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Orchidaceae is one of the largest families of angiosperms. Considering the large number of species in this family and its symbiotic relationship with fungi, Orchidaceae provide an ideal model to study the evolution of plant mitogenomes. However, to date, there is only one draft mitochondrial genome of this family available. Here, we present a fully assembled and annotated sequence of the mitochondrial genome (mitogenome) of Paphiopedilum micranthum, a species with high economic and ornamental value. The mitogenome of P. micranthum was 447,368 bp in length and comprised 26 circular subgenomes ranging in size from 5973 bp to 32,281 bp. The genome encoded for 39 mitochondrial-origin, protein-coding genes; 16 tRNAs (three of plastome origin); three rRNAs; and 16 ORFs, while rpl10 and sdh3 were lost from the mitogenome. Moreover, interorganellar DNA transfer was identified in 14 of the 26 chromosomes. These plastid-derived DNA fragments represented 28.32% (46,273 bp) of the P. micranthum plastome, including 12 intact plastome origin genes. Remarkably, the mitogenome of P. micranthum and Gastrodia elata shared 18% (about 81 kb) of their mitochondrial DNA sequences. Additionally, we found a positive correlation between repeat length and recombination frequency. The mitogenome of P. micranthum had more compact and fragmented chromosomes compared to other species with multichromosomal structures. We suggest that repeat-mediated homologous recombination enables the dynamic structure of mitochondrial genomes in Orchidaceae.

Epipogium roseum, commonly known as one of the ghost orchids due to its rarity and almost transparent color, is a non-photosynthetic and fully mycoheterotrophic plant. Given its special nutritional strategies and evolutionary significance, the mitogenome was first characterized, and three plastomes sampled from Asia were assembled. The plastomes were found to be the smallest among Orchidaceae, with lengths ranging from 18,339 to 19,047 bp, and exhibited high sequence variety. For the mitogenome, a total of 414,552 bp in length, comprising 26 circular chromosomes, were identified. A total of 54 genes, including 38 protein-coding genes, 13 tRNA genes, and 3 rRNA genes, were annotated. Multiple repeat sequences spanning a length of 203,423 bp (45.47%) were discovered. Intriguingly, six plastid regions via intracellular gene transfer and four plastid regions via horizontal gene transfer to the mitogenome were observed. The phylogenomics, incorporating 90 plastomes and 56 mitogenomes, consistently revealed the sister relationship of Epipogium and Gastrodia, with a bootstrap percentage of 100%. These findings shed light on the organelle evolution of Orchidaceae and non-photosynthetic plants.

Across eukaryotes, mitochondria exhibit staggering diversity in genomic architecture, including the repeated evolution of multichromosomal structures. Unlike in the nucleus, where mitosis and meiosis ensure faithful transmission of chromosomes, the mechanisms of inheritance in fragmented mitochondrial genomes remain mysterious. Multichromosomal mitochondrial genomes have recently been found in multiple species of flowering plants, including Silene noctiflora , which harbors an unusually large and complex mitochondrial genome with more than 50 circular-mapping chromosomes totaling ∼7 Mb in size. To determine the extent to which such genomes are stably maintained, we analyzed intraspecific variation in the mitochondrial genome of S. noctiflora . Complete genomes from two populations revealed a high degree of similarity in the sequence, structure, and relative abundance of mitochondrial chromosomes. For example, there are no inversions between the genomes, and there are only nine SNPs in 25 kb of protein-coding sequence. Remarkably, however, these genomes differ in the presence or absence of 19 entire chromosomes, all of which lack any identifiable genes or contain only duplicate gene copies. Thus, these mitochondrial genomes retain a full gene complement but carry a highly variable set of chromosomes that are filled with presumably dispensable sequence. In S. noctiflora , conventional mechanisms of mitochondrial sequence divergence are being outstripped by an apparently nonadaptive process of whole-chromosome gain/loss, highlighting the inherent challenge in maintaining a fragmented genome. We discuss the implications of these findings in relation to the question of why mitochondria, more so than plastids and bacterial endosymbionts, are prone to the repeated evolution of multichromosomal genomes.

