Symbiotic ß-Proteobacteria beyond Legumes: <em>Burkholderia</em> in <em>Rubiaceae</em>

<div><p>Symbiotic ß-proteobacteria not only occur in root nodules of legumes but are also found in leaves of certain <em>Rubiaceae</em>. The discovery of bacteria in plants formerly not implicated in endosymbiosis suggests a wider occurrence of plant-microbe interactions. Several ß-proteobacteria of the genus <em>Burkholderia</em> are detected in close association with tropical plants. This interaction has occurred three times independently, which suggest a recent and open plant-bacteria association. The presence or absence of <em>Burkholderia</em> endophytes is consistent on genus level and therefore implies a predictive value for the discovery of bacteria. Only a single <em>Burkholderia</em> species is found in association with a given plant species. However, the endophyte species are promiscuous and can be found in association with several plant species. Most of the endophytes are part of the plant-associated beneficial and environmental group, but others are closely related to <em>B. glathei</em>. This soil bacteria, together with related nodulating and non-nodulating endophytes, is therefore transferred to a newly defined and larger PBE group within the genus <em>Burkholderia</em>.</p> </div

Optimization of the <i>Vanguerieae</i>-<i>Burkholderia</i> interaction on a simplified representation of the host plant tree.

<p>In <i>Vanguerieae</i>, the association with <i>Burkholderia</i> endophytes has occurred three times independently (blue clades). The circles are pie charts that represent the chance of presence and absence of endophytes per node. However, because the Bayesian posterior probabilities are so high, they are or completely yellow (absence of endophytes) or completely blue (presence of endophytes). Support values for either absence (yellow) or presence (blue) are indicated below the nodes.</p

Phylogenetic tree of <i>Vanguerieae</i> with presence (blue) and absence (yellow) of endophytic <i>Burkholderia</i> bacteria.

<p>This phylogenetic dendrogram of the plant tribe <i>Vanguerieae</i> is adapted from Verstraete et al. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0055260#pone.0055260-Verstraete2" target="_blank">[11]</a>. Bacterial endosymbiosis is found in three groups of the tribe (arrows). Four species were not investigated because we did not have the specimens, and for seven other species we did not find <i>Burkholderia</i> endophytes although their phylogenetic position suggests so (not coloured). Bayesian posterior probabilities/bootstrap values are indicated below the branches.</p

SEM micrographs of the host plant <i>Fadogia homblei</i>.

<p>A. The upper leaf surface before sterilization with a lot of undefined debris. B. The lower leaf surface before sterilization with some debris present. C. After sterilization the upper leaf surface is almost entirely devoid of particles and no epiphytes are visible. D. On the lower leaf surface there are no epiphytes visible after sterilization. E. Cross section through a leaf showing the mesophyll cells with scattered bacterial endophytes. F. Detail of the endophytes in the intercellular space. G. The host plant <i>Fadogia homblei</i> does not have dark bacterial galls on its leaf blades.</p

<i>Burkholderia</i> tree with the phylogenetic position of the newly discovered endophytes.

<p>This phylogenetic dendrogram is based on the concatenated dataset of the 16S rRNA gene, <i>gyrB</i> gene and <i>recA</i> gene. Several clades are evident: the pathogenic group, a clade with <i>B. glathei</i> and related endophytes, and the plant-associated beneficial and environmental (PBE) group sensu Suárez-Moreno et al. (arrows). Most of the new endophytes fall in the latter group, except for the bacteria in <i>Globulostylis</i>, which are more related to the previously described <i>Candidatus</i> species. We therefore combine the <i>B. glathei</i> clade with the PBE group sensu Suárez-Moreno et al., to form a broadly defined PBE group. Bayesian posterior probabilities/bootstrap values are indicated below the branches.</p

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