Microbial Volatile Organic Compounds Profiles Research Articles

Multi-strain volatile profiling of pathogenic and commensal cutaneous bacteria

The detection of volatile organic compounds (VOC) emitted by pathogenic bacteria has been proposed as a potential non-invasive approach for characterising various infectious diseases as well as wound infections. Studying microbial VOC profiles in vitro allows the mechanisms governing VOC production and the cellular origin of VOCs to be deduced. However, inter-study comparisons of microbial VOC data remains a challenge due to the variation in instrumental and growth parameters across studies. In this work, multiple strains of pathogenic and commensal cutaneous bacteria were analysed using headspace solid phase micro-extraction coupled with gas chromatography–mass spectrometry. A kinetic study was also carried out to assess the relationship between bacterial VOC profiles and the growth phase of cells. Comprehensive bacterial VOC profiles were successfully discriminated at the species-level, while strain-level variation was only observed in specific species and to a small degree. Temporal emission kinetics showed that the emission of particular compound groups were proportional to the respective growth phase for individual S. aureus and P. aeruginosa samples. Standardised experimental workflows are needed to improve comparability across studies and ultimately elevate the field of microbial VOC profiling. Our results build on and support previous literature and demonstrate that comprehensive discriminative results can be achieved using simple experimental and data analysis workflows.

VOC fingerprints: metabolomic signatures of biothreat agents with and without antibiotic resistance

Category A and B biothreat agents are deemed to be of great concern by the US Centers for Disease Control and Prevention (CDC) and include the bacteria Francisella tularensis, Yersinia pestis, Burkholderia mallei, and Brucella species. Underscored by the impact of the 2020 SARS-CoV-2 outbreak, 2016 Zika pandemic, 2014 Ebola outbreak, 2001 anthrax letter attacks, and 1984 Rajneeshee Salmonella attacks, the threat of future epidemics/pandemics and/or terrorist/criminal use of pathogenic organisms warrants continued exploration and development of both classic and alternative methods of detecting biothreat agents. Volatile organic compounds (VOCs) comprise a large and highly diverse group of carbon-based molecules, generally related by their volatility at ambient temperature. Recently, the diagnostic potential of VOCs has been realized, as correlations between the microbial VOC metabolome and specific bacterial pathogens have been identified. Herein, we describe the use of microbial VOC profiles as fingerprints for the identification of biothreat-relevant microbes, and for differentiating between a kanamycin susceptible and resistant strain. Additionally, we demonstrate microbial VOC profiling using a rapid-throughput VOC metabolomics method we refer to as ‘simultaneous multifiber headspace solid-phase microextraction’ (simulti-hSPME). Finally, through VOC analysis, we illustrate a rapid non-invasive approach to the diagnosis of BALB/c mice infected with either F. tularensis SCHU S4 or Y. pestis CO92.

Response of Wild Spotted Wing Drosophila (Drosophila suzukii) to Microbial Volatiles.

The olfactory cues used by various animals to detect and identify food items often include volatile organic compounds (VOCs) produced by food-associated microorganisms. Microbial VOCs have potential as lures to trap animal pests, including insect crop pests. This study investigated microorganisms whose VOCs are attractive to natural populations of the spotted wing drosophila (SWD), an invasive insect pest of ripening fruits. The microorganisms readily cultured from wild SWD and SWD-infested fruits included yeasts, especially Hanseniaspora species, and various bacteria, including Proteobacteria (especially Acetobacteraceae and Enterobacteriaceae) and Actinobacteria. Traps in a raspberry planting that were baited with cultures of Hanseniaspora uvarum, H. opuntiae and the commercial lure Scentry trapped relatively high numbers of both SWD and non-target drosophilids. The VOCs associated with these baits were dominated by ethyl acetate and, for yeasts, other esters. By contrast, Gluconobacter species (Acetobacteraceae), whose VOCs were dominated by acetic acid and acetoin and lacked detectable ethyl acetate, trapped 60-75% fewer SWD but with very high selectivity for SWD. VOCs of two other taxa tested, the yeast Pichia sp. and Curtobacterium sp. (Actinobacteria), trapped very few SWD or other insects. Our demonstration of among-microbial variation in VOCs and their attractiveness to SWD and non-pest insects under field conditions provides the basis for improved design of lures for SWD management. Further research is required to establish how different microbial VOC profiles may function as reliable cues of habitat suitability for fly feeding and oviposition, and how this variation maps onto among-insect species differences in habitat preference.

Monitoring MVOC Profiles over Time from Isolates of Aspergillus flavus Using SPME GC-MS

Fungi produce a variety of microbial volatile organic compounds (MVOCs) during primary and secondary metabolism. The fungus, Aspergillus flavus, is a human, animal and plant pathogen which produces aflatoxin, one of the most carcinogenic substances known. In this study, MVOCs were analyzed using solid phase microextraction (SPME) combined with GCMS from two genetically different A. flavus strains, an aflatoxigenic strain, NRRL 3357, and a non-aflatoxigenic strain, NRRL 21882. A PDMS/CAR SPME fiber was used over 30 days to observe variations in MVOCs over time. The relative percentage of individual chemicals in several chemical classes (alcohols, aldehydes, esters, furans, hydrocarbons, ketones, and organic acids) was shown to change considerably during the varied fungal growth stages. This changing chemical profile reduces the likelihood of finding a single chemical that can be used consistently as a biomarker for fungal strain identification. In our study, discriminant analysis techniques were successfully conducted using all identified and quantified MVOCs enabling discrimination of the two A. flavus strains over the entire 30-day period. This study underscores the potential of using SPME GCMS coupled with multivariate analysis for fungi strain identification.

