A rare melioidosis case: from pathogen to commensal

Granuloma formation observed in the lung of a chronically ill C57BL/6 mouse infected by Burkholderia pseudomallei.
Granuloma formation observed in the lung of a chronically ill C57BL/6 mouse infected by Burkholderia pseudomallei. One of Tuanyok’s studies aims to characterize host proteins that are involved with a chronic infection, such as programmed cell death protein 1, which is stained dark brown in this image. (Image by Treenate Jiranantasak, a doctoral student in Tuanyok’s lab)

In a case study that spans sixteen years, a UF assistant professor in the College of Veterinary Medicine’s infectious diseases and immunology department helped investigate a rare and unusual case of melioidosis. The findings offer an exceptional window into how a highly lethal and pathogenic bacteria can adapt to life within a host.

Apichai Tuanyok, who is a member of UF’s Emerging Pathogens Institute, has spent his career studying Burkholderia pseudomallei, the causative agent of melioidosis. Even with proper treatment, the survival rate for melioidosis is only about 60 percent. Without treatment, it is always fatal. Melioidosis is a tropical disease traditionally found in Southeast Asia and Australia, although its range is expanding into the Americas.

When someone becomes infected, they require a unique cocktail of antibiotics administered over 14 days within a hospital and then continued for 20 weeks at home. There is no vaccine, although Tuanyok is involved in efforts to develop one. B. pseudomallei is innately resistant to most antibiotics. 

But an Australian woman who contracted the pathogen got a lucky break when mutations occurred in the bacteria she carried, which rendered them less and less virulent. Named “patient 314” to protect her privacy, researchers obtained 118 samples from her sputum over 16 years. Tuanyok was involved in studying the samples collected from this patient’s case, which offers extraordinary insight into how a highly pathogenic bacteria can transition to a commensal state within a single host. The research study was published this spring in PLOS-Pathogens.

“In my whole career, I’ve never seen anything like this,” Tuanyok says. “Around two years after her initial infection, we found a mutation in bacteria collected from this patient and, we saw that it had begun to adapt to its host.” 

The only other chronic infection cases with this particular bacterium tend to involve people who also have the lung disease cystic fibrosis; patient 314 did not. However, the patient did have permanently damaged bronchial tubes, a condition known as bronchiectasis, due to past lung infections. Despite receiving treatment, and the patient’s symptoms improving, researchers are still able to culture B. pseudomallei from the patient’s sputum.

Tuanyok’s research team performed whole-genome sequencing on the patient’s 16-year collection of samples. Because they had samples spanning an extended period, they were able to read the story of the bacteria’s mutation and adaptation to its host over time.

The story had striking similarities to a clonal relative of B. pseudomalleiB. mallei, which has adapted to life as a chronic respiratory pathogen of horses. Many of the gene deletions that occurred in B. pseudomallei within patient 314 also occurred within B. mallei in horses, Tuanyok notes.

Burkholderia produce something called O-antigen, which is a structural component of the bacteria’s outer membrane. O-antigen is a known virulence factor, and it tends to create a robust humoral immune response from an infected host. But as the B. pseudomallei inside patient 314 adapted to its new environment, Tuanyok’s team identified that the bacteria underwent “genomic reduction,” which is a process that deletes portions of an organism’s genome. In this case, it lost the genes responsible for producing O-antigen, thereby allowing the bacteria to persist inside patient 314 by not triggering her immune system. Genome reduction has also occurred with B. mallei within horses. It lost genes that confer resistance to certain antibiotics and genes known to assist in survival outside a host within the environment.

The research team sampled the patient’s home environment and found six of 21 samples from its well-water source that closely matched early samples taken from the patient’s sputum. Comparing the environmental isolates to the patient-derived samples showed that the bacteria began changing rather quickly after infecting its host; these changes are likely attributable to the randomness of genetic drift.

Other changes, such as the loss of O-antigen and loss of antibiotic resistance, took longer to emerge and might be the result of selective pressures.

“We think it probably has something to do with how this bacterium adapts to life within a host’s lungs,” Tuanyok says.

The patient has not been on antibiotics for the past five years, yet continues to test culture-positive for B. pseudomallei. Patient 314 is the only known case where anti-B. pseudomallei therapy was stopped even though the bacteria persist in their living host. The patient’s entire population of B. pseudomallei is thought to now be nonpathogenic.

“Most patients die from acute infections with melioidosis, not many researchers study the patients who develop chronic infections,” Tuanyok says. “This reveals a way we can obtain samples noninvasively from the patient’s airway, which may be helpful for taking a second look at chronic infections.”

Tuanyok plans to use the mutated strains from patient 314 for vaccine development because he suspects these unique virulence attenuated strains provoke cell-mediated immunity without producing disease symptoms.

“We have the strains here at UF,” Tuanyok says. “We’d like to look into doing some genetic engineering with them to advance our work searching for a melioidosis vaccine.”

Written by: DeLene Beeland