Methylobacterium is one of the most prevalent bacterial genera of the phyllosphere, present on the leaves of nearly every plant. Despite its ubiquity and its importance for host plant function, little is known about whether diversity over space and time within the genus reflects neutral processes like migration and drift, or environmental filtering of life history strategies and adaptations to temperature and host tree species. Here, we investigated how phylogenetic diversity within the genus Methylobacterium is structured by biogeography, seasonality, and growth strategies. Using deep, culture-independent barcoded marker gene sequencing coupled with culture-based approaches, we quantified seasonal shifts in Methylobacterium diversity over a year from early summer to fall in two temperate forests in Quebec, Canada. As an alternative to the 16S rRNA gene, we used rpoB, a highly polymorphic marker gene, which has not experienced detectable horizontal gene transfer nor copy number variation in Methylobacterium genomes. We cultured very different subsets of Methylobacterium diversity from the same leaf, depending upon the temperature of isolation and growth (20 C or 30 C). For instance, one the most abundant Methylobacterium lineages was almost exclusively isolated at 20 C, suggesting long-term adaptation to temperature. Both culture and barcoding approaches revealed that a considerable and previously underestimated diversity of Methylobacterium colonize the surface of leaves in temperate forests. This diversity was strongly structured according to both large (>100km; between forests) and small geographical scales (<1.2km within forests), and among host tree species, and was dynamic over seasonal time scales. To determine if these seasonal effects were driven by temperature, we measured growth of 79 isolates at different temperature treatments. Different lineages showed subtle and significant differences in growth performance when subject to temperature increase or decrease, but most have higher yield and slower growth rate at 20 than 30C, with strong lineage- and season-dependant variation in their overall growth strategies. We proposed that the homogenization of Methylobacterium community structure observed over a growing season resulted from the progressive replacement of isolates with high yield strategy typical of cooperative, structured communities, by isolates with rapid growth. Together our results show how Methylobacterium is phylogenetically structured into lineages with distinct growth strategies, which helps explain their differential abundance across regions, host tree species, and time. This works paves the way for further investigation of adaptive strategies and traits within a ubiquitous phyllosphere genus.
