A tale of two lineages: how the strains of the earliest divergent symbiotic Frankia clade spread over the world

Abstract

It is currently assumed that around 100 million years ago, the common ancestor to the Fabales, Fagales, Rosales and Cucurbitales in Gondwana, developed a root nodule symbiosis with a nitrogen-fixing bacterium. The symbiotic trait evolved first in Frankia cluster-2; thus, strains belonging to this cluster are the best extant representatives of this original symbiont. Most cluster-2 strains could not be cultured to date, except for Frankia coriariae, and therefore many aspects of the symbiosis are still elusive. Based on phylogenetics of cluster-2 metagenome-assembled genomes (MAGs), it has been shown that the genomes of strains originating in Eurasia are highly conserved. These MAGs are more closely related to Frankia cluster-2 in North America than to the single genome available thus far from the southern hemisphere, i.e., from Papua New Guinea.To unravel more biodiversity within Frankia cluster-2 and predict routes of dispersal from Gondwana, we sequenced and analysed the MAGs of Frankia cluster-2 from Coriaria japonica and Coriaria intermedia growing in Japan, Taiwan and the Philippines. Phylogenetic analyses indicate there is a clear split within Frankia cluster-2, separating a continental from an island lineage. Presumably, these lineages already diverged in Gondwana.Based on fossil data on the host plants, we propose that these two lineages dispersed via at least two routes. While the continental lineage reached Eurasia together with their host plants via the Indian subcontinent, the island lineage spread towards Japan with an unknown host plant.

Taxogenomic status of phylogenetically distant Frankia clusters warrants their elevation to the rank of genus: A description of Protofrankia gen. nov., Parafrankia gen. nov., and Pseudofrankia gen. nov. as three novel genera within the family Frankiaceae

“…Cluster 1 contains as-yet-uncultured Frankia microsymbionts, which are characterised by profuse sporulation within nodules tissues (Sp + types; Schwintzer, 1990 ), and a very higher proportion of cluster 2 strains are as-yet-uncultured. The Candidatus status has been used to accommodate some of these uncultured Frankia which were defined based on genome sequences ( Persson et al, 2011 , 2015 ; Nguyen et al, 2016 , 2019 ; Normand et al, 2017 ; Herrera-Belaroussi et al, 2020 ; Pozzi et al, 2020 ; Berckx et al, 2022 ). Two strains from cluster 2 have been successfully cultivated following a dual approach consisting of comparative genomics and direct physiological assay on nodule tissues ( Gtari et al, 2015 ; Gueddou et al, 2019 ).…”

Genome analysis and biogeographic distribution of the earliest divergent Frankia clade in the southern hemisphere

“…The association of each cluster with its group of host plants has been discussed in several publications (Benson and Dawson 2007 , Svistoonoff et al 2014 , Nguyen et al 2016 ). Frankia cluster-2 is the earliest divergent symbiotic clade within the genus (Nguyen et al 2016 , 2019 , Berckx et al 2022a ), and its host plants include Ceanothus (Rhamnaceae, Rosales), the Dryadoideae except for Dryas octopetala (Rosaceae, Rosales), Datiscaceae (Cucurbitales), and Coriariaceae (Cucurbitales).…”

“…Several more metagenome-assembled genomes (MAGs) of Frankia cluster-2 were sequenced from whole nodules obtained from both greenhouse cross-inoculation experiments and field samples. Here it was found that several closely related strains not only occupy a single nodule (Normand et al 2017 , Nguyen et al 2019 , Berckx et al 2022a ), but that some strains can persist as nodule-associated bacteria under unfavourable conditions and become the main symbiont in a more suitable host plant (Berckx et al 2022a ). Based on the analysis of the core genomes of Frankia cluster-2 strains from greenhouse samples, it could be concluded that the strains occurring in Eurasia, namely from France to Japan, form a single group with very low diversity (Nguyen et al 2019 ).…”

“…These strains were found to be the direct sister group of the Frankia cluster-2 strains occurring in North America, presumably by spreading via the Bering Strait (Nguyen et al 2019 ). Further analysis of cluster-2 termed the Eurasian and North American strains the ‘continental’ lineage, to contrast with the ‘island’ lineage: those Frankia cluster-2 strains which occur on islands in the Pacific Ocean, specifically Japan, Taiwan, the Philippines, and Papua New Guinea (Nguyen et al 2019 , Berckx et al 2022a ). Our previous study highlighted the importance of sequencing field nodules: while studies using cross-inoculation studies or isolation of greenhouse nodules of D. glomerata and C. myrtifolia found continental Frankia cluster-2 strains in Japan (Gtari et al 2015 , Nguyen et al 2019 ), sequencing field samples directly identified island lineage Frankia to be the main strains present in C. japonica nodules (Berckx et al 2022a ).…”

“…In light of the recent, whole genome analysis of Frankia cluster-2 (Berckx et al 2022a ), we were interested to see if Frankia strains occurring in New Zealand form a third lineage within cluster-2, or if they group with the strains making up the island lineage. Since Coriaria spp.…”