Methylotrophic Metabolism Is Advantageous for Methylobacterium extorquens during Colonization of Medicago truncatula under Competitive Conditions

Abstract

Facultative methylotrophic bacteria of the genus Methylobacterium are commonly found in association with plants. Inoculation experiments were performed to study the importance of methylotrophic metabolism for colonization of the model legume Medicago truncatula. Competition experiments with Methylobacterium extorquens wild-type strain AM1 and methylotrophy mutants revealed that the ability to use methanol as a carbon and energy source provides a selective advantage during colonization of M. truncatula. Differences in the fitness of mutants defective in different stages of methylotrophic metabolism were found; whereas approximately 25% of the mutant incapable of oxidizing methanol to formaldehyde (deficient in methanol dehydrogenase) was recovered, 10% or less of the mutants incapable of oxidizing formaldehyde to CO 2 (defective in biosynthesis of the cofactor tetrahydromethanopterin) was recovered. Interestingly, impaired fitness of the mutant strains compared with the wild type was found on leaves and roots. Single-inoculation experiments showed, however, that mutants with defects in methylotrophy were capable of plant colonization at the wild-type level, indicating that methanol is not the only carbon source that is accessible to Methylobacterium while it is associated with plants. Fluorescence microscopy with a green fluorescent protein-labeled derivative of M. extorquens AM1 revealed that the majority of the bacterial cells on leaves were on the surface and that the cells were most abundant on the lower, abaxial side. However, bacterial cells were also found in the intercellular spaces inside the leaves, especially in the epidermal cell layer and immediately underneath this layer.

Metaproteogenomic analysis of microbial communities in the phyllosphere and rhizosphere of rice

“…They were assigned to the genus Methylobacterium, a dominant member of the bacterial phyllosphere community. This finding is in agreement with observations on other plant species and underlines the importance of one-carbon metabolism for this genus upon phyllosphere colonization (Sy et al, 2005;Delmotte et al, 2009;Schmidt et al, 2010). Furthermore, a methanol:NDMA oxidoreductase was identified and assigned to Amycolatopsis (von Ophem et al, 1993), suggesting that other bacteria may also benefit from plant-released methanol in the rice phyllosphere, even though they are apparently less numerous.…”

A proteomic study of Methylobacterium extorquens reveals a response regulator essential for epiphytic growth

“…Among the proteins induced during bacterial growth in the phytosphere (leaf and root surface environments), key markers of methylotrophic metabolism (17) were found to be up-regulated (e.g., MxaF and Fae) with respect to growth on synthetic medium containing succinate as a carbon source ( Table 1). The induction of these enzymes upon epiphytic growth is in agreement with a previous study in which an advantage of wild-type (WT) M. extorquens cells in competition with methylotrophy-minus mutants was demonstrated, suggesting methanol utilization by the methylotroph (14). Another protein induced during phyllosphere colonization was PhaA, which initiates synthesis of the reserve polyhydroxy butyrate (PHB) (ref.…”

“…(19). The observed induction of PhaA might thus represent part of a general adaptation to nutrientlimiting conditions as would be expected for phyllospheric growth where the carbon source might not be a growth-limiting factor (14). The nature of this limiting factor is possibly suggested by the phyllosphere-specific induction of two putative periplasmic ABC transporter components predicted to be involved in iron and sulfate uptake (Table 1).…”

“…on plant surfaces benefit from methanol produced by plants (14) by means of methylotrophy (10,15). However, methanol is not the only carbon substrate that these bacteria are able to consume in the phyllosphere (14). The presence of Methylobacterium may be beneficial to plants through the production of plant hormones (13,16).…”

Methylobacterium phyllosphaerae sp. nov., a pink-pigmented, facultative methylotroph from the phyllosphere of rice

“…Their association with plants extends from free-living to epiphytic, endophytic and symbiotic (Pirttilä et al, 2000;Sy et al, 2001;Kutschera, 2007;Schauer & Kutschera, 2008) and their presence has been detected by cultivation-independent methods (Araú jo et al, 2002;Idris et al, 2004;Jackson et al, 2006). Interand intracellular colonization of plant tissues by Methylobacterium species and their mode of transmission from seed to aerial parts have been illustrated (Pirttilä et al, 2000;Sy et al, 2005;Poonguzhali et al, 2008).…”