Mitochondrial genomes (mitogenomes) in higher plants can induce cytoplasmic male sterility and be somehow involved in nuclear-cytoplasmic interactions affecting plant growth and agronomic performance. They are larger and more complex than in other eukaryotes, due to their recombinogenic nature. For most plants, the mitochondrial DNA (mtDNA) can be represented as a single circular chromosome, the so-called master molecule, which includes repeated sequences that recombine frequently, generating sub-genomic molecules in various proportions. Based on the relevance of the potato crop worldwide, herewith we report the complete mtDNA sequence of two S. tuberosum cultivars, namely Cicero and Désirée, and a comprehensive study of its expression, based on high-coverage RNA sequencing data. We found that the potato mitogenome has a multi-partite architecture, divided in at least three independent molecules that according to our data should behave as autonomous chromosomes. Inter-cultivar variability was null, while comparative analyses with other species of the Solanaceae family allowed the investigation of the evolutionary history of their mitogenomes. The RNA-seq data revealed peculiarities in transcriptional and post-transcriptional processing of mRNAs. These included co-transcription of genes with open reading frames that are probably expressed, methylation of an rRNA at a position that should impact translation efficiency and extensive RNA editing, with a high proportion of partial editing implying frequent mis-targeting by the editing machinery.

Background Orchidaceae is one of the largest groups of angiosperms, and most species have high economic value and scientific research value due to their ornamental and medicinal properties. In China, Chinese Cymbidium is a popular ornamental orchid with high economic value and a long history. However, to date, no detailed information on the mitochondrial genome of any species of Chinese Cymbidium has been published. Results Here, we present the complete assembly and annotation of the mitochondrial genome of Cymbidium ensifolium (L.) Sw. The mitogenome of C. ensifolium was 560,647 bp in length and consisted of 19 circular subgenomes ranging in size from 21,995 bp to 48,212 bp. The genome encoded 35 protein-coding genes, 36 tRNAs, 3 rRNAs, and 3405 ORFs. Repeat sequence analysis and prediction of RNA editing sites revealed a total of 915 dispersed repeats, 162 simple repeats, 45 tandem repeats, and 530 RNA editing sites. Analysis of codon usage showed a preference for codons ending in A/T. Interorganellar DNA transfer was identified in 13 of the 19 chromosomes, with plastid-derived DNA fragments representing 6.81% of the C. ensifolium mitochondrial genome. The homologous fragments of the mitochondrial genome and nuclear genome were also analysed. Comparative analysis showed that the GC content was conserved, but the size, structure, and gene content of the mitogenomes varied greatly among plants with multichromosomal mitogenome structure. Phylogenetic analysis based on the mitogenomes reflected the evolutionary and taxonomic statuses of C. ensifolium . Interestingly, compared with the mitogenomes of Cymbidium lancifolium Hook. and Cymbidium macrorhizon Lindl., the mitogenome of C. ensifolium lost 8 ribosomal protein-coding genes. Conclusion In this study, we assembled and annotated the mitogenome of C. ensifolium and compared it with the mitogenomes of other Liliidae and plants with multichromosomal mitogenome structures. Our findings enrich the mitochondrial genome database of orchid plants and reveal the rapid structural evolution of Cymbidium mitochondrial genomes, highlighting the potential for mitochondrial genes to help decipher plant evolutionary history.

Gentiana rigescens, an alpine plant with significant medicinal value, possesses a complex genetic background. However, comprehensive genomic research on G. rigescens is still lacking, particularly concerning its organelle genome. In this study, G. rigescens was studied to sequence the mitochondrial genome (mitogenome) and ascertain the assembly, informational content, and developmental expression of the mitogenome. The mitogenome of G. rigescens was 393,595 bp in length and comprised four circular chromosomes ranging in size from 6646 bp to 362,358 bp. The GC content was 43.73%. The mitogenome featured 30 distinct protein-coding genes, 26 tRNA genes, and 3 rRNA genes. The mitogenome of G. rigescens also revealed 70 SSRs, which were mostly tetra-nucleotides. In addition, 48 homologous fragments were found between the mitogenome and the chloroplast genome, with the longest measuring 23,330 bp. The documentation of the mitochondrial genome of G. rigescens is instrumental in advancing the understanding of its physiological development. Decoding the G. rigescens mitogenome will offer valuable genetic material for phylogenetic research on Gentianaceae and enhance the use of species germplasm resources.

