Complete chloroplast genome of Solanum viarum (Solanaceae)
Article information
Abstract
Solanum viarum Dunal, a perennial herb or subshrub belonging to the family Solanaceae, has been naturalized in Korea, forming a sustainable population that has grown naturally with native plants for more than ten years. However, its complete chloroplast genome sequence remains unreported. Therefore, we determined the complete chloroplast genome sequence of S. viarum Dunal via genome sequencing, assembly, and annotation of DNA extracted from fresh S. viarum leaves. The S. viarum genome is 156,209 bp in total length (large single-copy region, 86,877 bp; small single-copy region, 18,494 bp; two inverted repeat regions, 25,419 bp each) with a GC content of 37.8% and 130 genes, including 84 coding DNA sequences, 37 tRNA genes, eight rRNA genes, and one pseudogene (infA). To investigate phylogenetic relationships, a maximum likelihood tree was constructed based on 78 protein-coding genes extracted from the chloroplast genomes of 35 Solanum species. Calystegia soldanella and Calystegia hederacea (Convolvulaceae) were used as the outgroups. The phylogenetic analysis revealed that S. viarum is located within the Acanthophora clade and closely related to S. aculeatissimum. Our results provide genetic information pertaining to S. viarum and elucidate its relationship with related species, thereby offering a molecular foundation for future phylogenetic studies of Solanum.
INTRODUCTION
As the largest genus of the family Solanaceae, Solanum L. comprises approximately 1,230 species (POWO, 2025) and is one of the top 10 most species-rich genera in seed plants (Frodin, 2004; Bohs, 2005; Weese and Bohs, 2007). The Solanum genus includes globally important agricultural crops such as potato (Solanum tuberosum L.), tomato (Solanum lycopersicum L.), and eggplant (Solanum melongena L.) (Vorontsova et al., 2013). The subgenus Leptostemonum within the genus Solanum comprises approximately 350–450 species, accounting for approximately one-third of the entire genus (Bohs, 2005; Levin et al., 2005, 2006; Weese and Bohs, 2007). Leptostemonum species are also known as “spiny solanums” because of the common presence of epidermal prickles (Levin et al., 2005).
Solanum viarum is a perennial herb or subshrub native to South America (Brazil, Paraguay, Uruguay, and Argentina) (Nee, 1999) and is widely distributed across the tropical and subtropical regions of Asia, particularly northern India and China (Aubriot and Knapp, 2022). In Korea, S. viarum is distributed along roadsides, pastures, vacant lots, and grasslands on Jejudo Island. It was first introduced to Jejudo Island in 2000 as an alien species and has since grown naturally with native plants for over 10 years, forming a persistent naturalized plant population (Kang et al., 2020). S. viarum has potential medicinal uses owing to the presence of compounds with strong antioxidant activities (Wu et al., 2012). In addition, S. viarum can absorb and accumulate heavy metals through its roots, making it a potential candidate for the phytoremediation and plant-based treatment of heavy metal-contaminated tailing areas (Afonso et al., 2019). S. viarum is most similar to Solanum aculeatissimum; both species have simple trichomes on their adaxial (upper) leaf surfaces (Whalen, 1984) but S. viarum can be distinguished by deltate calyx lobes (vs. long acuminate in S. aculeatissimum) and puberulent ovary (vs. minutely stipitate glandular in S. aculeatissimum) (Aubriot and Knapp, 2022). Bohs (2005) conducted the first chloroplast-based phylogenetic analysis of the genus Solanum, which was followed by numerous molecular studies on both the genus Solanum and the subgenus Leptostemonum (Levin et al., 2005, 2006; Weese and Bohs, 2007; Stern et al., 2011; Särkinen et al., 2013; Tepe et al., 2016; Gagnon et al., 2021). However, the complete chloroplast genome of S. viarum has not yet been sequenced. In addition, it has not been compared at the clade level within the subgenus Leptostemonum using complete chloroplast genome data.
Therefore, the aim of this study was to present the first complete chloroplast genome of S. viarum, thereby providing crucial genetic information and improving our understanding of the phylogenetic relationships between this species and related taxa. The results of this study are expected to provide a molecular framework for future phylogenetic studies on Solanum and its subgenus Leptostemonum.
