Research Article
Print
Research Article
A new species of Semisulcospira O. Boettger, 1886 (Gastropoda, Cerithoidea, Semisulcospiridae) from Fujian, China with mitochondrial genome and its phylogenetic implications
expand article infoYi-Bin Xu‡, Yuan-Zheng Meng§, Sheng Zeng§, Hang-Jun Wang|, Shen Zhong#, De-Yuan Yang¤§, Xi-Ping Zhou«, Christopher J. Glasby»˄
‡ Fisheries Research Institute of Fujian, Xiamen, China
§ Xiamen University, Xiamen, China
| Ministry of Natural Resources of the People’s Republic of China, Wenzhou, China
¶ Marine Ecosystem Observation and Research Station on the Yangtze River Estuary, Wenzhou, China
# Xiamen Ocean Vocational College, Xiamen, China
¤ National Taiwan Ocean University, Keelung, China
« Xiamen University Tan Kah Kee College, Zhangzhou, China
» Natural Sciences, Museum & Art Gallery Northern Territory, Darwin, Australia
˄ Australian Museum Research Institute, Sydney, Australia

Corresponding author: De-Yuan Yang ( deyuanyang92@163.com )

Corresponding author: Xi-Ping Zhou ( zoeyzhou@xujc.com )

Corresponding author: Christopher J. Glasby ( glasby93@gmail.com )

Academic editor: Frank Köhler
© 2025 Yi-Bin Xu, Yuan-Zheng Meng, Sheng Zeng, Hang-Jun Wang, Shen Zhong, De-Yuan Yang, Xi-Ping Zhou, Christopher J. Glasby.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation: Xu Y-B, Meng Y-Z, Zeng S, Wang H-J, Zhong S, Yang D-Y, Zhou X-P, Glasby CJ (2025) A new species of Semisulcospira O. Boettger, 1886 (Gastropoda, Cerithoidea, Semisulcospiridae) from Fujian, China with mitochondrial genome and its phylogenetic implications. Zoosystematics and Evolution 101(1): 17-34. https://doi.org/10.3897/zse.101.136882
ZooBank: urn:lsid:zoobank.org:pub:F686B6D9-0DE3-4BB4-94E5-26FEA80E64CD
Open Access

Abstract

During our investigation of the diversity and phylogeny of the family Semisulcospiridae in China, we have found a presumed new species in mountain streams of Xiamen, Fujian Province on both morphology and molecular evidence. This new species, Semisulcospira egretta sp. nov., is characterized by its thin and smooth shell, retaining about 0.5 mm long embryonic shells in its brood pouch, and by distinctive features of its radular morphology. Phylogenetic analyses of partial sequences of the mitochondrial markers 16S ribosomal RNA (16S) and cytochrome c oxidase subunit I (COI) and complete sequences of the nuclear ribosomal internal transcribed spacer (ITS, including internal transcribed spacer 1, 5.8S ribosomal RNA gene, and internal transcribed spacer 2) supported the distinctiveness of S. egretta sp. nov. from other Semisulcospira species. These analyses consistently placed S. egretta sp. nov. in Semisulcospira confirming its monophyly. Phylogenetic analyses based on just COI and 16S exhibited low resolution rendering the phylogenetic position of the new species within the genus uncertain. In contrast, phylogenetic tree analyses of complete mitochondrial genomes, which include 13 protein-coding genes (PCGs), were better resolved. However, the mitogenome dataset included a more restricted sampling of taxa (16), impeding a detailed exploration of the phylogenetic position of the new species. To address this deficit, we provide seven complete mitochondrial genomes for S. egretta sp. nov., obtained using the genome skimming sequencing method. This study contributes important molecular and genomic data, aiding in the understanding of semisulcospirid phylogeny, and highlights the need for further taxonomic sampling and phylogenomic methods to resolve the evolutionary relationships within Semisulcospiridae.

Key Words

16S rRNA, COI, mitogenome, phylogenetic analysis, semisulcospirid gastropods

Introduction

Semisulcospiridae Morrison, 1952 is a family of freshwater gastropods comprising more than 90 species from four genera (Lydeard and Cummings 2019; Strong et al. 2022; Du and Yang 2023; MolluscaBase eds. 2024). This family is predominantly found in East Asia and North America, with the majority (over 60 species from three genera) documented in China (Du et al. 2019a, 2019b; Du and Yang 2023; He et al. 2024).

Semisulcospira O. Boettger is the type genus of the family, characterized by its viviparous reproductive strategy, while the other three genera are oviparous (Köhler 2017; Du et al. 2019b; Strong et al. 2022). Members of Semisulcospira are mainly distributed in East Asia and prefer living in freshwater streams, rivers, and lakes (Lydeard and Cummings 2019; Strong et al. 2022; Du and Yang 2023). They are important to environmental studies and public health (Köhler 2016); however, our understanding of their taxonomy and phylogenetic systematics remains limited. Due to significant morphological variation in shell characteristics, Semisulcospira contains numerous nominal species. Recently, anatomical and molecular studies have demonstrated that many species of Semisulcospiridae are misidentifications, even at the genus level (Du et al. 2019a, 2019b; Strong et al. 2022; Sawada et al. 2024). For example, the genera Namrutua Abbott, 1948 and Senckenbergia Yen, 1939 have been proven synonyms of Semisulcospira, and Melania dulcis Fulton, 1904, M. lauta Fulton, 1904, and Semisulcospira inflata S. Tchang, 1949 have been regarded as synonyms of Hua textrix (Heude, 1889) (Du et al. 2019a; Du et al. 2019b).

Currently, the genus Semisulcospira comprises more than 36 valid species, with 18 endemic to Lake Biwa in Japan. Approximately 10 species have been documented from China, with Semisulcospira libertina (A. Gould, 1859) as the only species recorded in Fujian Province (Liu et al. 1979; Liu et al. 1994; Du et al. 2019a, 2019b; Du and Yang 2023; MolluscaBase eds. 2024; Sawada et al. 2024). As molecular methods have been introduced in the systematic study of semisulcospirids, numerous cryptic species have been discovered in China, Japan, and the U. S. (Du et al. 2019a; Sawada and Nakano 2021; Strong et al. 2022; Chen et al. 2023; Du and Yang 2023; Sawada et al. 2024). Previous research has demonstrated high diversity within this family, indicating the potential for more cryptic species (Strong et al. 2022; Du and Yang 2023; He et al. 2024).

During the investigation of the diversity and phylogeny of Semisulcospiridae in China, we found a presumed new species from mountain streams of Xiamen, Fujian Province, based on both morphological and molecular data. Additionally, we provided seven complete mitochondrial genome (mitogenome) sequences of the putative new species using the genome skimming sequencing method (Yang et al. 2024). Phylogenetic analysis was performed using two partial mitochondrial genes, cytochrome oxidase subunit 1 (COI) and 16S ribosomal RNA (16S), as well as the complete mitogenomes.

Material and methods

Specimens and sampling

Samples were collected from streams in Xiamen, Fujian Province, China from 2022 to 2023. Examined specimens comprised dry shells and ethanol-preserved samples deposited in the Marine Biological Sample Museum, Third Institute of Oceanography, Ministry of Natural Resources (Xiamen, China) and De-Yuan Yang’s laboratory, College of the Environment and Ecology, Xiamen University, Xiamen, Fujian Province, China (Fig. 1).

Figure 1. 

The habitat and sampling localities of S. egretta sp. nov. in Xiamen City, Fujian Province, China. A. Type locality, 24°36'29"N, 117°53'39"E, in Haicang District; B. 24°52'35"N, 118°03'01"E, in Tong’ an District; C. A simplified map of Xiamen, locality A and B are marked with red points; D. A living specimen of S. egretta sp. nov. All photos are taken by Yuan-Zheng Meng.

Morphological methods

Living specimens for dissecting were soaked in hot water for 1 minute to separate soft parts from the shells. Anatomical techniques and terminology follow Strong (2011) and Strong and Köhler (2009). Specimens were photographed with an Olympus E-M1 Mark II camera with a 60 mm macro lens, and image stacks were obtained using Helicon Focus 7 (https://www.heliconsoft.com/heliconsoft-products/helicon-focus/) and post-processed using Adobe Photoshop CC 2019. Shells were photographed from two different perspectives, apertural view (viewed from aperture with shell parallel to the coiling axis) and abapertural view (viewed opposite from the apertural view). The morphological parameters of the shells were measured using calipers with a precision of 0.1 mm, following Du et al. (2019a). The radulae and embryos were digested by Proteinase K at 55 °C for 2 hours, and then cleaned, dried, coated with gold, and imaged under SEM (Phenom Prox). For more information about the specimens used in this study (Suppl. material 1: table S1).

