Comparative analysis of organelle genomes in three Terminalia species reveals structure evolution and phylogenetic position

Comparative analysis of organelle genomes in three Terminalia species reveals structure evolution and phylogenetic position | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Comparative analysis of organelle genomes in three Terminalia species reveals structure evolution and phylogenetic position xingyu Liu, Yingmin Zhang, Pengrui Luo, Zemin Yang, Yingmei Zuo, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8163245/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 10 You are reading this latest preprint version Abstract Background Plant organelle genomes are critical for evolutionary biology research, yet their characteristics in the Terminalia genus (Combretaceae, ~ 200 tropical species of which some possess medicinal and edible value) remain unstudied. This study systematically analyzes the mitochondrial genomes (mitogenomes) and chloroplast genomes (plastomes) of three Terminalia species ( T. chebula , T. franchetii , T. intricata ). Results Using high-throughput sequencing, we successfully obtained sequences of plastomes and mitogenomes for three plants, enabling comparative and evolutionary genomic analysis. Plastomes of the three species shared a typical quadripartite structure, with similar sizes (159,700–159,971 bp) and gene compositions (103–107 genes), but differed in specific regions. In contrast, mitogenomes showed significant structural size variation (350,904–365,711 bp), with T. chebula and T. intricata having three circular and one linear molecule, and T. franchetii has two linear molecules, reflecting extensive recombination. Ka/Ks analysis indicated positive selection on mitogenome genes (e.g., rps1 , nad4 ) linked to adaptation. RNA editing (all C-to-U) enhanced protein hydrophobicity. Phylogenetic trees clarified Terminalia ’s position. Conclusions This study provides crucial genomic data for phylogenetic research and molecular ecology within the Terminalia , holding significant value for future research. Terminalia plastomes mitogenomes organelle genome comparative genomics Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Background Terminalia belongs to the Combretaceae family, comprising approximately 200 species widely distributed across tropical and subtropical regions, particularly in Asia, Africa, and Australia. Most Terminalia species are deciduous or evergreen trees or shrubs, commonly found in forests, riverbanks, and wetlands [ 1 ]. Numerous Terminalia plants have extensive applications in traditional ethnopharmacology, serving as herbal remedies or formulations for treating various ailments such as headaches, fever, pneumonia, influenza, geriatric diseases, and cancer [ 2 – 4 ]. Additionally, they are employed to alleviate symptoms including memory loss, abdominal pain, back pain, cough, colds, conjunctivitis, diarrhea, heart disease, leprosy, sexually transmitted diseases, and urinary tract infections [ 5 – 7 ]. Research indicates these plants exhibit diverse biological activities, encompassing antibacterial, antifungal, anti-inflammatory, antiviral, antiretroviral, and antioxidant properties [ 8 – 10 ]. Current research on Terminalia primarily focuses on pharmacological effects [ 11 ], active constituents [ 12 , 13 ], processing methods, and molecular identification. However, no studies have reported on its mitochondrial genome (mitogenome) or chloroplast genome (plastome). Unraveling its mitogenomes and plastomes is crucial for the phylogenetic studies of Terminalia , providing vital clues for species origins, evolutionary relationships, and taxonomy. With the advancement of high-throughput sequencing, the genomes of organelles from an increasing number of plants have been sequenced, offering significant data for studying plant physiological functions, evolutionary relationships, and germplasm resource conservation. Both mitochondria and chloroplasts are believed to have evolved from ancient free-living bacteria through endosymbiosis [ 14 ]. Mitochondria, serving as the “powerhouse” of the cell, generate ATP through oxidative phosphorylation and function as the central hub of plant energy metabolism [ 15 ]. Compared to animal mitogenomes, plant mitogenomes contain not only genes related to mitochondrial function but also repetitive sequences. While plastomes typically maintain a relatively uniform circular structure [ 16 , 17 ], mitogenomes are characterized by a high incidence of repetitive sequences and exhibit significant structural heterogeneity, including polycyclic [ 18 ], linear [ 19 ], and branched forms [ 20 ]. Studying plant mitogenomes can reveal mechanisms of plant energy metabolism [ 21 ], providing new theoretical foundations for agricultural production, such as enhancing plant stress resistance and advancing breeding efforts [ 22 ]. In this study, we successfully assembled the plastomes and mitogenomes of three previously unreported Terminalia species: Terminalia chebula , Terminalia franchetii , and Terminalia intricata . We analyzed the organelle genomes of these Terminalia species by comparing the newly obtained sequences with existing organelle genome data. Analyses included GC content, codon usage bias, repetitive sequences, Ka/Ks ratios, prediction of RNA editing events, and nucleotide polymorphism analysis. Homologous sequences found in both chloroplast and mitogenomes were also analyzed. Furthermore, maximum likelihood phylogenetic trees were constructed using PCGs from 17 mitogenomes and 18 plastomes, respectively. We reconstructed phylogenetic relationships within the Terminalia , exploring connections and taxonomic boundaries between different species groups, and identified key regions of genetic variation. These findings will provide important references and valuable insights for molecular marker development and phylogenetic studies in the Terminalia . Results Genome organization of three Terminalia organelles The plastomes sizes of T. chebula , T. franchetii , and T. intricata were 159,915 bp, 159,700 bp, and 159,971 bp, respectively (Figure 1). All three genomes exhibited a typical quadripartite structure, consisting of a pair of inverted repeats (IRs) with lengths of 26,339 bp in T. chebula , 26,278 bp in T. franchetii , and 26,345 bp in T. intricata . The small single-copy region (18,888 bp in T. chebula , 18,734 bp in T. franchetii , and 18,908bp in T. intricata ) and a large single-copy region (71,568 bp in T. chebula , 71,290 bp in T. franchetii , and 71,600 bp in T. intricata ). In the plastomes of T. chebula , T. franchetii , and T. intricata , 76, 76, and 73 PCGs were annotated, respectively, along with 27, 26, and 26 tRNA genes. All three species contained 4 rRNA genes. Interestingly, the mitogenome structures of the three Terminalia species showed significant differences and were highly complex. The mitogenomes structure of T. chebula and T. intricata consisted of three circular molecules and one linear molecule, while T. franchetii had two linear molecules. (Figure 1). The sizes of the mitogenomes for T. chebula , T. franchetii , and T. intricata were 354,989 bp, 365,711 bp, and 350,904 bp, respectively. In these genomes, 35, 34, and 38 PCGs were annotated for T. chebula , T. franchetii , and T. intricata , respectively. Additionally, T. chebula contained 17 tRNA genes, T. franchetii had 18, and T. intricata had 17. The rRNA gene counts were 3 for T. chebula , 4 for T. franchetii , and 3 for T. intricata . The GC contents of the mitogenomes for T. chebula , T. franchetii , and T. intricata were 45.5%, 44.3%, and 44.7%, respectively. The plastomes of the three Terminalia species exhibited a circular structure, with sizes ranging from 159,700bp to 159,971 bp, showing a maximum size difference of 271 bp. The number of genes in the plastomes varied between 106 and 108 across species. In contrast, the mitogenomes displayed significant structural variations, with sizes ranging from 350,904 bp to 365,711 bp, resulting in a maximum size difference of 14,807 bp. The number of genes in the mitogenomes varied between 55 and 58 across species (Table 1). These results indicate that the mitogenomes exhibit substantial fluctuations in structure, genome size, and GC content, whereas the plastomes appear to be more conserved throughout evolution (Additional file 1). Table 1 Genomic characteristics of three Terminalia species Species Genome Type Genome Size (bp) Protein Genes tRNA Genes rRNA Genes Total Genes T. chebula plastome 159,915 76 27 4 107 T. franchetii plastome 159,700 76 26 4 106 T. intricata plastome 159,971 73 26 4 103 T. chebula mitogenome 354,989 35 17 3 55 T. franchetii mitogenome 365,711 34 18 4 56 T. intricata mitogenome 350,904 38 17 3 58 Repeat sequence analysis of three Terminalia organelles In the plastomes of T. chebula , T. franchetii , and T. intricata , 73, 70, and 70 simple sequence repeats (SSRs) were identified, respectively (Figure 2A). All SSRs in T. chebula were haploid, T. franchetii only contained one diploid SSR, while T. intricata had one diploid SSR and one triploid SSR, with no higher ploidy detected. The plastomes of all three Terminalia species contained complementary, palindromic, forward, reverse, and tandem repeats, with tandem and palindromic repeats being the predominant types. Among these species, T. chebula exhibited the highest number of tandem repeats, with 41 in total. In the mitogenomes of T. chebula , T. franchetii , and T. intricata , 57, 56, and 57 SSRs were identified, respectively. Among these, haploid repeats predominated, followed by diploid and triploid repeats, with no higher ploidy detected. The mitogenomes of three Terminalia species contained complementary, palindromic, forward, reverse, and tandem repeats. In T. chebula , forward (42.1%, 86 repeats) and palindromic (28.9%, 59 repeats) repeats were the most common. In T. franchetii , forward (41%, 41 repeats) and palindromic (38%, 38 repeats) repeats were also dominant. Notably, T. franchetii exhibited the highest number of tandem repeats among the three species, with 42 repeats. In T. intricata , forward (42.7%, 85 repeats) and palindromic (31.1%, 62 repeats) repeats were similarly predominant (Figure 2B). In the plastomes and mitogenomes of the three Terminalia species, several types of repeat sequences were present, including complementary repeats, palindromic repeats, forward repeats, reverse repeats, and tandem repeats. The distribution of simple sequence repeats (SSRs) across the two types of genomes showed similarities, with haploid repeats predominating. Although diploid and triploid repeats were also found in the mitogenomes, haploid repeats formed the highest proportion (Additional file 2). Codon usage analysis of three Terminalia organelles This study compared the codon usage preferences in the mitochondrial and plastomes of T. chebula , T. franchetii , and T. intricata , and analyzed their relative synonymous codon usage frequencies (Figure 3A). Arginine and Leucine were the most frequently used amino acids in the organellar genomes of the three Terminalia species, accounting for 9.8% of mitogenomes and 9.6% of plastomes, respectively. In comparative studies of plastomes across species, the TTA codon exhibited a strong preference (2.12) in T. chebula , while AGA showed the highest codon usage preference in T. franchetii and T. intricata , with values of 1.9269 and 1.9988, respectively. Conversely, codons such as CTG and GCG exhibited significantly lower RSCU values across all three species, indicating their infrequent usage. In the mitogenomes of the three Terminalia species, the ATG and TGG codons did not have synonymous codons, and their RSCU values were both 1 (Figure 3B). There were differences in codon usage preferences among the species. In T. chebula , the most preferred codon was GCT (1.57), followed by TTA (1.55), while the least preferred codon was GCG, with an RSCU value of 0.47. In contrast, T. franchetii and T. intricata showed highly consistent codon usage preferences, with the most commonly used codon being AGA, with RSCU values of 1.77 and 1.73, respectively. The lowest RSCU value for both species was CGT, with values of 0.59 and 0.56, respectively (Additional file 3). RNA editing site analysis of mitogenomes RNA editing involves the addition, deletion, or alteration of bases in the coding region of the transcribed RNA [23], and it occurs in all eukaryotes, including plants [24]. The results showed that 311, 364, and 349 RNA editing sites were predicted in mitogenomes of T. chebula , T. franchetii , and T. intricata , respectively, with all editing events involving a C-to-U base conversion (Additional file 4). This editing pattern is consistent with that observed in other plants. In all three species of the Terminalia , genes such as mttB , rps4 , and cob have a higher number of editing sites, while genes like atp8 , atp9 , and sdh3 have fewer editing sites. In the effective amino acid editing events of the three species, hydrophobic amino acids dominated, accounting for approximately 71.3% to 71.5%, while hydrophilic amino acids accounted for around 28.5%. The high-frequency codons were biased toward hydrophobic amino acid coding, with the codons for leucine (L) and phenylalanine (F) being more frequent. Specifically, UUA and UUC appeared frequently across the three species (Figure 4A). Substitution rates of mitochondrial PCGs Different species face varying ecological pressures during evolution, leading to genomic changes. The Ka/Ks ratio measures selection pressure on PCGs, helping infer species adaptation and evolutionary trajectories. When the Ka/Ks ratio is > 1, genes are undergoing positive selection, conversely, Ka/Ks < 1 denotes negative selection, and Ka/Ks = 1 represents neutral selection. To calculate the selective pressure on mitogenomes PCGs in three species of Terminalia , we computed the Ka/Ks values for the PCGs of 37 mitogenomes (Additional file 5). The Ka/Ks values of most PCGs were found to be below 1, while genes like nad4 , rps1 , rps4 , and rps7 had Ka/Ks values exceeding 1(Figure 4B). Homology analysis of genome sequences We searched for homologous sequences between the plastomes and mitogenomes in three Terminalia species to identify potential gene transfer events. We identified 17-26 homologous sequences between the plastomes and mitogenomes in the three species, with lengths ranging from 28bp to 7035bp (Figure 4C). Interestingly, we identified some complete plastid genome-derived PCGs in the mitogenomes, including petB , petG , psaA , psbJ , petB , psbM , ycf2 , and psb , among others. Notably, psbJ , psaA , petB , and petG were shared by all three species. Collinearity analysis of the mitogenomes Mauve was used to analyze the synteny and rearrangements of the mitogenomes across the three species. The three genomes exhibited a large number of locally collinear blocks (LCBs) with consistent coloring, indicating that they share a substantial number of conserved gene segments, which reflects the close evolutionary relationship within the genus. These homologous regions (LCBs) had different relative positions, suggesting the presence of extensive rearrangements within the mitogenomes (Figure 5A). Nucleotide polymorphism (Pi) analysis of organelle genomes The Pi analysis was conducted on 77 plastome PCGs from five species of Combretaceae ( Terminalia chebula , Terminalia franchetii , Terminalia intricata , Terminalia phillyreifolia, Terminalia guyanensis ). The results revealed that among the 77 PCGs, gene rpl22 exhibited the highest Pi value (0.11406), followed by rps19 (0.104353). In contrast, the lowest Pi values were observed for ndhB (0.009983). We also performed the Pi analysis on 37 mitochondrial PCGs from three species of Terminalia , and the results revealed that the highest Pi value was observed for rps10 (0.117162), followed by nad9 (0.1079), while the lowest Pi values, both 0, were found for rpl16 and rpl19 (Figure S1. Additional file 6). Expansion and contraction of the IR region A comparative analysis of the IR/SSC boundary regions in the plastomes of 6 Combretaceae species revealed subtle differences at the IR/SSC junction. In this study, the rps19 gene in Lumnitzera racemosa , Lumnitzera littorea spanned 106 bp of the LSC/IRb region, while in Terminalia franchetii and Terminalia intricata and Terminalia chebula it spanned 16-24 bp of the LSC/IRb region. In contrast, the rps19 gene in Laguncularia