Keywords
Malpighiales, Phyllanthaceae, phylogenetic relationships, plastome, star gooseberry
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Nguyen HD, Nguyen TDQ, Vu MT and Do HDK. The complete chloroplast genome of Phyllanthus acidus (L.) Skeels (Phyllanthaceae) [version 2; peer review: 1 approved with reservations]. F1000Research 2025, 12:1059 (https://doi.org/10.12688/f1000research.140134.2)
NOTE: If applicable, it is important to ensure the information in square brackets after the title is included in all citations of this article.
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Revised
The complete chloroplast genome of Phyllanthus acidus (L.) Skeels (Phyllanthaceae)
[version 2; peer review: 1 approved with reservations]
PUBLISHED 16 Oct 2025
OPEN PEER REVIEW
REVIEWER STATUS
This article is included in the Genomics and Genetics gateway.
Abstract
Corresponding author:
Hoang Dang Khoa Do
Competing interests:
No competing interests were disclosed.
Grant information:
This research is funded by Nguyen Tat Thanh University, Ho Chi Minh city, Vietnam to Nguyen Hoang Danh.
The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Copyright:
© 2025 Nguyen HD et al.
This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
How to cite: Nguyen HD, Nguyen TDQ, Vu MT and Do HDK. The complete chloroplast genome of Phyllanthus acidus (L.) Skeels (Phyllanthaceae) [version 2; peer review: 1 approved with reservations]. F1000Research 2025, 12:1059 (https://doi.org/10.12688/f1000research.140134.2)
First published: 31 Aug 2023, 12:1059 (https://doi.org/10.12688/f1000research.140134.1)
Latest published: 16 Oct 2025, 12:1059 (https://doi.org/10.12688/f1000research.140134.2)
The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Revised Amendments from Version 1
Based on the comments of the reviewer, we added the new information for phylogenetic analysis with Bayesian inference method, coverage depth, and cis/trans-splicing genes. In addition, we added the reason for conducting the current study with Phyllanthus acidus target.
See the authors' detailed response to the review by Jessica D. Rey
Introduction
Phyllanthus acidus (L.) Skeels, also known as star gooseberry, belongs to the Phyllanthaceae family and is commonly distributed in the wet tropical regions, including South Asia, Southeast Asia, Central Africa, the Caribbean region, Central America, and South America (POWO 2022). P. acidus has been traditionally used to treat various diseases, including inflammation, gastrointestinal problems, rheumatism, bronchitis, Alzheimer’s, and hepatic diseases (Jain et al. 2011; Chakraborty et al. 2012; Srirama et al. 2012; Uddin et al. 2016). The leaves and roots of P. acidus also possess antidotal properties against viper venom (Jayvir 1998). Moreover, P. acidus could potentially alleviate hypertension (Leeya et al. 2010).
The chloroplast (cp) genome is highly effective at inferring phylogeny since it is predominantly maternally inherited, has a conversed structure and gene content, and has a slow mutation frequency (Palmer et al. 1988). Additionally, the cp genomes provide essential data for examining population genetics, molecular identification, and genetic engineering (Powell et al. 1995; Daniell et al. 2016; Cao et al. 2022). The current study explored the characteristics of the P. acidus cp genome which has not been studied and its phylogenetic implication to gain more information about the evolution and phylogenetic relationships within the Phyllanthaceae family and closely related taxa.
Methods
Sample collection
The P. acidus sample (young branches with leaves) was collected from Can Tho, Viet Nam (9°56′55.7″N, 105°30′16.0″E) and labeled with voucher number: NTT-2022.12.CR (contact person: Dr. Do Hoang Dang Khoa, dhdkhoa@ntt.edu.vn). It was deposited at the NTT Hi-tech Institute, Nguyen Tat Thanh University. No specific permit was required to collect and study the species in Vietnam. The leaf sample was dried with silica gel and stored in a -80°C freezer until DNA extraction was conducted.
