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Abstract
Direct-infusion mass spectrometry (MS) was applied to study the metabolic effects of the symbiosis between the endophytic fungus Neotyphodium lolii and its host perennial ryegrass (Lolium perenne) in three different tissues (immature leaf, blade, and sheath). Unbiased direct-infusion MS using a linear ion trap mass spectrometer allowed metabolic effects to be determined free of any preconceptions and in a high-throughput fashion. Not only the full MS(1) mass spectra (range 150-1,000 mass-to-charge ratio) were obtained but also MS(2) and MS(3) product ion spectra were collected on the most intense MS(1) ions as described previously (Koulman et al., 2007b). We developed a novel computational methodology to take advantage of the MS(2) product ion spectra collected. Several heterogeneous MS(1) bins (different MS(2) spectra from the same nominal MS(1)) were identified with this method. Exploratory data analysis approaches were also developed to investigate how the metabolome differs in perennial ryegrass infected with N. lolii in comparison to uninfected perennial ryegrass. As well as some known fungal metabolites like peramine and mannitol, several novel metabolites involved in the symbiosis, including putative cyclic oligopeptides, were identified. Correlation network analysis revealed a group of structurally related oligosaccharides, which differed significantly in concentration in perennial ryegrass sheaths due to endophyte infection. This study demonstrates the potential of the combination of unbiased metabolite profiling using ion trap MS and advanced data-mining strategies for discovering unexpected perturbations of the metabolome, and generating new scientific questions for more detailed investigations in the future.
Highlights
Direct-infusion mass spectrometry (MS) was applied to study the metabolic effects of the symbiosis between the endophytic fungus Neotyphodium lolii and its host perennial ryegrass (Lolium perenne) in three different tissues
Much less is known about the impacts of fungal endophytes on general host plant performance and metabolism, and recent publications indicate that the effects of fungal endophytes on plant metabolism might be of importance to the understanding of ecosystem-wide impacts of the grassendophyte symbiosis (Hunt et al, 2005; Cheplick, 2007; Krauss et al, 2007; Rasmussen et al, 2007, 2008)
We present exploratory data analysis approaches to investigate how metabolites analyzed by DIMSn differ between endophyte-infected and uninfected ryegrass plants in three tissue samples corresponding to three developmental stages of the symbiosis of perennial ryegrass and N. lolii
Summary
Direct-infusion mass spectrometry (MS) was applied to study the metabolic effects of the symbiosis between the endophytic fungus Neotyphodium lolii and its host perennial ryegrass (Lolium perenne) in three different tissues (immature leaf, blade, and sheath). The two most widely studied associations are the perennial ryegrass-N. lolii symbiosis in Australasia and tall fescue (Lolium arundinaceum)Neotyphodium coenophialum in North America (Christensen et al, 1993) These two associations are of particular interest to agricultural pastoral systems because the fungal endophytes have been implicated in the toxicity of grazing livestock including ryegrass staggers and fescue foot, and found to confer a range of agronomic benefits to their grass hosts, mainly through toxicity and feeding deterrent activities toward invertebrate herbivores. We present exploratory data analysis approaches to investigate how metabolites analyzed by DIMSn differ between endophyte-infected and uninfected ryegrass plants in three tissue samples corresponding to three developmental stages (immature leaf, blade, and sheath) of the symbiosis of perennial ryegrass and N. lolii. Computational tools for harnessing the raw data from DIMSn experiments have been developed in this study and will be made available upon request
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