racemosa was entirely located within the LSC region. Significant differences were observed at the IRb/SSC junction (JSB). It is noteworthy that the position of the rpl2 gene in Terminalia intricata and Terminalia chebula differed significantly from that in other species. This difference may arise from the dynamic evolution of the plastome IR region boundaries. At this junction, only Lumnitzera racemosa , Lumnitzera littorea , and Laguncularia racemosa contained the ycf1 gene, while the other 3 species lacked it. Additionally, the ndhF gene was located at the IRb/SSC junction. In Lumnitzera racemosa , Lumnitzera littorea , Laguncularia racemosa , and Terminalia chebula , 15-18 bp of the ndhF gene are located in the IRb region, while in Terminalia franchetii and Terminalia intricata , 70-75 bp were located in the IRb region. Notably, the ycf1 gene spanned the SSC/IRa junction in all 6 species. The psbA and trnH genes were completely located in the LSC region (Figure 5B). Phylogenetic analysis of the organelle genom e A phylogenetic tree of 17 angiosperm species from five orders was constructed by extracting mitogenome PCGs sequences, with Vitis vinifera as the outgroup (Figure 6A). T. chebula was confirmed as a species of the genus Terminalia , T. intricata clustering with T. franchetii . The phylogenetic tree shows that the bootstrap support values for most nodes were above 95%. The overall topology of the tree is consistent with the classification results of the Angiosperm Phylogeny Group IV (APG IV) system. To clarify the phylogenetic position and evolutionary relationships of the Combretaceae family within the Myrtales order, a phylogenetic analysis was conducted using plastome sequences from eight species of Combretaceae and ten other plant species, with R. solosa from the Malpighiales as the outgroup. The results indicated that species within the Terminalia of Combretaceae exhibited a high degree of phylogenetic clustering, with strong branch support, revealing close genetic relationships among the species in this genus. T. franchetii and T. intricata were closely clustered in one group with a bootstrap support of 100%. The Onagraceae family, represented by O. biennis and O. parviflora , formed a separate clade, while the Lythraceae family, represented by P. granatum , appeared as an independent branch. The Myrtaceae family, which includes A. costata , C. henryi , C. gummifera , E. aromaphloia , E. deglupta , and P. trunciflora , formed a stable cluster R. stylosa from the Rhizophoraceae family clustered separately within the order of the Combretaceae, displaying significant evolutionary divergence from Myrtales species. Furthermore, within the Myrtales, the Onagraceae family exhibited a closer phylogenetic relationship with the Lythraceae family (Figure 6B). Discussion With the advent of advanced sequencing technologies, and the successful assembly of an increasing number of genomes, the complexity of mitogenomes has become more evident [ 25 ]. The genome structure of mitochondria also exhibits diversity, including linear, circular, highly branched, and networked forms, with such diversity even observed within a single individual, possibly due to frequent recombination [ 26 – 28 ]. The earliest mitogenome sequencing from animals was performed on humans and Drosophila, where the genomes were found to be small, circular molecules with highly conserved sequences [ 29 , 30 ]. As a result, it was once assumed that all mitogenomes were single, circular molecules. However, recent studies have shown that mitogenomes are not always single circular structures. For example, Quercus acutissima (Fagaceae) consists of one linear and two circular molecules [ 31 ]. Colocasia esculenta (Araceae) consists of five circular molecules [ 32 ], while Echinacanthus longipes (Acanthaceae) consists of two circular and three linear molecules [ 33 ]. The mitogenomes of three species in the Terminalia exhibit significant structural differences. The mitogenomes of T. chebula and T. intricata consist of three circular molecules and one linear molecule, while T. franchetii has two linear molecules. Mitogenomes in plants can show considerable variation between species. The size of plant mitogenomes also varies widely, typically ranging from 66 kb to 11,000 kb. The mitogenome sizes of the three Terminalia species fall between 350,904 and 354,989 bp. Studies have shown that plant mitogenomes can exhibit significant differences between species [ 34 ]. The structural and size variations of the mitogenomes in Terminalia species are considerable, which may be related to species evolution and phylogenetic affinity. Furthermore, the plastomes of three Terminalia species possess a unique quadripartite structure, consisting of two IR regions, one LSC region, and one SSC region, which is similar to the plastomes structure of other angiosperms [ 35 ]. Meanwhile, GC content significantly affects amino acid composition and the structural integrity of genomes during evolutionary processes [ 36 ]. Our analysis of the GC content in the plastomes and mitogenomes of the three Terminalia species indicates that plastomes exhibit more stable GC content (36.8%-37.0%), while the mitogenomes show greater variability in GC content (44.3%-45.5%). This result supports the conclusion that GC content is highly conserved in higher plants. Interestingly, among the three Terminalia species, the plastome lengths of the three Terminalia showed a maximum size difference of 271 bp, which suggests that variability in cp genome size is closely related to phylogenetic relationships and evolutionary history [ 37 ]. RNA editing is a post-transcriptional process in which nucleotides within mRNA molecules are inserted, deleted, or substituted, resulting in changes to the genetic code [ 38 ]. Previous studies have shown that the number of editing sites and the gene with the highest number of editing sites differ across species. For example, there are 455 RNA editing sites in 37 genes in Primulina hunanensis [ 39 ], 460 RNA editing sites in 34 genes in Glycyrrhiza glabra [ 40 ], and 441 RNA editing sites in 36 genes in Arabidopsis thaliana [ 41 ]. In this study, we predicted the RNA editing sites of PCGs in the mitogenomes of three Terminalia species and identified 311–369 editing sites, all of which involve the conversion of (C) to (U). Further analysis indicates that these C-U editing events exhibited a significant preference for specific codon positions, primarily concentrated at the first and second codon positions [ 42 ]. This process enhances the interspecies homogeneity of the mitochondrial proteome and promotes adaptive development in plants. Most RNA editing events in the three Terminalia species involve the conversion of amino acids to hydrophobic ones, which helps alter their physicochemical properties and enhance protein folding [ 43 ]. Therefore, widespread RNA editing leads to an increase in the number of hydrophobic amino acids, which may contribute to the translation of mRNA into polypeptides with the correct folded structure. This finding forms the structural basis for the specific functions of proteins [ 44 ]. The Ka/Ks ratio can reveal the evolutionary pathways of genes and their underlying genetic mechanisms [ 45 ]. We conducted Ka/Ks analysis on 34 shared mtDNA protein-coding genes (PCGs) across three Terminalia species. The results indicate signs of positive selection in rps1 , rps4 , rps7 , and nad4 . Specifically, rps1 , rps4 , and rps7 may be associated with plant adaptation to environmental changes, while nad4 is closely linked to mitochondrial energy metabolism. These findings likely reflect adaptive variation in redox reactions and environmental stress responses. As a valuable analytical tool, Ka/Ks analysis not only aids in understanding species adaptability and selective pressures but also offers new perspectives for assessing adaptive potential and studying genetic evolution [ 46 ]. A phylogenetic tree of mitochondrial and plastomes was constructed using maximum likelihood (ML) analysis. The results showed that the topology of the ML tree constructed with chloroplast PCGs was largely consistent with that of the ML tree constructed with mitochondrial PCGs. T. chebula , T. franchetii , and T. intricata were all classified within Terminalia , with T. intricata being a variety of T. franchetii , these two species showed a closer relationship. Due to the limited number of published mitogenomes for Myrtales, the mitogenomes tree encompasses a broader range of order-level taxa, reflecting evolutionary signals among distantly related groups. The overall structure of this mitochondrial DNA-based phylogeny is consistent with the latest classification of the Angiosperm Phylogeny Group (APG). In contrast, the plastomes tree focuses on interfamilial relationships within Myrtaceae. The organelle genomes of these three Terminalia species will provide valuable and important genetic information for further improving the phylogenetic resolution of angiosperms. Conclusion In this study, we successfully assembled the mitogenomes and plastomes of three Terminalia species, revealing unique branched conformations in the mitochondrial DNA structure. The results showed that the plastomes of the three Terminalia species were similar in GC content and genome size. The ka/ks ratio analysis of the mitogenomes indicated that genes rps1 , rps4 , rps7 , and nad4 are under positive selection. The presence of gene transfer and conserved syntenic blocks between the three Terminalia species reflects gene rearrangements and recombination, providing important genetic insights. RNA editing events revealed that most hydrophilic amino acids were converted to hydrophobic ones, enhancing the stability of protein structures. The construction of the phylogenetic tree deepens our understanding of the evolutionary relationships among Terminalia species, particularly highlighting the close relationship between T. franchetii and T. intricata . Overall, our findings suggest that organellar genomic data can effectively reveal phylogenetic relationships between plants and can be applied to molecular marker development and genetic evolution studies. Materials and Methods Plant materials, DNA extraction and sequencing Fresh leaves of T. chebula were collected from Longling County, Baoshan City, Yunnan Province, China (Sample location: 98°E, 24°N). This specimen was identified by Professor Li Guodong of Yunnan University of Chinese Medicine based on morphological characteristics. T. franchetii was collected from Baima Town, Miyi County, Panzhihua City, Sichuan Province, China, with coordinates (Sample Location: 102.2°E, 27.0°N). T. intricata was collected from Dongwang Township, Shangri-La City, Diqing Tibetan Autonomous Prefecture, Yunnan Province, China (Sample location: 99.5°E, 28.5°N). The specimen was identified by P.R. Luo based on morphological characteristics. The leaves were immediately frozen in liquid nitrogen, and total DNA was extracted using a modified CTAB method. The high-quality DNA samples that passed the test were sent to Novogen Biotechnology Co. for second- and third-generation sequencing, respectively. Selected high-quality DNA samples that passed the test (main band > 30kb); were randomly fragmented into 15-18kb segments using a G-Tube tube; enriched and purified using magnetic beads; damage repair and end repair was performed on the fragmented DNA; stem-loop sequencing adapters were lighted to both ends of the DNA fragments and exonucleases used to remove failed ligation fragments. The constructed library was then sequenced using the PacBio Revio/Sequel II/IIe platform. The raw sequencing reads contained a dumbbell-shaped structure with adapters at both ends, referred to as Polymerase reads. The raw data was fragmented at the adapter sites, and the adapter sequences filtered out to obtain subreads. The subreads were filtered according to the standard "Filtering subreads by minimum length = 50". Based on the subreads, high-accuracy HiFi reads were generated using CCS software. All reads had a quality value greater than Q20. Organelle genome assembly and annotation The plastomes of T. franchetii and T. intricata were de novo assembled using GetOrganelle v1.7.5.3 [ 47 ]. Using PacBio HiFi sequencing data, the mitogenomes of three Terminalia species and the plastomes of T. chebula was assembled with PMAT v1.5.3 [ 48 ]. The assembly parameters were configured as follows: “-st HiFi -tp mt -ml 70 -mi 90”. The mitogenomes was visualized using Bandage v0.8.1 [ 49 ]. The mitogenomes were annotated using IPMGA ( http://www.1kmpg.cn/ipmga/ ), with RNA-seq data used to precisely correct 29 representative angiosperm mitogenomes sequences as the reference genome. Annotation errors for each mitogenome were manually corrected using Geneious Prime [ 50 ]. Additionally, the plastomes of T. chebula, T. franchetii and T. intricata were annotated using Geseq[ 51 ]. Finally, the organelle genome was visualised using OGDraw [ 52 ]. Analysis of Repeat Elements MISA (v2.1) ( https://webblast.ipk-gatersleben.de/misa/ ), TRF (v4.09) ( https://tandem.bu.edu/trf/trf.unix.help.html ), and REPuter ( https://bibiserv.cebitec.uni-bielefeld.de/reputer/ ) were used to identify simple sequence repeats (SSRs), tandem repeats, and dispersed repeats, respectively. The results were visualized using Excel (2021) software and the Circos package (v0.69.9). Analysis of Codon Usage The protein-coding sequences of mitogenomes and plastomes were extracted with Phylosuite(v1.1.16) [ 53 ]. To analyze the codon bias of the protein-coding genes within the mitogenomes, MEGA v7.0 was used [ 54 ], and the Relative Synonymous Codon Usage (RSCU) values were subsequently calculated. Prediction of RNA editing sites The RNA editing sites were predicted via PREPACT3 [ 55 ]. The analysis employed reference sequences from Vitisvinifera and Millettia pinnata . The predicted RNA editing sites were subsequently visualized using R (4.4.0) to facilitate interpretation and analysis of the results. Homologous sequences identification Homogeneous sequences between the plastomes and mitogenomes of three Terminalia were identified using BLAST (v2.16.0) [ 56 ], with parameters set to an identity greater than 90% and an e-value of less than 1e − 5. We then analyzed the homologous sequence regions to determine the length, quantity, and gene type of the mitochondrial plastomes sequences (MTPTs). the results were visualized using the Circos package (v0.69.9) [ 57 ]. Substitution rates of PCGs First, the mitogenomes of three Terminalia species were aligned using MAFFT (v7.427) [ 58 ]. Subsequently, the Ka/Ks ratio was calculated for each gene pair using the YN method in KaKs_Calculator (v2.0) [ 59 ]. The aggregated Ka/Ks ratios were visualized as box plots generated with the ggplot2 package in R. Nucleotide diversity Sequences of protein-coding genes (PCGs) from organelle genomes were aligned using MAFFT (v7.477) [ 60 ], while the Pi value for each gene was calculated using DnaSP (v5.0) [ 61 ]. Analysis of Plastomes Boundaries Based on annotations, the plastome boundaries between the IR, LSC, and SSC regions were analyzed, and a schematic diagram was constructed using Irscope [ 62 ]. Collinearity Analysis and Phylogenetic Analysis Mauve was employed to analyze homology and sequence rearrangements within the mitogenomes of three Terminalia species [ 63 ]. All conserved PCGs from mitogenomes and plastomes were extracted using Phylosuite [ 64 ] and aligned with MAFFT (v7.427) [ 65 ]. The results were then imported into MACSE for optimization. The optimal model was selected via ModelFinder (v2.2.2) [ 66 ]. Bayesian analysis was performed on the PCGs concatenated data matrix using MrBayes (v.3.2) [ 67 ]. The final phylogenetic tree was constructed in IQ-tree and visualized with iTOL [ 68 ] for aesthetic refinement. Abbreviations NCBI National Center for Biotechnology Information. PCGs Protein-coding genes. RSCU Relative synonymous codon usage. SSRs Simple sequence repeats. cpDNA Plastomes. mtDNA Mitogenomes. MTPTs Mitochondrial plastid DNAs. Ka Nonsynonymous substitutions. Ks Synonymous substitutions. Pi Nucleotide Diversity Declarations Authorship contribution statement Xingyu Liu: Data curation, Visualization, Writing – original draft& editing. Pengrui Luo: Data curation, Writing – review & editing, Resources, Validation.Yingmei Zou: Data curation, Writing – original draft, Validation. M. James C. Crabbe: Data curation, Visualization, Writing – original draft. Guodong Li: Conceptualization, Supervision, Writing – review & editing, Funding acquisition. Ticao Zhang: Conceptualization, Resources, Supervision, Writing – review & editing, Funding acquisition. Funding This work is supported by grants from National Key R&D Program of China (2024YFF1306700), Strategic Priority Research Program of the Chinese Academy of Sciences (XDB1230000), National Natural Science Foundation of China (82260739, 32570434), Yunnan Fundamental Research Projects (202501AS070177 and 202101AZ070001-005). Ethics approval and consent to participate Not applicable. This study did not use animal or human materials. Our experimental research, including the collection of plant materials, complies with institutional, national, or international guidelines. The voucher specimen of T. chebula is deposited in the Herbarium of Yunnan University of Chinese Medicine, voucher numbers 2503. The voucher specimens of T. franchetii and T. intricata are deposited in the Kunming Institute of Botany Chinese Academy of Sciences. Voucher numbers 1651612 ( T. franchetii ) and 1651613 ( T. intricata ). Consent for publication Not applicable Declaration of Competing Interest We declare that we have no conflicts of interest. Data availability The dataset generated and analyzed by this graduate student has been submitted to the NCBI database. The current provisional accession number is SUB15777329. The final permanent accession number will be provided prior to the publication of the thesis. References Das G, Kim D-Y, Fan C, Gutiérrez-Grijalva EP, Heredia JB, Nissapatorn V, et al. Plants of the Genus Terminalia : An Insight on Its Biological Potentials, Pre-Clinical and Clinical Studies. Front Pharmacol. 