Evolution of antibiotic resistance through multicopy plasmids

It’s lab journal club day tomorrow! So in honor of that, I’ll take it as a chance to work on writing general summaries of papers. SciComm! -Ryan

By now, you’ve probably heard that antimicrobial resistance is a major clinical problem. The ‘golden age’ of antimicrobials is coming to an end as the existing repertoire of clinically used antibiotics is becoming less effective and more bacteria are becoming resistant to common antibiotics. Resistance typically arises through ‘target mutation’ (modifying the target of an antibiotic inside the cell), preventing entry of the antibiotic altogether, or directly degrading or modifying the antibiotic so it’s no longer active.

Genes associated with antibiotic resistance are often found on plasmids, circular pieces of DNA that are distinct from the bacterial chromosome. Plasmids are small compared to chromosomal DNA –  on the scale of 1000s of base pairs rather than millions. The amount of a plasmid within a single cell, referred to as a plasmid’s ‘copy number’, can vary from one to hundreds of copies.

Through a process called ‘horizontal gene transfer’ (HGT), plasmids can be passed between bacteria – notably, from a bacteria that is antibiotic-resistant to one that is antibiotic-sensitive. While this sounds like a good thing (for bacteria), maintenance of a plasmid comes at the price of fitness, the ability to produce progeny cells (or in this case, for the bacterium to keep dividing). Antibiotic resistance genes can find their way into the bacterial chromosome, getting rid of the need for a cell to keep the plasmid (and the fitness cost of doing so), yet plasmids still persist in the bacterial population. A recent study by San Millan et al. published in Nature Ecology and Evolution investigated the advantage for bacteria to maintain antibiotic resistance on plasmids versus the chromosome.

The authors used a strain of a common lab bacterium, Escherichia coli, that had a gene encoding for penicillin resistance on the bacterial chromosome or on a multicopy plasmid. When cells that had the plasmid and cells without the plasmid were grown together, cells without the plasmid were able to grow better. This showed that maintenance of a plasmid had an associated fitness cost. When exposed to penicillin, the plasmid-bearing bacterium was more resistant to the antibiotic.

Then, the bacteria were grown in the presence of a second antibiotic called ceftazidime. Normally, the gene they used for penicillin resistance does not lead to ceftazidime resistance. However, exposure to increasing amounts of ceftazidime over several days eventually led to antibiotic resistance when the gene was on the plasmid but not the chromosome. This experiment showed something important – where an antibiotic resistance gene is located (on a plasmid versus the chromosome) can provide an additional benefit.

The next thing they did was to look at why the plasmid encoded gene did this and the chromosome encoded one did not. They saw that when the bacteria gained ceftazidime resistance, there were key mutations in the DNA sequence of the plasmid. The antibiotic resistance gene itself gained a mutation that made the bacterium 128-times more resistant to ceftazidime than without that mutation. It is hypothesized that the presence of multiple copies of the plasmid can increase the rate at which beneficial mutations can occur, and this is why resistance is less likely to arise when the antibiotic resistance gene is located in one copy on the chromosome.

Even though this is already a huge increase in resistance, another set of mutations was found in a region called the ‘origin’ of the plasmid that could increase resistance by upwards of 256-times. The origin is a region that codes for factors that control the copy number and replication of a plasmid. Mutations in this region could increase copy number of the plasmid by upwards of 6 times, multiplying the effects of the original mutation in the antibiotic resistance gene.

So what does all this mean? Well, bring it back to the original question of ‘what is the advantage of encoding antibiotic resistance on a plasmid?’ First, having the resistance gene for a given antibiotic on a multicopy plasmid means you can produce more of that gene product. Second, the multicopy plasmid helps the antibiotic resistance gene itself to evolve and propagate. Because the many copies of the plasmid can be present, the rate that new mutations can appear increases as well. Once a beneficial mutation happens, its beneficial effects can be amplified because there are many copies of the plasmid.

When a cell divides, it gives rise to two ‘daughter cells’. The plasmids carrying the antibiotic resistance gene are randomly divided between the daughter cells. Between this and natural replication of the plasmid, the daughter cells will have more copies of the plasmids with beneficial mutations relative to the parent cell. Over many generations of bacteria, this leads to effective propagation and amplification of the antibiotic resistance gene.

So why is all this important? In the big picture of antimicrobial resistance, it helps us understand why maintaining plasmids can be advantageous for bacteria. It also demonstrates a potential way for bacteria to promote the rise of mutations that lead to antibiotic resistance.


Other items of interest:


San Millan, A., Escudero, J.A., Gifford, D.R., Mazel, D., and MacLean, R.C. 2016. Multicopy plasmids potentiate the evolution of antibiotic resistance in bacteria. Nat. Ecol. and Evol. doi:10.1038/s41559-016-0010

Podcasts I’ve been listening to:


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