Diagnosing Melioidosis Through Breath Analysis Using GCxGC-TOF MS

The rapid diagnosis and appropriate treatment of melioidosis, a life-threatening infectious disease caused by Burkholderia pseudomallei (Bp) are critical to reduce mortality, yet diagnosis is hindered by diverse clinical manifestations, mimicry with other diseases, and reliance on slow culture-based methods. As the detection of volatile compounds offer a non-invasive approach for rapid infection detection, a joint study conducted by The University of British Columbia (Canada) and the Charles Darwin University Menzies School of Health Research (Australia) aimed to identify volatile compounds in patients’ breath that can aid in diagnosing melioidosis and indicating response to treatment. Breath samples were analyzed using comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (GCxGC-TOF MS). A paper based on this work was published in the Journal of Breath Research.¹

The clinical presentation of melioidosis can range from single skin abscesses at an inoculation site to pneumonia, bacteremia without evident focus, and severe sepsis with multiple internal organ abscesses, which makes its clinical diagnosis a challenge.^2-4Treatment for the disease usually involves an initial intensive phase with ceftazidime or a carbapenem for a minimum of 14 days, which is then followed by an eradication therapy with trimethoprim and sulfamethoxazole for at least three months.⁵

The research team collected breath samples from 17 patients with culture-confirmed melioidosis and eight patients with other febrile illnesses. Longitudinal samples were collected from five of the 17 melioidosis patients over approximately one month of antibiotic treatment, and data analysis involved statistical comparison and machine learning–based feature selection.¹

Three breath markers —camphene, 1-butanol, and 3-methylheptyl acetate —were identified that discriminated melioidosis (n=7) from febrile controls (n=6) with an area under the receiver operating characteristic curve of 1.00. These three markers correctly classified 11 additional samples from 11 melioidosis patients, with one febrile control misclassified. Separately, the researchers selected four breath markers, three of which were hydrocarbons, that differentiated samples associated with a positive Bp culture from those with a negative Bp culture, with a random forest model developed upon these four markers showing a sensitivity of 98% and specificity of 95%. In addition, the researchers were able to identify a set of 16 volatile compounds that significantly correlated (correlation coefficient > 0.6) with blood C-reactive protein levels. Finally, a panel of 144 volatile compounds was identified that corresponded to treatment time, which the researchers state may reflect treatment response or shifts in disease severity.¹

“This pilot study,” write the authors,¹ “reports candidate breath-based markers for diagnosing melioidosis and assessing treatment outcome, supporting further validation in larger studies.”

The researchers recommend future research involving multi-site clinical trials with increased enrolment and balanced representation between the melioidosis and control groups. Further serial breath studies in cohorts of patients reflecting the diversity of melioidosis presentations and severity can determine if breath profile changes towards the designated end point of intensive intravenous therapy can assist by further refining the optimum timing of switch from intravenous phase to oral eradication phase. In addition, the researchers propose that samples be analyzed within three months and including internal and external standards to account for potential storage- and analysis- related variability. Finally, employing high-resolution mass spectrometry for the accurate biomarker identification of biomarkers and the exploration of the markers' biological origins are, in their opinion, also essential next steps for future clinical applications.¹

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References

1. Gao, A.; Mayo, M.; Woerle, C. et al. Diagnosing Melioidosis and Tracking Treatment Outcomes Using Breath. J Breath Res. 2026.DOI: 10.1088/1752-7163/ae4bfd
2. Wiersinga, W.J.; Virk, H.S.; Torres, A.G. et al. Melioidosis. Nat. Rev. Dis. Primer 2018, 4, 17107, DOI: 10.1038/nrdp.2017.107
3. Currie, B. Melioidosis: Evolving Concepts in Epidemiology, Pathogenesis, and Treatment. Semin. Respir. Crit. Care Med. 2015, 36, 111–125. DOI: 10.1055/s-0034-1398389
4. Hoffmaster, A.R.; AuCoin, D.; Baccam, P. et al. Melioidosis Diagnostic Workshop, 20131. Emerg. Infect. Dis. 2015, 21.DOI: 10.3201/eid2102.141045
5. Meumann, E.M.; Limmathurotsakul, D.; Dunachie, S.J. et al. Burkholderia Pseudomallei and Melioidosis. Nat. Rev. Microbiol. 2024,22 (3), 155-169. DOI: 10.1038/s41579-023-00972-5