Background and Aims Zanthoxylum 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. Conclusions 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.

中文翻译：

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目标富集提高了花椒属（芸香科）的系统发育分辨率，并表明不完整的谱系分类和杂交事件

背景和目的 花椒是芸香科中唯一的泛热带属，少数物种原产于温带东亚和北美洲。使用 Sanger 测序的努力未能解决花椒的骨干系统发育。在这项研究中，我们采用目标富集高通量测序来提高分辨率。检查基因树的一致性并评估花椒的截面分类。研究了脱靶读数以识别用于诱饵细化的推定单拷贝标记，以及用于推定杂交事件证据的低拷贝标记。方法针对花椒设计了一个针对354个基因、中位数为321 bp的定制诱饵集，并应用于44个花椒属植物和一种作为外群的花椒属植物。Illumina 读数是通过 HybPhyloMaker 管道处理的。系统发育推断是使用基于连接数据集的合并和最大似然方法进行的。使用四重采样评估一致性。对从脱靶读数中提取的假定单拷贝和低拷贝基因进行了额外的系统发育分析。主要结果 花椒属中支持四个主要进化枝：非洲进化枝、Z. asiaticum 进化枝、亚太-澳大利亚进化枝和美洲-东亚进化枝。虽然总体支持有所改善，但冲突地区与之前观察到的地区相似。基因树不一致表明夏威夷血统的祖先发生了杂交事件，而美国骨干中的血统分类不完整。脱靶推定的单拷贝基因在很大程度上证实了在靶结果，和推定的低拷贝基因为夏威夷谱系中的杂交提供了额外的证据。花椒的五个部分中只有两个被确定为单系。结论 靶点富集适用于评价花椒的系统发育关系。我们的系统发育分析表明，当前的分区分类需要修订。四重树一致性表明网状进化的几个实例。脱靶读数被证明可用于识别额外的系统发育信息区域，以进行诱饵细化或基于基因树的方法。我们的系统发育分析表明，当前的分区分类需要修订。四重树一致性表明网状进化的几个实例。脱靶读数被证明可用于识别额外的系统发育信息区域，以进行诱饵细化或基于基因树的方法。我们的系统发育分析表明，当前的分区分类需要修订。四重树一致性表明网状进化的几个实例。脱靶读数被证明可用于识别额外的系统发育信息区域，以进行诱饵细化或基于基因树的方法。