Microbial volatile organic compounds in moldy interiors: A long‐term climate chamber study

The present study simulated large-scale indoor mold damage in order to test the efficiency of air sampling for the detection of microbial volatile organic compounds (MVOCs). To do this, a wallpaper damaged by condensation was stored in a climate chamber (representing a hypothetical test room of 40 m(3) volume) and was inoculated with 14 typical indoor fungal strains. The chamber ventilation conditions were adjusted to common values found in moldy homes, and the mold growth was allowed to continue to higher than average values. The MVOC content of the chamber air was analyzed daily for a period of 105 days using coupled gas chromatography/mass spectrometry (GC-MS). This procedure guarantees MVOC profiling without external factors such as outdoor air, building materials, furniture, and occupants. However, only nine MVOCs could be detected during the sampling period, which indicates that the very low concentrated MVOCs are hardly accessible, even under these favorable conditions. Furthermore, most of the MVOCs that were detected cannot be considered as reliable indicators of mold growth in indoor environments.

Identification of volatile markers for indoor fungal growth and chemotaxonomic classification of Aspergillus species

Microbial volatile organic compounds (MVOCs) were collected in water-damaged buildings to evaluate their use as possible indicators of indoor fungal growth. Fungal species isolated from contaminated buildings were screened for MVOC production on malt extract agar by means of headspace solid-phase microextraction followed by gas chromatography-mass spectrometry (GC–MS) analysis. Some sesquiterpenes, specifically derived from fungal growth, were detected in the sampled environments and the corresponding fungal producers were identified. Statistical analysis of the detected MVOC profiles allowed the identification of species-specific MVOCs or MVOC patterns for Aspergillus versicolor group, Aspergillus ustus, and Eurotium amstelodami. In addition, Chaetomium spp. and Epicoccum spp. were clearly differentiated by their volatile production from a group of 76 fungal strains belonging to different genera. These results are useful in the chemotaxonomic discrimination of fungal species, in aid to the classical morphological and molecular identification techniques.

Identification and profiling of volatile metabolites of the biocontrol fungus Trichoderma atroviride by HS-SPME-GC-MS

In the present study we describe a method, which is based on solid phase microextraction (SPME) coupled to gas chromatography–mass spectrometry (GC-MS) and which can be used for the profiling of microbial volatile organic compounds (MVOCs) in the headspace (HS) of cultures of filamentous fungi. The method comprises the following successive steps: 1. growth of the fungus on a solid culture medium directly in headspace vials, 2. measurement of volatiles by HS-SPME-GC-MS, 3. deconvolution of mass spectra, 4. identification of volatiles by comparison of measured, deconvoluted mass spectra and linear temperature programmed retention indices (LTPRI) on two stationary GC phases with database entries and LTPRI published in the literature, and 5. profiling of the identified MVOCs. The developed method was successfully applied to cultures of the biocontrol fungus Trichoderma atroviride. An in-house library consisting of mass spectra and LTPRI values of fungal VOCs was established and used to study the profiles of MVOCs of this fungus. In total, 25 different MVOCs were identified by applying strict criteria (spectral match factor at least 90% and a maximum relative deviation of LTPRI of ± 2% from literature values). The MVOCs were assigned to the compound classes of alcohols, ketones, alkanes, furanes, pyrones (mainly the bioactive 6-pentyl-alpha-pyrone), mono- and sesquiterpenes, 13 of which have never been reported to be produced by Trichoderma spp. before. Eleven of these volatiles have been additionally confirmed using authentic standards. Finally, time course experiments and cultivation of T. atroviride in the presence of the mycotoxin fusaric acid demonstrated the potential of the method to study the dynamics of MVOC profiles as well as the effect of different environmental/biological conditions on the expression of MVOCs of filamentous fungi.

Detection of Gaseous Effluents and By-Products of Fungal Growth that Affect Environments (RP-1243)

Musty odors, often associated with damp or water-damaged buildings, originate from the release of microbial volatile organic compounds (MVOCs) from mold growing on building materials and construction substrates. Chemical analysis of air samples is a feasible way to supplement conventional bioaerosol techniques during building investigations. Analytical methodologies for MVOCs are straightforward; however, development of a scientifically validated method to measure unique MVOCs that indicate with high confidence the presence of hidden mold regardless of the amount of mold present remains a challenge. Laboratory studies identified and quantified specific MVOCs associated with various mold species and MVOCs generated by specific molds growing on selected building materials in simulated, realistic conditions. This research determined that numerous MVOCs are released from active mold growth and are dependent on both the type of mold and the host substrate. MVOC profiles generated by 32 combinations of various molds and materials were determined, but only a few of these compounds demonstrated effectiveness in field/building studies. Certain mold-selective MVOCs were identified as potential indicators for specific mold, including methoxybenzene for Stachybotrys chartarum and benzothiazole and menthol for Chaetomium globosum. These studies provided a firm foundation for continued research of mold-specific MVOC markers as indicators of hidden mold and as predictors of potential mold sources in problem buildings.