2020;11:561248. Kim HJ, Song H-K, Park SH, Jang S, Park K-S, Song KH, et al. Terminalia chebula Retz. extract ameliorates the symptoms of atopic dermatitis by regulating anti-inflammatory factors in vivo and suppressing STAT1/3 and NF-ĸB signaling in vitro. Phytomedicine Int J Phytother Phytopharm. 2022;104:154318. Ramesh P, Palaniappan A. Terminalia arjuna , a Cardioprotective Herbal Medicine-Relevancy in the Modern Era of Pharmaceuticals and Green Nanomedicine-A Review. Pharm Basel Switz. 2023;16:126. Gupta A, Kumar R, Bhattacharyya P, Bishayee A, Pandey AK. Terminalia bellirica (Gaertn.) roxb. (Bahera) in health and disease: A systematic and comprehensive review. Phytomedicine Int J Phytother Phytopharm. 2020;77:153278. Mahendra RVNASR. Antimicrobial and anti-inflammatory activity of Terminalia arjuna . Bioinformation. 2023;19:184–9. Cheesman MJ, Shivashekaregowda NKH, Cock IE. Bacterial Foodborne Illness in Malaysia: Terminalia spp. as a Potential Resource for Treating Infections and Countering Antibiotic Resistance. Malays J Med Sci MJMS. 2023;30:42–54. Liu H, Chen Z, Liu M, Li E, Shen J, Wang J, et al. The Terminalia chebula Retz extract treats hyperuricemic nephropathy by inhibiting TLR4/MyD88/NF-κB axis. J Ethnopharmacol. 2024;322:117678. Hassan Bulbul MR, Uddin Chowdhury MN, Naima TA, Sami SA, Imtiaj MS, Huda N, et al. A comprehensive review on the diverse pharmacological perspectives of Terminalia chebula Retz. Heliyon. 2022;8:e10220. Wang C, Zhang H, Wang X, Wang X, Li X, Li C, et al. Comprehensive Review on Fruit of Terminalia chebula : Traditional Uses, Phytochemistry, Pharmacology, Toxicity, and Pharmacokinetics. Mol Basel Switz. 2024;29:5547. Zai MJ, Cheesman MJ, Cock IE. Terminalia petiolaris A.Cunn ex Benth. Extracts Have Antibacterial Activity and Potentiate Conventional Antibiotics against β-Lactam-Drug-Resistant Bacteria. Antibiot Basel Switz. 2023;12:1643. Gao H, Lu H, Fang N, Su J, Li R, Wang W, et al. The potential of Terminalia chebula in alleviating mild cognitive impairment: a review. Front Pharmacol. 2024;15:1484040. Cock IE. The medicinal properties and phytochemistry of plants of the genus Terminalia (Combretaceae). Inflammopharmacology. 2015;23:203–29. Kumar V, Sharma N, Saini R, Mall S, Zengin G, Sourirajan A, et al. Therapeutic potential and industrial applications of Terminalia arjuna bark. J Ethnopharmacol. 2023;310:116352. Gray MW. The endosymbiont hypothesis revisited. Int Rev Cytol. 1992;141:233–357. Shtolz N, Mishmar D. The Mitochondrial Genome–on Selective Constraints and Signatures at the Organism, Cell, and Single Mitochondrion Levels. Front Ecol Evol. 2019;7. Tian S, Lu P, Zhang Z, Wu JQ, Zhang H, Shen H. Chloroplast genome sequence of Chongming lima bean ( Phaseolus lunatus L.) and comparative analyses with other legume chloroplast genomes. BMC Genomics. 2021;22:194. Yanfei N, Tai S, Chunhua W, Jia D, Fazhong Y. Complete chloroplast genome sequences of the medicinal plant Aconitum transsectum (Ranunculaceae): comparative analysis and phylogenetic relationships. BMC Genomics. 2023;24:90. Sloan DB. One ring to rule them all? Genome sequencing provides new insights into the master circle model of plant mitochondrial DNA structure. New Phytol. 2013;200:978–85. Gualberto JM, Mileshina D, Wallet C, Niazi AK, Weber-Lotfi F, Dietrich A. The plant mitochondrial genome: dynamics and maintenance. Biochimie. 2014;100:107–20. Jackman SD, Coombe L, Warren RL, Kirk H, Trinh E, MacLeod T, et al. Complete Mitochondrial Genome of a Gymnosperm, Sitka Spruce ( Picea sitchensis ), Indicates a Complex Physical Structure. Genome Biol Evol. 2020;12:1174–9. Nass MM, Mitochondrial DNA. Advances, Problems, and Goals. Science. 1969;165:25–35. Liberatore KL, Dukowic-Schulze S, Miller ME, Chen C, Kianian SF. The role of mitochondria in plant development and stress tolerance. Free Radic Biol Med. 2016;100:238–56. Brennicke A, Marchfelder A, Binder S. RNA editing. FEMS Microbiol Rev. 1999;23:297–316. Covello PS, Gray MW. RNA editing in plant mitochondria. Nature. 1989;341:662–6. Yang H, Ni Y, Zhang X, Li J, Chen H, Liu C. The mitochondrial genomes of Panax notoginseng reveal recombination mediated by repeats associated with DNA replication. Int J Biol Macromol. 2023;252:126359. Chen Z, Nie H, Grover CE, Wang Y, Li P, Wang M, et al. Entire nucleotide sequences of Gossypium raimondii and G.arboreum mitochondrial genomes revealed A-genome species as cytoplasmic donor of the allotetraploid species. Plant Biol Stuttg Ger. 2017;19:484–93. Feng L, Wang Z, Wang C, Yang X, An M, Yin Y. Multichromosomal mitochondrial genome of Punica granatum : comparative evolutionary analysis and gene transformation from chloroplast genomes. BMC Plant Biol. 2023;23:512. Wang H, Wu Z, Li T, Zhao J. Highly active repeat-mediated recombination in the mitogenome of the aquatic grass Hygroryza aristata . BMC Plant Biol. 2024;24:644. Anderson S, Bankier AT, Barrell BG, de Bruijn MH, Coulson AR, Drouin J, et al. Sequence and organization of the human mitochondrial genome. Nature. 1981;290:457–65. Clary DO, Wolstenholme DR. The Drosophila mitochondrial genome. Oxf Surv Eukaryot Genes. 1984;1:1–35. Liu D, Guo H, Zhu J, Qu K, Chen Y, Guo Y, et al. Complex Physical Structure of Complete Mitochondrial Genome of Quercus acutissima (Fagaceae): A Significant Energy Plant. Genes. 2022;13:1321. Li H, Liu L, Qiu Z, He F, Dong W. Complete mitochondrial genome assembly and comparative analysis of Colocasia esculenta . BMC Plant Biol. 2025;25:67. Gao C, Wang S, Huang Y, Deng Y. Assembly and comparative analysis of the complete mitochondrial genome of Echinacanthus longipes (Acanthaceae), endemic to the Sino-Vietnamese karst flora. BMC Genomics. 2025;26:251. Wu Z-Q, Liao X-Z, Zhang X-N, Tembrock LR, Broz A. Genomic architectural variation of plant mitochondria—A review of multichromosomal structuring. J Syst Evol. 2022;60:160–8. Zhang Y, Li H-L, Zhong J-D, Wang Y, Yuan C-C. Chloroplast Genome Sequences and Comparative Analyses of Combretaceae Mangroves with Related Species. BioMed Res Int. 2020;2020:5867673. Wang Y, Sun N, Shi W, Ma Q, Sun L, Hao M, et al. Assembly and Comparative Analysis of the Complete Mitochondrial Genome of Ilex macrocarpa . Forests. 2023;14:2372. Wang L, Liu X, Xu Y, Zhang Z, Wei Y, Hu Y, et al. Assembly and comparative analysis of the first complete mitochondrial genome of a traditional Chinese medicine Angelica biserrata (Shan et Yuan) Yuan et Shan. Int J Biol Macromol. 2024;257:128571. Qu Y, Zhou P, Tong C, Bi C, Xu L. Assembly and analysis of the Populus deltoides mitochondrial genome: the first report of a multicircular mitochondrial conformation for the genus Populus. J Res. 2023;34:717–33. Chen L, Dong X, Huang H, Xu H, Rono PC, Cai X, et al. Assembly and comparative analysis of the initial complete mitochondrial genome of Primulina hunanensis (Gesneriaceae): a cave-dwelling endangered plant. BMC Genomics. 2024;25:322. Zhou G, Qin M, Liu X, Qi Y, Ou X, Tang M. De novo assembly of the mitochondrial genome of Glycyrrhiza glabra and identification of two types of homologous recombination configurations caused by repeat sequences. BMC Genomics. 2025;26:13. Unseld M, Marienfeld JR, Brandt P, Brennicke A. The mitochondrial genome of Arabidopsis thaliana contains 57 genes in 366,924 nucleotides. Nat Genet. 1997;15:57–61. Zhang Y, Shen Z, Meng X, Zhang L, Liu Z, Liu M, et al. Codon usage patterns across seven Rosales species. BMC Plant Biol. 2022;22:65. Lloyd Evans D, Hlongwane TT, Joshi SV, Riaño Pachón DM. The sugarcane mitochondrial genome: assembly, phylogenetics and transcriptomics. PeerJ. 2019;7:e7558. Yura K, Go M. Correlation between amino acid residues converted by RNA editing and functional residues in protein three-dimensional structures in plant organelles. BMC Plant Biol. 2008;8:79. Wang L, Liu X, Xu Y, Zhang Z, Wei Y, Hu Y, et al. Assembly and comparative analysis of the first complete mitochondrial genome of a traditional Chinese medicine Angelica biserrata (Shan et Yuan) Yuan et Shan. Int J Biol Macromol. 2024;257:128571. Xie D-F, Yu H-X, Price M, Xie C, Deng Y-Q, Chen J-P et al. Phylogeny of Chinese Allium Species in Section Daghestanica and Adaptive Evolution of Allium (Amaryllidaceae, Allioideae) Species Revealed by the Chloroplast Complete Genome. Front Plant Sci. 2019;10. Jin J-J, Yu W-B, Yang J-B, Song Y, dePamphilis CW, Yi T-S, et al. GetOrganelle: a fast and versatile toolkit for accurate de novo assembly of organelle genomes. Genome Biol. 2020;21:241. Bi C, Shen F, Han F, Qu Y, Hou J, Xu K, et al. PMAT: an efficient plant mitogenome assembly toolkit using low-coverage HiFi sequencing data. Hortic Res. 2024;11:uhae023. Wick RR, Schultz MB, Zobel J, Holt KE. Bandage: interactive visualization of de novo genome assemblies. Bioinforma Oxf Engl. 2015;31:3350–2. Kearse M, Moir R, Wilson A, Stones-Havas S, Cheung M, Sturrock S, et al. Geneious Basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinforma Oxf Engl. 2012;28:1647–9. Tillich M, Lehwark P, Pellizzer T, Ulbricht-Jones ES, Fischer A, Bock R, et al. GeSeq - versatile and accurate annotation of organelle genomes. Nucleic Acids Res. 2017;45:W6–11. Greiner S, Lehwark P, Bock R. OrganellarGenomeDRAW (OGDRAW) version 1.3.1: expanded toolkit for the graphical visualization of organellar genomes. Nucleic Acids Res. 2019;47:W59–64. Zhang D, Gao F, Jakovlić I, Zou H, Zhang J, Li WX, et al. PhyloSuite: An integrated and scalable desktop platform for streamlined molecular sequence data management and evolutionary phylogenetics studies. Mol Ecol Resour. 2020;20:348–55. Kumar S, Stecher G, Tamura K. MEGA7: Molecular Evolutionary Genetics Analysis Version 7.0 for Bigger Datasets. Mol Biol Evol. 2016;33:1870–4. Lenz H, Hein A, Knoop V. Plant organelle RNA editing and its specificity factors: enhancements of analyses and new database features in PREPACT 3.0. BMC Bioinformatics. 2018;19:255. Camacho C, Coulouris G, Avagyan V, Ma N, Papadopoulos J, Bealer K, et al. BLAST+: architecture and applications. BMC Bioinformatics. 2009;10:421. https://doi.org/10.1186/1471-2105-10-421 . Krzywinski M, Schein J, Birol İ, Connors J, Gascoyne R, Horsman D, et al. Circos: An information aesthetic for comparative genomics. Genome Res. 2009;19:1639–45. Katoh K, Standley DM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 2013;30:772–80. Wang D, Zhang Y, Zhang Z, Zhu J, Yu J. KaKs_Calculator 2.0: a toolkit incorporating gamma-series methods and sliding window strategies. Genomics Proteom Bioinf. 2010;8:77–80. Katoh K, Standley DM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 2013;30:772–80. Rozas J, Ferrer-Mata A, Sánchez-DelBarrio JC, Guirao-Rico S, Librado P, Ramos-Onsins SE, et al. DnaSP 6: DNA Sequence Polymorphism Analysis of Large Data Sets. Mol Biol Evol. 2017;34:3299–302. Amiryousefi A, Hyvönen J, Poczai P. IRscope: an online program to visualize the junction sites of chloroplast genomes. Bioinforma Oxf Engl. 2018;34:3030–1. Darling ACE, Mau B, Blattner FR, Perna NT. Mauve: multiple alignment of conserved genomic sequence with rearrangements. Genome Res. 2004;14:1394–403. Zhang D, Gao F, Jakovlić I, Zou H, Zhang J, Li WX, et al. PhyloSuite: An integrated and scalable desktop platform for streamlined molecular sequence data management and evolutionary phylogenetics studies. Mol Ecol Resour. 2020;20:348–55. Katoh K, Standley DM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 2013;30:772–80. Kalyaanamoorthy S, Minh BQ, Wong TKF, von Haeseler A, Jermiin LS. ModelFinder: fast model selection for accurate phylogenetic estimates. Nat Methods. 2017;14:587–9. Ronquist F, Teslenko M, van der Mark P, Ayres DL, Darling A, Höhna S et al. MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Syst Biol. Letunic I, Bork P. Interactive Tree Of Life (iTOL) v4: recent updates and new developments. Nucleic Acids Res. 2019;47:W256–9. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Revision requested 06 Mar, 2026 Reviews received at journal 05 Mar, 2026 Reviewers agreed at journal 25 Feb, 2026 Reviews received at journal 16 Feb, 2026 Reviewers agreed at journal 26 Jan, 2026 Reviewers invited by journal 09 Jan, 2026 Editor assigned by journal 02 Dec, 2025 Editor invited by journal 01 Dec, 2025 Submission checks completed at journal 28 Nov, 2025 First submitted to journal 28 Nov, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-8163245","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":572865759,"identity":"882cc3e3-ba83-4a35-9431-28da0c906878","order_by":0,"name":"xingyu Liu","email":"","orcid":"","institution":"Yunnan University of Chinese Medicine","correspondingAuthor":false,"prefix":"","firstName":"xingyu","middleName":"","lastName":"Liu","suffix":""},{"id":572865760,"identity":"ed9750af-8b04-4c91-9b0e-1ab929f20be7","order_by":1,"name":"Yingmin Zhang","email":"","orcid":"","institution":"Yunnan University of Chinese Medicine","correspondingAuthor":false,"prefix":"","firstName":"Yingmin","middleName":"","lastName":"Zhang","suffix":""},{"id":572865761,"identity":"af15b7c1-9464-42a1-9a41-25c157677234","order_by":2,"name":"Pengrui Luo","email":"","orcid":"","institution":"Kunming Institute of Botany, Chinese Academy of Sciences","correspondingAuthor":false,"prefix":"","firstName":"Pengrui","middleName":"","lastName":"Luo","suffix":""},{"id":572865762,"identity":"17466be7-203f-42d8-97af-cb5b88f2b91a","order_by":3,"name":"Zemin Yang","email":"","orcid":"","institution":"Kunming Institute of Botany, Chinese Academy of Sciences","correspondingAuthor":false,"prefix":"","firstName":"Zemin","middleName":"","lastName":"Yang","suffix":""},{"id":572865763,"identity":"514f425b-8774-43f2-b38e-87ea47fe0d30","order_by":4,"name":"Yingmei Zuo","email":"","orcid":"","institution":"Yunnan Academy of Agricultural Sciences","correspondingAuthor":false,"prefix":"","firstName":"Yingmei","middleName":"","lastName":"Zuo","suffix":""},{"id":572865764,"identity":"f9175ace-35f7-45ef-8c9f-936e5a3108b1","order_by":5,"name":"M. James C. Crabbe","email":"","orcid":"","institution":"University of Oxford","correspondingAuthor":false,"prefix":"","firstName":"M.","middleName":"James C.","lastName":"Crabbe","suffix":""},{"id":572865765,"identity":"b3f5baf1-544d-4e8c-a657-cec309057bcf","order_by":6,"name":"Guodong Li","email":"","orcid":"","institution":"Yunnan University of Chinese Medicine","correspondingAuthor":false,"prefix":"","firstName":"Guodong","middleName":"","lastName":"Li","suffix":""},{"id":572865766,"identity":"3c0775f7-32d6-4b5c-9cf8-c3e17541dfd0","order_by":7,"name":"Ticao