Jean-Paul Latgé originally wanted to know if he could test the breath of patients with Aspergillus infections for volatile compounds produced by the fungus. His group at the Pasteur Institute in Paris thought this might be a new way of diagnosing fungal culprits like Aspergillus fumigatus that often colonize the lungs of cystic fibrosis (CF) and immunocompromised patients.
His team began investigating the volatile compounds produced by Aspergillus but the work soon overlapped with another part of the lab, where Benoit Briard, a graduate student at the time, was characterizing molecules released by the bacterium Pseudomonas aeruginosa—another common opportunistic lung pathogen found in the same vulnerable patients. The overlap led Latgé’s team to veer into completely unexpected territory—showing that a ‘scent’ released by the bacteria can spur the growth of fungus at a distance.
The finding, published this week in mBio was surprising because other researchers including Briard had already shown that Pseudomonas releases inhibitory compounds that stunt the growth of its pathogenic rival when the two are in contact with each other. “In the literature, most papers are looking at inhibition of growth by other molecules and volatiles,” says Latgé. “To our big surprise, the volatiles produced by Pseudomonas were promoting the growth of the fungus.” [Image: The lung pathogen Aspergillus fumigatus fungus (small plate, upper right) grows rapidly toward Pseudomonas aeruginosa bacteria (large plate, lower right) when co-cultured compared to when grown alone (left panel)]
It was so counterintuitive, that Latgé instructed postdoctoral fellows Christoph Heddergott and Briard to repeat the experiments several times over. They easily did so, growing the Aspergillus in a smaller Petri dish situated to the side of a larger dish containing an already-established culture of Pseudomonas. Physically separated by the plastic dishes, the microbes shared the airspace above the dish surface.
“It was nicely reproducible,” says Heddergott. “When you simply put these two organisms together and wait a couple of days, the fungus grows faster and grows toward the bacteria.” Clearly, the bacteria were emitting something stimulatory. But what?
“I have to confess it was a rather stinky story,” says Heddergott. He placed an absorptive fiber into the airspace of the co-cultures to capture all volatile compounds for analysis by gas chromatography/mass spectrometry.
Next, he went shopping online and purchased all of the individual volatile compounds made by Pseudomonas to test on Aspergillus cultures. The smelliest one, dimethyl sulfide, stimulated the fungus to grow in the same way and at the same levels as co-culturing did.
“That microbes ‘smell’ other microbes is not new,” says Latgé. “But that one species of bacteria is stimulating the growth of a fungus by ‘smell’—this is totally new.” Even more impressive, the fungus actually uses the essential nutrient sulfur from the volatile dimethyl sulfide as fuel to grow.
Fungus grown on plates lacking sulfur grew better when dimethyl sulfide was pumped into the airspace. Also, Heddergott measured how much of the dimethyl sulfide was consumed from the air by the fungus. “In some cases, even if there are other sulfur sources on the plate, the fungus prefers the sulfur in the volatile compounds,” he says.
The finding opens up the possibilities for how these two specific pathogens—and more generally how all microbes—interact and communicate across distances. Heddergott says that figuring out how microbiota inside the human host interact through volatile-based signaling is a new research frontier.
As for these particular pulmonary microbes, knowing that bacteria can promote fungal growth adds another layer of understanding to how these co-infections progress. “If a patient has a bacterial colonization first, then the volatiles being produced could help the fungus germinate and grow, giving it a huge advantage in establishing itself,” says Latgé.
He says the simplest explanation for why the bacteria might be stimulating fungal growth at a distance, but inhibiting it after contact could be that when the two pathogens meet they are competing directly for food. But from afar, “Pseudomonas, stupidly, is producing something that Aspergillus likes and uses.”
He notes that it’s unknown if these signals operate the same way in the natural environment as they do in the laboratory. “Pseudomonas and Aspergillus are often present in the same ecological niches, but we haven’t looked yet to see what volatiles are doing between the bacteria and fungi in the soil where they live.”
Investigating how volatiles may influence interactions between bacteria, fungi, and amoebas in the soil is another area ripe for discovery. Heddergott’s labmates would surely be happy to see him take his work outside. When testing his smelly sulfur-containing compounds, he says he tried to run his experiments over the weekends “for the good of all.”