Abbreviations

MBSM: the Marine Biological Sample Museum, Third Institute of Oceanography, Ministry of Natural Resources (Xiamen, China). XMU_DYY: Laboratory of De-Yuan Yang, College of the Environment and Ecology, Xiamen University (Xiamen, Fujian Province, China). ZMB: Museum für Naturkunde, Berlin. NHMUK: Natural History Museum, London. MNHN: Muséum national d’Histoire naturelle, Paris. YZM: Collection by Yuan-Zheng Meng. LWL: Collection by Li-Wen Lin. SEM: Scanning electron microscope. 16S: 16S ribosomal RNA. COI: Cytochrome oxidase subunit I. ITS: Internal transcribed spacer complete sequence, including internal transcribed spacer 1, 5.8S ribosomal RNA gene, and internal transcribed spacer 2. PCGs: Protein-coding genes. ML: Maximum likelihood. BI: Bayesian inference. PP: Posterior probability. BS: Ultrafast bootstrap support. H: Shell height. B: Shell width. BW: Height of the last whorl. WA: Width of aperture. LA: Length of aperture.

DNA extraction, amplification, and sequencing

Genomic DNA was extracted from a 1 mm3 sample of muscle tissue from the foot of specimens using a TIANamp Genomic DNA Kit (TIANGEN, Beijing, China) according to the manufacturer protocols.

Two partial mitochondrial markers:16S rRNA (16S, 467 bp) and cytochrome oxidase subunit I (COI, 655 bp) were sequenced from three specimens of the new species (XMU_DYY_XM02, XM04, and XM05). For detailed information on protocols and primers of PCR (polymerase chain reaction), see Suppl. material 1: table S2.

Mitogenome sequences were obtained from seven specimens (XMU_DYY_XMA03, XMA05, XMA06, XMB04, XMC01, XMU_DYY_RTM05, and RTM25). Genome skimming for the mitogenome sequences was conducted using next-generation sequencing (NGS) on the Illumina NovaSeq X Plus and DNBSEQ-T7 platforms, generating paired-end reads of 150 bp in length. The sequencing was performed by Novogene Bioinformatics Technology Co., Ltd. in Beijing, China.

Mitogenome assembly and annotation

The assembly and annotation strategy of mitogenomes followed the methods of Yang et al. (2024) and Xu et al. (2024). In summary, we initially assembled the data using Getorganelle v1.7.6.1 (Jin et al. 2020). If assembly failed, we used other Semisulcospiridae mitogenomes as seeds for reassembly.

Nuclear genes assembly and annotation

To evaluate the p-distance of internal transcribed spacer (ITS) sequences, we extracted these sequences from genome skimming data of the seven newly sequenced specimens, following the protocol outlined by Yang et al. (2024). Briefly, we employed Getorganelle v1.7.6.1 (Jin et al. 2020), using all publicly available 18S ribosomal RNA (18S rRNA), 28S ribosomal RNA (28S rRNA), and ITS sequences from the family Semisulcospiridae as seed sequences. In addition, we assembled all publicly available genome skimming data from the NCBI Sequence Read Archive (SRA accession numbers: SRR26074378, SRR26107127, SRR12518401) to obtain ITS gene sequences, and successful assembly was achieved for the data SRR26107127 and SRR12518401 (Suppl. material 1: table S3).

Molecular analyses

The phylogenetic analysis was conducted in four parts: 1) three separate datasets—a concatenated 16S+COI dataset integrated from previous studies containing 132 individuals, each with both 16S and COI sequences, as well as separate 16S and COI datasets (Suppl. material 1: table S3); 2) the extensive COI dataset (COI-ex), comprising 877 sequences (Suppl. material 1: table S4); 3) the ITS dataset, including 9 newly obtained ITS sequences in this study and two existing ITS sequences in NCBI (Suppl. material 1: table S4); 4) the mitogenome dataset, including the seven newly sequenced mitogenomes of S. egretta sp. nov. and the dataset from Xu et al. (2024) (Suppl. material 1: table S5). Hua jacqueti (Dautzenberg & H. Fischer, 1906) and Juga plicifera (I. Lea, 1838) were used as the outgroups of datasets 1 & 2, following Strong and Köhler (2009) and Köhler (2016). Sequences used in this study were integrated from previously published studies (Strong and Köhler 2009; Zeng et al. 2015; Hilgers et al. 2016; Köhler 2016; Whelan and Strong 2016; Zeng et al. 2016; Chiu et al. 2017; Köhler 2017; Kim and Lee 2018; Nguyen et al. 2018; Du et al. 2019a, 2019b; Guo et al. 2019; Hartnell College Genomics Group et al. 2019; Lee et al. 2019; Stelbrink et al. 2019; Xu et al. 2019; Lee et al. 2020; Miura et al. 2020; Yan et al. 2020a, b; Choi et al. 2021; Gim et al. 2021; Kato et al. 2022; Ling et al. 2022; Strong et al. 2022; Yang and Deng 2022; Yin et al. 2022; He et al. 2024; Xu et al. 2024).

Most subsequent analyses were conducted in PhyloSuite v1.2.3 (Zhang et al. 2020), including multiple sequence alignment, trimming, model selection, and phylogenetic tree construction.

For the 16S and COI sequences, we initially extracted the full-length 16S and COI sequences from our newly assembled mitochondrial genomes and used them as reference sequences for alignments. The alignments were performed using the online version of MAFFT (https://mafft.cbrc.jp/alignment/server/, accessed October 2024) with default settings. The following parameters were adjusted: UPPERCASE/lowercase: same as input; Direction of nucleotide sequences: adjusted according to the first sequence (sufficient for most cases); Output order: same as input.

For the mitogenomes dataset, we only focused on the 13 protein-coding genes (PCGs). These sequences were aligned using the MAFFT v.7.313 (Katoh and Standley 2013) plugin in PhyloSuite under the codon alignment mode. All aligned sequences were inspected in Geneious Prime 2024.0.4 (https://www.geneious.com) before and after trimming.

Considering the COI and 16S sequences mainly from Köhler (2016, 2017) and Du et al. (2019a, 2019b), which were amplified using different primers, leading to differences in the amplified regions (Fig. 2), we applied five trimming methods for dataset 1 to to assess the impact of trimming on phylogenetic reconstruction: (1) using TrimAl v.1.2 (Capella-Gutiérrez et al. 2009) with the automated1 setting (referred to as auto); (2) trimming sequences based on the length of sequences from Du et al. (2019b, 2019b) as a reference (referred to as ‘du’); (3) trimming sequences based on the length of sequences from Köhler (2016, 2017) as a reference (referred to as ‘ko’); (4) using the shared overlapping region between Köhler (2016, 2017) and Du et al. (2019b, 2019b) as a reference (‘du ∩ ko’); and (5) using the combined region from both Köhler (2016, 2017) and Du et al. (2019b, 2019b) as a reference (‘du ∪ ko’) (Fig. 2). As a result, the dataset 1 generated 15 sub-datasets (see Suppl. materials 2–4). Datasets 2–4 were trimmed under the auto mode (Suppl. material 1: table S6).

Figure 2. 

A schematic diagram of the COI gene trimming methods.

ModelFinder v.2.2.0 (Kalyaanamoorthy et al. 2017) was used to select the best substitution models under the Bayesian Information Criterion (BIC) for maximum likelihood (ML) analysis and the corrected Akaike Information Criterion (AICc) for Bayesian inference (BI) analysis (Suppl. material 1: table S7). Maximum likelihood phylogenetic reconstruction was conducted in IQ-TREE v.2.2.2 (Nguyen et al. 2015), using the best partition scheme and under an edge-linked partition model for 20000 ultrafast bootstraps (UFBoot2). Bayesian Inference phylogenies were inferred using MrBayes v.3.2.7a (Ronquist et al. 2012) under a partition model (2 parallel runs, 2 000 000 generations), ensuring the average standard deviation of split frequencies (ASDSF) value was below 0.01 (Ronquist et al. 2012). If convergence was not achieved after 2,000,000 generations (ASDSF value > 0.01), additional generations were added to continue the analysis.

Pairwise tree structure comparison was conducted using the all.equal.phylo function in the ape v.5.7.1 package (Paradis and Schliep 2019), and topological differences were assessed using TreeSpace (Jombart et al. 2017), both implemented in R v.4.3.1 (R Core Team 2023). Finally, iTOL v.6 (Letunic and Bork 2024) was used to visualize the trees.