Zhang","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABD0lEQVRIiWNgGAWjYDACCTjJfICBwcCCQYKZhxgtCSCSLQGoRYJoLSCCxwDMlWAgoEV+dvOzh19/WMjzt/d83fCjQIJBsp33AMOPGgZ5cxxaGOccMzeWSZAwnHHm7LabPUCHSTPzJTD2HGMw3NmAXQuzRIKZtESCRALDjdxtN3iAWuSYgS7kbWBIMDiAXQubRPo3sBb5GznPbv6BamH8i0cLj0SOmeQHoBaDGzlst3nADuMxYMZni4RETpk0Q5qE4cYzx8xuyxhI8Eg28yUcljkmYbgBhxb5GenbJH/Y1MnLHW9+dvPNHxs5ifNnDz58U2Mjj8sWcBAgRwSYfQCWKnABxh94pUfBKBgFo2DEAwCfRE7v+r/NOQAAAABJRU5ErkJggg==","orcid":"","institution":"Kunming Institute of Botany, Chinese Academy of Sciences","correspondingAuthor":true,"prefix":"","firstName":"Ticao","middleName":"","lastName":"Zhang","suffix":""}],"badges":[],"createdAt":"2025-11-20 10:16:56","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-8163245/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8163245/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":100179015,"identity":"4f5395f5-6b5c-4067-b278-7b341c41cecd","added_by":"auto","created_at":"2026-01-13 18:46:11","extension":"docx","order_by":0,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":6114138,"visible":true,"origin":"","legend":"","description":"","filename":"ComparativeanalysisoforganellegenomesinthreeTerminaliaspeciesrevealstructureevolutionandphylogeneticposition.docx","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/18dee25dd5de220175e27e2c.docx"},{"id":100179007,"identity":"e7eb9adb-207a-44eb-8a79-3ab35b4df203","added_by":"auto","created_at":"2026-01-13 18:46:11","extension":"json","order_by":1,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":9152,"visible":true,"origin":"","legend":"","description":"","filename":"2efcdf89d2ca4f6e8231beec6a78812e.json","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/eed9d5bb1a4518b261a1ba12.json"},{"id":100369617,"identity":"cd24888f-5676-4468-865e-f9bec3964620","added_by":"auto","created_at":"2026-01-16 07:59:11","extension":"xml","order_by":2,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":160779,"visible":true,"origin":"","legend":"","description":"","filename":"2efcdf89d2ca4f6e8231beec6a78812e1enriched.xml","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/fbb356f5ab49297c30c5f1d8.xml"},{"id":100179011,"identity":"e58e6b69-fd50-4465-82cb-62c6894b3718","added_by":"auto","created_at":"2026-01-13 18:46:11","extension":"png","order_by":3,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":1963067,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/da12f4b0a3c45062522a384b.png"},{"id":100179008,"identity":"832e7635-d927-4e49-b0a6-76f0964627ef","added_by":"auto","created_at":"2026-01-13 18:46:11","extension":"png","order_by":4,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":341409,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/b07793b42f44b59a1015bf61.png"},{"id":100370037,"identity":"92fb9ef9-0845-4e33-8da9-5ff9e21520fb","added_by":"auto","created_at":"2026-01-16 07:59:49","extension":"png","order_by":5,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":439973,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/d95a2b7ff94e5963e7969799.png"},{"id":100179019,"identity":"457c578a-cb0d-4963-b11b-e00bd257a074","added_by":"auto","created_at":"2026-01-13 18:46:11","extension":"png","order_by":6,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":963252,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/1c6a12d90c47b0b1c481e8eb.png"},{"id":100179013,"identity":"70a0739e-a858-46fa-b101-219adeaf916b","added_by":"auto","created_at":"2026-01-13 18:46:11","extension":"png","order_by":7,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":2209173,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/20259b2d3db4df26f20d0fe2.png"},{"id":100369466,"identity":"2632a35f-e6f0-416b-8b96-436e7d0c4f32","added_by":"auto","created_at":"2026-01-16 07:59:04","extension":"png","order_by":8,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":463102,"visible":true,"origin":"","legend":"","description":"","filename":"floatimage6.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/eb744b0ecc425f986b4eed4e.png"},{"id":100369352,"identity":"29cc4191-db87-48ee-b76a-83a24c56bfbd","added_by":"auto","created_at":"2026-01-16 07:58:57","extension":"png","order_by":9,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":498701,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/2db8ab88db48d842353eb6d2.png"},{"id":100368796,"identity":"8254ab8e-9224-4329-aad2-bf18582fbedf","added_by":"auto","created_at":"2026-01-16 07:58:22","extension":"png","order_by":10,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":138024,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/bb242df75a3a08063bc109b0.png"},{"id":100370040,"identity":"253d4be2-c250-4fb6-b6cb-979609ca58b7","added_by":"auto","created_at":"2026-01-16 07:59:50","extension":"png","order_by":11,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":196876,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/f2121f6675fc4876d071a1a9.png"},{"id":100179021,"identity":"f5572d05-356e-4042-a2cd-860bf7b329b4","added_by":"auto","created_at":"2026-01-13 18:46:11","extension":"png","order_by":12,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":311435,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/1325e7a26e71df65f37287a1.png"},{"id":100179024,"identity":"44b98c27-6ad0-46eb-ae09-fc524542f353","added_by":"auto","created_at":"2026-01-13 18:46:11","extension":"png","order_by":13,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":358892,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/055cc25656ed539842933bf8.png"},{"id":100368701,"identity":"9027aac4-1c36-4627-9c3f-76a1cfb1cb72","added_by":"auto","created_at":"2026-01-16 07:58:17","extension":"png","order_by":14,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":184152,"visible":true,"origin":"","legend":"","description":"","filename":"Onlinefloatimage6.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/ff05622cb2938f87104179cc.png"},{"id":100179026,"identity":"26d00b60-6019-4925-91c4-c35bde44a0b5","added_by":"auto","created_at":"2026-01-13 18:46:11","extension":"xml","order_by":15,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":155794,"visible":true,"origin":"","legend":"","description":"","filename":"2efcdf89d2ca4f6e8231beec6a78812e1structuring.xml","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/53c3d5d85d95be1db2fbda9c.xml"},{"id":100368685,"identity":"1ef01024-58a0-495d-a702-bd7cd4eb59b4","added_by":"auto","created_at":"2026-01-16 07:58:16","extension":"html","order_by":16,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":179219,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/650f19b5d24af8347be4ca2a.html"},{"id":100369624,"identity":"5e8a716b-c0be-4d6f-9ad4-fd5d915ac513","added_by":"auto","created_at":"2026-01-16 07:59:11","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":1963067,"visible":true,"origin":"","legend":"\u003cp\u003eGenome maps of the plastomes of three \u003cem\u003eTerminalia\u003c/em\u003e Species. (A-C) plastomes, (D-F) mitogenome\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/c843c3460801af9d3ea0648d.png"},{"id":100179003,"identity":"9bf2ccb2-c929-47dc-9762-4cca6ee747ce","added_by":"auto","created_at":"2026-01-13 18:46:10","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":341409,"visible":true,"origin":"","legend":"\u003cp\u003eThe repeat sequences in the plastomes of the three \u003cem\u003eTerminalia\u003c/em\u003e species: (A) plastome (B) mitogenome\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/8a99ced6502259f35ae86f04.png"},{"id":100369348,"identity":"971e0061-5bcd-47e3-9212-7644381adf26","added_by":"auto","created_at":"2026-01-16 07:58:57","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":439973,"visible":true,"origin":"","legend":"\u003cp\u003eRelative synonymous codon usage (RSCU) of 3 \u003cem\u003eTerminalia\u003c/em\u003e ganelle genes. (A) mitogenome, (B) plastome.\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/9ccefe41c3c22c676747aebb.png"},{"id":100179010,"identity":"d240ebf6-f404-42ce-9dee-1454d2b49ef9","added_by":"auto","created_at":"2026-01-13 18:46:11","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":963252,"visible":true,"origin":"","legend":"\u003cp\u003e(A). The RNA editing sites of mitogenomes of three \u003cem\u003eTerminalia\u003c/em\u003e species. (B). Boxplots of the pairwise Ka/Ks values among every shared mitogenomes of the 37 PCGs. The x-axis represents gene names, and the y-axis represents the values of Ka and Ks. The red dashed line represents 1. In the box plot, the vertical lines above and below the box indicate the upper and lower quartiles of the data, while the thick line inside the box represents the median. (C). Homology analysis between mitogenomes and plastomes of three \u003cem\u003eTerminalia\u003c/em\u003e species. The blue and the green arcs denote the mtDNA and cpDNA, respectively. The grey lines signify the homologous sequence\u003c/p\u003e","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/1b046abb9a8e0c3768cc5d6b.png"},{"id":100179006,"identity":"8cd080cc-d548-4fd8-9367-50deb09463ac","added_by":"auto","created_at":"2026-01-13 18:46:11","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":2209173,"visible":true,"origin":"","legend":"\u003cp\u003e(A). Mauve visualization of synteny analysis among three \u003cem\u003eTerminalia\u003c/em\u003especies: Colored blocks (LCBs) represent homologous gene segments, with consistent coloring indicating conserved sequences (homologous relationships). Connecting lines show the syntenic associations of LCBs across different genomes (indicating the order and direction of arrangement). (B) Comparison of the borders of the LSC, SSC, and IR regions among the six plastomes of Combretaceae species. (JLB: junction of the LSC and IRB; JSB: junction of the SSC and IRB; JSA: junction of the SSC and IRA; and JLA: junction of the LSC and IRA).\u003c/p\u003e","description":"","filename":"floatimage5.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/b68d662128ccddd04776a367.png"},{"id":100369587,"identity":"cdbe14f8-4fdc-44e7-b19d-a664622d6273","added_by":"auto","created_at":"2026-01-16 07:59:09","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":463102,"visible":true,"origin":"","legend":"\u003cp\u003e(A) Phylogenetic tree of the mitogenomes constructed using the ML method. (B) Phylogenetic tree of the plastomes constructed using the ML method.\u003c/p\u003e","description":"","filename":"floatimage6.png","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/e71cd75ced2074b003dfe744.png"},{"id":100382775,"identity":"5fc3b2ee-dcfb-4bc6-a2b4-4d1915550416","added_by":"auto","created_at":"2026-01-16 10:44:10","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":6389111,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8163245/v1/6375fc0a-52dd-42b2-81b9-a6b28eeac306.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Comparative analysis of organelle genomes in three Terminalia species reveals structure evolution and phylogenetic position","fulltext":[{"header":"Background","content":"\u003cp\u003e \u003cem\u003eTerminalia\u003c/em\u003e belongs to the Combretaceae family, comprising approximately 200 species widely distributed across tropical and subtropical regions, particularly in Asia, Africa, and Australia. Most \u003cem\u003eTerminalia\u003c/em\u003e species are deciduous or evergreen trees or shrubs, commonly found in forests, riverbanks, and wetlands [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. Numerous \u003cem\u003eTerminalia\u003c/em\u003e plants have extensive applications in traditional ethnopharmacology, serving as herbal remedies or formulations for treating various ailments such as headaches, fever, pneumonia, influenza, geriatric diseases, and cancer [\u003cspan additionalcitationids=\"CR3\" citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. Additionally, they are employed to alleviate symptoms including memory loss, abdominal pain, back pain, cough, colds, conjunctivitis, diarrhea, heart disease, leprosy, sexually transmitted diseases, and urinary tract infections [\u003cspan additionalcitationids=\"CR6\" citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Research indicates these plants exhibit diverse biological activities, encompassing antibacterial, antifungal, anti-inflammatory, antiviral, antiretroviral, and antioxidant properties [\u003cspan additionalcitationids=\"CR9\" citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Current research on \u003cem\u003eTerminalia\u003c/em\u003e primarily focuses on pharmacological effects [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e], active constituents [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e], processing methods, and molecular identification. However, no studies have reported on its mitochondrial genome (mitogenome) or chloroplast genome (plastome). Unraveling its mitogenomes and plastomes is crucial for the phylogenetic studies of \u003cem\u003eTerminalia\u003c/em\u003e, providing vital clues for species origins, evolutionary relationships, and taxonomy.\u003c/p\u003e \u003cp\u003eWith the advancement of high-throughput sequencing, the genomes of organelles from an increasing number of plants have been sequenced, offering significant data for studying plant physiological functions, evolutionary relationships, and germplasm resource conservation. Both mitochondria and chloroplasts are believed to have evolved from ancient free-living bacteria through endosymbiosis [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. Mitochondria, serving as the \u0026ldquo;powerhouse\u0026rdquo; of the cell, generate ATP through oxidative phosphorylation and function as the central hub of plant energy metabolism [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. Compared to animal mitogenomes, plant mitogenomes contain not only genes related to mitochondrial function but also repetitive sequences. While plastomes typically maintain a relatively uniform circular structure [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e, \u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e], mitogenomes are characterized by a high incidence of repetitive sequences and exhibit significant structural heterogeneity, including polycyclic [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e], linear [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e], and branched forms [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. Studying plant mitogenomes can reveal mechanisms of plant energy metabolism [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e], providing new theoretical foundations for agricultural production, such as enhancing plant stress resistance and advancing breeding efforts [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eIn this study, we successfully assembled the plastomes and mitogenomes of three previously unreported \u003cem\u003eTerminalia\u003c/em\u003e species: \u003cem\u003eTerminalia chebula\u003c/em\u003e, \u003cem\u003eTerminalia franchetii\u003c/em\u003e, and \u003cem\u003eTerminalia intricata\u003c/em\u003e. We analyzed the organelle genomes of these \u003cem\u003eTerminalia\u003c/em\u003e species by comparing the newly obtained sequences with existing organelle genome data. Analyses included GC content, codon usage bias, repetitive sequences, Ka/Ks ratios, prediction of RNA editing events, and nucleotide polymorphism analysis. Homologous sequences found in both chloroplast and mitogenomes were also analyzed. Furthermore, maximum likelihood phylogenetic trees were constructed using PCGs from 17 mitogenomes and 18 plastomes, respectively. We reconstructed phylogenetic relationships within the \u003cem\u003eTerminalia\u003c/em\u003e, exploring connections and taxonomic boundaries between different species groups, and identified key regions of genetic variation. These findings will provide important references and valuable insights for molecular marker development and phylogenetic studies in the \u003cem\u003eTerminalia\u003c/em\u003e.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cstrong\u003eGenome organization of three \u003cem\u003eTerminalia\u003c/em\u003e organelles\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe plastomes sizes of \u003cem\u003eT. chebula\u003c/em\u003e, \u003cem\u003eT. franchetii\u003c/em\u003e, and \u003cem\u003eT. intricata\u003c/em\u003e were 159,915 bp, 159,700 bp, and 159,971 bp, respectively (Figure 1). All three genomes exhibited a typical quadripartite structure, consisting of a pair of inverted repeats (IRs) with lengths of 26,339 bp in \u003cem\u003eT. chebula\u003c/em\u003e, 26,278 bp in \u003cem\u003eT. franchetii\u003c/em\u003e, and 26,345 bp in \u003cem\u003eT. intricata\u003c/em\u003e. The small single-copy region (18,888 bp in \u003cem\u003eT. chebula\u003c/em\u003e, 18,734 bp in \u003cem\u003eT. franchetii\u003c/em\u003e, and 18,908bp in \u003cem\u003eT. intricata\u003c/em\u003e) and a large single-copy region (71,568 bp in \u003cem\u003eT. chebula\u003c/em\u003e, 71,290 bp in \u003cem\u003eT. franchetii\u003c/em\u003e, and 71,600 bp in \u003cem\u003eT. intricata\u003c/em\u003e). In the plastomes of \u003cem\u003eT. chebula\u003c/em\u003e, \u003cem\u003eT. franchetii\u003c/em\u003e, and \u003cem\u003eT. intricata\u003c/em\u003e, 76, 76, and 73 PCGs were annotated, respectively, along with 27, 26, and 26 tRNA genes. All three species contained 4 rRNA genes.