Chloroplast and nuclear ribosomal cistron phylogenomics in a group of closely related sections in Salvia subg. Calosphace

Abstract

Sage phylogenies have not resolved many shallow level clades within problematic Salvia subgenus Calosphace, hindered by the difficulty in finding sufficiently variable loci. We assembled fifteen chloroplast genomes and nuclear ribosomal cistrons from non-target reads of a nuclear hybrid enrichment project to assess the phylogenetic utility for a closely related group of mainly Mexican sections. Chloroplast synteny is confirmed with 10 Lamiales chloroplasts, supporting a map to reference assembly. Salvia miltiorrhiza Bunge chloroplast was used as a reference for assemblies, averaging 25,970 reads with mean depth of 20.71 reads; genomes ranged from 141,451 to 150,339 bp. The S. carduacea Benth. nuclear ribosomal cistron (18S, ITS1, 5.8S, ITS2 and 26S) was used as a reference for assembly, averaging 66,387 reads and mean depth of 1508 reads. We evaluated several partitioning schemes for plastid, plastid and nrDNA and reduced sampling, to assess whether these would render the same phylogenetic inferences. Maximum likelihood inferences resulted with high bootstrap support and two main clades with interspersed species from Salvia sect. Scorodoniae: clade A with species from S. sects. Atratae and Mitratae and clade B from S. sects. Sigmoideae and Uricae. Only S. sect. Uricae is monophyletic in every analysis supporting its identity, not merged into S. sect. Scorodoniae; S. sect. Sigmoideae is inferred monophyletic only if the ribosomal DNA sequences are included. Greater resolution and higher branch support are obtained with the entire plastome and nrDNA, rather than subsampling highly variable regions; thus we recommend this approach with expanded taxon sampling, coupled with a morphological review to better solve sectional circumscription in closely related S. subgenus Calosphace species.

Phylogenomics of Salvia L. subgenus Calosphace (Lamiaceae)

“…Here we support the monophyly of eight of the 13 Salvia sections sampled (Table 2): Biflorae, Curtiflorae, Hastatae, Incarnatae, Lavanduloideae, Sigmoideae, and also S. sections Cardinales and Uricae (as in Olvera-Mendoza et al, 2020). Although our tree is well-resolved, our Calosphace sample is <15% of the estimated species diversity in the subgenus, undoubtedly having an effect on clade resolution, and unsampled species could potentially be inserted in future phylogenetic studies to further resolve finescale relationships with each clade.…”

“…Chloroplasts were assembled in HPM using S. miltiorrhiza genome as a pseudoreference, obtaining a 92,461 bp assembly for the 90 Salvia samples evaluated (Supplementary Table 5). A map to reference approach was previously tested (Olvera-Mendoza et al, 2020) on 15 samples from these same data to FIGURE 1 | Exon recovery heat map for 527 putative exons targeted by our baits. Each column represents an exon, and each row is each species.…”

“…The "Core Calosphace" clade was initially described by and refers to a clade of "core radiation" that is "difficult to characterize morphologically but is well-supported in the molecular analyses...". It has been hypothesized that recent divergence events are clouding the phylogenetic signal, which could be further tested with expanded taxon sampling and additional phylogenetically informative sequence data (Olvera-Mendoza et al, 2020;Villaverde et al, 2020). This was attempted by Fragoso-Martínez et al (2017) and Kriebel et al (2019) using hybrid enrichment protocols across Salvia and to test sectional monophyly of the Calosphace.…”

A Cautionary Note on the Use of Genotype Callers in Phylogenomics

“…Yan et al, 2020), and plants (e.g. Nobre et al, 2018;Kreuzer et al, 2019;Helmstetter et al, 2020;Olvera-Mendoza et al, 2020;Brandrud et al, 2020;Wang et al, 2020).…”

“…For instance, countless studies over the past few years had access to only one single reference genome, if any, despite analyzing multiple species within entire genera. Such is the case for fish (e.g., Chakrabarty et al 2017 ; Burress et al 2018 ; Hulsey et al 2017 , 2018 ; Marcionetti et al 2019 ), mammals (e.g., Kumar et al 2017 ; Lima et al 2018 ; Moura et al 2020 ), birds (e.g., Ottenburghs et al 2016 ), reptiles (e.g., Bragg et al 2016 ), amphibians (e.g., Portik et al 2016 ), insects (e.g., Yan et al 2020 ), and plants (e.g., Nobre et al 2018 ; Kreuzer et al 2019 ; Helmstetter et al 2020 ; Olvera-Mendoza et al 2020 ; Brandrud et al 2020 ; Wang et al 2020 ). The situation can sometimes be even more dramatic with a single reference genome available for entire tribes or subfamilies, encompassing multiple genera (e.g., Hulsey et al 2017 ; Wang et al 2017 ; Hulsey et al 2018 ; Heckenhauer et al 2019 ; Loiseau et al 2019 ).…”