Summary statistics for multiple alignments were generated using a custom Python script, which utilizes the alignment summary function in BioKIT (Steenwyk et al. 2022) (Table 1).

Table 1.
Download as
CSV
XLSX

Summary statistics for multiple alignment of various datasets.

Dataset Gene Alignment length Trimal methods Trimal length Percentage (%) Constant sites Number of taxa Parsimony informative sites Variable sites
Mitogenome ATP6 705 auto 693 98.3 267 32 355 426
ATP8 177 auto 159 89.8 39 32 113 120
COX1 1533 auto 1512 98.6 882 32 563 630
COX2 687 auto 666 96.9 327 32 281 339
COX3 777 auto 759 97.7 354 31 335 405
CYTB 1137 auto 1134 99.7 526 31 539 608
ND1 942 auto 933 99 390 32 467 543
ND2 1077 auto 1062 98.6 298 32 663 764
ND3 351 auto 324 92.3 130 32 172 194
ND4 1380 auto 1359 98.5 442 32 810 917
ND4L 321 auto 288 89.7 101 31 144 187
ND5 1719 auto 1689 98.3 599 32 933 1090
ND6 549 auto 492 89.6 136 32 307 356
Concatenation 11355 auto 11070 97.5 4470 32 5702 6600
COI+16S COI 1538 auto 1533 99.7 1018 132 469 515
du 724 47.1 429 132 277 295
ko 658 42.8 368 132 272 290
du ∩ ko 550 35.8 312 132 225 238
du ∪ ko 832 54.1 485 132 324 347
16S 1459 auto 1345 92.2 791 132 460 554
du 544 37.3 333 132 176 211
ko 817 56.0 410 132 348 407
du ∩ ko 527 36.1 324 132 171 203
du ∪ ko 834 57.2 419 132 353 415
COI+16S 2997 auto 2878 96.0 1809 132 929 1069
du 1268 42.3 762 132 453 506
ko 1475 49.2 778 132 620 697
du ∩ ko 1077 35.9 636 132 396 441
du ∪ ko 1666 55.6 904 132 677 762
COI-ex COI 1533 auto 548 35.7 279 8877 247 270
ITS ITS 2657 auto 964 36.3 804 11 69 160

Genetic distances (p-distance and Kimura 2 Parameters, K2P) were computed using MEGA X with default settings (Kumar et al. 2018). A heatmap of the p-distances between selected specimens (clade D of Fig. 3) was generated by TBtools (Chen et al. 2020) (Fig. 4, Suppl. material 1: table S8).

Figure 3. 

Maximum likelihood tree (left) and Bayesian inference tree (right) inferred from COI and 16S gene sequences. Bootstrap supports (BS) /posterior probabilities (PP) are shown on the nodes. Notes: Semisulcospira egretta sp. nov. is highlighted in red.

Figure 4. 

A. A heatmap of p-distances between selected congeners. The p-distances based on COI and 16S are shown at the bottom left and upper right, respectively; B. Box plots of interspecific p-distances between S. egretta sp. nov. and other species based on COI (upper) and 16S (bottom). The specific name of the congeners is indicated by the first three letters; C. ML and BI phylogenetic trees of ITS genes; D. p-distance of S. egretta sp. nov. and other semisulcospirids.

Results

DNA barcoding

The average interspecific genetic distances (p-distance) for COI and 16S sequences between Semisulcospira egretta sp. nov. and other congeneric species ranged from 11.9% to 14.4%, and 5.0% to 7.4%, respectively. This value is significantly higher than the average intraspecific genetic distances within S. egretta sp. nov., which varied from 0% to 3.1% (0.84% on average for COI) and 0% to 2.2% (0.73% on average for 16S). The p-distance of ITS sequences between Semisulcospira egretta sp. nov ranged from 0% to 1% (0.56% on average) (Fig. 4, Suppl. material 1: table S8).

Phylogenetic analyses

A summary of the characteristics of the data matrices can be found in Table 1. The mitogenome dataset (13 PCGs) includes 16 Semisulcospiridae sequences, with a total length of 11070 bp and 5702 parsimony informative sites. In the COI+16S dataset, we selected the sub-dataset (du ∪ ko) for further analysis. This sub-dataset includes sequences from 132 individuals, with a total length of 1666 bp and 677 parsimony informative sites. The trimmed COI dataset (du ∪ ko) is 832 bp in length, containing 324 parsimony informative sites, and the 16S dataset (du ∪ ko) is 834 bp in length, containing 353 parsimony informative sites. The trimmed COI-ex dataset consists of sequences from 863 taxa that are 548 bp in length and include 247 parsimony informative sites.

Phylogenetic reconstruction of the 16S+COI dataset, using different trimming methods and employing Maximum Likelihood (ML) and Bayesian Inference (BI), resulted in 30 phylogenetic trees. TreeSpace (Jombart et al. 2017) was used to assess potential discrepancies among these trees. Analyzing all 30 trees in TreeSpace showed that the concatenated 16S+COI dataset exhibited fewer discrepancies compared to the individual 16S or COI datasets (Suppl. materials 2–4). For the 16S dataset, inferred trees based on different trimming methods did not cluster together (Suppl. material 5). In the COI dataset, three tree clusters were identified. Phylogenetic trees inferred using ML from different trimmed sub-datasets formed one cluster, whereas trees inferred using BI were divided into two distinct clusters (Suppl. material 5).

In the combined 16S+COI dataset, three tree clusters were identified. Generally, phylogenetic trees derived from different trimming methods using the same inference approach clustered together, with the exception of the ML tree from the ‘du’ sub-dataset (Suppl. material 5).

Based on this evidence, we can conclude that the phylogenetic tree inferred from the combined 16S+COI dataset is more reliable compared to those inferred from the individual 16S or COI datasets. Additionally, we propose that different trimming methods have an influence on the phylogenetic reconstruction.

Given that the ML and BI trees of the sub-dataset (du ∪ ko) cluster together closely, we present these two trees in more detail.

Phylogenetic trees (16S+COI) using maximum likelihood (ML) and Bayesian Inference (BI) are both divided into four primary clades. Clades A, B, and D only include Semisulcospira species, while clade C only includes Koreoleptoxis species (except NC_023364, see below). S. egretta sp. nov. forms a distinct lineage inside clade D (PP = 0.547, BS = 92.4%).

Phylogenetic analyses (ML and BI) inferred from the COI datasets have different topologies (Suppl. material 4), but Semisulcospira egretta sp. nov. has a close relationship with a clade containing 31 sequences of eight species (S. gottschei, S. coreana, S. forticosta, S. libertina, S. calculus, S. multigranosa, S. reiniana, and S. dorlosa, PP = 0.58; BS = 92.8%). Meanwhile, phylogenetic trees inferred from the 16S dataset show that S. egretta sp. nov. have a close relationship with Semisulcospira multigranosa (O. Boettger, 1886) in the ML and BI tree (PP = 0%, BI = 0.36, Suppl. material 4). Phylogenetic analyses inferred from the COI-ex (ML) dataset and the ITS dataset (ML and BI) all showed that S. egretta formed a monophyletic clade (BS = 100% in COI-ex, PP = 1, BS = 98.6% in ITS, Suppl. material 6 and Fig. 4C).

The topologies of the phylogenetic trees inferred from 13 PCGs are broadly consistent with the phylogenetic tree in Xu et al. (2024) (Fig. 5A). The tree contains 30 Cerithoidea mitogenome sequences, 16 of which are Semisulcospiridae sequences: 10 Semisulcospira (including the 7 newly reported ones in this study), five Koreoleptoxis, and one Hua. Nine Semisulcospira sequences clustered in a clade, including the 7 newly reported sequences of S. egretta sp. nov. The new species is closely related to S. gottschei and S. coreana, with high support values (BS = 100%, PP = 1). Another clade, a sister group of the former, contains five Koreoleptoxis sequences and a likely misidentified ‘Semisulcospira libertina’ sequence (see Xu et al. 2024).

Figure 5. 

A. Phylogenetic tree of Cerithioidea species inferred from 13 PCGs. The numbers at the internodes represent maximum likelihood (ML) bootstrap (BS) and Bayesian inference (BI) posterior probabilities (PP). The BS equal to 100% and PP equal to 1 were hidden; B. Mitochondrial gene map of Semisulcospira egretta sp. nov.; C. The gene map of the mitochondrion of S. egretta sp. nov. XMU_DYY_RTM25 (PQ165125) (A) and the gene order of 13 PCGs and 2 rRNAs for Cerithioidea, to show the complete synteny between them (B). The GenBank accession numbers used are listed after the species names. Notes: Semisulcospira egretta sp. nov. is highlighted in red (A).