\u003c/p\u003e\n\u003cp\u003eInterestingly, the mitogenome structures of the three \u003cem\u003eTerminalia\u003c/em\u003e species showed significant differences and were highly complex. The mitogenomes structure of \u003cem\u003eT. chebula\u003c/em\u003e and \u003cem\u003eT. intricata\u003c/em\u003e consisted of three circular molecules and one linear molecule, while \u003cem\u003eT. franchetii\u003c/em\u003e had two linear molecules. (Figure 1). The sizes of the mitogenomes for \u003cem\u003eT. chebula\u003c/em\u003e, \u003cem\u003eT. franchetii\u003c/em\u003e, and \u003cem\u003eT. intricata\u003c/em\u003e were 354,989 bp, 365,711 bp, and 350,904 bp, respectively. In these genomes, 35, 34, and 38 PCGs were annotated for \u003cem\u003eT. chebula\u003c/em\u003e, \u003cem\u003eT. franchetii\u003c/em\u003e, and \u003cem\u003eT. intricata\u003c/em\u003e, respectively. Additionally, \u003cem\u003eT. chebula\u003c/em\u003e contained 17 tRNA genes, \u003cem\u003eT. franchetii\u003c/em\u003e had 18, and \u003cem\u003eT. intricata\u003c/em\u003e had 17. The rRNA gene counts were 3 for \u003cem\u003eT. chebula\u003c/em\u003e, 4 for \u003cem\u003eT. franchetii\u003c/em\u003e, and 3 for \u003cem\u003eT. intricata\u003c/em\u003e. The GC contents of the mitogenomes for \u003cem\u003eT. chebula\u003c/em\u003e, \u003cem\u003eT. franchetii\u003c/em\u003e, and \u003cem\u003eT. intricata\u003c/em\u003e were 45.5%, 44.3%, and 44.7%, respectively.\u003c/p\u003e\n\u003cp\u003eThe plastomes of the three\u003cem\u003e\u0026nbsp;Terminalia\u0026nbsp;\u003c/em\u003especies exhibited a circular structure, with sizes ranging from 159,700bp to 159,971 bp, showing a maximum size difference of 271 bp. The number of genes in the plastomes varied between 106 and 108 across species. In contrast, the mitogenomes displayed significant structural variations, with sizes ranging from 350,904 bp to 365,711 bp, resulting in a maximum size difference of 14,807 bp. The number of genes in the mitogenomes varied between 55 and 58 across species (Table 1). These results indicate that the mitogenomes exhibit substantial fluctuations in structure, genome size, and GC content, whereas the plastomes appear to be more conserved throughout evolution (Additional file 1).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 1 Genomic characteristics of three \u003cem\u003eTerminalia\u0026nbsp;\u003c/em\u003especies\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"506\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 102px;\"\u003e\n \u003cp\u003eSpecies\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 98px;\"\u003e\n \u003cp\u003eGenome Type\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 72px;\"\u003e\n \u003cp\u003eGenome Size (bp)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 64px;\"\u003e\n \u003cp\u003eProtein Genes\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003etRNA Genes\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003erRNA Genes\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003eTotal Genes\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 102px;\"\u003e\n \u003cp\u003e\u003cem\u003eT. chebula\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 98px;\"\u003e\n \u003cp\u003eplastome\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 72px;\"\u003e\n \u003cp\u003e159,915\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 64px;\"\u003e\n \u003cp\u003e76\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e27\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e107\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 102px;\"\u003e\n \u003cp\u003e\u003cem\u003eT. franchetii\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 98px;\"\u003e\n \u003cp\u003eplastome\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 72px;\"\u003e\n \u003cp\u003e159,700\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 64px;\"\u003e\n \u003cp\u003e76\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e26\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e106\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 102px;\"\u003e\n \u003cp\u003e\u003cem\u003eT. intricata\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 98px;\"\u003e\n \u003cp\u003eplastome\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 72px;\"\u003e\n \u003cp\u003e159,971\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 64px;\"\u003e\n \u003cp\u003e73\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e26\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e103\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 102px;\"\u003e\n \u003cp\u003e\u003cem\u003eT. chebula\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 98px;\"\u003e\n \u003cp\u003emitogenome\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 72px;\"\u003e\n \u003cp\u003e354,989\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 64px;\"\u003e\n \u003cp\u003e35\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e55\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 102px;\"\u003e\n \u003cp\u003e\u003cem\u003eT. franchetii\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 98px;\"\u003e\n \u003cp\u003emitogenome\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 72px;\"\u003e\n \u003cp\u003e365,711\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 64px;\"\u003e\n \u003cp\u003e34\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e18\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e4\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e56\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 102px;\"\u003e\n \u003cp\u003e\u003cem\u003eT. intricata\u003c/em\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 98px;\"\u003e\n \u003cp\u003emitogenome\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 72px;\"\u003e\n \u003cp\u003e350,904\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 64px;\"\u003e\n \u003cp\u003e38\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e17\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e3\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 57px;\"\u003e\n \u003cp\u003e58\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003e\u003cstrong\u003eRepeat sequence analysis of three \u003cem\u003eTerminalia\u003c/em\u003e organelles\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn the plastomes of \u003cem\u003eT. chebula\u003c/em\u003e, \u003cem\u003eT. franchetii\u003c/em\u003e, and \u003cem\u003eT. intricata\u003c/em\u003e, 73, 70, and 70 simple sequence repeats (SSRs) were identified, respectively (Figure 2A). All SSRs in \u003cem\u003eT. chebula\u003c/em\u003e were haploid, \u003cem\u003eT. franchetii\u0026nbsp;\u003c/em\u003eonly contained one diploid SSR, while \u003cem\u003eT. intricata\u003c/em\u003e had one diploid SSR and one triploid SSR, with no higher ploidy detected. The plastomes of all three \u003cem\u003eTerminalia\u0026nbsp;\u003c/em\u003especies contained complementary, palindromic, forward, reverse, and tandem repeats, with tandem and palindromic repeats being the predominant types. Among these species, \u003cem\u003eT. chebula\u003c/em\u003e exhibited the highest number of tandem repeats, with 41 in total.\u003c/p\u003e\n\u003cp\u003eIn the mitogenomes of \u003cem\u003eT. chebula\u003c/em\u003e, \u003cem\u003eT. franchetii\u003c/em\u003e, and \u003cem\u003eT. intricata\u003c/em\u003e, 57, 56, and 57 SSRs were identified, respectively. Among these, haploid repeats predominated, followed by diploid and triploid repeats, with no higher ploidy detected. The mitogenomes of three \u003cem\u003eTerminalia\u003c/em\u003e species contained complementary, palindromic, forward, reverse, and tandem repeats. In \u003cem\u003eT. chebula\u003c/em\u003e, forward (42.1%, 86 repeats) and palindromic (28.9%, 59 repeats) repeats were the most common. In \u003cem\u003eT. franchetii\u003c/em\u003e, forward (41%, 41 repeats) and palindromic (38%, 38 repeats) repeats were also dominant. Notably, \u003cem\u003eT. franchetii\u003c/em\u003e exhibited the highest number of tandem repeats among the three species, with 42 repeats. In \u003cem\u003eT. intricata\u003c/em\u003e, forward (42.7%, 85 repeats) and palindromic (31.1%, 62 repeats) repeats were similarly predominant (Figure 2B).\u003c/p\u003e\n\u003cp\u003eIn the plastomes and mitogenomes of the three \u003cem\u003eTerminalia\u003c/em\u003e species, several types of repeat sequences were present, including complementary repeats, palindromic repeats, forward repeats, reverse repeats, and tandem repeats. The distribution of simple sequence repeats (SSRs) across the two types of genomes showed similarities, with haploid repeats predominating. Although diploid and triploid repeats were also found in the mitogenomes, haploid repeats formed the highest proportion (Additional file 2).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCodon usage analysis of three \u003cem\u003eTerminalia\u003c/em\u003e organelles\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study compared the codon usage preferences in the mitochondrial and plastomes of \u003cem\u003eT. chebula\u003c/em\u003e, \u003cem\u003eT. franchetii\u003c/em\u003e, and \u003cem\u003eT. intricata\u003c/em\u003e, and analyzed their relative synonymous codon usage frequencies (Figure 3A). Arginine and Leucine were the most frequently used amino acids in the organellar genomes of the three \u003cem\u003eTerminalia\u0026nbsp;\u003c/em\u003especies, accounting for 9.8% of mitogenomes and 9.6% of plastomes, respectively. In comparative studies of plastomes across species, the TTA codon exhibited a strong preference (2.12) in \u003cem\u003eT. chebula\u003c/em\u003e, while AGA showed the highest codon usage preference in\u003cem\u003e\u0026nbsp;T. franchetii\u0026nbsp;\u003c/em\u003eand \u003cem\u003eT. intricata\u003c/em\u003e, with values of 1.9269 and 1.9988, respectively. Conversely, codons such as CTG and GCG exhibited significantly lower RSCU values across all three species, indicating their infrequent usage.\u003c/p\u003e\n\u003cp\u003eIn the mitogenomes of the three\u003cem\u003e\u0026nbsp;Terminalia\u0026nbsp;\u003c/em\u003especies, the ATG and TGG codons did not have synonymous codons, and their RSCU values were both 1 (Figure 3B). There were differences in codon usage preferences among the species. In \u003cem\u003eT. chebula\u003c/em\u003e, the most preferred codon was GCT (1.57), followed by TTA (1.55), while the least preferred codon was GCG, with an RSCU value of 0.47. In contrast, \u003cem\u003eT. franchetii\u0026nbsp;\u003c/em\u003eand \u003cem\u003eT. intricata\u0026nbsp;\u003c/em\u003eshowed highly consistent codon usage preferences, with the most commonly used codon being AGA, with RSCU values of 1.77 and 1.73, respectively. The lowest RSCU value for both species was CGT, with values of 0.59 and 0.56, respectively (Additional file 3).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eRNA editing site analysis of mitogenomes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eRNA editing involves the addition, deletion, or alteration of bases in the coding region of the transcribed RNA [23], and it occurs in all eukaryotes, including plants [24]. The results showed that 311, 364, and 349 RNA editing sites were predicted in mitogenomes of \u003cem\u003eT. chebula\u003c/em\u003e, \u003cem\u003eT. franchetii\u003c/em\u003e, and \u003cem\u003eT. intricata\u003c/em\u003e, respectively, with all editing events involving a C-to-U base conversion (Additional file 4). This editing pattern is consistent with that observed in other plants. In all three species of the \u003cem\u003eTerminalia\u003c/em\u003e, genes such as \u003cem\u003emttB\u003c/em\u003e, \u003cem\u003erps4\u003c/em\u003e, and \u003cem\u003ecob\u003c/em\u003e have a higher number of editing sites, while genes like \u003cem\u003eatp8\u003c/em\u003e, \u003cem\u003eatp9\u003c/em\u003e, and \u003cem\u003esdh3\u003c/em\u003e have fewer editing sites. In the effective amino acid editing events of the three species, hydrophobic amino acids dominated, accounting for approximately 71.3% to 71.5%, while hydrophilic amino acids accounted for around 28.5%. The high-frequency codons were biased toward hydrophobic amino acid coding, with the codons for leucine (L) and phenylalanine (F) being more frequent. Specifically, UUA and UUC appeared frequently across the three species (Figure 4A).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eSubstitution rates of mitochondrial PCGs\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eDifferent species face varying ecological pressures during evolution, leading to genomic changes. The Ka/Ks ratio measures selection pressure on PCGs, helping infer species adaptation and evolutionary trajectories. When the Ka/Ks ratio is \u0026gt;\u0026thinsp;1, genes are undergoing positive selection, conversely, Ka/Ks\u0026thinsp;\u0026lt;\u0026thinsp;1 denotes negative selection, and Ka/Ks\u0026thinsp;=\u0026thinsp;1 represents neutral selection. To calculate the selective pressure on mitogenomes PCGs in three species of \u003cem\u003eTerminalia\u003c/em\u003e, we computed the Ka/Ks values for the PCGs of 37 mitogenomes (Additional file 5). The Ka/Ks values of most PCGs were found to be below 1, while genes like \u003cem\u003enad4\u003c/em\u003e,\u003cem\u003e\u0026nbsp;rps1\u003c/em\u003e, \u003cem\u003erps4\u003c/em\u003e, and\u003cem\u003e\u0026nbsp;rps7\u003c/em\u003e had Ka/Ks values exceeding 1(Figure 4B).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eHomology analysis of genome sequences\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe searched for homologous sequences between the\u0026nbsp;plastomes and mitogenomes in three \u003cem\u003eTerminalia\u003c/em\u003e species to identify potential gene transfer events. We identified 17-26 homologous sequences between the plastomes and mitogenomes in the three species, with lengths ranging from 28bp to 7035bp (Figure 4C). Interestingly, we identified some complete plastid genome-derived PCGs in the mitogenomes, including \u003cem\u003epetB\u003c/em\u003e, \u003cem\u003epetG\u003c/em\u003e, \u003cem\u003epsaA\u003c/em\u003e, \u003cem\u003epsbJ\u003c/em\u003e, \u003cem\u003epetB\u003c/em\u003e, \u003cem\u003epsbM\u003c/em\u003e, \u003cem\u003eycf2\u003c/em\u003e, and \u003cem\u003epsb\u003c/em\u003e, among others. Notably, \u003cem\u003epsbJ\u003c/em\u003e,\u003cem\u003e\u0026nbsp;psaA\u003c/em\u003e, \u003cem\u003epetB\u003c/em\u003e, and \u003cem\u003epetG\u003c/em\u003e were shared by all three species.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCollinearity analysis of the mitogenomes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMauve was used to analyze the synteny and rearrangements of the mitogenomes across the three species. The three genomes exhibited a large number of locally collinear blocks (LCBs) with consistent coloring, indicating that they share a substantial number of conserved gene segments, which reflects the close evolutionary relationship within the genus. These homologous regions (LCBs) had different relative positions, suggesting the presence of extensive rearrangements within the mitogenomes (Figure 5A).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eNucleotide polymorphism (Pi) analysis of organelle genomes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe Pi analysis was conducted on 77 plastome PCGs from five species of Combretaceae (\u003cem\u003eTerminalia chebula\u003c/em\u003e, \u003cem\u003eTerminalia franchetii\u003c/em\u003e, \u003cem\u003eTerminalia intricata\u003c/em\u003e,\u003cem\u003e\u0026nbsp;\u003c/em\u003e\u003cem\u003eTerminalia phillyreifolia,\u003c/em\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003cem\u003eTerminalia guyanensis\u003c/em\u003e). The results revealed that among the 77 PCGs, gene\u003cem\u003e\u0026nbsp;rpl22\u003c/em\u003e exhibited the highest Pi value (0.11406), followed by \u003cem\u003erps19\u003c/em\u003e (0.104353). In contrast, the lowest Pi values were observed for \u003cem\u003endhB\u003c/em\u003e (0.009983). We also performed the Pi analysis on 37 mitochondrial PCGs from three species of \u003cem\u003eTerminalia\u003c/em\u003e, and the results revealed that the highest Pi value was observed for \u003cem\u003erps10\u003c/em\u003e (0.117162), followed by \u003cem\u003enad9\u003c/em\u003e (0.1079), while the lowest Pi values, both 0, were found for\u003cem\u003e\u0026nbsp;rpl16\u0026nbsp;\u003c/em\u003eand \u003cem\u003erpl19\u003c/em\u003e (Figure S1. Additional file 6).