Research Article |
Corresponding author: Yuchun Li ( li_yuchun@sdu.edu.cn ) Academic editor: Jesus Maldonado
© 2019 Yaoyao Li, Haotian Li, Masaharu Motokawa, Yi Wu, Masashi Harada, Huimei Sun, Xinmin Mo, Jing Wang, Yuchun Li.
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:
Li Y, Li H, Motokawa M, Wu Y, Harada M, Sun H, Mo X, Wang J, Li Y (2019) A revision of the geographical distributions of the shrews Crocidura tanakae and C. attenuata based on genetic species identification in the mainland of China. ZooKeys 869: 147-160. https://doi.org/10.3897/zookeys.869.33858
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The Taiwanese gray shrew (Crocidura tanakae) and Asian gray shrew (C. attenuata) are so similar in size and morphology that the taxonomic status of the former has changed several times since its description; C. tanakae has also been regarded as an endemic species of Taiwan Island. In recent years, molecular identification has led to several reports of C. tanakae being distributed in the mainland of China. In this study, we determine the geographical distribution of C. attenuata and C. tanakae based on more than one hundred specimens collected during 2000 to 2018 over a wide area covering the traditional ranges of the two species in the mainland of China, and show a substantial revision of their distributions. Among 110 individuals, 33 C. attenuata and 77 C. tanakae were identified by Cytb gene and morphologies. Our results show, (1) C. attenuata and C. tanakae are distributed sympatrically in the mainland of China; (2) contrary to the previous reports, the distribution range of C. attenuata is restricted and much smaller than that of C. tanakae in the mainland of China; (3) Hainan Island, like Taiwan Island, is inhabited by C. tanakae only according to the present data.
Crocidura attenuata, Crocidura tanakae, geographical distribution, mainland of China, Taiwan Island
The Taiwanese gray shrew (Crocidura tanakae Kuroda, 1938) and Asian gray shrew (C. attenuata Milne Edwards, 1872) are distinct species with very similar morphological characters and measurements, such that the taxonomic status of C. tanakae has been changed several times by taxonomists. Crocidura tanakae was originally described from Taiwan as a new species by
In recent years, the application of molecular identification techniques led to reports of C. tanakae populating the mainland of China.
We accumulated more than one hundred specimens from 19 areas of C. attenuata and C. tanakae in our field surveys in the mainland of China from 2000 to 2018, which expands the previous distributions from the aforementioned reports from a few localities. A re-evaluation of geographical distributions of the two species is important to a range of studies and practical needs, such as zoogeography, geophylogeny, agriculture animal management, health and epidemic prevention. Here we report the wide geographical distributions of C. attenuata and C. tanakae in the mainland of China.
A total of 110 specimens of the C. attenuata complex were collected from 11 provinces in the mainland of China including Anhui (2 specimens), Fujian (4), Guangdong (23), Guangxi (4), Hubei (4), Hunan (3), Jiangxi (13), Sichuan (Chongqing is included) (30), Yunnan (1) and Zhejiang (23) as well as the Hainan Island (2) using Sherman live cages during 18 years from Aug. 2000 to Sep. 2018. The geographical position (longitude and latitude) of each specimen was recorded by GPS (Suppl. material
DNA was extracted from muscle samples using the Ezup Column Animal Genomic DNA Purification Kit (Sangon Biotech Co., Ltd., Shanghai, China). The complete mitochondrial cytochrome b gene (Cytb, 1140 bp) was amplified by PCR with the primers as in
Cytb gene sequences were aligned using BioEdit v.7.2.5 (
We also included Cytb sequence data from several earlier studies (
In order to attribute these genetic lineages to taxonomically correct species names, we photographed the dorsal, ventral, lateral of skull and lateral view of the mandible of C. attenuata from type locality – Baoxing (Moupin), Sichuan – and also photographed the corresponding teeth, and marked the characteristic features on the pictures for this species. We repeated the same procedure with the only sample of C. tanakae from the same locality (Baoxing) for interspecific comparisons.
We conducted a morphological investigation of the specimens sampled to identify the two species by determining three external measurements: total body length (TBL), head and body length (HBL), ear length (EL); and 10 skull measurements: greatest length of skull (GLS), cranial base length (GBL), median palatal length (MPL), length of teeth row (LUTR), greatest palatal breadth (GPB), breadth of occipital condyles (BOC), greatest breath of braincase (BBC), interorbital breadth (IOB), height of the braincase (HB), length of mandible (LM) according to
We calculated the mean and standard deviation of external and skull morphological indices. The pairwise differences between the two species were tested by independent sample t-tests or Mann-Whitney U tests according to results of the Kolmogorov-Smirnov test for their normality of distribution. Principal component analysis (PCA) was used to test the general appropriateness of the groupings supplied by assessment of overall variation in the skull characters. These analyses were performed using SPSS Statistics 24.0 (SPSS, Chicago, IL, USA).