In all phylogenetic trees of this study (16S+COI, 16S, COI, COI-ex, ITS, and mitognomes), the sequences of S. egretta sp. nov. clustered in a monophyletic group.

Mitochondrial genome analysis

Seven complete mitochondrial genomes of Semisulcospira egretta sp. nov. were sequenced, resulting in circular molecules with lengths ranging from 15,469 bp to 15,677 bp, and have been deposited in NCBI (accession numbers listed in Suppl. material 1: table S5). For clarity in the description, PQ165124 was selected for detailed analysis.

As with other Semisulcospiridae mitogenomes, the nucleotide composition of S. egretta sp. nov. exhibits a negative AT-skew (Suppl. material 1: table S9). The newly sequenced mitogenome contains 37 genes (13 PCGs, 2 rRNAs, and 22 tRNAs) (Fig. 5B). Nine of the PCGs and seven tRNAs are encoded on the positive strand (forward strand) and the remaining four PCGs, 15 tRNAs, and two rRNAs are located on the negative strand (reverse strand) (Fig. 5B).

The arrangement of the 13 PCGs and 2 rRNA genes is consistent with all other publicly available cerithioidean mitochondrial genomes (Fig. 5C). The general composition of the GC/AT ratio of the S. egretta sp. nov. mitogenome is 37.1/62.9%, PCGs 38.2/61.8%, rRNAs 34.4/65.6%, and tRNAs 36.6/63.4%, with negative GC skew (-0.028) and AT skew (-0.069) (Suppl. material 1: table S9). The AT skew and GC skew of 13 PCGs in S. egretta sp. nov. range from -0.126 (COX2) to −0.330 (ATP6) and 0.073 (ND2) to −0. 164 (ND5) (Suppl. material 1: table S9).

The total length of the 13 PCGs of S. egretta sp. nov. is 11,286 bp and begins with the typical mitogenome codon ATG, except for the ND4 gene which begins with GTG. All stop codons of the 13 PCGs were TAA/TAG. The large ribosomal RNA (16S rRNA) gene of S. egretta sp. nov. is 1,344 bp in size, with an A + T content of 66.8%, and its small ribosomal RNA (12S rRNA) is 890 bp, with an A + T content of 63.6% (Suppl. material 1: table S9). The two rRNA genes are located between trnL1 and trnV and trnT and trnS1. The 22 tRNAs have a size range of 63 bp (trnC) to 70 bp (trnN) and are 1476 bp in total length (Suppl. material 1: table S9).

The relatively synonymous codon usage (RSCU) values for the PCGs in the mitogenome of the S. egretta sp. nov. were analyzed and compared here with S. gottschei and S. coreana, and showed very high similarity (Supplementary File 7). The most frequently found codons of S. egretta sp. nov. include UCU(S), UUA(L), AAA(K), and GUU(V), whereas GCG(A), UCG(S), AAG(K), and ACG(T) have the lowest frequencies (Suppl. material 1: table S9).

Systematics

Semisulcospira egretta Y.-Z. Meng, sp. nov.

https://zoobank.org/A2840C7D-A9ED-48F0-B042-FA0D2DB21F76
Table 2, Figs 1, 6, 7, 8

Material examined

Holotype. China • Haicang District [海沧区], Xiamen City [厦门市], Fujian Province [福建省], China; 24°36'29"N, 117°53'39"E; 13 January 2024; Yuan-Zheng Meng leg.; MBSM-2024-XM-001.

Paratypes. China • 20; the same data as holotype; MBSM-2024-XM-002 to 004, XMU_DYY_TZS_001 to 002 and 005 to 017, YZM/1 (empty shell) • 16; the same locality as the holotype; 16 April 2022; Yuan-Zheng Meng and Li-Wen Lin leg.; XMU_DYY_XMA01 to A15, LWL/1 (empty shell).

Other material

China • 13; Tong’an District [同安区], Xiamen City [厦门市], Fujian Province [福建省], China; 24°52'35"N, 118°03'01"E; 26 February 2023; Yuan-Zheng Meng and Li-Wen Lin leg.; XMU_DYY_XMB01 to B04, XMU_DYY_XMC01 to C06; XMU_DYY_RTM05 and RTM25, YZM/1 (empty shell) • 3; the same locality as holotype; 16 April 2022; Yuan-Zheng Meng and Li-Wen Lin leg.; XMU_DYY_RTM05, XMU_DYY_RTM02, 04 to 05.

Diagnosis

Viviparous. Shell medium size, up to 25 mm, yellow-brownish, elongate, smooth, thin, comprising seven to nine whorls. Pregnant female bears about 150 embryos. Embryonic shell about 0.5 mm in length. Only known in Xiamen, Fujian Province, China.

Description

Shell (Fig. 6A–F, Table 2, n = 45). Shell height 10.7–24.1 mm, width 5.7–11.7 mm, body whorl height 7.1–16.6 mm, aperture width 3.2–6.2 mm, aperture height 4.8–12.2 mm. Shell elongate, conical, thin, brown to yellowish-brown, sometimes with 1–3 brown bands and black sediment, five to seven whorls. Shell surface smooth, without any spiral ridges or ribs. Apex always eroded. Aperture ovate. Measurements and counts are shown in Table 1.

Table 2.
Download as
CSV
XLSX

Shell measurements of Semisulcospira egretta sp. nov. (n = 45). The holotype is highlighted in red.

Voucher No. H/mm B/mm BW/mm WA/mm LA/mm Whorls B/H (%) BW/H (%) B/BW (%)
MBSM-2024-XM-001 23.0 10.7 15.7 5.2 11.0 6 46.5 68.3 68.2
MBSM-2024-XM-002 16.2 7.6 10.6 3.9 7.5 6 46.9 65.4 71.7
MBSM-2024-XM-003 18.4 9.8 13.1 4.8 9.6 6 53.3 71.2 74.8
MBSM-2024-XM-004 19.4 9.1 12.8 4.5 10.3 6 46.9 66 71.1
MBSM-2024-XM-005 16.2 7.9 11.1 3.8 8.7 6 48.8 68.5 71.2
XMU_DYY_TZS_001 18.8 10.0 12.4 5.1 9.5 7 53.2 66 80.6
XMU_DYY_TZS_002 24.1 11.1 16.6 5.8 12.2 6 46.1 68.9 66.9
XMU_DYY_TZS_005 19.6 9.4 13.8 4.9 10.1 5 48 70.4 68.1
XMU_DYY_TZS_006 17.5 9.5 12.7 4.2 9.2 6 54.3 72.6 74.8
XMU_DYY_TZS_007 15.9 8.7 11.6 4.3 9.6 6 54.7 73 75
XMU_DYY_TZS_008 18.4 8.9 12.7 4.3 9.6 6 48.4 69 70.1
XMU_DYY_TZS_009 21.1 10.2 14.1 4.7 10.5 5 48.3 66.8 72.3
XMU_DYY_TZS_010 20.7 10.1 14.7 4.7 11.0 6 48.8 71 68.7
XMU_DYY_TZS_011 21.4 9.3 14.6 4.7 10.7 6 43.5 68.2 63.7
XMU_DYY_TZS_012 19.9 9.0 13.4 4.4 9.7 5 45.2 67.3 67.2
XMU_DYY_TZS_013 15.6 8.0 10.9 4.1 8.7 6 51.3 69.9 73.4
XMU_DYY_TZS_014 14.0 7.3 10.3 3.5 7.7 5 52.1 73.6 70.9
XMU_DYY_TZS_015 18.7 8.9 13.0 4.4 9.5 6 47.6 69.5 68.5
XMU_DYY_TZS_016 20.1 8.7 13.4 4.3 9.5 6 43.3 66.7 64.9
XMU_DYY_TZS_017 18.2 9.1 12.5 4.3 9.3 6 50 68.7 72.8
XMU_DYY_XMA01 11.9 5.9 8.1 3.5 6.1 6 49.6 68.1 72.8
XMU_DYY_XMA02 11.7 5.7 7.7 3.7 5.9 6 48.7 65.8 74
XMU_DYY_XMA03 11.1 6.0 7.9 3.5 6.0 5 54.1 71.2 75.9
XMU_DYY_XMA04 10.7 6.2 7.1 4.0 5.9 5 57.9 66.4 87.3
XMU_DYY_XMA05 13.1 7.1 9.1 4.3 8.2 6 54.2 69.5 78
XMU_DYY_XMA06 14.0 6.9 9.9 4.4 7.6 6 49.3 70.7 69.7
XMU_DYY_XMA07 13.5 6.5 8.8 3.6 6.5 6 48.1 65.2 73.9
XMU_DYY_XMA08 14.5 7.0 10.0 4.7 7.4 5 48.3 69 70
XMU_DYY_XMA09 15.6 7.6 10.7 4.5 8.2 6 48.7 68.6 71
XMU_DYY_XMA10 15.6 7.6 10.9 4.9 8.3 5 48.7 69.9 69.7
XMU_DYY_XMA11 16.4 8.0 11.3 5.2 8.0 5 48.8 68.9 70.8
XMU_DYY_XMA12 16.6 8.0 11.3 5.2 8.0 5 48.2 68.1 70.8
XMU_DYY_XMA13 17.9 8.0 11.7 4.7 8.3 6 44.7 65.4 68.4
XMU_DYY_XMA14 20.3 9.2 12.8 5.7 9.4 5 45.3 63.1 71.9
XMU_DYY_XMA15 23.1 10.5 15.6 6.2 10.7 5 45.5 67.5 67.3
XMU_DYY_XMB01 11.8 6.0 8.2 3.5 4.8 6 50.8 69.5 73.2
XMU_DYY_XMB02 12.0 6.4 9.1 3.9 7.5 6 53.3 75.8 70.3
XMU_DYY_XMB03 13.4 6.7 9.7 3.4 7.2 6 50 72.4 69.1
XMU_DYY_XMB04 15.8 7.5 11.0 4.4 7.8 7 47.5 69.6 68.2
XMU_DYY_XMC01 14.3 6.2 8.7 3.6 6.6 7 43.4 60.8 71.3
XMU_DYY_XMC02 15.2 6.9 10.0 4.7 7.8 6 45.4 65.8 69
XMU_DYY_XMC03 16.0 6.8 10.2 4.1 6.9 7 42.5 63.8 66.7
XMU_DYY_XMC04 11.3 6.3 7.9 3.2 6.2 6 55.8 69.9 79.7
XMU_DYY_XMC05 14.9 7.2 10.5 3.7 7.6 7 48.3 70.5 68.6
XMU_DYY_XMC06 18.3 7.8 11.2 4.4 8.3 7 42.6 61.2 69.6
Statistical data of the shell measurements (n = 45)
minimum 10.7 5.7 7.1 3.2 4.8 5.0 42.5 60.8 63.7
maximum 24.1 11.1 16.6 6.2 12.2 7.0 57.9 75.8 87.3
average 16.6 8.0 11.3 4.4 8.4 5.9 48.8 68.4 71.4
standard deviation 3.46 1.46 2.32 0.67 1.65 0.63 3.68 3.05 4.26
Figure 6. 