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eExpansion and contraction of the IR region\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA comparative analysis of the IR/SSC boundary regions in the plastomes of 6 Combretaceae species revealed subtle differences at the IR/SSC junction. In this study, the \u003cem\u003erps19\u003c/em\u003e gene in \u003cem\u003eLumnitzera racemosa\u003c/em\u003e, \u003cem\u003eLumnitzera littorea\u003c/em\u003e spanned 106 bp of the LSC/IRb region, while in \u003cem\u003eTerminalia franchetii\u003c/em\u003e and \u003cem\u003eTerminalia intricata\u003c/em\u003e and \u003cem\u003eTerminalia chebula\u003c/em\u003e it spanned 16-24 bp of the LSC/IRb region. In contrast, the \u003cem\u003erps19\u003c/em\u003e gene in\u003cem\u003e\u0026nbsp;Laguncularia racemosa\u003c/em\u003e was entirely located within the LSC region. Significant differences were observed at the IRb/SSC junction (JSB). It is noteworthy that the position of the \u003cem\u003erpl2\u003c/em\u003e gene in \u003cem\u003eTerminalia intricata\u003c/em\u003e and \u003cem\u003eTerminalia chebula\u003c/em\u003e differed significantly from that in other species. This difference may arise from the dynamic evolution of the plastome IR region boundaries. At this junction, only\u003cem\u003e\u0026nbsp;Lumnitzera racemosa\u003c/em\u003e, \u003cem\u003eLumnitzera littorea\u003c/em\u003e, and \u003cem\u003eLaguncularia racemosa\u003c/em\u003e contained the \u003cem\u003eycf1\u003c/em\u003e gene, while the other 3 species lacked it. Additionally, the \u003cem\u003endhF\u003c/em\u003e gene was located at the IRb/SSC junction. In \u003cem\u003eLumnitzera racemosa\u003c/em\u003e, \u003cem\u003eLumnitzera littorea\u003c/em\u003e, \u003cem\u003eLaguncularia racemosa\u003c/em\u003e, and \u003cem\u003eTerminalia chebula\u003c/em\u003e, 15-18 bp of the\u003cem\u003e\u0026nbsp;ndhF\u003c/em\u003e gene are located in the IRb region, while in \u003cem\u003eTerminalia franchetii\u003c/em\u003e and \u003cem\u003eTerminalia intricata\u003c/em\u003e, 70-75 bp were located in the IRb region. Notably, the \u003cem\u003eycf1\u003c/em\u003e gene spanned the SSC/IRa junction in all 6 species. The \u003cem\u003epsbA\u003c/em\u003e and \u003cem\u003etrnH\u003c/em\u003e genes were completely located in the LSC region (Figure 5B).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003ePhylogenetic analysis of the organelle genom\u003c/strong\u003e\u003cstrong\u003ee\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA phylogenetic tree of 17 angiosperm species from five orders was constructed by extracting mitogenome PCGs sequences, with \u003cem\u003eVitis vinifera\u003c/em\u003e as the outgroup (Figure 6A). \u003cem\u003eT. chebula\u003c/em\u003e was confirmed as a species of the genus \u003cem\u003eTerminalia\u003c/em\u003e,\u0026nbsp;\u003cem\u003eT. intricata\u003c/em\u003e clustering with\u003cem\u003e\u0026nbsp;T. franchetii\u003c/em\u003e. The phylogenetic tree shows that the bootstrap support values for most nodes were above 95%. The overall topology of the tree is consistent with the classification results of the Angiosperm Phylogeny Group IV (APG IV) system.\u003c/p\u003e\n\u003cp\u003eTo clarify the phylogenetic position and evolutionary relationships of the Combretaceae family within the Myrtales order, a phylogenetic analysis was conducted using\u0026nbsp;plastome sequences from eight species of Combretaceae and ten other plant species, with \u003cem\u003eR. solosa\u003c/em\u003e from the Malpighiales as the outgroup. The results indicated that species within the \u003cem\u003eTerminalia\u0026nbsp;\u003c/em\u003eof Combretaceae exhibited a high degree of phylogenetic clustering, with strong branch support, revealing close genetic relationships among the species in this genus. \u003cem\u003eT. franchetii\u003c/em\u003e and \u003cem\u003eT. intricata\u003c/em\u003e were closely clustered in one group with a bootstrap support of 100%. The Onagraceae family, represented by \u003cem\u003eO. biennis\u003c/em\u003e and \u003cem\u003eO. parviflora\u003c/em\u003e, formed a separate clade, while the Lythraceae family, represented by\u003cem\u003e\u0026nbsp;P. granatum\u003c/em\u003e, appeared as an independent branch. The Myrtaceae family, which includes \u003cem\u003eA. costata\u003c/em\u003e, \u003cem\u003eC. henryi\u003c/em\u003e, \u003cem\u003eC. gummifera\u003c/em\u003e, \u003cem\u003eE. aromaphloia\u003c/em\u003e, \u003cem\u003eE. deglupta\u003c/em\u003e, and\u003cem\u003e\u0026nbsp;P. trunciflora\u003c/em\u003e, formed a stable cluster \u003cem\u003eR. stylosa\u003c/em\u003e from the Rhizophoraceae family clustered separately within the order of the Combretaceae, displaying significant evolutionary divergence from Myrtales species. Furthermore, within the Myrtales, the Onagraceae family exhibited a closer phylogenetic relationship with the Lythraceae family (Figure 6B).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eWith the advent of advanced sequencing technologies, and the successful assembly of an increasing number of genomes, the complexity of mitogenomes has become more evident [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. The genome structure of mitochondria also exhibits diversity, including linear, circular, highly branched, and networked forms, with such diversity even observed within a single individual, possibly due to frequent recombination [\u003cspan additionalcitationids=\"CR27\" citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. The earliest mitogenome sequencing from animals was performed on humans and Drosophila, where the genomes were found to be small, circular molecules with highly conserved sequences [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e, \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e]. As a result, it was once assumed that all mitogenomes were single, circular molecules. However, recent studies have shown that mitogenomes are not always single circular structures. For example, \u003cem\u003eQuercus acutissima\u003c/em\u003e (Fagaceae) consists of one linear and two circular molecules [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e]. \u003cem\u003eColocasia esculenta\u003c/em\u003e (Araceae) consists of five circular molecules [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e], while \u003cem\u003eEchinacanthus longipes\u003c/em\u003e (Acanthaceae) consists of two circular and three linear molecules [\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e]. The mitogenomes of three species in the \u003cem\u003eTerminalia\u003c/em\u003e exhibit significant structural differences. The mitogenomes of \u003cem\u003eT. chebula\u003c/em\u003e and \u003cem\u003eT. intricata\u003c/em\u003e consist of three circular molecules and one linear molecule, while \u003cem\u003eT. franchetii\u003c/em\u003e has two linear molecules. Mitogenomes in plants can show considerable variation between species. The size of plant mitogenomes also varies widely, typically ranging from 66 kb to 11,000 kb. The mitogenome sizes of the three \u003cem\u003eTerminalia\u003c/em\u003e species fall between 350,904 and 354,989 bp. Studies have shown that plant mitogenomes can exhibit significant differences between species [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e]. The structural and size variations of the mitogenomes in \u003cem\u003eTerminalia\u003c/em\u003e species are considerable, which may be related to species evolution and phylogenetic affinity.\u003c/p\u003e \u003cp\u003eFurthermore, the plastomes of three \u003cem\u003eTerminalia\u003c/em\u003e species possess a unique quadripartite structure, consisting of two IR regions, one LSC region, and one SSC region, which is similar to the plastomes structure of other angiosperms [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e]. Meanwhile, GC content significantly affects amino acid composition and the structural integrity of genomes during evolutionary processes [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e]. Our analysis of the GC content in the plastomes and mitogenomes of the three \u003cem\u003eTerminalia\u003c/em\u003e species indicates that plastomes exhibit more stable GC content (36.8%-37.0%), while the mitogenomes show greater variability in GC content (44.3%-45.5%). This result supports the conclusion that GC content is highly conserved in higher plants. Interestingly, among the three \u003cem\u003eTerminalia\u003c/em\u003e species, the plastome lengths of the three \u003cem\u003eTerminalia\u003c/em\u003e showed a maximum size difference of 271 bp, which suggests that variability in cp genome size is closely related to phylogenetic relationships and evolutionary history [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eRNA editing is a post-transcriptional process in which nucleotides within mRNA molecules are inserted, deleted, or substituted, resulting in changes to the genetic code [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e]. Previous studies have shown that the number of editing sites and the gene with the highest number of editing sites differ across species. For example, there are 455 RNA editing sites in 37 genes in \u003cem\u003ePrimulina hunanensis\u003c/em\u003e [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e], 460 RNA editing sites in 34 genes in \u003cem\u003eGlycyrrhiza glabra\u003c/em\u003e [\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e], and 441 RNA editing sites in 36 genes in \u003cem\u003eArabidopsis thaliana\u003c/em\u003e [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e]. In this study, we predicted the RNA editing sites of PCGs in the mitogenomes of three \u003cem\u003eTerminalia\u003c/em\u003e species and identified 311\u0026ndash;369 editing sites, all of which involve the conversion of (C) to (U). Further analysis indicates that these C-U editing events exhibited a significant preference for specific codon positions, primarily concentrated at the first and second codon positions [\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e]. This process enhances the interspecies homogeneity of the mitochondrial proteome and promotes adaptive development in plants. Most RNA editing events in the three \u003cem\u003eTerminalia\u003c/em\u003e species involve the conversion of amino acids to hydrophobic ones, which helps alter their physicochemical properties and enhance protein folding [\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e]. Therefore, widespread RNA editing leads to an increase in the number of hydrophobic amino acids, which may contribute to the translation of mRNA into polypeptides with the correct folded structure. This finding forms the structural basis for the specific functions of proteins [\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe Ka/Ks ratio can reveal the evolutionary pathways of genes and their underlying genetic mechanisms [\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e]. We conducted Ka/Ks analysis on 34 shared mtDNA protein-coding genes (PCGs) across three \u003cem\u003eTerminalia\u003c/em\u003e species. The results indicate signs of positive selection in \u003cem\u003erps1\u003c/em\u003e, \u003cem\u003erps4\u003c/em\u003e, \u003cem\u003erps7\u003c/em\u003e, and \u003cem\u003enad4\u003c/em\u003e. Specifically, \u003cem\u003erps1\u003c/em\u003e, \u003cem\u003erps4\u003c/em\u003e, and \u003cem\u003erps7\u003c/em\u003e may be associated with plant adaptation to environmental changes, while \u003cem\u003enad4\u003c/em\u003e is closely linked to mitochondrial energy metabolism. These findings likely reflect adaptive variation in redox reactions and environmental stress responses. As a valuable analytical tool, Ka/Ks analysis not only aids in understanding species adaptability and selective pressures but also offers new perspectives for assessing adaptive potential and studying genetic evolution [\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eA phylogenetic tree of mitochondrial and plastomes was constructed using maximum likelihood (ML) analysis. The results showed that the topology of the ML tree constructed with chloroplast PCGs was largely consistent with that of the ML tree constructed with mitochondrial PCGs. \u003cem\u003eT. chebula\u003c/em\u003e, \u003cem\u003eT. franchetii\u003c/em\u003e, and \u003cem\u003eT. intricata\u003c/em\u003e were all classified within \u003cem\u003eTerminalia\u003c/em\u003e, with \u003cem\u003eT. intricata\u003c/em\u003e being a variety of \u003cem\u003eT. franchetii\u003c/em\u003e, these two species showed a closer relationship. Due to the limited number of published mitogenomes for Myrtales, the mitogenomes tree encompasses a broader range of order-level taxa, reflecting evolutionary signals among distantly related groups. The overall structure of this mitochondrial DNA-based phylogeny is consistent with the latest classification of the Angiosperm Phylogeny Group (APG). In contrast, the plastomes tree focuses on interfamilial relationships within Myrtaceae. The organelle genomes of these three \u003cem\u003eTerminalia\u003c/em\u003e species will provide valuable and important genetic information for further improving the phylogenetic resolution of angiosperms.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eIn this study, we successfully assembled the mitogenomes and plastomes of three \u003cem\u003eTerminalia\u003c/em\u003e species, revealing unique branched conformations in the mitochondrial DNA structure. The results showed that the plastomes of the three \u003cem\u003eTerminalia\u003c/em\u003e species were similar in GC content and genome size. The ka/ks ratio analysis of the mitogenomes indicated that genes \u003cem\u003erps1\u003c/em\u003e, \u003cem\u003erps4\u003c/em\u003e, \u003cem\u003erps7\u003c/em\u003e, and \u003cem\u003enad4\u003c/em\u003e are under positive selection. The presence of gene transfer and conserved syntenic blocks between the three \u003cem\u003eTerminalia\u003c/em\u003e species reflects gene rearrangements and recombination, providing important genetic insights. RNA editing events revealed that most hydrophilic amino acids were converted to hydrophobic ones, enhancing the stability of protein structures. The construction of the phylogenetic tree deepens our understanding of the evolutionary relationships among \u003cem\u003eTerminalia\u003c/em\u003e species, particularly highlighting the close relationship between \u003cem\u003eT. franchetii\u003c/em\u003e and \u003cem\u003eT. intricata\u003c/em\u003e. Overall, our findings suggest that organellar genomic data can effectively reveal phylogenetic relationships between plants and can be applied to molecular marker development and genetic evolution studies.\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003ePlant materials, DNA extraction and sequencing\u003c/h2\u003e \u003cp\u003eFresh leaves of \u003cem\u003eT. chebula\u003c/em\u003e were collected from Longling County, Baoshan City, Yunnan Province, China (Sample location: 98\u0026deg;E, 24\u0026deg;N). This specimen was identified by Professor Li Guodong of Yunnan University of Chinese Medicine based on morphological characteristics. \u003cem\u003eT. franchetii\u003c/em\u003e was collected from Baima Town, Miyi County, Panzhihua City, Sichuan Province, China, with coordinates (Sample Location: 102.2\u0026deg;E, 27.0\u0026deg;N). \u003cem\u003eT. intricata\u003c/em\u003e was collected from Dongwang Township, Shangri-La City, Diqing Tibetan Autonomous Prefecture, Yunnan Province, China (Sample location: 99.5\u0026deg;E, 28.5\u0026deg;N). The specimen was identified by P.R. Luo based on morphological characteristics. The leaves were immediately frozen in liquid nitrogen, and total DNA was extracted using a modified CTAB method. The high-quality DNA samples that passed the test were sent to Novogen Biotechnology Co. for second- and third-generation sequencing, respectively.