| Summary: | <i>Anchusa officinalis</i> (L.) interacts with various microorganisms including arbuscular mycorrhizal fungi (AMF). Recently, the AMF <i>Rhizophagus irregularis</i> MUCL 41833 has been shown to modulate the metabolome of <i>A. officinalis</i>. However, little information is available on the impact that different AMF species may have on primary and secondary plant metabolites. In this study, four AMF species belonging to the genus <i>Rhizophagus</i> (<i>R. irregularis</i> MUCL 41833, <i>R. intraradices</i> MUCL 49410, <i>R. clarus</i> MUCL 46238, <i>R. aggregatus</i> MUCL 49408), were evaluated for their potential to modulate <i>A. officinalis</i> metabolome under controlled semi-hydroponic cultivation conditions. An untargeted metabolomic analysis was performed using UHPLC-HRMS followed by a multivariate data analysis. Forty-two compounds were reported to be highly modulated in relation to the different AMF associations. Among them, six new secondary metabolites were tentatively identified including two acetyl- and four malonyl- phenylpropanoid and saponin derivatives, all presenting a common substitution at position C-6 of the glycosidic moiety. In addition, an enhanced accumulation of primary and secondary metabolites was observed for <i>R. irregularis</i> and <i>R. intraradices</i>, showing a stronger effect on <i>A. officinalis</i> metabolome compared to <i>R. clarus</i> and <i>R. aggregatus</i>. Therefore, our data suggest that different AMF species may specifically modulate <i>A. officinalis</i> metabolite production.
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