# Characteristic analysis of the chloroplast genome of Cyathula capitata and comparative analysis with Cyathula officinalis and their hybrid Cyathula officinalis × Cyathula capitata

## TL;DR

This study analyzes the chloroplast genomes of Cyathula capitata, Cyathula officinalis, and their hybrid to understand genetic variation and phylogenetic relationships, aiding in species identification and conservation.

## Contribution

The study provides novel insights into chloroplast genome structure and variation in Cyathula species and their hybrid, offering markers for species identification and quality control.

## Key findings

- Chloroplast genomes of Cyathula species show typical quadripartite structure with notable intraspecific diversity in C. capitata.
- Hexanucleotide SSRs, a unique insertion in rpl22, and a forward repeat in rps16_1-trnQ-UUG distinguish C. officinalis and hybrids.
- Phylogenetic analysis shows hybrid Z4 groups within C. capitata, supporting maternal inheritance of chloroplast genomes.

## Abstract

Cyathula capitata (Wall.) Moq., a widely used medicinal herb in Yi medicine, is often combined with other herbs to treat traumatic injuries, rheumatism, and rheumatoid arthritis. This study aims to characterize the chloroplast genomes, assess variation levels, and elucidate the phylogenetic relationships among Cyathula capitata, Cyathula officinalis, and their hybrid (Cyathula officinalis × Cyathula capitata). The findings provide valuable references for species identification, genetic background analysis, and quality control of Cyathula medicinal materials.

We employed second-generation high-throughput sequencing technology to sequence the chloroplast genomes of Cyathula capitata, Cyathula officinalis, and their hybrid (Cyathula officinalis × Cyathula capitata). Comparative genomic analyses were conducted to examine their genomic structures, simple sequence repeats (SSRs), codon usage preferences, and inverted repeat (IR) regions. Additionally, a phylogenetic tree based on chloroplast genome sequences was constructed to clarify their evolutionary relationships.

The chloroplast genomes of all three taxa displayed the typical quadripartite structure, comprising a large single-copy region, a small single-copy region, and two IR regions (IRa and IRb). The Cyathula capitata genomes ranged from 151,428 to 151,436 bp, showing notable intraspecific diversity likely influenced by geographic factors. However, two C. capitata samples shared an identical genome length of 151,518 bp and exhibited consistent genotypes across locations, indicating relative genomic conservation. Repeat sequence analysis identified hexanucleotide SSRs, a unique 16-bp single nucleotide insertion in the rpl22 gene, and a 30–40 bp forward repeat within the rps16_1-trnQ-UUG intergenic region as distinguishing markers for Cyathula officinalis and its hybrids. Codon usage analysis revealed no strong bias at the third codon position among the three species, although codons ending with thymine (T) were used more frequently. IR boundary analysis showed variation only among isolated C. capitata samples. Comparative genomics highlighted psbI-trnS-GCU and rps16_1-trnQ-UUG as highly variable regions. Phylogenetic analysis indicated that the hybrid (Z4) grouped within C. capitata, consistent with the maternal inheritance pattern of chloroplast genomes. Given the potential for variations in maternal parents among different hybrid batches, the positions of these elements in the phylogenetic tree may accordingly vary.

This study elucidated the chloroplast genome features and phylogenetic relationships of C. capitata, C. officinalis, and their hybrid. The findings offer significant molecular insights that facilitate species identification, genetic analysis, and quality assessment of Cyathula medicinal resources. These insights support the sustainable utilization and conservation of these resources.