Bacterial Leaf Symbiosis in Angiosperms: Host Specificity without Co-Speciation

Abstract

Bacterial leaf symbiosis is a unique and intimate interaction between bacteria and flowering plants, in which endosymbionts are organized in specialized leaf structures. Previously, bacterial leaf symbiosis has been described as a cyclic and obligate interaction in which the endosymbionts are vertically transmitted between plant generations and lack autonomous growth. Theoretically this allows for co-speciation between leaf nodulated plants and their endosymbionts. We sequenced the nodulated Burkholderia endosymbionts of 54 plant species from known leaf nodulated angiosperm genera, i.e. Ardisia, Pavetta, Psychotria and Sericanthe. Phylogenetic reconstruction of bacterial leaf symbionts and closely related free-living bacteria indicates the occurrence of multiple horizontal transfers of bacteria from the environment to leaf nodulated plant species. This rejects the hypothesis of a long co-speciation process between the bacterial endosymbionts and their host plants. Our results indicate a recent evolutionary process towards a stable and host specific interaction confirming the proposed maternal transmission mode of the endosymbionts through the seeds. Divergence estimates provide evidence for a relatively recent origin of bacterial leaf symbiosis, dating back to the Miocene (5–23 Mya). This geological epoch was characterized by cool and arid conditions, which may have triggered the origin of bacterial leaf symbiosis.

Endosymbiont Transmission Mode in Bacterial Leaf Nodulation as Revealed by a Population Genetic Study of Psychotria leptophylla

“…A previous study (13), however, rejected a history of strict cospeciation in leaf-nodulated Psychotria species and suggested a recent shift toward a host-specific association and ongoing cospeciation. To test this hypothesis, we analyzed a five-gene endosymbiont and twogene host data set, including 15 and 8 lineages and populations of Psychotria leptophylla, respectively.…”

“…Recent studies using culture-independent PCR methods revealed the identity of the endophytes of all leaf-nodulated plants as Burkholderia (10)(11)(12)24). More interestingly, it was also demonstrated that high host specificity occurs as a result of vertical symbiont transfer and that frequent host switches are observed, rejecting a long-term cospeciation (13). These results suggest a diffuse bacterium-plant interaction as an ancestral state of bacterial leaf symbiosis, followed by an evolution toward a more specific one-to-one interaction, where the bacterial associate became an obligate and vertically inherited endosymbiont of the host.…”

“…All plant species with voucher and GenBank accession numbers are listed in Table S1. DNA extraction, PCR amplification, and sequencing follow the method of Lemaire et al (13). Primer and temperature profiles are listed in Table S2.…”

“…The congruency between DNA markers was tested with the parsimony-based incongruence length difference test (ILD test) (4) as implemented in the software program PAUP* v.4b10 (23). ML and BI analyses were conducted as described in previous studies (12,13). For the BI analyses, the software program MrModeltest v.3.06 (20) selected an HKYϩI model for the gltB, recA, and external transcribed sequence (ETS) data sets, a GTRϩI model for the gyrB, lepA, and internal transcribed spacer (ITS) data sets, and a GTRϩG model for the trpB data set.…”

“…This result supports findings of previous morphological studies and the hypothesis that the symbionts are vertically transmitted. In Psychotria leptophylla, the presence of endosymbionts in every plant part is also expected because different populations of either P. kirkii or P. leptophylla are characterized by a host-specific endosymbiont (13). More important, the small amount of bacterial DNA within the seeds might explain why it remains so difficult to observe endosymbionts within the seed stage of the plant life cycle (16).…”