate of translation, which means slower growth).

Other modes of antibiotic resistance involve the pumping of the offending compound out of the cell, so that it never reaches its target. In these cases (e.g., tetracycline, erythromycin) the pumping is carried out by a special enzyme (an ATP-dependent efflux pump), and it costs the bacterial cell energy (in the form of ATP) to do the work of pumping out the offending compound.

There are other methods of achieving antibiotic resistance, but I won't go into them here, because I think that these two will suffice.

So, now you're probably asking, "if antibiotics have been around for billions of years, why aren't all bacteria resistant to them?". Well, as you can see with the methods of drug resistance I've described, resistance comes at a cost to the cell. If you resist through the use of a mutated rRNA, then you sacrifice efficiency of your ribosomes for drug resistance. If you are using a protein to pump drugs out of your cytoplasm, then (in addition to the ATP that it costs to make the pump work), you have to make sufficient amounts of the efflux pump to be effective, and you have to replicate the gene that codes for the protein.

These costs are worth the extra trouble when you are in the presence of the antibiotic, but for hundreds of millions of years, the only time a particular bacterial strain came into contact with a particular antibiotic was when it was trying to grow near a particular fungus, and different fungal species developed different antibiotic drugs. In cases where there are no antibiotic drugs present, a bacterial strain that is maintaining drug resistant genes will be out-competed by strains that have more efficient ribosomes and that aren't spending extra time and energy replicating the genes for efflux proteins and synthesizing copious efflux pumps. So, much of the time, there was no use for antibiotic resistance, and natural selection favored those bacteria that could replicate efficiently (i.e., without maintaining antibiotic resistance genes).

The final wrinkle is that different bacterial strains/species can exchange genes in a process known as congugation. I'm not going to go into this in detail, because it is already described in our archives (984777600.Mb). But, this means that bacterial species can pass resistance genes (e.g., rRNA or efflux pump genes) to each other. Its like bacterial file-sharing. :) This is why many antibiotic resistance genes are found on small, circular chromosomes known as plasmids, because plasmids can be transmitted more rapidly than the large main chromosome, and they can be lost when there are no antibiotics present without involving the rest of the crucial genes on the main chromosome.

However in today's environment, where human activity has spread antibiotic (and other antibacterial) drugs throughout the environment, there are antibiotic challenges to bacterial survival everywhere. This has resulted in a strong selection pressure for bacterial strains that maintain multiple resistance genes. From the bacterial point of view, there are potentially toxic fungi lurking around every corner.

So, what about question (2); why haven't bacteria been able to resist being cooked? Well, when we cook our food, we are very rapidly (over the course of minutes) raising it to high temperatures (>100 degrees C) that will denature proteins. Bacterial resistance to anything relies on the resistant cell's ability to replicate itself, which occurs on the order of a score of minutes. When a bacterial cell is cooked, all of it's proteins are denatured at the same time. There are organisms (primarily Archaea and Bacteria) that live quite well at high temperatures, known as thermophiles, but these organisms don't do so well at low temperatures (like 37 degrees C). The proteins in a bacterium that you would find on your food and that would grow in you and make you ill will function best at about your body temperature (which is why we culture so many of these at nice warm 37 degrees C). In order to function at temperatures >100 degrees C, proteins have to be modified at a large number of sites to make them much more stable at these high temperatures, and it seems likely that these proteins would not be functional at 37 degrees C.

For example, Cambillau and Claverie (below) compared the genomes of a variety of thermophilic organisms, and found that these organisms all had some protein characteristics in common; these organisms all have very large differences between the fraction of polar (noncharged) and charged residues in their proteins, with very high proportions of charged residues on the surface of proteins, which allows thermostable proteins to be stabilized at high temperatures via ionic bonds.

In addition, there are numerous examples of proteins found in thermophiles that are stable and active at high temperatures (70 - 100 degrees C), but which function poorly, or not at all, at 37 degrees C. I've cited three examples below; Zoldak et al describe NADH oxidase from Thermus thermophilus, which has optimal activity at 70 degrees C, Malhotra et al describe alpha-amylase from Bacillus thermooleovorans NP54, which has optimal activity at 100 degrees C, and Mandal et al describe a P-type Ag+/Cu+-ATPase from Archaeoglobus fulgidus that is active at 75 degrees C and inactive at 37 degrees C.

In addition, denatured proteins tend to aggregate, so that the cytoplasm sort of precipitates out of solution, which is something of a disaster. Bacteria do have mechanisms (Heat Shock proteins) that protect themselves from sudden, brief elevations in temperature by refolding denatured proteins, but these mechanisms are not effective for elevations as drastic as those that occur when we cook food.

So, antibiotic drugs target specific metabolic functions in a cell, and resistance to drugs like these can be achieved through single point mutations, or through the expression of a single protein coding gene. Cooking targets all of the functions of a cell, and causes them all to "crash" simultaneously. There's no single point mutation or single gene product that could protect a cell from being cooked.

I guess one could invent a scenario in which cooking developed gradually, with food first being cooked at 42 degrees C for a few hundred thousand years, and then at 52 degrees C for another hundred thousand or so, and then at 62, 72, 82, and 92 degrees C, until after 1.8 million years food was cooked at 102 degrees C, fostering the evolution of a thermophile, but at that point the organism wouldn't function at 37 degrees C, because that temperature would be too "cold" for its thermostable proteins.


You can find more specifics and information about bacterial resistance to antibiotics in any college- level Biochemistry textbook, such as Biochemistry by L. Stryer. In addition, you might want to read this paper about a mult-drug resistant ATP-Dependent efflux pump:

H Bolhuis, D Molenaar, G Poelarends, H W van Veen, B Poolman, A J Driessen, and W N Konings Proton motive force-driven and ATP-dependent drug extrusion systems in multidrug-resistant Lactococcus lactis. J Bacteriol. 1994 November; 176(22): 69576964.

These papers are advanced, but they are useful examples of the study of thermostable proteins and thermophiles.

Cambillau C, Claverie JM. Structural and genomic correlates of hyperthermostability. J Biol Chem. 2000 Oct 20;275 (42):32383-6.

Malhotra R, Noorwez SM, Satyanarayana T Production and partial characterization of thermostable and calcium-independent alpha-amylase of an extreme thermophile Bacillus thermooleovorans NP54. Lett Appl Microbiol. 2000 Nov;31(5):378-84.

Mandal AK, Cheung WD, Arguello JM Characterization of a thermophilic P-type Ag+/Cu+-ATPase from the extremophile Archaeoglobus fulgidus J Biol Chem, 2002 Mar 1, 277(9), 7201 - 7208.

Zoldak G, Sut'ak R, Antalik M, Sprinzl M, Sedlak E Role of conformational flexibility for enzymatic activity in NADH oxidase from Thermus thermophilus Eur. J. Biochem. 270, 4887-4897 (2003).

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