is a perennial herb valued for both its ornamental and medicinal properties. Despite its significance, no comprehensive analysis of its mitochondrial genome has been previously reported. Plant mitochondrial genomes are known for their large size, structural complexity, and frequent recombination events. This study aims to provide the first complete assembly and characterization of the mitochondrial genome of , with a focus on its structure, gene content, repeat elements, and RNA editing sites. We used GetOrganelle and Unicycler software to hybrid assemble Nanopore and Illumina data to obtain the mitochondrial genome of . The mitochondrial genome of consists of four contigs: contig1 (222,498 bp), contig2 (90,780 bp), contig3 (42,563 bp), and contig4 (39,247 bp). Two repeat sequences, R1 (7,784 bp) and R2 (3,519 bp), facilitate the conformation of three circular chromosomes, suggesting a complex multi-chromosomal structure. A total of 34 protein-coding genes, including 24 core genes and 10 non-core genes were identified. Analysis of tandem repeat elements revealed significant variability, with Chromosome 1 showing the highest diversity of SSRs and scattered repeats. Additionally, 20 homologous fragments were identified between the mitochondrial and chloroplast genomes, accounting for 1.10% of the mitochondrial genome. Phylogenetic analysis based on 24 conserved mitochondrial genes placed in close relation to and . Furthermore, 545 RNA editing sites were identified, with notable variations across genes, suggesting that RNA editing plays a significant role in regulating mitochondrial gene expression. The complete assembly of the mitochondrial genome reveals a complex multichromosomal structure characterized by recombination events. The high number of RNA editing sites and the presence of transferred plastid DNA highlight the dynamic nature of the genome, contributing to its adaptability and evolution. These findings provide a genetic foundation into the plant's medicinal properties, adaptive mechanisms, and potential for environmental resilience.

This study reports for the first time the complete sequence characteristics of the mitochondrial genome of the tea plant cultivar Camellia sinensis cv. 'Baihaozao'. It systematically unveils its multi-chromosomal structure, RNA editing patterns, and adaptive evolutionary mechanisms, providing critical theoretical insights into the structural complexity and evolutionary mechanisms of the tea plant mitochondrial genome. The mitochondrial genome was fully analyzed using genome sequencing and annotation techniques. RNA editing sites were predicted to evaluate editing patterns. Codon usage bias analysis was conducted to identify high-frequency codons. Repeat sequence analysis was used to characterize dispersed and tandem repeats. Adaptive evolutionary analysis, based on Ka/Ks ratios, was performed to investigate gene selection pressures. The mitochondrial genome consists of 11 linear chromosomes, with a total length of 909,843 bp and a GC content of 45.62%. A total of 73 functional genes were annotated, among which 14 variable genes (e.g., ribosomal protein coding genes) retain intact functions without pseudogenization, which is rare among Theaceae plants. RNA editing site prediction revealed significant spatial heterogeneity, with the cox1 gene being a hotspot containing 19 editing sites. Approximately 58.49% of editing events were concentrated on the second base of codons, and 48.61% of the sites resulted in amino acid changes from hydrophilic to hydrophobic. Codon usage bias analysis showed significant enrichment of high-frequency codons, including UUU (phenylalanine), AUU (isoleucine), and UUC (phenylalanine). The genome's repeat sequences were predominantly dispersed repeats (70.6%), with forward and palindromic repeats of 30-40 bp being dominant. Tandem repeats exhibited significant distribution heterogeneity among chromosomes. Adaptive evolution analysis showed that most PCGs (protein-coding genes) had Ka/Ks ratios below 1 (ranging from 0.07 to 0.78), with the atp9 gene showing the lowest ratio (0.07), while the mttB gene exhibited a significantly higher Ka/Ks ratio of 3.48. Additionally, 1.62% of the mitochondrial genome sequence was homologous to the chloroplast genome, carrying 26 complete functional genes, including 15 tRNA and 2 rRNA genes. Codon usage bias may be related to mutation pressure due to the high AT content of the genome or reflect adaptive selection pressures for translational efficiency. The Ka/Ks results align with the widespread purifying selection observed in mitochondrial genomes, while the high Ka/Ks ratio of the mttB gene suggests it might be under positive selection to adapt to environmental pressures. The evolutionary evidence of inter-organelle gene transfer highlights the homologous sequences between mitochondria and chloroplasts. Overall, these findings systematically elucidate the adaptive evolutionary mechanisms and functional regulation of the tea plant mitochondrial genome.