\u003c/p\u003e \u003cp\u003eSelected high-quality DNA samples that passed the test (main band\u0026thinsp;\u0026gt;\u0026thinsp;30kb); were randomly fragmented into 15-18kb segments using a G-Tube tube; enriched and purified using magnetic beads; damage repair and end repair was performed on the fragmented DNA; stem-loop sequencing adapters were lighted to both ends of the DNA fragments and exonucleases used to remove failed ligation fragments. The constructed library was then sequenced using the PacBio Revio/Sequel II/IIe platform.\u003c/p\u003e \u003cp\u003eThe raw sequencing reads contained a dumbbell-shaped structure with adapters at both ends, referred to as Polymerase reads. The raw data was fragmented at the adapter sites, and the adapter sequences filtered out to obtain subreads. The subreads were filtered according to the standard \"Filtering subreads by minimum length\u0026thinsp;=\u0026thinsp;50\". Based on the subreads, high-accuracy HiFi reads were generated using CCS software. All reads had a quality value greater than Q20.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003eOrganelle genome assembly and annotation\u003c/h2\u003e \u003cp\u003eThe plastomes of \u003cem\u003eT. franchetii\u003c/em\u003e and \u003cem\u003eT. intricata\u003c/em\u003e were \u003cem\u003ede novo\u003c/em\u003e assembled using GetOrganelle v1.7.5.3 [\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e]. Using PacBio HiFi sequencing data, the mitogenomes of three \u003cem\u003eTerminalia\u003c/em\u003e species and the plastomes of \u003cem\u003eT. chebula\u003c/em\u003e was assembled with PMAT v1.5.3 [\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e]. The assembly parameters were configured as follows: \u0026ldquo;-st HiFi -tp mt -ml 70 -mi 90\u0026rdquo;. The mitogenomes was visualized using Bandage v0.8.1 [\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e].\u003c/p\u003e \u003cp\u003eThe mitogenomes were annotated using IPMGA (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttp://www.1kmpg.cn/ipmga/\u003c/span\u003e\u003cspan address=\"http://www.1kmpg.cn/ipmga/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e), with RNA-seq data used to precisely correct 29 representative angiosperm mitogenomes sequences as the reference genome. Annotation errors for each mitogenome were manually corrected using Geneious Prime [\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e]. Additionally, the plastomes of \u003cem\u003eT. chebula, T. franchetii\u003c/em\u003e and \u003cem\u003eT. intricata\u003c/em\u003e were annotated using Geseq[\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e]. Finally, the organelle genome was visualised using OGDraw [\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003eAnalysis of Repeat Elements\u003c/h2\u003e \u003cp\u003eMISA (v2.1) (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://webblast.ipk-gatersleben.de/misa/\u003c/span\u003e\u003cspan address=\"https://webblast.ipk-gatersleben.de/misa/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e), TRF (v4.09) (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://tandem.bu.edu/trf/trf.unix.help.html\u003c/span\u003e\u003cspan address=\"https://tandem.bu.edu/trf/trf.unix.help.html\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e), and REPuter (\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://bibiserv.cebitec.uni-bielefeld.de/reputer/\u003c/span\u003e\u003cspan address=\"https://bibiserv.cebitec.uni-bielefeld.de/reputer/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e) were used to identify simple sequence repeats (SSRs), tandem repeats, and dispersed repeats, respectively. The results were visualized using Excel (2021) software and the Circos package (v0.69.9).\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003eAnalysis of Codon Usage\u003c/h2\u003e \u003cp\u003eThe protein-coding sequences of mitogenomes and plastomes were extracted with Phylosuite(v1.1.16) [\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e]. To analyze the codon bias of the protein-coding genes within the mitogenomes, MEGA v7.0 was used [\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e], and the Relative Synonymous Codon Usage (RSCU) values were subsequently calculated.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003ePrediction of RNA editing sites\u003c/h2\u003e \u003cp\u003eThe RNA editing sites were predicted via PREPACT3 [\u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e]. The analysis employed reference sequences from \u003cem\u003eVitisvinifera\u003c/em\u003e and \u003cem\u003eMillettia pinnata\u003c/em\u003e. The predicted RNA editing sites were subsequently visualized using R (4.4.0) to facilitate interpretation and analysis of the results.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec18\" class=\"Section2\"\u003e \u003ch2\u003eHomologous sequences identification\u003c/h2\u003e \u003cp\u003eHomogeneous sequences between the plastomes and mitogenomes of three \u003cem\u003eTerminalia\u003c/em\u003e were identified using BLAST (v2.16.0) [\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e], with parameters set to an identity greater than 90% and an e-value of less than 1e\u0026thinsp;\u0026minus;\u0026thinsp;5. We then analyzed the homologous sequence regions to determine the length, quantity, and gene type of the mitochondrial plastomes sequences (MTPTs). the results were visualized using the Circos package (v0.69.9) [\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec19\" class=\"Section2\"\u003e \u003ch2\u003eSubstitution rates of PCGs\u003c/h2\u003e \u003cp\u003eFirst, the mitogenomes of three \u003cem\u003eTerminalia\u003c/em\u003e species were aligned using MAFFT (v7.427) [\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e]. Subsequently, the Ka/Ks ratio was calculated for each gene pair using the YN method in KaKs_Calculator (v2.0) [\u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e]. The aggregated Ka/Ks ratios were visualized as box plots generated with the ggplot2 package in R.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec20\" class=\"Section2\"\u003e \u003ch2\u003eNucleotide diversity\u003c/h2\u003e \u003cp\u003eSequences of protein-coding genes (PCGs) from organelle genomes were aligned using MAFFT (v7.477) [\u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e60\u003c/span\u003e], while the Pi value for each gene was calculated using DnaSP (v5.0) [\u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec21\" class=\"Section2\"\u003e \u003ch2\u003eAnalysis of Plastomes Boundaries\u003c/h2\u003e \u003cp\u003eBased on annotations, the plastome boundaries between the IR, LSC, and SSC regions were analyzed, and a schematic diagram was constructed using Irscope [\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e].\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec22\" class=\"Section2\"\u003e \u003ch2\u003eCollinearity Analysis and Phylogenetic Analysis\u003c/h2\u003e \u003cp\u003eMauve was employed to analyze homology and sequence rearrangements within the mitogenomes of three \u003cem\u003eTerminalia\u003c/em\u003e species [\u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e]. All conserved PCGs from mitogenomes and plastomes were extracted using Phylosuite [\u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e64\u003c/span\u003e] and aligned with MAFFT (v7.427) [\u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e]. The results were then imported into MACSE for optimization. The optimal model was selected via ModelFinder (v2.2.2) [\u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e66\u003c/span\u003e]. Bayesian analysis was performed on the PCGs concatenated data matrix using MrBayes (v.3.2) [\u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e67\u003c/span\u003e]. The final phylogenetic tree was constructed in IQ-tree and visualized with iTOL [\u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e68\u003c/span\u003e] for aesthetic refinement.\u003c/p\u003e \u003c/div\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eNCBI \u0026nbsp; \u0026nbsp; \u0026nbsp; National Center for Biotechnology Information.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003ePCGs \u0026nbsp; \u0026nbsp; \u0026nbsp; Protein-coding genes.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eRSCU \u0026nbsp; \u0026nbsp; \u0026nbsp;Relative synonymous codon usage.\u003c/p\u003e\n\u003cp\u003eSSRs \u0026nbsp; \u0026nbsp; \u0026nbsp; Simple sequence repeats.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003ecpDNA \u0026nbsp; \u0026nbsp; Plastomes.\u003c/p\u003e\n\u003cp\u003emtDNA \u0026nbsp; \u0026nbsp; Mitogenomes.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eMTPTs \u0026nbsp; \u0026nbsp; Mitochondrial plastid DNAs.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eKa \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; Nonsynonymous substitutions.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eKs \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; Synonymous substitutions.\u003c/p\u003e\n\u003cp\u003ePi \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp; \u0026nbsp;Nucleotide Diversity\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAuthorship contribution statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eXingyu Liu: Data curation, Visualization, Writing \u0026ndash; original draft\u0026amp; editing. Pengrui Luo: Data curation, Writing \u0026ndash; review \u0026amp; editing, Resources, Validation.Yingmei Zou: Data curation, Writing \u0026ndash; original draft, Validation. M. James C. Crabbe: Data curation, Visualization, Writing \u0026ndash; original draft. Guodong Li: Conceptualization, Supervision, Writing \u0026ndash; review \u0026amp; editing, Funding acquisition. Ticao Zhang: Conceptualization, Resources, Supervision, Writing \u0026ndash; review \u0026amp; editing, Funding acquisition.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis work is supported by grants from National Key R\u0026amp;D Program of China (2024YFF1306700), Strategic Priority Research Program of the Chinese Academy of Sciences (XDB1230000), National Natural Science Foundation of China (82260739, 32570434), Yunnan Fundamental Research Projects (202501AS070177 and 202101AZ070001-005).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable. This study did not use animal or human materials. Our experimental research, including the collection of plant materials, complies with institutional, national, or international guidelines. The voucher specimen of \u003cem\u003eT. chebula\u003c/em\u003e is deposited in the Herbarium of Yunnan University of Chinese Medicine, voucher numbers 2503. The voucher specimens of \u003cem\u003eT. franchetii\u003c/em\u003e and \u003cem\u003eT. intricata\u0026nbsp;\u003c/em\u003eare deposited in the Kunming Institute of Botany Chinese Academy of Sciences. Voucher numbers 1651612 (\u003cem\u003eT. franchetii\u003c/em\u003e) and 1651613 (\u003cem\u003eT. intricata\u003c/em\u003e).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDeclaration of Competing Interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe declare that we have no conflicts of interest.\u003c/p\u003e\u003cp\u003e\u003cstrong\u003eData availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe dataset generated and analyzed by this graduate student has been submitted to the NCBI database. The current provisional accession number is SUB15777329. The final permanent accession number will be provided prior to the publication of the thesis.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eDas G, Kim D-Y, Fan C, Guti\u0026eacute;rrez-Grijalva EP, Heredia JB, Nissapatorn V, et al. Plants of the Genus \u003cem\u003eTerminalia\u003c/em\u003e: An Insight on Its Biological Potentials, Pre-Clinical and Clinical Studies. Front Pharmacol. 2020;11:561248.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKim HJ, Song H-K, Park SH, Jang S, Park K-S, Song KH, et al. \u003cem\u003eTerminalia chebula\u003c/em\u003e Retz. extract ameliorates the symptoms of atopic dermatitis by regulating anti-inflammatory factors in vivo and suppressing STAT1/3 and NF-ĸB signaling in vitro. Phytomedicine Int J Phytother Phytopharm. 2022;104:154318.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRamesh P, Palaniappan A. \u003cem\u003eTerminalia arjuna\u003c/em\u003e, a Cardioprotective Herbal Medicine-Relevancy in the Modern Era of Pharmaceuticals and Green Nanomedicine-A Review. Pharm Basel Switz. 2023;16:126.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGupta A, Kumar R, Bhattacharyya P, Bishayee A, Pandey AK. \u003cem\u003eTerminalia bellirica\u003c/em\u003e (Gaertn.) roxb. (Bahera) in health and disease: A systematic and comprehensive review. Phytomedicine Int J Phytother Phytopharm. 2020;77:153278.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMahendra RVNASR. Antimicrobial and anti-inflammatory activity of \u003cem\u003eTerminalia arjuna\u003c/em\u003e. Bioinformation. 2023;19:184\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCheesman MJ, Shivashekaregowda NKH, Cock IE. Bacterial Foodborne Illness in Malaysia: \u003cem\u003eTerminalia\u003c/em\u003e spp. as a Potential Resource for Treating Infections and Countering Antibiotic Resistance. Malays J Med Sci MJMS. 2023;30:42\u0026ndash;54.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLiu H, Chen Z, Liu M, Li E, Shen J, Wang J, et al. The \u003cem\u003eTerminalia chebula\u003c/em\u003e Retz extract treats hyperuricemic nephropathy by inhibiting TLR4/MyD88/NF-κB axis. J Ethnopharmacol. 2024;322:117678.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHassan Bulbul MR, Uddin Chowdhury MN, Naima TA, Sami SA, Imtiaj MS, Huda N, et al. A comprehensive review on the diverse pharmacological perspectives of \u003cem\u003eTerminalia chebula\u003c/em\u003e Retz. Heliyon. 2022;8:e10220.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang C, Zhang H, Wang X, Wang X, Li X, Li C, et al. Comprehensive Review on Fruit of \u003cem\u003eTerminalia chebula\u003c/em\u003e: Traditional Uses, Phytochemistry, Pharmacology, Toxicity, and Pharmacokinetics. Mol Basel Switz. 2024;29:5547.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZai MJ, Cheesman MJ, Cock IE. \u003cem\u003eTerminalia petiolaris\u003c/em\u003e A.Cunn ex Benth. Extracts Have Antibacterial Activity and Potentiate Conventional Antibiotics against β-Lactam-Drug-Resistant Bacteria. Antibiot Basel Switz. 2023;12:1643.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGao H, Lu H, Fang N, Su J, Li R, Wang W, et al. The potential of \u003cem\u003eTerminalia chebula\u003c/em\u003e in alleviating mild cognitive impairment: a review. Front Pharmacol. 2024;15:1484040.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCock IE. The medicinal properties and phytochemistry of plants of the genus \u003cem\u003eTerminalia\u003c/em\u003e (Combretaceae). Inflammopharmacology. 2015;23:203\u0026ndash;29.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKumar V, Sharma N, Saini R, Mall S, Zengin G, Sourirajan A, et al. Therapeutic potential and industrial applications of \u003cem\u003eTerminalia arjuna\u003c/em\u003e bark. J Ethnopharmacol. 2023;310:116352.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGray MW. The endosymbiont hypothesis revisited. Int Rev Cytol. 1992;141:233\u0026ndash;357.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eShtolz N, Mishmar D. The Mitochondrial Genome\u0026ndash;on Selective Constraints and Signatures at the Organism, Cell, and Single Mitochondrion Levels. Front Ecol Evol. 2019;7.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTian S, Lu P, Zhang Z, Wu JQ, Zhang H, Shen H. Chloroplast genome sequence of Chongming lima bean (\u003cem\u003ePhaseolus lunatus\u003c/em\u003e L.) and comparative analyses with other legume chloroplast genomes. BMC Genomics. 2021;22:194.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYanfei N, Tai S, Chunhua W, Jia D, Fazhong Y. Complete chloroplast genome sequences of the medicinal plant \u003cem\u003eAconitum transsectum\u003c/em\u003e (Ranunculaceae): comparative analysis and phylogenetic relationships. BMC Genomics. 2023;24:90.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSloan DB. One ring to rule them all? Genome sequencing provides new insights into the master circle model of plant mitochondrial DNA structure. New Phytol. 2013;200:978\u0026ndash;85.