Data collection and analysis
The total genomic DNA extraction from the dried leaves was carried out using the Cetyltrimethylammonium bromide (CTAB) protocol (Doyle & Doyle 1987). The quality of genomic DNA samples was checked using gel electrophoresis and the NanoDrop OneC Spectrophotometer. The DNA samples that showed a clear band on agarose gel and had a 260/280 ratio between 1,8-2 and a 260/230 ratio between 2.0-2.2 were selected for conducting the next-generation sequencing step. Subsequently, the library was prepared with the NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB, USA). The library was sequenced using the Illumina MiSeq platform to generate paired-end reads of 150 bp (Ktest Science Co. Ltd., Vietnam). The raw reads were qualified and filtered for low-quality reads (Q score < 20 and length < 100 bp) and reads containing primers or adapters using FastQC v0.12.1 and Trimmomatic v0.39 programs (Andrews 2010; Bolger et al. 2014). For the assembly of the cp genome, the NOVOPlasty v4.3.1 program was used (Dierckxsens et al. 2017). Preliminary annotation was conducted by Geseq with default parameters (Tillich et al. 2017). The complete annotation genome was illustrated using OrganellarGenomeDRAW v1.3.1 (Greiner et al. 2019). All 79 protein-coding regions in the cp genomes of P. acidus and 16 related taxa from the Phyllanthaceae were extracted and aligned for phylogenetic analysis using MUSCLE v5 program (Edgar 2004). The chloroplast genome of Acalypha hispida (Euphorbiaceae; Genbank accession no. NC_070339) was selected as an outgroup. A maximum likelihood phylogenetic tree was reconstructed using IQTREE with 1000 bootstrap replicates and GTRGAMMA substitution model (Nguyen et al. 2015). In addition, Bayesian inference method was applied to the aligned sequnces under GTRGAMMA substitution model using MrBayes v3.2.7a with 1,000,000 generations and a discard of 25% sample (Ronquist et al. 2012).
Results
Approximately 349.8 MB of clean reads were obtained and used for completing the cp genome of P. acidus. The assembly process utilized 1,166,034 paired-end reads, resulting in an average coverage depth of 2,234.3X (Nguyen, Nguyen, Do, and Vu, 2023; Nguyen, Do, and Vu, 2023) (Supplementary Figure S1, Do 2025). The quadripartite cp genome of P. acidus (GenBank accession number OR050568) had a length of 156,331 bp and consisted of an LSC region of 85,807 bp, a SSC region of 19,262 bp, and a pair of IR regions of 25,631 bp ( Figure 1). The overall GC contents of the genome was 36.9%, and the GC content of the LSC, SSC, and IR regions were 34.6%, 30.6%, and 43.1%, respectively. The cp genome of P. acidus contained a total of 130 genes, including 85 protein-coding regions, 37 tRNA genes, and eight rRNA genes (Table 1). Among 85 protein-coding genes, 17 genes contained introns, of which ycf3 and clpP contained two introns (Supplementary Figure S2, Do 2025). In IR regions, a total of 19 genes were duplicated, including eight protein-coding regions (i.e., rps19, rpl2, rpl23, ycf1, ycf2, ndhB, rps12, and rps7), seven tRNAs (trnI_CAU, trnL_CAA, trnV_GAC, trnI_GAU, trnA_UGC, trnR_ACG, and trnN_GUU), and four rRNAs (rrn16S, rrn23S, rrn4.5S, and rrn5S). Notably, rps19 and ycf1 duplications were incomplete. The phylogenetic analysis revealed a paraphyly of Phyllanthus species, in which Breynia futicosa and Glochidion chodoense formed a clade with Phyllanthus amarus ( Figure 2). Therefore, more genomic data and samples of Phyllanthaceae species are required for further phylogenetic studies.
Figure 1. Map of the cp genome of Phyllanthus acidus.
Genes located inside the circle are transcribed in a clockwise direction, while genes outside the circle are transcribed counterclockwise. The inner circle depicted in dark gray represents the GC content, while the light-gray circle represents the AT content of the genome. LSC: large single copy; SSC: small single copy; IRA/IRB: inverted repeat regions.
Table 1. List of genes in the chloroplast genome of Phyllanthus acidus.