Tea ( ) is a significant economic crop, and investigations into the structure and function of its mitochondrial genome are crucial for understanding the evolutionary history and genetic characteristics of this species. This study presents the first comprehensive analysis of the mitochondrial genome of the tea cultivar 'Zhuyeqi' ( cv. 'Zhuyeqi'), aiming to elucidate its genomic structural features, gene composition, and evolutionary patterns. The findings provide a theoretical foundation for genetic breeding and molecular biology research in tea plants. High-throughput sequencing was employed to sequence the mitochondrial genome of 'Zhuyeqi'. Bioinformatics approaches were utilized for genome assembly and annotation. Various analytical strategies, including identification of RNA editing sites, codon usage bias analysis, repeat sequence recognition, calculation of non-synonymous substitution rates (Ka) and synonymous substitution rates (Ks), comparative genomics, and collinearity analysis, were applied to comprehensively analyze the structural features and evolutionary dynamics of the mitochondrial genome. The mitochondrial genome of 'Zhuyeqi' consists of one circular chromosome and six linear chromosomes, with a total length of 911,255 bp and a GC content of 46%. Genome annotation identified 77 functional genes, including 38 protein-coding genes (PCGs). The study revealed heterogeneously distributed introns within genes such as trnM-CAT (5 copies) and . RNA editing analysis identified 556 C-to-U editing sites, notably enriched in (38 sites) and (34 sites). Codon usage bias analysis indicated that leucine (Leu, 10%) and arginine (Arg, 7%) were the most frequently used amino acids. Repeat sequence analysis showed that dispersed repeats (780, 72%) dominated, with satellite DNA exhibiting significant distribution biases on (11) and (5). Ka/Ks analysis revealed that 37 PCGs were under varying selective pressures (0.09-2.70), with (Pi=0.09) and (Pi=0.09) showing exceptionally high variability, while (Pi=0) was completely conserved. Comparative genomics uncovered 66 homologous segments (25,656 bp) between the mitochondrial and chloroplast genomes, containing 27 intact genes such as , confirming horizontal gene transfer events. Collinearity analysis demonstrated a high degree of conservation in genomic structures between 'Zhuyeqi' and closely related species. This study lays an important theoretical foundation for further elucidating the structural characteristics and evolutionary mechanisms of the tea plant mitochondrial genome.

Background Brucella spp. are Gram-negative, facultative intracellular pathogens that cause brucellosis in both humans and animals. The B. abortus vaccine strain 104 M is the only vaccine available in China for the prevention of brucellosis in humans. Although the B. abortus 104 M genome has been fully sequenced, the current genome annotations are not yet complete. In addition, the main mechanisms underpinning its residual toxicity and vaccine-induced immune protection have yet to be elucidated. Mapping the proteome of B. abortus 104 M will help to improve genome annotation quality, thereby facilitating a greater understanding of its biology. Results In this study, we utilized a proteogenomic approach that combined subcellular fractionation and peptide fractionation to perform a whole-proteome analysis and genome reannotation of B. abortus 104 M using high-resolution mass spectrometry. In total, 1,729 proteins (56.3% of 3,072) including 218 hypothetical proteins were identified using the culture conditions that were employed this study. The annotations of the B. abortus 104 M genome were also refined following identification and validation by reverse transcription-PCR. In addition, 14 pivotal virulence factors and 17 known protective antigens known to be involved in residual toxicity and immune protection were confirmed at the protein level following induction by the 104 M vaccine. Moreover, a further insight into the cell biology of multichromosomal bacteria was obtained following the elucidation of differences in protein expression levels between the small and large chromosomes. Conclusions The work presented in this report used a proteogenomic approach to perform whole-proteome analysis and genome reannotation in B. abortus 104 M; this work helped to improve genome annotation quality. Our analysis of virulence factors, protective antigens and other protein effectors provided the basis for further research to elucidate the mechanisms of residual toxicity and immune protection induced by the 104 M vaccine. Finally, the potential link between replication dynamics, gene function, and protein expression levels in this multichromosomal bacterium was detailed.