Modelling Phagocytosis

Modelling Phagocytosis
Why Typhoid Bacteria Live Through It - a Project

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Elucidating the Strategy of the Bacteria which Cause Typhoid Fever

In the early 1990's, and recently joined to Washington University in St. Louis, Eduardo Groisman was the keynote speaker at an annual meeting of the Amer Soc for Microbiology. He showed a fascinating video clip his lab had made of macrophages' separately "eating" two different kinds of bacteria - Salmonella and benign E.coli. Here's what happened:

When the macrophage ingested the E.coli into a food vacuole, the cytoplasm churned and suddenly the E.coli cells inside the vacuole burst, and the remnants soon disappeared.
In the case where the macrophage ingested the Salmonella, the cytoplasm churned, and nothing happened for awhile. Then there were two Salmonella cells; then four; then eight; then the macrophage started to disintegrate.

Recognize that there are many different strains of Salmonella. The group that cause typhoid fever in humans is Salmonella typhi (not to be confused with typhus, which is a disease caused by an obligately intracellular prokaryot Rickettsia typhi. For obvious reasons human researchers shy away from working with S. typhi if less virulent relatives can be found. One of those is S. typhimurium (literally: mouse typhoid). All that were used by Groisman and reported on here has dealt with S. typhimurium. Also, for your edification and added confusion(!), most bacterial taxonomists include all the Salmonella species as really being virulent strains of Escherichia coli. Similarly, the Shigella species, which cause dysentery, are often spoken of as really being extremely virulent strains of otherwise benign Escherichia coli.

The means by which Salmonella escape being digested is not known. What is known, and which Groisman also determined, is that Salmonella possess approximately 28 virulence genes, and if any one of those is inactivated, the bacteria cannot cause a lethal infection in mice.

Because Groisman had moved on to other research interests by the time he showed his video, the problem is wide open for further study. He agrees that what he should have done as a follow up was to simultaneously feed Salmonella and E.coli to the macrophages, and then search for a cell in which there was a vacuole in which both types of bacteria were seen to be engulfed. In private conversation with him, we envisioned three different scenarios, and each would tell a lot about the strategy by which the Salmonella survive inside of the vacuole and eventually kill the white blood cell:

Three Scenarios for Survival

Both types of bacteria survive and multiply. This would mean that the Salmonella is putting out some sort of protective substance that either inactivates the chlorine containing digestion system, or perhaps prevents the merging of the lysosomes with the vacuole.

The Salmonolla survive and the E.coli burst. This means that the Salmonella are like Superman and the digestive enzymes bounce off, but the E.coli is killed.

Both types of bacteria die. This would indicate that Salmonella use stealth technology. The vacuole doesn't detect its presence, and perhaps the lysosomes are not beckoned to merge and spill the digestive enzymes into the vacuole. You might liken this to Salmonella's using deodorant, while E.coli stinks.

Three Scenarios for Survival
Both types of bacteria survive and multiply. This would mean that the Salmonella is putting out some sort of protective substance that either inactivates the chlorine containing digestion system, or perhaps prevents the merging of the lysosomes with the vacuole. The Salmonolla survive and the E.coli burst. This means that the Salmonella are like Superman and the digestive enzymes bounce off, but the E.coli is killed. Both types of bacteria die. This would indicate that Salmonella use stealth technology. The vacuole doesn't detect its presence, and perhaps the lysosomes are not beckoned to merge and spill the digestive enzymes into the vacuole. You might liken this to Salmonella's using deodorant, while E.coli stinks.

Now don't go gleefully running off thinking you can do this experiment with impunity! I can personally atest to the fact that S. typhimurium can make you VERY sick. Thus you would want to look for a safer way of solving which scenario is THE one used. You cannot reliably use an avirulent strain of S. typhimurium because it might not act normally in this setup. So you might look for a model system of completely different organisms which are completely harmless to you. For example, use amoeba and two other protozoans. One of those 'other' protozoans must be an insidious one which the amoeba phagocytoses, and once inside the amoeba, the sneaky protozoan starts digesting the amoeba. Paramecium, on the other hand is a common, easily digested food of amoeba.

What's neat about this protozoan triangle is that all the participating cells are huge compared to macrophages and bacteria. Indeed, you can just barely see them with the unaided eye. You thus will easily find low power microscopes that can "handle" what you need to look at. And if you get the system to work, get back to me and we can plan its presentation to the ASM. AND try to get your prof interested in it to be a coauthor. There is a very good reason for getting the prof involved - you won't have a deadline for getting it done.

Oh, and if you ever actually contact Eduardo, tell him that I still have his tennis racket, which he left behind in Milton Saier's lab in California.

So, invite your prof to lunch to talk this over. Never underestimate the importance of eating places in the advancement of science! Real science is a social activity.

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