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGualberto JM, Mileshina D, Wallet C, Niazi AK, Weber-Lotfi F, Dietrich A. The plant mitochondrial genome: dynamics and maintenance. Biochimie. 2014;100:107\u0026ndash;20.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJackman SD, Coombe L, Warren RL, Kirk H, Trinh E, MacLeod T, et al. Complete Mitochondrial Genome of a Gymnosperm, Sitka Spruce (\u003cem\u003ePicea sitchensis\u003c/em\u003e), Indicates a Complex Physical Structure. Genome Biol Evol. 2020;12:1174\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNass MM, Mitochondrial DNA. Advances, Problems, and Goals. Science. 1969;165:25\u0026ndash;35.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLiberatore KL, Dukowic-Schulze S, Miller ME, Chen C, Kianian SF. The role of mitochondria in plant development and stress tolerance. Free Radic Biol Med. 2016;100:238\u0026ndash;56.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBrennicke A, Marchfelder A, Binder S. RNA editing. FEMS Microbiol Rev. 1999;23:297\u0026ndash;316.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCovello PS, Gray MW. RNA editing in plant mitochondria. Nature. 1989;341:662\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYang H, Ni Y, Zhang X, Li J, Chen H, Liu C. The mitochondrial genomes of \u003cem\u003ePanax notoginseng\u003c/em\u003e reveal recombination mediated by repeats associated with DNA replication. Int J Biol Macromol. 2023;252:126359.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChen Z, Nie H, Grover CE, Wang Y, Li P, Wang M, et al. Entire nucleotide sequences of \u003cem\u003eGossypium raimondii\u003c/em\u003e and \u003cem\u003eG.arboreum\u003c/em\u003e mitochondrial genomes revealed A-genome species as cytoplasmic donor of the allotetraploid species. Plant Biol Stuttg Ger. 2017;19:484\u0026ndash;93.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eFeng L, Wang Z, Wang C, Yang X, An M, Yin Y. Multichromosomal mitochondrial genome of \u003cem\u003ePunica granatum\u003c/em\u003e: comparative evolutionary analysis and gene transformation from chloroplast genomes. BMC Plant Biol. 2023;23:512.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang H, Wu Z, Li T, Zhao J. Highly active repeat-mediated recombination in the mitogenome of the aquatic grass \u003cem\u003eHygroryza aristata\u003c/em\u003e. BMC Plant Biol. 2024;24:644.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAnderson S, Bankier AT, Barrell BG, de Bruijn MH, Coulson AR, Drouin J, et al. Sequence and organization of the human mitochondrial genome. Nature. 1981;290:457\u0026ndash;65.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eClary DO, Wolstenholme DR. The Drosophila mitochondrial genome. Oxf Surv Eukaryot Genes. 1984;1:1\u0026ndash;35.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLiu D, Guo H, Zhu J, Qu K, Chen Y, Guo Y, et al. Complex Physical Structure of Complete Mitochondrial Genome of \u003cem\u003eQuercus acutissima\u003c/em\u003e (Fagaceae): A Significant Energy Plant. Genes. 2022;13:1321.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLi H, Liu L, Qiu Z, He F, Dong W. Complete mitochondrial genome assembly and comparative analysis of \u003cem\u003eColocasia esculenta\u003c/em\u003e. BMC Plant Biol. 2025;25:67.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGao C, Wang S, Huang Y, Deng Y. Assembly and comparative analysis of the complete mitochondrial genome of \u003cem\u003eEchinacanthus longipes\u003c/em\u003e (Acanthaceae), endemic to the Sino-Vietnamese karst flora. BMC Genomics. 2025;26:251.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWu Z-Q, Liao X-Z, Zhang X-N, Tembrock LR, Broz A. Genomic architectural variation of plant mitochondria\u0026mdash;A review of multichromosomal structuring. J Syst Evol. 2022;60:160\u0026ndash;8.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhang Y, Li H-L, Zhong J-D, Wang Y, Yuan C-C. Chloroplast Genome Sequences and Comparative Analyses of Combretaceae Mangroves with Related Species. BioMed Res Int. 2020;2020:5867673.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang Y, Sun N, Shi W, Ma Q, Sun L, Hao M, et al. Assembly and Comparative Analysis of the Complete Mitochondrial Genome of \u003cem\u003eIlex macrocarpa\u003c/em\u003e. Forests. 2023;14:2372.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang L, Liu X, Xu Y, Zhang Z, Wei Y, Hu Y, et al. Assembly and comparative analysis of the first complete mitochondrial genome of a traditional Chinese medicine Angelica biserrata (Shan et Yuan) Yuan et Shan. Int J Biol Macromol. 2024;257:128571.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eQu Y, Zhou P, Tong C, Bi C, Xu L. Assembly and analysis of the Populus deltoides mitochondrial genome: the first report of a multicircular mitochondrial conformation for the genus Populus. J Res. 2023;34:717\u0026ndash;33.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eChen L, Dong X, Huang H, Xu H, Rono PC, Cai X, et al. Assembly and comparative analysis of the initial complete mitochondrial genome of \u003cem\u003ePrimulina hunanensis\u003c/em\u003e (Gesneriaceae): a cave-dwelling endangered plant. BMC Genomics. 2024;25:322.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhou G, Qin M, Liu X, Qi Y, Ou X, Tang M. De novo assembly of the mitochondrial genome of \u003cem\u003eGlycyrrhiza glabra\u003c/em\u003e and identification of two types of homologous recombination configurations caused by repeat sequences. BMC Genomics. 2025;26:13.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eUnseld M, Marienfeld JR, Brandt P, Brennicke A. The mitochondrial genome of \u003cem\u003eArabidopsis thaliana\u003c/em\u003e contains 57 genes in 366,924 nucleotides. Nat Genet. 1997;15:57\u0026ndash;61.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhang Y, Shen Z, Meng X, Zhang L, Liu Z, Liu M, et al. Codon usage patterns across seven Rosales species. BMC Plant Biol. 2022;22:65.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLloyd Evans D, Hlongwane TT, Joshi SV, Ria\u0026ntilde;o Pach\u0026oacute;n DM. The sugarcane mitochondrial genome: assembly, phylogenetics and transcriptomics. PeerJ. 2019;7:e7558.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYura K, Go M. Correlation between amino acid residues converted by RNA editing and functional residues in protein three-dimensional structures in plant organelles. BMC Plant Biol. 2008;8:79.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang L, Liu X, Xu Y, Zhang Z, Wei Y, Hu Y, et al. Assembly and comparative analysis of the first complete mitochondrial genome of a traditional Chinese medicine \u003cem\u003eAngelica biserrata\u003c/em\u003e (Shan et Yuan) Yuan et Shan. Int J Biol Macromol. 2024;257:128571.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eXie D-F, Yu H-X, Price M, Xie C, Deng Y-Q, Chen J-P et al. Phylogeny of Chinese Allium Species in Section Daghestanica and Adaptive Evolution of Allium (Amaryllidaceae, Allioideae) Species Revealed by the Chloroplast Complete Genome. Front Plant Sci. 2019;10.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJin J-J, Yu W-B, Yang J-B, Song Y, dePamphilis CW, Yi T-S, et al. GetOrganelle: a fast and versatile toolkit for accurate de novo assembly of organelle genomes. Genome Biol. 2020;21:241.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBi C, Shen F, Han F, Qu Y, Hou J, Xu K, et al. PMAT: an efficient plant mitogenome assembly toolkit using low-coverage HiFi sequencing data. Hortic Res. 2024;11:uhae023.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWick RR, Schultz MB, Zobel J, Holt KE. Bandage: interactive visualization of de novo genome assemblies. Bioinforma Oxf Engl. 2015;31:3350\u0026ndash;2.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKearse M, Moir R, Wilson A, Stones-Havas S, Cheung M, Sturrock S, et al. Geneious Basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinforma Oxf Engl. 2012;28:1647\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTillich M, Lehwark P, Pellizzer T, Ulbricht-Jones ES, Fischer A, Bock R, et al. GeSeq - versatile and accurate annotation of organelle genomes. Nucleic Acids Res. 2017;45:W6\u0026ndash;11.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGreiner S, Lehwark P, Bock R. OrganellarGenomeDRAW (OGDRAW) version 1.3.1: expanded toolkit for the graphical visualization of organellar genomes. Nucleic Acids Res. 2019;47:W59\u0026ndash;64.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhang D, Gao F, Jakovlić I, Zou H, Zhang J, Li WX, et al. PhyloSuite: An integrated and scalable desktop platform for streamlined molecular sequence data management and evolutionary phylogenetics studies. Mol Ecol Resour. 2020;20:348\u0026ndash;55.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKumar S, Stecher G, Tamura K. MEGA7: Molecular Evolutionary Genetics Analysis Version 7.0 for Bigger Datasets. Mol Biol Evol. 2016;33:1870\u0026ndash;4.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLenz H, Hein A, Knoop V. Plant organelle RNA editing and its specificity factors: enhancements of analyses and new database features in PREPACT 3.0. BMC Bioinformatics. 2018;19:255.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCamacho C, Coulouris G, Avagyan V, Ma N, Papadopoulos J, Bealer K, et al. BLAST+: architecture and applications. BMC Bioinformatics. 2009;10:421. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://doi.org/10.1186/1471-2105-10-421\u003c/span\u003e\u003cspan address=\"10.1186/1471-2105-10-421\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKrzywinski M, Schein J, Birol İ, Connors J, Gascoyne R, Horsman D, et al. Circos: An information aesthetic for comparative genomics. Genome Res. 2009;19:1639\u0026ndash;45.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKatoh K, Standley DM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 2013;30:772\u0026ndash;80.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWang D, Zhang Y, Zhang Z, Zhu J, Yu J. KaKs_Calculator 2.0: a toolkit incorporating gamma-series methods and sliding window strategies. Genomics Proteom Bioinf. 2010;8:77\u0026ndash;80.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKatoh K, Standley DM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 2013;30:772\u0026ndash;80.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRozas J, Ferrer-Mata A, S\u0026aacute;nchez-DelBarrio JC, Guirao-Rico S, Librado P, Ramos-Onsins SE, et al. DnaSP 6: DNA Sequence Polymorphism Analysis of Large Data Sets. Mol Biol Evol. 2017;34:3299\u0026ndash;302.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAmiryousefi A, Hyv\u0026ouml;nen J, Poczai P. IRscope: an online program to visualize the junction sites of chloroplast genomes. Bioinforma Oxf Engl. 2018;34:3030\u0026ndash;1.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eDarling ACE, Mau B, Blattner FR, Perna NT. Mauve: multiple alignment of conserved genomic sequence with rearrangements. Genome Res. 2004;14:1394\u0026ndash;403.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eZhang D, Gao F, Jakovlić I, Zou H, Zhang J, Li WX, et al. PhyloSuite: An integrated and scalable desktop platform for streamlined molecular sequence data management and evolutionary phylogenetics studies. Mol Ecol Resour. 2020;20:348\u0026ndash;55.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKatoh K, Standley DM. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol Biol Evol. 2013;30:772\u0026ndash;80.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKalyaanamoorthy S, Minh BQ, Wong TKF, von Haeseler A, Jermiin LS. ModelFinder: fast model selection for accurate phylogenetic estimates. Nat Methods. 2017;14:587\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRonquist F, Teslenko M, van der Mark P, Ayres DL, Darling A, H\u0026ouml;hna S et al. MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Syst Biol.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLetunic I, Bork P. Interactive Tree Of Life (iTOL) v4: recent updates and new developments. Nucleic Acids Res. 2019;47:W256\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"bmc-genomics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"gics","sideBox":"Learn more about [BMC Genomics](http://bmcgenomics.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/gics","title":"BMC Genomics","twitterHandle":"#BMCGenomics","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"Terminalia, plastomes, mitogenomes, organelle genome, comparative genomics","lastPublishedDoi":"10.21203/rs.3.rs-8163245/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8163245/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground\u003c/h2\u003e \u003cp\u003ePlant organelle genomes are critical for evolutionary biology research, yet their characteristics in the \u003cem\u003eTerminalia\u003c/em\u003e genus (Combretaceae, ~\u0026thinsp;200 tropical species of which some possess medicinal and edible value) remain unstudied. This study systematically analyzes the mitochondrial genomes (mitogenomes) and chloroplast genomes (plastomes) of three \u003cem\u003eTerminalia\u003c/em\u003e species (\u003cem\u003eT. chebula\u003c/em\u003e, \u003cem\u003eT. franchetii\u003c/em\u003e, \u003cem\u003eT. intricata\u003c/em\u003e).\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e \u003cp\u003eUsing high-throughput sequencing, we successfully obtained sequences of plastomes and mitogenomes for three plants, enabling comparative and evolutionary genomic analysis. Plastomes of the three species shared a typical quadripartite structure, with similar sizes (159,700\u0026ndash;159,971 bp) and gene compositions (103\u0026ndash;107 genes), but differed in specific regions. In contrast, mitogenomes showed significant structural size variation (350,904\u0026ndash;365,711 bp), with \u003cem\u003eT. chebula\u003c/em\u003e and \u003cem\u003eT. intricata\u003c/em\u003e having three circular and one linear molecule, and \u003cem\u003eT. franchetii\u003c/em\u003e has two linear molecules, reflecting extensive recombination. Ka/Ks analysis indicated positive selection on mitogenome genes (e.g., \u003cem\u003erps1\u003c/em\u003e, \u003cem\u003enad4\u003c/em\u003e) linked to adaptation. RNA editing (all C-to-U) enhanced protein hydrophobicity. Phylogenetic trees clarified \u003cem\u003eTerminalia\u003c/em\u003e\u0026rsquo;s position.\u003c/p\u003e\u003ch2\u003eConclusions\u003c/h2\u003e \u003cp\u003eThis study provides crucial genomic data for phylogenetic research and molecular ecology within the \u003cem\u003eTerminalia\u003c/em\u003e, holding significant value for future research.\u003c/p\u003e","manuscriptTitle":"Comparative analysis of organelle genomes in three Terminalia species reveals structure evolution and phylogenetic position","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-01-13 18:46:06","doi":"10.21203/rs.3.rs-8163245/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2026-03-06T09:48:54+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-05T06:32:51+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"155949604353084860893416376833438490713","date":"2026-02-26T04:47:34+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-02-16T06:39:53+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"48860737158918288015684371841962822227","date":"2026-01-26T05:57:55+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-01-09T11:14:35+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-12-03T00:51:51+00:00","index":"","fulltext":""},{"type":"editorInvited","content":"","date":"2025-12-01T18:46:14+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-11-28T08:56:26+00:00","index":"","fulltext":""},{"type":"submitted","content":"BMC Genomics","date":"2025-11-28T08:48:16+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"bmc-genomics","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"gics","sideBox":"Learn more about [BMC Genomics](http://bmcgenomics.biomedcentral.com/)","snPcode":"","submissionUrl":"https://www.editorialmanager.com/gics","title":"BMC Genomics","twitterHandle":"#BMCGenomics","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"em","reportingPortfolio":"BMC Series","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"ed171f4e-0b80-42c6-91b1-2e294afb71cf","owner":[],"postedDate":"January 13th, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-04-22T08:38:17+00:00","versionOfRecord":[],"versionCreatedAt":"2026-01-13 18:46:06","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8163245","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8163245","identity":"rs-8163245","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}