| Groups of genes | Name of genes |
|---|---|
| Ribosomal RNAs | rrn4.5(2x), rrn5(2x), rrn16(2x), rrn23(2x) |
| Transfer RNAs | trnA_UGC *(2x), trnC_GCA, trnD_GUC, trnE_UUC, trnF_GAA, trnG_UCC *, trnG_GCC, trnH_GUG, trnI_GAU *(2x), trnK_UUU *, trnL_CAA(2x), trnL_UAA *, trnL_UAG, trnfM_CAU, trnM_CAU(2x), trnM_CAU, trnN_GUU(2x), trnP_UGG, trnQ_UUG, trnR_ACG(2x), trnR_UCU, trnS_GCU, trnS_GGA, trnS_UGA, trnT_GGU, trnT_UGU, trnV_GAC(2x), trnV_UAC *, trnW_CCA, trnY_GUA |
| Photosystem I | psaA, psaB, psaC, psaI, psaJ |
| Photosystem II | psbA, psbB, psbC, psbD, psbE, psbF, psbH, psbI, psbJ, psbK, psbL, psbM, psbN, psbT, psbZ |
| Cytochrome | petA, petB *, petD *, petG, petL, petN |
| ATP synthases | atpA, atpB, atpE, atpF *, atpH, atpI |
| Large unit of Rubisco | rbcL |
| NADH dehydrogenase | ndhA *, ndhB *(2x), ndhC, ndhD, ndhE, ndhF, ndhG, ndhH, ndhI, ndhJ, ndhK |
| ATP-dependent protease subunit P | clpP * |
| Envelop membrane protein | cemA |
| Large units of ribosome | rpl2 *(2x), rpl14, rpl16 *, rpl20, rpl22, rpl23(2x), rpl32, rpl33, rpl36 |
| Small units of ribosome | rps2, rps3, rps4, rps7(2x), rps8, rps11, rps12 *(2x), rps14, rps15, rps16 *, rps18, rps19(2xa) |
| RNA polymerase | rpoA, rpoB, rpoC1 *, rpoC2 |
| Initiation factor | infA |
| Other genes | accD, ccsA, matK |
| Hypothetical proteins and conserved reading frames | ycf1(2xa), ycf2(2x), ycf3 *, ycf4 |
Figure 2. The phylogenetic tree of Phyllanthus acidus and related species based on the 79 protein-coding sequences of cp genome.
The asterisk indicates P. acidus sequenced in this study. The numbers next to each node are bootstrap values and posterior probability inferred from maximum likelihood and Bayesian inference methods.
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how to cite this article
Nguyen HD, Nguyen TDQ, Vu MT and Do HDK. The complete chloroplast genome of Phyllanthus acidus (L.) Skeels (Phyllanthaceae) [version 2; peer review: 1 approved with reservations]. F1000Research 2025, 12:1059 (https://doi.org/10.12688/f1000research.140134.2)
NOTE: If applicable, it is important to ensure the information in square brackets after the title is included in all citations of this article.
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Version 1
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PUBLISHED 31 Aug 2023
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How to cite this report:
Rey JD. Reviewer Report For: The complete chloroplast genome of Phyllanthus acidus (L.) Skeels (Phyllanthaceae) [version 2; peer review: 1 approved with reservations]. F1000Research 2025, 12:1059 (https://doi.org/10.5256/f1000research.153464.r202362)
The direct URL for this report is:
https://f1000research.com/articles/12-1059/v1#referee-response-202362
https://f1000research.com/articles/12-1059/v1#referee-response-202362
NOTE: it is important to ensure the information in square brackets after the title is included in this citation.
Reviewer Report 22 Jan 2024
Approved with Reservations
VIEWS 67
This is a good genome note on the sequencing and assembly of the plastome of Phyllanthus acidus. However, the paper lacks enough details on the reasons behind investigating the species (i.e., limited genetic and genomic resources? problems with identification?).
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HOW TO CITE THIS REPORT
Rey JD. Reviewer Report For: The complete chloroplast genome of Phyllanthus acidus (L.) Skeels (Phyllanthaceae) [version 2; peer review: 1 approved with reservations]. F1000Research 2025, 12:1059 (https://doi.org/10.5256/f1000research.153464.r202362)
The direct URL for this report is:
https://f1000research.com/articles/12-1059/v1#referee-response-202362
https://f1000research.com/articles/12-1059/v1#referee-response-202362
NOTE: it is important to ensure the information in square brackets after the title is included in all citations of this article.
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Author Response 15 Oct 2025Hoang Dang Khoa Do, NTT Hi-Tech Institute, Nguyen Tat Thanh University, Ho Chi Minh City, Vietnam15 Oct 2025Author ResponseDear Reviewers,
Thank you very much for your helpful comments.
1/ We added the reason for conducting genomic research about Phyllanthus acidus in the Introduction as “which has ... Continue reading
COMMENTS ON THIS REPORT
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Author Response 15 Oct 2025Hoang Dang Khoa Do, NTT Hi-Tech Institute, Nguyen Tat Thanh University, Ho Chi Minh City, Vietnam15 Oct 2025Author ResponseDear Reviewers,
Thank you very much for your helpful comments.
1/ We added the reason for conducting genomic research about Phyllanthus acidus in the Introduction as “which has ... Continue reading
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