Semisulcospira egretta sp. nov. and congeners. A-F, S. egretta sp. nov. A. Holotype, with operculum, XMU_DYY_TZS_003; B. Paratype 6, XMU_DYY_TZS_002; C. Paratype 34, XMU_DYY_XMA15; D. Paratype 32, XMU_DYY_XMA13; E. XMU_DYY_XMB01; F. XMU_DYY_XMC01; G–J. S. kurodai; G. ZMB 114713a; H. ZMB 114713b; I. ZMB 114754a; J. ZMB 114754b; K–M. S. libertina, syntypes of Melania japonica, NHMUK ZOO 20210249; N. S. pleuroceroides, lectotype of Melania pleurocerides, MNHN-IM-2000-21289; O. S. guilinensis, holotype, NNUH20240901; P. S. tigra, holotype, NNUH20240101; Q. S. sangpuensis, holotype, NNUH20241001. A–F. Photographed by Yuan-Zheng Meng; G–J. from Köhler (2016); O–Q. from He et al. (2024).

External morphology (Fig. 1, n = 20). Snout, neck, sides of foot black in color with golden spots on tentacles. Snout broad, squarish, with long cephalic tentacles, tentacles significantly longer than snout in length. Viviparous. Mantle edge smooth.

Operculum (Fig. 6A, n = 17). Corneous, ovate, brown, with approximately 3 whorls. Nucleus of operculum in lower one third of the operculum.

Radula (Fig. 7A, B, n = 9). Central teeth with one large central triangular cusp, and three to four conical denticles on each side, lateral teeth with single prominent triangular cusp, with two to three small denticles on each side, inner marginal teeth with four to five and outer with five to seven flattened, rounded denticles.

Figure 7. 

The radulae and embryonic shells of S. egretta sp. nov. A. The radula image of XMU_DYY_TZS_008; B. The radula image of XMU_DYY_TZS_010; C. Different views of embryonic shells. (Photographed by Y.-Z. Meng).

Midgut (Fig. 8A, n = 4). Oesophagus opening under ledge on the left side of midgut floor. Marginal fold extending anteriorly from oesophageal aperture alongside major typhlosole, then turning posteriorly bordering right margin of sorting area. Sorting area triangular, posterior tip curving left around sorting area pad. Midgut roof left of sorting area coarsely folded and cuticularised. Gastric shield tongue-shaped and concave. Glandular pad small, triangular, overhanging lip forming shallow pocket behind gastric shield and curving right. Crescentic ridge long, shallow, deep crescentic groove. Caecal fold along midgut floor opposite caecum. Style sac U-shaped, in contact with intestinal groove.

Figure 8. 

Anatomy of S. egretta sp. nov. A. Midgut anatomy. B. Female reproductive system anatomy. Abbreviations: bp, brood pouch; c, caecum; cf, caecal fold; cr, cresentic ridge; cu, cuticula; eg, egg; emb, embryos; ep, epithelial tissue; gp, glandular pad; gs, gastric shield; mf, marginal fold; ovi, renal oviduct; rcs, seminal receptacle; sa, sorting area; sap, sorting area pad; sg, sperm gutter; spb, spermatophore bursa; ss, style sac aperture; t1, major typhlosole. (Drawn by Yuan-Zheng Meng).

Reproductive systems

Female (Fig. 8B, n = 20). Long narrow oviduct, emerging from ovary, entering near seminal receptacle without protrusion. Ventral edge of spermatophore bursa with groove with curved surface forming sperm gutter, extending toward mantle cavity. Brood pouch uterine, on dorsal side of spermatophore bursa, inflated dorsally, separated into many cells, filled with eggs or embryos, depending on level of sexual development.

Male. No male was observed.

Sex ratio. All 20 dissected individuals were female.

Embryonic shell (Fig. 7C, D, n = 150). Globose, smooth, possessing slight spiral striae, usually comprising about 1.5 whorls. About 0.5 mm in height.

Habitat and distribution

(Fig. 1). The species was collected in mountain streams of Haicang District and Tong’an District of Xiamen, Fujian Province.

Etymology

The word egretta means “egret” in Latin. Egret is the city bird of Xiamen, which also suggests the type locality of this species. We suggest the Chinese common name as “白鹭短沟蜷”.

Comparative remarks

Many species of Semisulcospira have sculptured shells, for example, S. ningpoensis (I. Lea, 1857), S. gredleri (O. Boettger, 1886), and S. reticulata Kajiyama & T. Habe, 1961, etc. But Semisulcospira egretta sp. nov. can be easily distinguished from them by its thin and smooth shell. This species is also characterized by the relatively small embryonic shells (about 0.5 mm in length), and the denticle numbers on the inner and outer marginal teeth of its radula (four to five on the inner and five to seven on the outer). For detailed comparisons with similar species (also with relatively smooth shells refer) to Table 3. Semisulcospira egretta sp. nov. is the only semisulcospirid species documented in Xiamen; the only region from which it is currently known.

Table 3.
Download as
CSV
XLSX

Summary of differences between Semisulcospira egretta sp. nov. and some other congners. Details from He et al. (2024), Davis (1969), Ko et al. (2001), Du et al. (2019a), Du and Yang (2023).