MATERIALS AND METHODS
Plant sampling
Fresh leaves of S. viarum were collected in July 2022 from Jejudo Island, Korea (33°19′40.8″N, 126°19′56.6″E) (Fig. 1). The voucher specimen was deposited at the Korea National Arboretum Herbarium (KH) under voucher number KHB1662054 (Fig. 2); the contact person is Dong Chan Son (E-mail: sdclym@korea.kr).
Photographs of Solanum viarum taken by Seong Gwon Lee and Sa Bum Jang in Jejudo Island (A–D). Permission to use the photographs was obtained from Seong Gwon Lee and Sa Bum Jang. (A) Solanum viarum, a naturalized species in Korea, has been confirmed to occur only on Jejudo Island. It grows in pastures, grasslands, and along roadsides. (B) Leaf of S. viarum, broadly ovate with deeply lobed margins and large prickles along the veins. (C) Flower of S. viarum, white to greenish, with lanceolate corolla lobes that are reflexed. (D) Fruit of S. viarum, globose and turning bright yellow when mature.
Korea National Arboretum herbarium collection specimens for Solanum viarum Dunal (voucher number: KHB1662054).
DNA extraction and genome sequencing, assembly, and annotation
Total DNA was extracted using a DNeasy Plant Mini Kit (Qiagen, Hilden, Germany) following the manufacturer’s protocol. The extracted DNA was detected on agarose gel (2%), and high-quality gDNA was sequenced using an Illumina MiSeq platform (Illumina Inc., San Diego, CA, USA) with a 301-bp insert size. In total, 7,061,852 reads were obtained and combined using Geneious v.8.0.5. (https://www.geneious.com), with reference to S. aculeatissimum Jacq. (OR381845.1), which is registered in the National Center for Biotechnology Information (NCBI) database. Thereafter, to verify the genome assembly, read coverage depth was assessed using the Draw_SequencingDepth.py script as described by Ni et al. (2023). Finally, GeSeq (Tillich et al., 2017) was used for genome sequence annotation. A circular map of the complete chloroplast genome was generated using OrganellarGenomeDRAW (Greiner et al., 2019). The complete sequenced chloroplast genome of S. viarum was submitted to GenBank (accession no. PV866731).
Phylogenetic analysis
Maximum likelihood (ML) analysis was performed with 34 subgenus of Leptostemonum (genus Solanum) and two outgroups (genus Calystegia in Convolvulaceae) to investigate the phylogenetic relationships. The chloroplast genome sequences of 36 species excluding S. viarum were downloaded from the NCBI GenBank database. In total, 78 coding DNA sequences (CDSs) were concatenated and aligned using MAFFT in PhyloSuite v.1.2.2 (Zhang et al., 2020); this process was performed using Geneious. The analysis model was tested using PhyloSuite v.1.2.3 (Zhang et al., 2020; Xiang et al., 2023). ML analysis was performed with IQ-tree v.2.2.0 (Nguyen et al., 2015) by setting the transversion model (TVM) + freerate model parameters with three of categories (R3) + empirical base frequency (F) model with 5,000 ultrafast bootstraps (Minh et al., 2013).
RESULTS AND DISCUSSION
Complete chloroplast genome of Solanum viarum
The complete chloroplast genome of S. viarum has a sequence length of 156,209 bp and a 37.8% GC content (Fig. 3), with an average read coverage depth of 165.79× (Fig. 4). The genome sequence was deposited in GenBank under the accession number PV866731, with associated BioProject, Sequence Read Archive, and Bio-Sample numbers of PRJNA1285601, SUB15430035, and SAMN49770012, respectively. The genome displayed a typical quadripartite structure comprising a pair of inverted repeats (IR, 25,419 bp each) separating a large single-copy region (LSC, 86,877 bp) from a small single-copy region (18,494 bp). The genome comprised 130 genes, including 84 CDSs, 37 tRNA genes, 8 rRNA genes, and 1 pseudogene (infA) (Table 1). The IR contained 18 genes, including 6 CDS genes, 7 tRNA genes, and 4 rRNA genes (CDS: ndhB, rpl2, rpl23, rps12, rps7, ycf2; tRNA: trnA-UGC, trnl-CAU, trnl-GAU, trnL-CAA, trnNGUU, trnR-ACG, trnV-GAC; rRNA: rrn16, rrn23, rrn4.5, rr5). Of the genes, 17 had one intron (10 CDS: atpF, ndhA, ndhB, petB, petD, rpl16, rpl2, rpoC1, rps12, rps16; 6 tRNA: trnAUGC, trnG-UCC, trnI-GAU, trnK-UUU, trnL-UAA, trnVUAC; 1 rRNA: rrn23) and two had two introns (clpP1 and pafI) (Fig. 5A). The rps12 gene is a trans-spliced gene with the 5′ end located in the LSC region and the duplicated 3′ end located in the IR regions (Fig. 5B).