We obtained 1140 bp of mitochondrial DNA sequences from 110 individuals in this study. The ML tree indicated that the specimens we collected were divided into two lineages, one was clustered with the C. attenuata download from GenBank which was distributed in its type locality, i.e., Baoxing of Sichuan Province, and the other was clustered with the C. tanakae download from GenBank which was exclusively distributed in its type locality, i.e., Taiwan Island (Fig.
Distributions of Crocidura attenuata and C. tanakae in the mainland of China,Laos and Vietnam. Black and white triangles represent the sampled sites of C. tanakae first presented in this study and in previous studies, respectively. White circles and squares represent the sampled sites of C. attenuata first presented in this study and in previous studies. Black triangles and white circles overlapped indicate sympatry sites.
By investigating our samples of C. attenuata lineage from Baoxing, Sichuan, we found some morphological features correlated with the holotype: the superior articular facets are more angular in dorsal view and the basioccipital region is narrow and ridged particularly anterior to the position of the basioccipital suture in C. attenuata (Fig.
Comparison of crania of Crocidura attenuata (S2576) and Crocidura tanakae (S2566) from Baoxing, Sichuan. Top row from left to right: dorsal views of the skulls of C. attenuata and C. tanakae (S2576 andS2566), ventral views of the skulls in the same order. Lower row: lateral view of skulls and mandibles from top to bottom of C. attenuata and C. tanakae (S2576 and S2566).
A total of 90 adult individuals were screened by age identification including 26 C. attenuata and 64 C. tanakae. The external and two skull measurements (BOC and GPB) were judged as a non-normal distribution by the Kolmogorov-Smirnov test (P<0.05), so we used the Mann-Whitney U Test for interspecific comparisons; for the others with normal distribution (P>0.05) the parametric independent sample t-test was used (Suppl. material
Scatter plot of C. attenuata (red) and C. tanakae (blue) sample distribution over PC1 and PC2 axes constructed based on external and skull morphological variables. Different symbols represent different populations. GD: Guangdong, FJ: Fujian, ZJ: Zhejiang, SCBX: Baoxing, Sichuan, HB: Hubei, SC: Sichuan, GX: Guangxi, AH: Anhui, HuN: Hunan, HN: Hainan, CQ: Chongqing, YN: Yunnan, JX: Jiangxi
External and cranial measurements of C. attenuata and C. tanakae (in millimeters). Mean ± standard deviation (SD), and range, with number of specimens in parentheses are presented for characters available.
Morphological indices | Crocidura attenuata | Crocidura tanakae | ||||
This study |
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This study |
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Total Body Length (TBL) | 133.92±6.22 | – | 132.95±6.84 | – | – | – |
120.00–144.00 (26) | – | 115.00–148.00 (64) | – | – | – | |
Head and Body Length (HBL) | 77.96±3.54 | 71.40±6.64 | 78.90±5.42 | 66.00 | 77.90±3.60 | 73.36±6.67 |
71.00–87.00 (26) | 63.00–82.00 (10) | 70.00–95.00 (64) | – | 69.70–86.00 | 64.5–84.5 | |
Ear Length (EL) | 8.32±1.05 | – | 8.79±1.00 | – | 8.96±0.39 | 9.05±0.91 |
6.05–10.16 (26) | 10.00 (2) | 5.97–11.06 (63) | – | 8.10–9.80 | 7.9–10.2 | |
Greatest Length of Skull (GLS) | 20.99±0.59 | 20.55±0.66 | 20.54±0.61 | 20.00 | 20.84±0.42 | 20.85±0.41 |
20.12–22.36 (23) | 19.60–21.70 (9) | 19.23–21.69 (61) | – | 20.03–21.82 | 19.94–21.31 | |
Cranial base Length (GBL) | 20.91±0.57 | – | 20.39±0.61 | – | 19.88±0.46 | – |
20.04–22.26 (24) | – | 18.85–21.68 (61) | – | 19.06–20.73 | – | |
Median palatal Length (MPL) | 9.07±0.41 | – | 8.89±0.31 | – | – | – |
8.35–10.13 (26) | – | 8.14–9.61 (63) | – | – | – | |
Length of tooth row (LUTR) | 9.36±0.26 | 8.72±0.38 | 9.16±0.34 | 9.00 | 9.29±0.16 | 9.33±0.19 |
8.97–9.96 (26) | 8.20–9.50 (11) | 8.19–9.91 (63) | – | 8.73–9.62 | 8.87–9.54 | |
Greatest palatal breadth (GPB) | 6.25±0.16 | 6.09±0.22 | 6.32±0.31 | – | 6.33±0.16 | 6.41±0.13 |
5.96–6.55 (26) | 5.80–6.50 (11) | 5.65–6.97 (64) | – | 5.98–6.68 | 6.20–6.58 | |
Breadth of Occipital Condyles (BOC) | 5.20±0.19 | – | 5.13±0.23 | – | – | – |
4.78–5.55 (22) | – | 4.48–5.64 (60) | – | – | – | |
Greatest Breath of Braincase (BBC) | 9.59±0.27 | 9.17±0.24 | 9.26±0.32 | – | 9.23±0.18 | 9.22±0.22 |
8.98–10.25 (26) | 8.70–9.40 (8) | 8.57–10.02 (64) | – | 8.00–9.66 | 8.87–9.50 | |
Interorbital Breadth (IOB) | 3.97±0.23 | 4.43±0.18 | 3.79±0.17 | 4.50 | 4.50±0.10 | 4.62±0.10 |
3.61–4.46 (26) | 4.10–4.70 (9) | 3.43–4.23 (64) | – | 4.29–4.68 | 4.47–4.74 | |
Height of the Braincase (HB) | 5.13±0.14 | 4.91±0.10 | 5.01±0.15 | – | – | – |
4.80–5.42 (26) | 4.80–5.10 (8) | 4.65–5.35 (64) | – | – | – | |
Length of mandible (LM) | 10.01±0.28 | 12.76±0.62 | 9.85±0.33 | – | 8.38±0.20 | – |
9.34–10.67 (26) | 11.7–13.9 (11) | 9.06–10.52 (64) | – | 7.88–8.91 | – |
Morphological comparisons and significant difference between C. attenuata and C. tanakae in this study. Values in bold show significant differences.