Species Shell Whorls Length of Embryo /mm Embryo Whorl Numbers Embryos Number Inner Marginal Tooth Cusp Number Outer Marginal Tooth Cusp Number Distribution
S. calculus No sculpture, whorls convex. 5-7 ~0.4 – 6 6 6 North Korea, and Liaoning Province, China
S. egretta sp. nov. No sculpture, thin. 5-7 ~0.5 1.5 ~200 4-5 5-7 Xiamen, Fujian Province, China
S. elonga No sculpture. 5-7 – 3-4 24-37 6 6 Ningbo, Zhejiang Province, China
S. guilinensis Growth lines not obvious, thick. 8-9 – – – 4 6-7 Guilin, Guangxi Zhuang Autonomous Region, China
S. kurodai With spiral ribs on apical whorls, fragile. 4-6 < 1.62 < 3.0 18-66 – – Japan
S. libertina With spiral cords, thick. 4-5 < 1.35 < 3.0 137-979 5 6 Japan, Korea, China (doubtful)
S. pleuroceroides With spiral ribs. 7-14 – 2 117-120 4-6 4-5 The middle and lower reaches of the Yangtze River, China
S. sangpuensis Surface with some horizontal ribs, thick. 5 – – – 7-8 4-5 Shantou, Guangdong Province, China
S. tigra Growth lines not obvious, thick. 7-8 – – – 4 9-10 Liuzhou, Guangxi Zhuang Autonomous Region, China

Discussion

The taxonomic status and phylogeny of Semisulcospira egretta sp. nov

Our species hypothesis for Semisulcospira egretta sp. nov. is supported by both morphological and molecular data, as follows:

  1. S. egretta sp. nov. is placed in the genus Semisulcospira due to its viviparity, a synapomorphy of the genus (Glaubrecht 2006; Strong and Köhler 2009; Köhler 2017).
  2. Some taxonomic characters in S. egretta sp. nov., such as the thin, smooth shell (n = 45); embryonic shell size (about 0.5 mm; n = 150); and radular morphology (n = 9) exhibit relative stability and can distinguish it from other congeners. These characters (e.g. reproductive organ, shell, and radula) have been widely adopted by previous studies for classifying semisulcospirid species (Strong and Köhler 2009; Du et al. 2019a, 2019b; He et al. 2024).
  3. Intraspecific uncorrected p-distances were 0% to 3.1% (0.84% on average) in COI, 0% to 2.2% (0.72% on average) in 16S and 0% to 1.0% (0.56% on average) in ITS among sequences of S. egretta sp. nov. Although the two populations show a relatively large COI p-distance with each other (2.7% to 3.1%, 2.8% on average), the distances within the same population are very low respectively (0.077% of location A and 0.12% of location B). In most situations, the COI p-distance between congeners of Semisulcospiridae is more than 4% except for Hua aubryana, H. jacqueti and H. tchangsii (Du et al. 2019a). Given the ITS p-distance between the two populations is not more than 1%, and there is no significant morphological difference between the two populations, we regard specimens from locations A and B as a single species. The higher distances between the two populations are possibly due to geographical isolation.
  4. All phylogenetic trees based on mitochondrial data (16S+ COI, COI, 16S, COI-ex, ITS, and mitogenomes) show all S. egretta sp. nov. sequences forming a monophyletic group.

Despite evidence from morphology, DNA barcoding, and molecular phylogenetic analysis based on mitochondrial data supporting our proposed new species, many species still lack detailed morphological studies and molecular data, which impedes comprehensive comparisons across the genus, including those reported in China. Furthermore, the putative new species has not been tested using nuclear data. Therefore, further validation of the proposed new species is essential.

In China, our new species may be confused in the literature with S. libertina. This species has been documented in many regions of China, including Jilin, Liaoning, Zhejiang, Anhui, Jiangxi, Hubei, Hunan, Fujian, Taiwan, Guangdong, Guizhou, and Yunnan provinces since the last century (Xu 2008). However, these records lack detailed morphological studies and do not record the reproductive mode. Zeng et al. (2015) reported the complete mitogenome of S. libertina, but we have suggested it is a misidentification of a Koreoleptoxis species (this study, Xu et al. 2024). This makes us question the validity of all S. libertina records in China.

We noticed that many individuals marked as the same species (e.g. S. libertina or S. reiniana) did not form a monophyletic group in the phylogenetic tree (Fig. 3), and appeared in different clades of the tree. Species of Semisulcospira did not cluster into a monophyletic group, either. This result has also been observed in some previous studies, such as Lee et al. (2007), Köhler (2016, 2017), and Morita et al. (2024). Some different hypotheses were proposed to explain these phenomena: retention of ancestral polymorphisms, introgression, paternal leakage, heteroplasmy, recombination, pseudogenes, non-neutral mitochondrial evolution, and reversion of reproduction strategies, etc. (Köhler 2016, 2017; Du et al. 2019b). Köhler (2017) suggested that clade A and B should be omitted from the phylogenetic reconstruction of the Semisulcospiridae due to their unusual evolutionary rates. However, it is beyond the focus and indeed the capacity of this study to explain these problems.

Our phylogenetic trees based on short sequences (16S, COI, 16S+COI, COI-ex, and ITS) exhibit low resolution (Fig. 3, Suppl. materials 2–4, 6), making the phylogenetic relationships of Semisulcospira egretta sp. nov. unclear. Although the phylogenetic trees based on mitogenomes exhibit higher resolution, they do not indicate the exact phylogenetic position of S. egretta sp. nov. due to limited data (Fig. 5A). We suggest phylogenomic methods including transcriptomes, ultraconserved elements (UCEs), whole genomes, and restriction association site DNA (RAD), etc. with broader taxon sampling in studying the phylogeny of Semisulcospiridae, is needed.

Unresolved problems in the phylogenetics of Semisulcospira

The phylogenetic trees based on 16S and COI sequences in this study suggested that Semisulcospira is polyphyletic (Fig. 3). This result is generally consistent with previous studies (Köhler 2016, 2017; Du et al. 2019a, 2019b). However, these results are only based on a few genes, especially partial mitochondrial genes. It remains to be tested whether Semisulcospira is truly polyphyletic when a larger number of nuclear genes and a broader taxon sampling are included. In this study, we do not solve this problem directly but provide transcriptome data for future research.

Authors’ contributions

Conceptualization and project administration: D.-Y. Y. and Y.-B. X.; Data curation: Y.-Z. M., S. Zeng and D.-Y. Y.; Formal analysis: S. Zeng, D.-Y. Y and Y.-Z. M.; Funding acquisition: Y.-B. X; Investigation: Y.-Z. M.; Methodology: D.-Y. Y. and Y.-Z. M.; Supervision: D.-Y. Y., C. J. G. and X.-P. Z.; Visualization: S. Zeng; Writing (original draft): Y.-Z. M., D.-Y. Y., Y.-B. X and S. Zeng.; Writing (review & editing): All authors.

Acknowledgments

This study was supported by the Basic Scientific Research Project of Fujian Provincial Public Welfare Research Institutes (2022R1013001). The authors are grateful to Dr. Li-Na Du’s advice in dissecting operations and DNA amplification. We are also grateful to Dr. Zhi Wang and Ms. Ying Lu’s help with SEM operations. We also thank Mr. Li-Wen Lin, Ying-Lin Shi, and Ms. Cheng Chen’s help in collecting specimens, and Mr. Bing-Peng Xing from TIO for accepting our type specimens. We sincerely thank the reviewers and editor for their constructive comments on this article.

