It’s been clear for awhile that we’re running out of options against bacteria that resist multiple antibiotics (“superbugs”). In fact, that’s one of the “top 10 public health threats facing humanity”, according to the WHO.
Sheesh, like we need another one of those.
I mean, what happens when you get a staph infection, and antibiotics don’t work anymore? (I’ll spare you the gruesome images.) The ultimate results of this can be sepsis (where invading bacteria get into your bloodstream) and death (where you don’t continue living anymore). The Mayo Clinic says:
Sometimes the bacteria remain confined to the skin. But they can also burrow deep into the body, causing potentially life-threatening infections in bones, joints, surgical wounds, the bloodstream, heart valves and lungs.
Eek.
You’ve probably heard of MRSA in particular. That stands for “methicillin-resistant Staphylococcus aureus”, but we really ought to say “multidrug-resistant Staphylococcus aureus”. These are bacteria that have developed resistance to most or all of the antibiotics we have in our arsenal.
Drug-resistant infections already kill 35,000 people in the U.S. and 700,000 people worldwide per year, and that’s expected to increase exponentially if we don’t figure out new ways to combat them. That means roughly 10 million deaths by 2050.
We still rely on traditional antibiotics that stop the growth of bacteria, or just plain kill them. That’s a good short-term solution, but that means the only survivors are mutants that are antibiotic-resistant. Overuse of traditional antibiotics is an especially great way to concentrate these “superbugs” in our environment and magnify the problem. We need to stop carelessly applying antibiotics where they aren’t needed, but we also need new approaches to the problem altogether.
Enter Cassandra Quave. She and her team at Emory University and the University of Colorado have found a way to halt the spread of MRSA infections without the need to kill the bacteria. Instead, they disarm the bacteria by interfering with the trigger that causes them to make toxins. This way, the immune system can deal with these bacteria like it does most others. Their latest study was published June 28 in Frontiers in Pharmacology.
Quave has been interested in finding useful compounds in plants for some time, and she found out by doing interviews several years ago that people in southern Italy have used chestnut tree leaves for treating skin problems for many years. Her lab sifts through lots of anecdotal examples like this, and they try to find the promising ones.
Well, the chestnut tree turned out to be one of those promising ones. Five years ago Quave and company found that an extract made of chestnut tree leaves could quell the toxicity of MRSA by blocking a process used by these bacteria to decide when it’s time to crank out toxins.
Their extract contained thousands of compounds, and of course if you tell the FDA that your new therapeutic is “something or other within this potion”, they kinda frown on that. So the Quave lab needed to isolate the single chemical species that was responsible.
And ... this is where LEGO comes in. (Really, you’ll see!)
Usually when you separate an extract into its components, you use some kind of chromatography. That is, you pass your extract through beads or powder that different chemicals stick to with different levels of enthusiasm. The chemicals that don’t stick come out the other end first, and the ones that stick really well get held up and come out last.
A nice simple example of that is paper chromatography to separate black ink into its components. We use a solvent that will dissolve the black ink and carry it along, let it crawl up some paper (which is our sticking agent), and voilà!
Once Quave’s team separated the chestnut tree leaf extract in a chromatograph, they had to collect a lot of tiny samples at the other end in a precise and reproducible way. You can certainly buy a device that does this — why, here’s a nice one for $11,335! A young professor’s lab, though, often doesn’t have the money to cover all the fancy equipment for everybody’s projects. But the Quave lab was undaunted!
They built their own custom programmable fraction collector out of *LEGO* for $500! You just can’t make this stuff up. LEGO has a subset of products called MINDSTORMS that let kids … err, also researchers at universities? … build robotic devices. Marco Caputo from Quave’s group did just that, and he thus solved the problem of an affordable device that could work with the needed precision. Published it on the side, too, in order to help other labs “git ‘er done” as well. I mean, hats off to this group for not letting a limited budget stop them.
