Cophylogeny of the anther smut fungi and their caryophyllaceous hosts: prevalence of host shifts and importance of delimiting parasite species for inferring cospeciation

Abstract

Background: Using phylogenetic approaches, the expectation that parallel cladogenesis should occur between parasites and hosts has been validated in some studies, but most others provided evidence for frequent host shifts. Here we examine the evolutionary history of the association between Microbotryum fungi that cause anther smut disease and their Caryophyllaceous hosts. We investigated the congruence between host and parasite phylogenies, inferred cospeciation events and host shifts, and assessed whether geography or plant ecology could have facilitated the putative host shifts identified. For cophylogeny analyses on microorganisms, parasite strains isolated from different host species are generally considered to represent independent evolutionary lineages, often without checking whether some strains actually belong to the same generalist species. Such an approach may mistake intraspecific nodes for speciation events and thus bias the results of cophylogeny analyses if generalist species are found on closely related hosts. A second aim of this study was therefore to evaluate the impact of species delimitation on the inferences of cospeciation.

Results: We inferred a multiple gene phylogeny of anther smut strains from 21 host plants from several geographic origins, complementing a previous study on the delimitation of fungal species and their host specificities. We also inferred a multi-gene phylogeny of their host plants, and the two phylogenies were compared. A significant level of cospeciation was found when each host species was considered to harbour a specific parasite strain, i.e. when generalist parasite species were not recognized as such. This approach overestimated the frequency of cocladogenesis because individual parasite species capable of infecting multiple host species (i.e. generalists) were found on closely related hosts. When generalist parasite species were appropriately delimited and only a single representative of each species was retained, cospeciation events were not more frequent than expected under a random distribution, and many host shifts were inferred.Current geographic distributions of host species seemed to be of little relevance for understanding the putative historical host shifts, because most fungal species had overlapping geographic ranges. We did detect some ecological similarities, including shared pollinators and habitat types, between host species that were diseased by closely related anther smut species. Overall, genetic similarity underlying the host-parasite interactions appeared to have the most important influence on specialization and host-shifts: generalist multi-host parasite species were found on closely related plant species, and related species in the Microbotryum phylogeny were associated with members of the same host clade.

Conclusion: We showed here that Microbotryum species have evolved through frequent host shifts to moderately distant hosts, and we show further that accurate delimitation of parasite species is essential for interpreting cophylogeny studies.

Figure 1

Bayesian 50% majority-rule consensus tree of the Caryophyllaceae hosts used in this study based on the concatenation of the ITS, intron trnL and spacer trnLF. Statistical supports indicate Bayesian Posterior Probabilities (BPP)/Maximum Parsimony Bootstraps/Neighbor-Joining Bootstraps. Only nodes supported by more than two methods are indicated, the significant statistical supports being considered as higher than respectively 0.9/70/70. The tree is rooted based on previous studies (see text).

Figure 2

Bayesian 50% majority-rule consensus tree of the Microbotryum strains analyzed in this study based on the concatenation of β-tub, γ-tub and EF1α sequences, and delimitation of the corresponding species. Statistical supports indicate Bayesian Posterior Probabilities (BPP)/Maximum Parsimony Bootstraps/Neighbor-Joining Bootstraps. Only nodes supported by more than two methods are indicated, the significant statistical supports being considered as higher than respectively 0.9/70/70. High support in individual gene trees was indicated: + for the γ-tub tree, × for the Ef1α tree, # for the β-tub tree. The tree is rooted based on previous studies (see text). Taxa labels correspond to the host plant on which fungal strains were collected. Brackets indicate clades and evolutionary units, i.e. cryptic fungal species, identified in the study using the three individual gene phylogenies (see text).

Figure 3

Comparison of plant and fungal phylogenies using one strain per host species. Representation of the associations between Caryophyllaceae (left) and Microbotryum (right) with the a priori least congruent combinations between all possible resolved topologies for host and parasite trees, using one strain per host species (with 'Min' topology for the plant tree, and 'A1' topology for fungal tree, see Table 3). The symbol * highlights resolution of a polytomy in the plant tree according to a previous study [33]; The symbol ¤ highlights resolutions in the plant tree and in the fungal tree that differed from the other topology tested (Max versus Min in plants, and 2 versus 1 for fungi); The symbol ^- highlights resolutions that had no impact on congruence between the two phylogenies. Dots on the nodes indicate where cospeciation events were inferred by TreeMap.

Figure 4

Comparison of plant and fungal phylogenies using a single representative per fungal species. Representation of the associations between Caryophyllaceae (left) and Microbotryum (right) with the a priori least congruent combinations between all possible resolved topologies for host and parasite trees, using a single representative per fungal species (with 'MinT' topology for the plant tree, and 'B1' topology for fungal tree, See Table 3). See Fig. 3 for symbol legend.

Figure 5

Ecology of the host plants of the different Microbotryum species. Phylogenetic tree of the Microbotryum species with indication, on the branches, of their plant host clade (see Fig. 1 for host clade delimitation). The numbers in brackets indicate the five terminal incongruences between the plant and fungal phylogenies. Here MvDsp1 and MvDsp2 were not distinguished (fused in MvDsp) because of poor resolution in this part of plant phylogeny and thus inability to identify any incongruence between the two phylogenies. Information about the ecology of the host plants is given: the most common clades of pollinators are indicated in black and type of habitat in grey (Fields from the U.S. were drawn apart to highlight the geographical barrier to host shift). Boxes indicate ecological similarities of host plants which parasite species are closely related.