References

  • Capella-Gutiérrez S, Silla-Martínez JM, Gabaldón T (2009) trimAl: a tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics 25: 1972–1973. https://doi.org/10.1093/bioinformatics/btp348
  • Chen C, Chen H, Zhang Y, Thomas HR, Frank MH, He Y, Xia R (2020) TBtools: an integrative toolkit developed for interactive analyses of big biological data. Molecular Plant 13: 1194–1202. https://doi.org/10.1016/j.molp.2020.06.009
  • Chen H, Shi B-Y, Du L-N, Sun H-Y (2023) Description of a New Species of Hua (Gastropoda: Semisulcospiridae) from Guizhou, China, Based on Morphology and Molecular Evidence. Zoological Science 40: 414–421. https://doi.org/10.2108/zs230025
  • Chiu Y-W, Bor H, Kuo P-H, Hsu K-C, Tan M-S, Wang W-K, Lin H-D (2017) Origins of Semisulcospira libertina (Gastropoda: Semisulcospiridae) in Taiwan. Mitochondrial DNA Part A 28: 518–525. https://doi.org/10.3109/24701394.2016.1149823
  • Choi EH, Choi NR, Hwang UW (2021) The mitochondrial genome of an Endangered freshwater snail Koreoleptoxis nodifila (Caenogastropoda: Semisulcospiridae) from South Korea. Mitochondrial DNA Part B 6: 1120–1123. https://doi.org/10.1080/23802359.2021.1901626
  • Davis GM (1969) A taxonomic study of some species of Semisulcospira in Japn (Mesogastropoda: Pleuroceridae). Malacologia 7: 211–294.
  • Du L-N, Yang J-X (2023) Colored Atlas of Chinese Melania. Henan Science and Technology Press, Zhengzhou, 225 pp.
  • Du L-N, Köhler F, Yu G-H, Chen X-Y, Yang J-X (2019a) Comparative morpho-anatomy and mitochondrial phylogeny of Semisulcospiridae in Yunnan, south-western China, with description of four new species (Gastropoda: Cerithioidea). Invertebrate Systematics 33: 825–848. https://doi.org/10.1071/IS18084
  • Du LN, Chen J, Yu GH, Yang JX (2019b) Systematic relationships of Chinese freshwater semisulcospirids (Gastropods, Cerithioidea) revealed by mitochondria) sequences. Zoological Research 40: 541–551. https://doi.org/10.24272/j.issn.2095-8137.2019.033
  • Gim J-A, Baek K-W, Hah Y-S, Choo HJ, Kim J-S, Yoo J-I (2021) Draft genome of Semisulcospira libertina, a species of freshwater snail. Genomics & Informatics 19: Glaubrecht M (2006) Independent evolution of reproductive modes in viviparous freshwater Cerithioidea (Gastropoda, Sorbeoconcha)-a brief review. Basteria 70: 23–28. https://doi.org/10.5808/gi.21039
  • Glaubrecht M (2006) Independent evolution of reproductive modes in viviparous freshwater Cerithioidea (Gastropoda, Sorbeoconcha)-a brief review. Basteria 70: 23–28.
  • Guo Y, Fu Z, Feng J, Ye Y, Li J, Guo B, Lv Z (2019) The complete mitochondrial genome and phylogenetic analysis of Epitonium scalare (Gastropoda, Epitoniidae). Mitochondrial DNA Part B 4: 1070–1071. https://doi.org/10.1080/23802359.2019.1586462
  • Hartnell College Genomics Group Andrade P, Arreola L, Belnas M, Bland E, Castillo A, Cisneros O, Contreras V, Diaz C, Do KT (2019) The complete mitogenome of the invasive Japanese mud snail Batillaria attramentaria (Gastropoda: Batillariidae) from Elkhorn Slough, California, USA. Mitochondrial DNA Part B 4: 4031–4032. https://doi.org/10.1080/23802359.2019.1688719
  • He YM, Lu YZ, Fu ZY, Xiang HQ, Chen H (2024) Description of 17 new species of Semisulcospiridae (Gastropoda: Cerithioidea) from southern China based on morphological and molecular evidence. Ecologica Montenegrina 75: 12–32. https://doi.org/10.37828/em.2024.75.2
  • Hilgers L, Grau JH, Pfaender J, von Rintelen T (2016) The complete mitochondrial genome of the viviparous freshwater snail Tylomelania sarasinorum (Caenogastropoda: Cerithioidea). Mitochondrial DNA Part B 1: 330–331. https://doi.org/10.1080/23802359.2016.1172046
  • Jin J-J, Yu W-B, Yang J-B, Song Y, DePamphilis CW, Yi T-S, Li D-Z (2020) GetOrganelle: a fast and versatile toolkit for accurate de novo assembly of organelle genomes. Genome Biology 21: 1–31. https://doi.org/10.1186/s13059-020-02154-5
  • Jombart T, Kendall M, Almagro‐Garcia J, Colijn C (2017) treespace: Statistical exploration of landscapes of phylogenetic trees. Molecular Ecology Resources 17: 1385–1392. https://doi.org/10.1111/1755-0998.12676
  • Kalyaanamoorthy S, Minh BQ, Wong TK, Von Haeseler A, Jermiin LS (2017) ModelFinder: fast model selection for accurate phylogenetic estimates. Nature Methods 14: 587–589. https://doi.org/10.1038/nmeth.4285
  • Kato S, Itoh H, Fukumori H, Nakajima N, Kanaya G, Kojima S (2022) The mitochondrial genome of the threatened tideland snail Pirenella pupiformis (Mollusca: Caenogastropoda: Potamididae) determined by shotgun sequencing. Mitochondrial DNA Part B 7: 632–634. https://doi.org/10.1080/23802359.2022.2060143
  • Katoh K, Standley DM (2013) MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Molecular Biology and Evolution 30: 772–780. https://doi.org/10.1093/molbev/mst010
  • Ko J, Lee J, Kwon O (2001) Study on radulae of seven species of the Family Pleuroceridae in Korea. The Korean Journal of Malacology 17: 105–115.
  • Köhler F (2016) Rampant taxonomic incongruence in a mitochondrial phylogeny of Semisulcospira freshwater snails from Japan (Cerithioidea: Semisulcospiridae). Journal of Molluscan Studies 82: 268–281. https://doi.org/10.1093/mollus/eyv057
  • Köhler F (2017) Against the odds of unusual mtDNA inheritance, introgressive hybridisation and phenotypic plasticity: systematic revision of Korean freshwater gastropods (Semisulcospiridae, Cerithioidea). Invertebrate Systematics 31: 249–268. https://doi.org/10.1071/IS16077
  • Kumar S, Stecher G, Li M, Knyaz C, Tamura K (2018) MEGA X: molecular evolutionary genetics analysis across computing platforms. Molecular Biology and Evolution 35: 1547. https://doi.org/10.1093/molbev/msy096
  • Lee SY, Lee HJ, Kim YK (2019) Comparative analysis of complete mitochondrial genomes with Cerithioidea and molecular phylogeny of the freshwater snail, Semisulcospira gottschei (Caenogastropoda, Cerithioidea). International Journal of Biological Macromolecules 135: 1193–1201. https://doi.org/10.1016/j.ijbiomac.2019.06.036
  • Lee W-K, Hou BK, Ju S-J, Kim S-J (2020) Complete mitochondrial genome of the hydrothermal vent provannid snail Alviniconcha boucheti (Gastropoda: Abyssochrysoidea) from the North Fiji Basin. Mitochondrial DNA Part B 5: 1848–1849. https://doi.org/10.1080/23802359.2020.1750983
  • Lee T, Hong HC, Kim JJ, Foighil DÓ (2007) Phylogenetic and taxonomic incongruence involving nuclear and mitochondrial markers in Korean populations of the freshwater snail genus Semisulcospira (Cerithioidea: Pleuroceridae). Molecular Phylogenetics and Evolution 43: 386–397. https://doi.org/10.1016/j.ympev.2007.02.018
  • Letunic I, Bork P (2024) Interactive Tree of Life (iTOL) v6: recent updates to the phylogenetic tree display and annotation tool. Nucleic Acids Research 52(W1): W78–W82. https://doi.org/10.1093/nar/gkae268
  • Ling Y, Zhang Y, Ngatia JN, Zhou H (2022) The complete mitochondrial genome of Melanoides tuberculata (Müller, 1774) in Guangdong, China. Mitochondrial DNA Part B 7: 1319–1320. https://doi.org/10.1080/23802359.2022.2054735
  • Liu Y, Zhang W, Wang Y, Wang E (1979) Economic Fauna of China: Freshwater Mollusca. Science Press, Beijing, 134 pp.
  • Liu Y-Y, Zhang W-Z, Wang Y-X, Duan Y-H (1994) Eight New Species of Freshwater Molluscs in Southwest China. Acta Zootaxonomica Sinica 19: 25–36.
  • Miura O, Urabe M, Mori H, Chiba S (2020) Ancient drainage networks mediated a large‐scale genetic introgression in the East Asian freshwater snails. Ecology and Evolution 10: 8186–8196. https://doi.org/10.1002/ece3.6523
  • Morita K, Saito T, Uechi T, Sawada N, Miura O (2024) Out of the ancient lake: Multiple riverine colonizations and diversification of the freshwater snails in the genus Semisulcospira around Lake Biwa. Molecular Phylogenetics and Evolution 191: 107987. https://doi.org/10.1016/j.ympev.2023.107987
  • Nguyen L-T, Schmidt HA, Von Haeseler A, Minh BQ (2015) IQ-TREE: a fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Molecular Biology and Evolution 32: 268–274. https://doi.org/10.1093/molbev/msu300
  • Nguyen DH, Lemieux C, Turmel M, Nguyen VD, Mouget J-L, Witkowski A, Tremblay R, Gastineau R (2018) Complete mitogenome of Cerithidea obtusa, the red chut-chut snail from the Cần Giờ Mangrove in Vietnam. Mitochondrial DNA Part B 3: 1267–1269. https://doi.org/10.1080/23802359.2018.1532832
  • R Core Team (2023) R: A language and environment for statistical computing. In: R Foundation for Statistical Computing (Ed.) Vienna, Austria.
  • Ronquist F, Teslenko M, Van Der Mark P, Ayres DL, Darling A, Höhna S, Larget B, Liu L, Suchard MA, Huelsenbeck JP (2012) MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Systematic Biology 61: 539–542. https://doi.org/10.1093/sysbio/sys029
  • Sawada N, Nakano T (2021) Revisiting a 135-year-old taxonomic account of the freshwater snail Semisulcospira multigranosa: designating its lectotype and describing a new species of the genus (Mollusca: Gastropoda: Semisulcospiridae). Zoological Studies 60: e7.
  • Sawada N, Fuke Y, Miura O, Toyohara H, Nakano T (2024) Redescription of Semisulcospira reticulata (Mollusca, Semisulcospiridae) with description of a new species from Lake Biwa, Japan. Evolutionary Systematics 8: 127–144. https://doi.org/10.3897/evolsyst.8.124491
  • Steenwyk JL, Buida III TJ, Gonçalves C, Goltz DC, Morales G, Mead ME, LaBella AL, Chavez CM, Schmitz JE, Hadjifrangiskou M (2022) BioKIT: a versatile toolkit for processing and analyzing diverse types of sequence data. Genetics 221: iyac079. https://doi.org/10.1093/genetics/iyac079
  • Stelbrink B, Kehlmaier C, Wilke T, Albrecht C (2019) The near-complete mitogenome of the critically endangered Pseudocleopatra dartevellei (Caenogastropoda: Paludomidae) from the Congo River assembled from historical museum material. Mitochondrial DNA Part B 4: 3229–3231. https://doi.org/10.1080/23802359.2019.1669081
  • Strong EE, Köhler F (2009) Morphological and molecular analysis of ‘Melania’ jacqueti Dautzenberg & Fischer, 1906: from anonymous orphan to critical basal offshoot of the Semisulcospiridae (Gastropoda: Cerithioidea). Zoologica Scripta 38: 483–502. https://doi.org/10.1111/j.1463-6409.2008.00385.x
  • Strong EE, Garner JT, Johnson PD, Whelan NV (2022) A systematic revision of the genus Juga from fresh waters of the Pacific Northwest, USA (Cerithioidea, Semisulcospiridae). European Journal of Taxonomy 848: 1–97. https://doi.org/10.5852/ejt.2022.848.1993
  • Whelan NV, Strong EE (2016) Morphology, molecules and taxonomy: extreme incongruence in pleurocerids (Gastropoda, Cerithioidea, Pleuroceridae). Zoologica Scripta 45: 62–87. https://doi.org/10.1111/zsc.12139
  • Xu X (2008) Classification and Morphology on Pleuroceridae (Gastropoda: Mesogastropoda) of Some Areas in Middle Reaches of Changjiang River. MA. Sc Thesis, Nanchang, Jiangxi Province, China: Nanchang University, 79 pp.
  • Xu Y, Luo P, Wang P, Zhu P, Zhang H, Wu H, Liao Y, Yu M, Fu J (2019) The complete mitochondrial genome of Cerithidea sinensis (Philippi, 1848). Mitochondrial DNA Part B 4: 2742–2743. https://doi.org/10.1080/23802359.2019.1644549
  • Xu Y, Zeng S, Meng Y, Yang D, Yang S (2024) The mitochondrial genome of Hua aristarchorum (Heude, 1889)(Gastropoda, Cerithioidea, Semisulcospiridae) and its phylogenetic implications. ZooKeys 1192: 237–255. https://doi.org/10.3897/zookeys.1192.116269
  • Yan C, Feng J, Ye Y, Li J, Guo B (2020a) The complete mitochondrial genome and phylogenetic analysis of Batillaria cumingi (Gastropoda: Batillariidae). Mitochondrial DNA Part B 5: 2355–2356. https://doi.org/10.1080/23802359.2020.1772685
  • Yan C, Feng J, Ye Y, Li J, Guo B (2020b) The complete mitochondrial genome and phylogenetic analysis of Batillaria zonalis (Gastropoda: Batillariidae). Mitochondrial DNA Part B 5: 2256–2257. https://doi.org/10.1080/23802359.2020.1772139
  • Yang D, Zeng S, Wang Z, Zhang Y, Yang D, Glasby CJ, Hwang JS, Cai L (2024) Molecular systematics of Perinereis and an investigation of the status and relationships of the cultured species Perinereis wilsoni Glasby & Hsieh, 2006 (Annelida, Nereididae). Zoosystematics and Evolution 100: 1297–1314. https://doi.org/10.3897/zse.100.127201
  • Yin N, Zhao S, Huang X-C, Ouyang S, Wu X-P (2022) Complete mitochondrial genome of the freshwater snail Tarebia granifera (Lamarck, 1816)(Gastropoda: Cerithioidea: Thiaridae). Mitochondrial DNA Part B 7: 259–261. https://doi.org/10.1080/23802359.2022.2026832
  • Zeng T, Yin W, Xia R, Fu C, Jin B (2015) Complete mitochondrial genome of a freshwater snail, Semisulcospira libertina (Cerithioidea: Semisulcospiridae). Mitochondrial DNA 26: 897–898. https://doi.org/10.3109/19401736.2013.861449
  • Zhang D, Gao F, Jakovlić I, Zou H, Zhang J, Li WX, Wang GT (2020) PhyloSuite: An integrated and scalable desktop platform for streamlined molecular sequence data management and evolutionary phylogenetics studies. Molecular Ecology Resources 20: 348–355. https://doi.org/10.1111/1755-0998.13096

