The discovery, made
in research laboratories at the University of Michigan Medical School and the
U.S. Food and Drug Administration, is published in the online journal PLoS
Pathogens. It helps solve a key mystery about C. diff: What triggers it to
germinate, or break its dormancy, from its hard spore form when it reaches the
gut.
Though the findings
were made in mice, not humans, the researchers say the crucial role of calcium
may help explain another mystery: Why some hospital patients and nursing home
residents have a much higher risk of contracting C. diff infections and the
resulting diarrhea that carries its spores out of the
body.
That group includes
people whose guts are flooded with extra calcium because they're taking certain
medications or supplements, have low levels of Vitamin D in their blood or have
gut diseases that keep them from absorbing calcium.
The new discovery
shows that C. diff can recognize this extra calcium, along with a substance
called bile salt produced in the liver, to trigger its awakening and the
breaking of its shell.
Previous research had
suggested it couldn't do this without another key component, an amino acid
called glycine. But the new findings show calcium and the bile salt called
taurochlorate alone are enough. Mouse gut contents that were depleted of gut
calcium had a 90 percent lower rate of C. diff spore germination.
"These spores
are like armored seeds, and they can pass through the gut's acidic environment
intact," says Philip Hanna, Ph.D., senior author of the new paper and a
professor of microbiology and immunology at U-M. "Much of the spore's own
weight is made of calcium, but we've shown that calcium from the gut can work
with bile salts to trigger the enzyme needed to activate the spore and start
the germination process."
Ironically, the
researchers say, one way to use this new knowledge in human patients might be
to add even more calcium to the system.
That could awaken all
the dormant C. diff spores in a patient's gut at once, and make them vulnerable
to antibiotics that can only kill the germinated form. That could also prevent
the transmission of more spores through diarrhea to the patient's room. That
could slow or stop the cycle of transmission that could threaten them or other
patients in the future.
Hanna's graduate
student, Travis Kochan, made a key observation that led to the discovery. He
noted that the fluid "growth medium" that the researchers typically
grow C. diff in for their studies had calcium in it. He realized this could
artificially alter the results of their experiments about what caused C. diff
spores to germinate.
So, he used a
chemical to remove the calcium while leaving all the other nutrients that keep
C. diff growing. The result: no new spore germination happened in the
calcium-free growth medium.
FDA's Center for
Biologics Evaluation and Research conducted further research in laboratory
dishes and in the guts of mice. FDA's Paul Carlson, Ph.D., a former U-M
research fellow, and his laboratory found that C. diff spores that were mutated
so that glycine couldn't act on them could still germinate and colonize mice.
This suggested that calcium, and not glycine, was critical for this process.
Both mutant and
regular forms of the bacteria could still activate an enzyme inside the C. diff
spore that led the bacteria to start dissolving their hard shell. This released
the store of calcium that the spore had been harboring inside itself, and
increases the local level of the nutrient even further.
"These spores
don't want to germinate in the wrong place," says Kochan, whose
grandfather suffered from a severe C. diff infection which ultimately led to
his death. "C. diff spores have specialized to germinate in the gut
environment, especially in the environment of the small intestine, where
calcium and the bile salt injection from the liver comes in."
Hanna notes that the
bile salt connection to C. diff spore germination was first discovered at U-M
in 1982 by a team led by Ken Wilson, M.D.
See:
Travis J. Kochan, Madeline J. Somers,
Alyssa M. Kaiser, Michelle S. Shoshiev, Ada K. Hagan, Jessica L. Hastie, Nicole
P. Giordano, Ashley D. Smith, Alyxandria M. Schubert, Paul E. Carlson, Philip
C. Hanna. Intestinal calcium and bile salts facilitate germination
of Clostridium difficile spores. PLOS Pathogens, 2017;
13 (7): e1006443 DOI: 10.1371/journal.ppat.1006443
Posted by Dr. Tim Sandle