Quantification of hydrogen cyanide and 2-aminoacetophenone in the headspace of Pseudomonas aeruginosa cultured under biofilm and planktonic conditions

Abstract

Hydrogen cyanide (HCN) and 2-aminoacetophenone (2-AA; H₂NC₆H₄COCH₃) are possible biomarkers of pulmonary Pseudomonas aeruginosa (PA) infection that could be used in an exhaled breath test. All factors affecting their production need to be investigated, including the culture conditions: planktonic (free-floating) or biofilm (non-motile communities attached to a solid surface). In vivo, the change from planktonic to biofilm growth is signalled when a certain population density is reached. Using selected ion flow tube mass spectrometry, SIFT-MS, we have analysed HCN and 2-AA produced by 12 genotyped PA samples, cultured under both planktonic and biofilm conditions after 24, 48, 72 and 96 hours of incubation. The 12 samples included 3 different strains (genotypes), 50% of which had a mucoid phenotype and 50% had a non-mucoid phenotype. All samples produced significant concentrations of HCN; median (25^th to 75^th percentiles, IQR) concentration: 144 (61–512) parts-per-billion by volume (ppbv). Multivariate analysis showed HCN production varied dependent on genotype (p = 0.0014), culture duration (p = 0.005) and phenotype (p < 0.001) but not culture conditions (planktonic/biofilm). Much smaller concentrations of 2-AA were detected, median (IQR) concentration 1.8 (1.3–3) ppbv, despite which, multivariate analysis showed production was affected by genotype (p < 0.001) and culture duration (p = 0.007) but not phenotype or culture conditions. These data show that biofilm formation does not affect HCN production by PA and supports its use as a biomarker of PA infection. The concentrations of 2-AA are much lower than previous studies have shown. The reason for this is unclear but it raises questions about its suitability as a biomarker of PA infection.