1 Yi-Bin Xu, Yuan-Zheng Meng and Sheng Zeng contributed equally to this work.

Supplementary materials

Supplementary material 1 

Additional tables

Yi-Bin Xu, Yuan-Zheng Meng, Sheng Zeng, Hang-Jun Wang, Shen Zhong, De-Yuan Yang, Xi-Ping Zhou, Christopher J. Glasby

Data type: xlsx

Explanation note: table S1. Specimen information of Semisulcospira egretta sp. nov. table S2. Protocols for extracting DNA sequences. table S3. Single gene dataset used for phylogenetic analysis. table S4. The COI gene dataset used for phylogenetic analysis in fig. S1. table S5. Mitochondrial genome dataset used for phylogenetic analysis. table S6. Original and trimAl lengths of the PCGs and PCGsAA sequences. table S7. Best partitioning schemes and models based on different datasets for Bayesian inference (BI) and maximum likelihood (ML) analysis. table S8. Pairwise distances and Kimura 2 Parameters between sequences of Clade D in Fig. 3. table S9. Mitochondrial genome analysis for mitogenomes of Semisulcospiridae.

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
Download file (170.46 kb)
Supplementary material 2 

Phylogenetic tree inferred from 16S and COI sequences, based on different trimming methods

Yi-Bin Xu, Yuan-Zheng Meng, Sheng Zeng, Hang-Jun Wang, Shen Zhong, De-Yuan Yang, Xi-Ping Zhou, Christopher J. Glasby

Data type: pdf

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
Download file (1.70 MB)
Supplementary material 3 

Phylogenetic tree inferred from 16S sequences, based on different trimming methods

Yi-Bin Xu, Yuan-Zheng Meng, Sheng Zeng, Hang-Jun Wang, Shen Zhong, De-Yuan Yang, Xi-Ping Zhou, Christopher J. Glasby

Data type: pdf

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
Download file (1.70 MB)
Supplementary material 4 

Phylogenetic tree inferred from COI sequences, based on different trimming methods

Yi-Bin Xu, Yuan-Zheng Meng, Sheng Zeng, Hang-Jun Wang, Shen Zhong, De-Yuan Yang, Xi-Ping Zhou, Christopher J. Glasby

Data type: pdf

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
Download file (1.68 MB)
Supplementary material 5 

The topological differences of 16S+COI, 16S, and COI datasets generated by TreeSpace

Yi-Bin Xu, Yuan-Zheng Meng, Sheng Zeng, Hang-Jun Wang, Shen Zhong, De-Yuan Yang, Xi-Ping Zhou, Christopher J. Glasby

Data type: pdf

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
Download file (240.24 kb)
Supplementary material 6 

Maximum Likelihood tree inferred from extensive COI sequences (COI-ex)

Yi-Bin Xu, Yuan-Zheng Meng, Sheng Zeng, Hang-Jun Wang, Shen Zhong, De-Yuan Yang, Xi-Ping Zhou, Christopher J. Glasby

Data type: pdf

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
Download file (744.38 kb)
Supplementary material 7 

Codon usage analysis in S. egretta sp. nov. and congeners

Yi-Bin Xu, Yuan-Zheng Meng, Sheng Zeng, Hang-Jun Wang, Shen Zhong, De-Yuan Yang, Xi-Ping Zhou, Christopher J. Glasby

Data type: pdf

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
Download file (2.28 MB)
login to comment