Basidiomycetes Are Particularly Sensitive to Bacterial Volatile Compounds: Mechanistic Insight Into the Case Study of Pseudomonas protegens Volatilome Against Heterobasidion abietinum

Abstract

Volatile organic compounds (VOCs) play an important role in the communication among organisms, including plants, beneficial or pathogenic microbes, and pests. In vitro, we observed that the growth of seven out of eight Basidiomycete species tested was inhibited by the VOCs of the biocontrol agent Pseudomonas protegens strain CHA0. In the Ascomycota phylum, only some species were sensitive (e.g., Sclerotinia sclerotiorum, Botrytis cinerea, etc.) but others were resistant (e.g., Fusarium oxysporum f. sp. cubense, Verticillium dahliae, etc.). We further discovered that CHA0 as well as other ten beneficial or phytopathogenic bacterial strains were all able to inhibit Heterobasidion abietinum, which was used in this research as a model species. Moreover, such an inhibition occurred only when bacteria grew on media containing digested proteins like peptone or tryptone (e.g., Luria-Bertani agar or LBA). Also, the inhibition co-occurred with a pH increase of the agar medium where the fungus grew. Therefore, biogenic ammonia originating from protein degradation by bacteria was hypothesized to play a major role in fungus inhibition. Indeed, when tested as a synthetic compound, it was highly toxic to H. abietinum (effective concentration 50% or EC50 = 1.18 M; minimum inhibitory concentration or MIC = 2.14 M). Using gas chromatography coupled to mass spectrometry (GC/MS), eight VOCs were found specifically emitted by CHA0 grown on LBA compared to the bacterium grown on potato dextrose agar (PDA). Among them, two compounds were even more toxic than ammonia against H. abietinum: dimethyl trisulfide had EC50 = 0.02 M and MIC = 0.2 M, and 2-ethylhexanol had EC50 = 0.33 M and MIC = 0.77 M. The fungus growth inhibition was the result of severe cellular and sub-cellular alterations of hyphae occurring as early as 15 min of exposure to VOCs, as evidenced by transmission and scanning electron microscopy observations. Transcriptome reprogramming of H. abietinum induced by CHA0’s VOCs pointed out that detrimental effects occurred on ribosomes and protein synthesis while the cells tried to react by activating defense mechanisms, which required a lot of energy diverted from the growth and development (fitness cost).

Designing a synthetic microbial community devoted to biological control: The case study of Fusarium wilt of banana

“…This was not a surprising outcome since different formae speciales of F. oxysporum have been earlier reported as resistant to microbial antagonists in similar in vitro studies. For instance, Foc R1 and TR4 showed resistance to volatile compounds emitted by Pseudomonas protegens CHA0, while several Basidiomycetes and Ascomycetes were sensitive (Prigigallo et al, 2021). Also, F. oxysporum f. sp.…”

“…The overlapping plate assay was performed according to Prigigallo et al (2021). A 6 mm-agar plug of Trichoderma spp.…”

“…Protease, catalase, phosphatase, phytase, β-glucosidase, 1aminocyclopropane-1-carboxylate (ACC) deaminase, and amylase activities, as well as the production of 2,3-butanediol, hydrogen cyanide (HCN), and siderophores, were assessed as previously described (Gómez-Lama Cabanás et al, 2018a and references therein). Ammonia (NH 3 ) production was determined using Nessler's reagent as described by Prigigallo et al (2021). Moreover, the capability of three Pseudomonas spp.…”

Mechanisms of Action of Microbial Biocontrol Agents against Botrytis cinerea

“…For example, P. chlororaphis ZL3 was evaluated for its antifungal activity, and 23 VOCs were found [140]. In the same way, P. protegens CHAo was evaluated for inhibition of several fungi, being positive for those related to B. cinerea, because of the production of two volatile molecules, ammonia and dimethyl trisulfide [141]. Most experiments of this type have been performed in vitro.…”

Pulcherrimin: a bacterial swiss army knife in the iron war

“…There is multiple evidence that volatiles play an important role in modulating microbial communities 41,42 . Notably, Pseudomonas species' volatiles are diverse, and they can antagonize other microorganisms, including fungal and bacterial plant pathogens [43][44][45] .…”