Gene map of the complete chloroplast genome of Solanum viarum. Genes belonging to different functional groups are shown in different colors. Inner circle represents different regions of the chloroplast genome. Inverted repeat region A (IRA); inverted repeat region B (IRB); large single-copy region (LSC); small single-copy region (SSC). Line-chart in gray shows the GC content along the genome.
Sequencing depth and coverage map of the chloroplast genome assembly of Solanum viarum. The horizontal axis represents the base position of the plastome, and the vertical axis indicates the sequencing depth corresponding to each base.
List of genes annotated in the chloroplast genome of Solanum viarum.
Schematic maps of cis-spliced genes (A) and the trans-spliced gene (B) (rps12) in the chloroplast genome of Solanum viarum.
Phylogenetic analysis
An ML phylogenetic tree was constructed from the chloroplast genome of 78 CDSs from 37 species, including 35 species from the subgenus Leptostemonum of the genus Solanum (family Solanaceae) and two species from the genus Calystegia (family Convolvulaceae) as outgroups (Fig. 6). ML analysis supported the monophyly of subgenus Leptostemonum (bootstrap value = 100). The resulting phylogenetic tree was structured according to the classification systems described by Levin et al. (2006) and Stern et al. (2011). However, S. hieronymi, S. multispinum, and S. crotonoides were treated as exceptions because their phylogenetic relationships have not been clearly resolved in previous studies (Whalen, 1984; Levin et al., 2006; Stern et al., 2011). Specifically, S. viarum, S. aculeatissimum, and S. capsicoides (Acanthophora clade) formed a strongly supported clade along with S. lasiocarpum (Lasiocarpa clade) (bootstrap value = 99), which was placed at the basal position in the tree. Within this clade, members of the Acanthophora clade were grouped into a well-supported subclade (bootstrap value = 100), with S. viarum and S. aculeatissimum showing a close relationship strongly supported by a high bootstrap value (Fig. 6). This result is consistent with previous phylogenetic analyses (Levin et al., 2006) based on two nuclear gene regions (internal transcribed spacer and granule-bound starch synthase gene [GBSSI or waxy]) and one chloroplast spacer region (trnS–trnG), which also identified S. viarum and S. aculeatissimum as sister taxa. S. viarum and S. aculeatissimum were previously treated as S. khasianum (Babu and Hepper, 1979), but a subsequent monographic revision by Nee (1979) clarified their status as distinct species. The results of this study confirmed that S. viarum is closely related to S. aculeatissimum but remains a separate taxon distinguished by phylogenetic analysis of the chloroplast genome. In addition to clarifying the phylogenetic placement of S. viarum, this study also revealed that the Androceras/Crinitum clade exhibits paraphyly. This finding contrasts with Stern et al. (2011), who reported weak support for the clade (bootstrap value = 63) based on limited nuclear and chloroplast markers. By contrast, a more recent chloroplast genome study (Zhang et al., 2024) produced results consistent with ours, and in our analyses S. wrightii and S. rostratum were clearly separated with strong support (bootstrap value = 100). These results underscore the utility of complete chloroplast genomes for resolving problematic relationships within Solanum. This study reports the complete chloroplast genome of S. viarum for the first time, as well as a phylogenetic analysis of the subgenus Leptostemonum. These findings provide a molecular basis for future phylogenetic studies within this subgenus.
Maximum likelihood phylogenetic tree of 35 species in the genus Solanum (subgenus Leptostemonum) with two outgroups (genus Calystegia in Convolvulaceae) based on concatenated 78 coding DNA sequences of chloroplast genomes. Numbers above the nodes indicate bootstrap values.
Notes
ACKNOWLEDGMENTS
This study was supported by research projects of the Korea National Arboretum [KNA-1-2-39, 21-2].
CONFLICTS OF INTEREST
The authors declare that there are no conflicts of interest.