Ion mobility spectrometry for microbial volatile organic compounds: a new identification tool for human pathogenic bacteria

Abstract

Presently, 2 to 4 days elapse between sampling at infection suspicion and result of microbial diagnostics. This delay for the identification of pathogens causes quite often a late and/or inappropriate initiation of therapy for patients suffering from infections. Bad outcome and high hospitalization costs are the consequences of these currently existing limited pathogen identification possibilities. For this reason, we aimed to apply the innovative method multi-capillary column-ion mobility spectrometry (MCC-IMS) for a fast identification of human pathogenic bacteria by determination of their characteristic volatile metabolomes. We determined volatile organic compound (VOC) patterns in headspace of 15 human pathogenic bacteria, which were grown for 24 h on Columbia blood agar plates. Besides MCC-IMS determination, we also used thermal desorption-gas chromatography-mass spectrometry measurements to confirm and evaluate obtained MCC-IMS data and if possible to assign volatile compounds to unknown MCC-IMS signals. Up to 21 specific signals have been determined by MCC-IMS for Proteus mirabilis possessing the most VOCs of all investigated strains. Of particular importance is the result that all investigated strains showed different VOC patterns by MCC-IMS using positive and negative ion mode for every single strain. Thus, the discrimination of investigated bacteria is possible by detection of their volatile organic compounds in the chosen experimental setup with the fast and cost-effective method MCC-IMS. In a hospital routine, this method could enable the identification of pathogens already after 24 h with the consequence that a specific therapy could be initiated significantly earlier.

Fig. 1

Summarized MCC-IMS topographic plot of bacterial volatiles, which can be used for bacterial discrimination. Fifteen (1–15) compounds have been determined in positive (a) and 16 (17–32) compounds have been determined in negative ion detection mode (b). All depicted compounds are listed in Table 1

Fig. 2

Region comparison image of MCC-IMS analyses of bacterial volatiles. Only monomeric MCC-IMS signals are shown. Red boxes indicate the presence of a VOC, which can be used for bacterial discrimination. Signals from 1 to 14 have been determined in positive ion mode, whereas signals from 17 to 29 have been determined in negative ion mode. An example of headspace VOCs of sheep blood agar and one example of headspace of every bacterial strain is displayed. All strains have been cultivated for 24 h. Signal intensities are illustrated by different colours (white = zero, blue = low, red = medium, yellow = high). The following signals have been used for further analyses: 1 ethanol, 2 indole, 3 phenethyl alcohol, 4 p_649_194, 5 p_675_194, 6 p_679_429, 7 p_700_423, 8 p_711_3, 9 p_749_26, 10 p_749_37, 11 p_771_54, 13 p_793_19, 14 p_794_12, 17 phenethyl alcohol, 18 p_493_17, 19 p_543_12, 20 p_563_24, 21 p_572_8, 22 2-(methylthio)-ethanol, 25 indole, 26 p_629_10, 27 p_654_20, 28 p_681_36 and 29 p_688_22

Fig. 3

MCC-IMS topographic plot of volatiles which have been detected in bacterial headspace but have not been chosen for discrimination of bacteria. Because either concentration of VOCs was too low in 10 mL headspace samples or high density of background signals of Columbia sheep blood agar, headspace prevents an unambiguous differentiation. For compound name, see Table 2

Fig. 4

Total ion chromatograms of bacterial headspace samples. For pre-concentration, VOCs of 2 L headspace volume have been adsorbed to Tenax GR. Total ion chromatograms are shown from 7 to 28 min. Identity of VOCs is summarized in Table 2. Cyclohexanol Rt = 12.17 min, styrene Rt = 12.35 min, cyclohexanone Rt = 12.45 min, benzaldehyde Rt = 14.14 min, 2-ethyl-1-hexanol Rt = 15.61 min and acetophenone Rt = 16.68 min have been detected as main volatile compounds over Columbia sheep blood agar. Benzothiazole—a contaminant of the operation gas—has been detected in every headspace sample at Rt = 20.37 min