In Vitro Systems for Toxicity Evaluation of Microbial Volatile Organic Compounds on Humans: Current Status and Trends

Microbial volatile organic compounds (mVOC) are metabolic products and by-products of bacteria and fungi. They play an important role in the biosphere: They are responsible for inter- and intra-species communication and can positively or negatively affect growth in plants. But they can also cause discomfort and disease symptoms in humans. Although a link between mVOCs and respiratory health symptoms in humans has been demonstrated by numerous studies, standardized test systems for evaluating the toxicity of mVOCs are currently not available. Also, mVOCs are not considered systematically at regulatory level. We therefore performed a literature survey of existing in vitro exposure systems and lung models in order to summarize the state-of-the-art and discuss their suitability for understanding the potential toxic effects of mVOCs on human health. We present a review of submerged cultivation, air-liquid-interface (ALI), spheroids and organoids as well as multi-organ approaches and compare their advantages and disadvantages. Furthermore, we discuss the limitations of mVOC fingerprinting. However, given the most recent developments in the field, we expect that there will soon be adequate models of the human respiratory tract and its response to mVOCs.

Volatile Organic Compound Profiles Associated with Microbial Development in Feedlot Pellets Inoculated with Bacillus amyloliquefaciens H57 Probiotic

Mould and bacterial contamination releases microbial volatile organic compounds (mVOCs), causing changes in the odour profile of a feed. Bacillus amyloliquefaciens strain H57 (H57) has the potential ability to inhibit microbial growth in animal feeds. This study tested the hypothesis that H57 influences the odour profile of stored feedlot pellets by impeding the production of mVOCs. The emission of volatile organic compounds (VOCs) of un-inoculated pellets and those inoculated with H57, stored either at ambient temperature (mean 22 °C) or at 5 °C, was monitored at 0, 1, 2, and 3 months by gas chromatography–mass spectrometry. Forty VOCs were identified in all the pellet samples analysed, 24 of which were potentially of microbial and 16 of non-microbial origin. A score plot of the principal component analysis (PCA) showed that the VOC profiles of the pellets stored at ambient temperature changed more rapidly over the 3 months than those stored at 5 °C, and that change was greater in the un-inoculated pellets when compared to the inoculated ones. The bi-plot and correlation loading plots of the PCA indicated that the separation of the un-inoculated pellets from the other treatments over the 3 months was primarily due to nine mVOCs. These mVOCs have been previously identified in grains spoiled by fungi, and could be considered potential markers of the types of fungi that H57 can protect pellets against. These data indicate the ability of H57 to maintain the odour profile and freshness of concentrated feed pellets. This protective influence can be detected as early as 3 months into ambient temperature storage.

Articulating Fragrant Agarwood Formation as an Outcome of the Interaction between the Insect Zeuzera conferta and Aquilaria trees – A Review

Agarwood is the resinous infected wood obtained from Aquilaria species, which is a highly priced product in the flavour and fragrance market. Its formation is a complex process of interaction between the plant, insect, and microorganisms. Multiple studies concerning the interaction of microorganisms with the Aquilaria tree have been reported. However, the significant interaction between the insect Zeuzera conferta Walker (Lepidoptera: Cossidae) with Aquilaria has been overlooked, and only exiguous studies have been accomplished. Considering the dearth of available literature on this interesting phenomenon a review has been attempted. The taxonomical and morphological descriptions proffered by researchers and the insect life cycle are discussed. The review lays emphasis on the chemical ecology of the interaction between Z. conferta, Aquilaria and associating microorganisms as a possible continuum operating in the form of complex chemical signalling via release and sensing of Volatile Organic Compounds (VOCs), Herbivore Induced Plant Volatiles (HIPVs) and Microbial Volatile Organic Compounds (MVOCs). The review also scrutinizes the future perspectives of understanding the interaction in devising suitable management strategies to prevent uncontrolled infestation and, simultaneously, develop artificial rearing technology for the insect Z. conferta as a strategy for ensuring sustainable livelihood of farmers dependent on agarwood production.

Microbial Volatile Organic Compounds from Tempered and Incubated Grain Mediate Attraction by a Primary but Not Secondary Stored Product Insect Pest in Wheat

There has been a dearth of research elucidating the behavioral effect of microbially-produced volatile organic compounds on insects in postharvest agriculture. Demonstrating attraction to MVOC’s by stored product insects would provide an additional source of unique behaviorally-relevant stimuli to protect postharvest commodities at food facilities. Here, we assessed the behavioral response of a primary (Rhyzopertha dominica) and secondary (Tribolium castaneum) grain pest to bouquets of volatiles produced by whole wheat that were untempered, or tempered to 12%, 15%, or 19% grain moisture and incubated for 9, 18, or 27 days. We hypothesized that MVOC’s may be more important for the secondary feeder because they signal that otherwise unusable, intact grains have become susceptible by weakening of the bran. However, contrary to our expectations, we found that the primary feeder, R. dominica, but not T. castaneum was attracted to MVOC’s in a wind tunnel experiment, and in a release-recapture assay using commercial traps baited with grain treatments. Increasing grain moisture resulted in elevated grain damage detected by near-infrared spectroscopy and resulted in small but significant differences in the blend of volatiles emitted by treatments detected by gas chromatography coupled with mass spectrometry (GC–MS). In sequencing the microbial community on the grain, we found a diversity of fungi, suggesting that an assemblage was responsible for emissions. We conclude that R. dominica is attracted to a broader suite of MVOC’s than T. castaneum, and that our work highlights the importance of understanding insect-microbe interactions in the postharvest agricultural supply chain.