After a lot of testing of individual fractions for their ability to squelch toxin production by MRSA, and then following up with a whole lot of analytical techniques, they arrived at the responsible compound — castaneroxy A — and its structure, which is not unlike that of cholesterol:
They isolated castaneroxy A so precisely — thanks to that LEGO device, of course — that they were even able to make pure crystals of it.
They then applied small amounts of it to MRSA infections on mouse skin. With this treatment, the infections were totally unable to spread, while without treatment they did spread out and lasted much longer.
In the graph below, the difference is easy to see. “Vehicle” means a cream with no drug in it, and “10 μg” and “50 μg” mean a cream with a mere 10 or 50 micrograms of castaneroxy A in it. “MRSA Δagr” means MRSA bacteria whose ability to infect has been removed genetically — that’s your negative control. You can see that the lesion size with 50 micrograms of castaneroxy A is the same as that with the genetically hobbled bacteria: basically zilch. No spread of infection at all. (Luckily for the untreated mice, their infections did go away, but it took about 3 weeks.)
So how does castaneroxy A actually do that?
It doesn’t kill or even slow down the bacteria. Instead, it inhibits their quorum sensing, a process they use to detect a crowd. It’s a lot like us humans being in a packed gym. If enough people are in there, it becomes pretty evident to us by our sense of smell.
Bacteria have a simplified version of that. They exude a short little protein called autoinducing peptide (AIP) — basically bacterial B.O. — and they also have sensors to detect it. If they detect enough of it — that is, if there are enough other bacteria around (call it “groupstink”) — that’s what triggers their toxin production. One of these toxins is δ-hemolysin, which pokes holes in your cells’ membranes and causes them to explode. That’s … not good.
Here’s a cartoonish view of quorum sensing in Staphylococcus aureus:
The gist of what’s going on here is this: The purple squiggle (the AgrC protein) is the “stink” detector. It sits out there in the cell membrane and senses AIP outside the cell. When the AIP level gets high enough, AgrC sticks a phosphate group onto the AgrA protein, turning it into AgrA-P. Only when AgrA gets that “-P” can it land on DNA (the colored arrows at the bottom) and cause that DNA to be used to make proteins.
Each colored arrow is a gene, and each gene makes one or more proteins. Among those genes are the ones encoding the “stink” machinery itself: AgrD, the “stink” protein (that is, the unfinished form of AIP), and AgrB, the protein that folds AgrD into AIP and ships it out of the cell. So the overall effect is a bonkers exponential response: If a cell smells AIP, it makes more AIP, and this snowballs. The cells know it must be party time!
That aqua-colored gene (called RNAIII), which also gets turned on this way, encodes, among other nasties, δ-hemolysin. So, the “stink” signal also means, “Let’s make lots of toxins!”
Castaneroxy A interferes with this whole process by a mechanism that isn’t yet clear. It might stop AgrA-P from reaching DNA. It might get stuck on AgrC and prevent it from detecting anything. But one thing we do know is that small amounts of castaneroxy A (between 1 and 30 micrograms per milliliter) cut MRSA’s δ-hemolysin production down to near zero, while still allowing the MRSA to grow at near-normal levels.
One promising aspect of all this is that after 15 days stewing in a solution of castaneroxy A, the MRSA bacteria don’t develop any resistance to it. They can still grow just fine in that solution, so there’s no particular reason for them to mutate and get around it. Very unlike traditional antibiotics.
Castaneroxy A certainly won’t be the only compound we find or synthesize to address this problem, because other groups are at work on this as well. For the Quave lab’s part, they’ve already got another lead from the Brazilian peppertree.
While no single molecule will be the magic elixir for all future infections, the Quave lab and others have learned how to go through this process of finding new treatments, and they’re getting good at it. We are finally enabling ourselves to do more than ride the one-trick pony of antibiotics. True, it has been a really, really good trick, but we’ve got to learn new ones before infectious agents are on to us completely.