Morphological indices | T test/Mann-Whitney U Test | |||||
F | Sig. | t | Df (N) | Z | P | |
Total Body Length (TBL) | (90) | -0.687 | 0.492 | |||
Head and Body Length (HBL) | (90) | -0.478 | 0.633 | |||
Ear Length (EL) | (89) | -2.08 | 0.038 | |||
Greatest Length of Skull (GLS) | 0.149 | 0.701 | -3.045 | 82 | 0.003 | |
Cranial base Length (GBL) | 0.133 | 0.717 | -3.541 | 83 | 0.001 | |
Median palatal Length (MPL) | 1.214 | 0.274 | -2.263 | 87 | 0.026 | |
Length of teeth row (LUTR) | 1.174 | 0.282 | -2.615 | 87 | 0.011 | |
Greatest palatal breadth (GPB) | (90) | -1.077 | 0.281 | |||
Breadth of Occipital Condyles (BOC) | (82) | -1.256 | 0.209 | |||
Greatest Breath of Braincase (BBC) | 1.416 | 0.237 | -4.666 | 88 | <0.001 | |
Interorbital Breadth (IOB) | 2.492 | 0.118 | -4.143 | 88 | <0.001 | |
Height of the Braincase (HB) | 1.131 | 0.29 | -3.61 | 88 | 0.001 | |
Length of mandible (LM) | 2.766 | 0.1 | 2.195 | 88 | 0.031 |
Principal component loadings as performed by analyses of 13 morphological measurements of C. attenuata and C. tanakae.
Variable | Component | ||
PC1 | PC2 | PC3 | |
Total Body Length (TBL) | 0.691 | 0.550 | 0.065 |
Head and Body Length (HBL) | 0.435 | 0.607 | 0.458 |
Ear Length (EL) | 0.101 | 0.706 | -0.314 |
Greatest Length of Skull (GLS) | 0.948 | -0.088 | -0.062 |
Cranial base Length (GBL) | 0.966 | -0.096 | -0.055 |
Median palatal Length (MPL) | 0.815 | 0.041 | -0.372 |
Length of teeth row (LUTR) | 0.867 | -0.139 | 0.017 |
Greatest palatal breadth (GPB) | 0.566 | 0.138 | 0.458 |
Breadth of Occipital Condyles (BOC) | 0.545 | -0.358 | 0.561 |
Greatest Breath of Braincase (BBC) | 0.837 | -0.140 | -0.022 |
Interorbital Breadth (IOB) | 0.470 | 0.035 | -0.368 |
Height of the Braincase (HB) | 0.590 | -0.223 | -0.252 |
Length of mandible (LM) | 0.820 | -0.126 | -0.045 |
% of total variance explained | 49.876 | 11.087 | 9.110 |
Eigenvalue | 6.484 | 1.441 | 1.184 |
Among the localities of our field surveys, C. tanakae was recorded at almost all sites investigated (Fig.
This study indicates that C. attenuata and C. tanakae are sympatrically distributed not only in continental Indochina (
Note that the map of C. attenuata (Fig.
Based on morphological features we found among our samples and the results of its comparisons with type materials of C. attenuata and C. tanakae (
There are many research reports listing C. attenuata in the mainland of China. For example,
This study is funded by the National Natural Science Foundation of China (NSFC, 31672254, 31110103910). We thank students from Shandong University and Guangzhou University for their participation in field surveys.
Tables S1–S3
Data type: species data
Explanation note: Table S1. The information of Crocidura tanakae and C. attenuata specimens used in present study. Table S2. The specimen information of genus Crocidura and outgroup used in the present study. Table S3. Normality test of external and skull morphological indices of C. attenuata and C. tanakae.