Learning about microbial language: possible interactions mediated by microbial volatile organic compounds (VOCs) and relevance to understanding Malassezia spp. metabolism

Background Microorganisms synthesize and release a large diversity of small molecules like volatile compounds, which allow them to relate and interact with their environment. Volatile organic compounds (VOCs) are carbon-based compounds with low molecular weight and generally, high vapor pressure; because of their nature, they spread easily in the environment. Little is known about the role of VOCs in the interaction processes, and less is known about VOCs produced by Malassezia, a genus of yeasts that belongs to the human skin mycobiota. These yeasts have been associated with several dermatological diseases and currently, they are considered as emerging opportunistic yeasts. Research about secondary metabolites of these yeasts is limited. The pathogenic role and the molecular mechanisms involved in the infection processes of this genus are yet to be clarified. VOCs produced by Malassezia yeasts could play an important function in their metabolism; in addition, they might be involved in either beneficial or pathogenic host-interaction processes. Since these yeasts present differences in their nutritional requirements, like lipids to grow, it is possible that these variations of growth requirements also define differences in the volatile organic compounds produced in Malassezia species. Aim of review We present a mini review about VOCs produced by microorganisms and Malassezia species, and hypothesize about their role in its metabolism, which would reveal clues about host-pathogen interaction. Key scientific concepts of review Since living organisms inhabit a similar environment, the interaction processes occur naturally; as a result, a signal and a response from participants of these processes become important in understanding several biological behaviors. The efforts to elucidate how living organisms interact has been studied from several perspectives. An important issue is that VOCs released by the microbiota plays a key role in the setup of relationships between living micro and macro organisms. The challenge is to determine what is the role of these VOCs produced by human microbiota in commensal/pathogenic scenarios, and how these allow understanding the species metabolism. Malassezia is part of the human mycobiota, and it is implicated in commensal and pathogenic processes. It is possible that their VOCs are involved in these behavioral changes, but the knowledge about this remains overlocked. For this reason, VOCs produced by microorganisms and Malassezia spp. and their role in several biological processes are the main topic in this review.

Monitoring the volatile language of fungi using gas chromatography-ion mobility spectrometry

Fusarium oxysporum is a plant pathogenic fungus leading to severe crop losses in agriculture every year. A sustainable way of combating this pathogen is the application of mycoparasites—fungi parasitizing other fungi. The filamentous fungus Trichoderma atroviride is such a mycoparasite that is able to antagonize phytopathogenic fungi. It is therefore frequently applied as a biological pest control agent in agriculture. Given that volatile metabolites play a crucial role in organismic interactions, the major aim of this study was to establish a method for on-line analysis of headspace microbial volatile organic compounds (MVOCs) during cultivation of different fungi. An ion mobility spectrometer with gas chromatographic pre-separation (GC-IMS) enables almost real-time information of volatile emissions with good selectivity. Here we illustrate the successful use of GC-IMS for monitoring the time- and light-dependent release of MVOCs by F. oxysporum and T. atroviride during axenic and co-cultivation. More than 50 spectral peaks were detected, which could be assigned to 14 volatile compounds with the help of parallel gas chromatography-mass spectrometric (GC-MS) measurements. The majority of identified compounds are alcohols, such as ethanol, 1-propanol, 2-methyl propanol, 2-methyl butanol, 3-methyl-1-butanol and 1-octen-3-ol. In addition to four ketones, namely acetone, 2-pentanone, 2-heptanone, 3-octanone, and 2-octanone; two esters, ethyl acetate and 1-butanol-3-methylacetate; and one aldehyde, 3-methyl butanal, showed characteristic profiles during cultivation depending on axenic or co-cultivation, exposure to light, and fungal species. Interestingly, 2-octanone was produced only in co-cultures of F. oxysporum and T. atroviride, but it was not detected in the headspace of their axenic cultures. The concentrations of the measured volatiles were predominantly in the low ppbv range; however, values above 100 ppbv were detected for several alcohols, including ethanol, 2-methylpropanol, 2-methyl butanol, 1- and 3-methyl butanol, and for the ketone 2-heptanone, depending on the cultivation conditions. Our results highlight that GC-IMS analysis can be used as a valuable analytical tool for identifying specific metabolite patterns for chemotaxonomic and metabolomic applications in near-to-real time and hence easily monitor temporal changes in volatile concentrations that take place in minutes.