# Antimicrobial Volatiles of the Insect Pathogen _Metarhizium brunneum_
## 4 Discussion
The current study showed that _Candidia_ yeasts and test bacteria exhibit similar patterns of sensitivity to _M. brunneum_ VOCs, with the cells undergoing autolysis, ultimately leading to total cell degradation since no cell wall fragments or any other cell debris was evident in the inhibition zone. Biotic and environmental stresses are known to trigger autolysis in bacteria [53]. The degree of inhibition or lysis is clearly dependent upon the VOC dose but there is some evidence that the VOC must make direct contact with the target since cells hidden by other cells often survived and even flourished, presumably utilising the nutrients released from the adjacent lysed cells. In the soil environment, the lysate may be utilised by other microbes. Thus, EPF VOCs may influence the microbial profile in the rhizosphere. Indeed, analysis of the plant microbiome after _Metarhizium_ amendment reveals an increase in the abundance of plant-growth-promoting organisms [54]. Only a tiny fraction of compounds produced by _M. brunneum_ were screened; it is possible that the blend produced in nature is more potent than the individual compounds. In other words, some combination of VOCs may act synergistically, as reported for several bacteria species [19; 55]. Synergies would conserve energy since fewer bioactive compounds need to be produced. Since _M. brunneum_ produces a wide range of VOCs, the synergies between different combinations of VOCs may allow the fungus to compete with a wide range of microbial antagonists. This study showed that cultural conditions can influence VOC profiles. This plasticity would allow _M. brunneum_ to adapt to its environment and compete with indigenous microbes. Indeed, it would also explain the success of this and other EPF as they recovered from disparate habitats (soils, crops, etc.) around the world. This confirmed that these organisms possess the appropriate armoury to survive in what is recognised as a highly competitive environment [2; 56]. There is much evidence showing that the microbial VOC profile is altered depending on environmental conditions, with novel compounds often being produced, especially under conditions of stress [19]. Stress factors may be environmental or biotic, such as exposure to antagonists, as illustrated by the mycophagous bacterium, i.e., _Collimonas fungiporans_, which only produces VOCs with strong antifungal activity in the presence of susceptible fungi [12]. The natural antimicrobial VOCs identified in this study have the potential for use as plant protection products. Their ephemeral nature and action through fumigation mean that they could be used to clean soils of harmful diseases or protect produce from post-harvest diseases. They would be relatively safer than many current chemical fumigants, which are highly toxic and often require long safety period. Indeed, many conventional chemical pesticides have been withdrawn due to their persistence and environmental pollution. The following supporting information can be downloaded at: [[https://www.mdpi.com/article/10.3390/jof8040326/s1](https://www.mdpi.com/article/10.3390/jof8040326/s1)]([https://www.mdpi.com/article/10.3390/jof8040326/s1](https://www.mdpi.com/article/10.3390/jof8040326/s1)), Table S1: Metarhizium brunneum volatile organic compounds (VOCs) tested for biocidal activity; Table S2: Bacterial, fungal and oomycete targets used to test VOC potency; Table S3: Metarhizium brunneum VOCs identified by NIST. The list shows compounds produced independent of strain, developmental stage (mycelium vs conidia), production in vivo or in vitro. The list includes compounds produced consistently as well as those produced intermittantly; Table S4: VOCs produced by Metarhizium brunneum ARSEF4556, ARSEF3297 and V275 on different substrates. **Author Contributions:** Conceptualisation, E.H.H. and T.M.B. ; Formal analysis, E.H.H. ; Funding acquisition, T.M.B. ; Investigation, E.H.H., T.M.B., E.D. and J.L. ; Methodology, E.H.H., Y.C., T.M.B., M.D., N.M.K., S.K., E.D., T.M. and J.L. ; Project administration, T.M.B.