Target enrichment improves phylogenetic resolution in the genus Zanthoxylum (Rutaceae) and indicates both incomplete lineage sorting and hybridization events

Abstract

Background and aims Zanthoxylum L. is the only pantropical genus within Rutaceae, with a few species native to temperate eastern Asia and North America. Efforts using Sanger sequencing failed to resolve the backbone phylogeny of Zanthoxylum. In this study, we employed target enrichment high-throughput sequencing to improve resolution. Gene trees were examined for concordance and sectional classifications of Zanthoxylum were evaluated. Off-target reads were investigated to identify putative single-copy markers for bait refinement, and low-copy markers for evidence of putative hybridization events. Methods A custom bait set targeting 354 genes, with a median of 321 bp, was designed for Zanthoxylum and applied to 44 Zanthoxylum species and one Tetradium species as the outgroup. Illumina reads were processed via the HybPhyloMaker pipeline. Phylogenetic inferences were conducted using coalescent and maximum likelihood methods based on concatenated datasets. Concordance was assessed using quartet sampling. Additional phylogenetic analyses were performed on putative single- and low-copy genes extracted from off-target reads. Key results Four major clades are supported within Zanthoxylum: the African clade, the Z. asiaticum clade, the Asian-Pacific-Australian clade, and the American-eastern Asian clade. While overall support has improved, regions of conflict are similar to those previously observed. Gene tree discordances indicate a hybridization event in the ancestor of the Hawaiian lineage, and incomplete lineage sorting in the American backbone. Off-target putative single-copy genes largely confirm on-target results, and putative low-copy genes provide additional evidence for hybridization in the Hawaiian lineage. Only two of the five sections of Zanthoxylum are resolved as monophyletic. Conclusion Target enrichment is suitable for assessing phylogenetic relationships in Zanthoxylum. Our phylogenetic analyses reveal that current sectional classifications need revision. Quartet tree concordance indicates several instances of reticulate evolution. Off-target reads are proven useful to identify additional phylogenetically informative regions for bait refinement or gene tree based approaches.

Plastome sequences fail to resolve shallow level relationships within the rapidly radiated genus Isodon (Lamiaceae)

“…Moreover, our results demonstrate that plastome data does not accurately reflect species relationships and is insufficient to resolve shallow level relationships within a rapidly radiated genus like Isodon . To achieve for a better resolved phylogeny of the genus, significant larger data sets derived from nuclear genome with next-generation sequencing technology might be helpful, such as targeted enrichment ( Gardner et al, 2020 ; Reichelt et al, 2021 ; Giaretta et al, 2022 ), deep genome skimming ( Liu et al, 2021 ), and RNA-seq ( Nevado et al, 2016 ; Jin et al, 2021 ; Kong et al, 2022 ; Xia et al, 2022 ), which are compatible with more comprehensive tools for disambiguating genetic lineages and causes of gene tree conflicts.…”

Ancient DNA extraction methods for herbarium specimens: When is it worth the effort?

“…Under these circumstances, assembling complete plastomes might be difficult. Instead, focusing on the most abundant plastid and nuclear ribosomal regions could be a valuable alternative (Reichelt et al, 2021 ; Šlenker et al, 2021 ). Nevertheless, our data support the potential to assemble entire plastid genomes from herbarium samples up to 200 years old using standard extraction and sequencing methods (Bakker, 2017 ; Alsos et al, 2020 ).…”

DNA barcoding of Cymbidium by genome skimming: Call for next‐generation nuclear barcodes

“…Therefore, in plant phylogenetics, low−/single‐copy nuclear gene markers from higher sequencing depths have been used to resolve the relationships of closely related species (Jiang et al, 2022; Pillon et al, 2013; Sang, 2002; Zhao et al, 2016). Currently, large‐scale low‐/single‐copy nuclear data have also been increasingly adopted, such as in Zanthoxylum (Rutaceae) (Reichelt et al, 2021) and Senecioneae (Asteraceae) (Ren et al, 2021), offering new opportunitites for elucidating the rapid radiation and the origin of hybridization in plant species (Berger et al, 2017; Ding et al, 2012; Granados Mendoza et al, 2015; Johnson et al, 2019; Pillon et al, 2013; Zhang et al, 2012; Zimmer & Wen, 2012). Thus, nuclear genome data, such as provided by low−/single‐copy nuclear genes, is expected to be used in plant DNA barcoding, particularly for recently radiated or reticulately evolved taxa, such as C .…”