Research Article |
Corresponding author: Tao Luo ( 18786413778@163.com ) Academic editor: Nina Bogutskaya
© 2023 Tao Luo, Zhi-Xia Chen, Xin-Rui Zhao, Jing Yu, Chang-Ting Lan, Jia-Jun Zhou, Ning Xiao, Jiang Zhou.
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:
Luo T, Chen Z-X, Zhao X-R, Yu J, Lan C-T, Zhou J-J, Xiao N, Zhou J (2023) Balitora anlongensis, the first cavefish species of the genus Balitora (Teleostei, Balitoridae) from Guizhou Province, southwest China. ZooKeys 1185: 21-42. https://doi.org/10.3897/zookeys.1185.108545
|
This work describes a new species, Balitora anlongensis sp. nov., collected from a cave at Xinglong Town, Anlong County, Guzihou, China. Phylogenetic trees reconstructed based on two mitochondrial and three nuclear genes show that the new species represents an independent evolutionary lineage with large genetic differences, 7.1%–12.0% in mitochondrial gene cytochrome b and 9.2%–12.1% in cytochrome oxidase subunit 1, from congeners. Morphologically, the new species can be distinguished from the 18 species currently assigned to the genus Balitora by a combination of characters, most clearly by having two pairs of maxillary barbels; 8½ branched dorsal-fin rays; 5½ branched anal-fin rays; pectoral fin not reaching pelvic fin origin; dorsal-fin origin in front of pelvic fin origin; eye small (eye diameter approximately equal to outer maxillary barbel length); and fins lacking pigment in live fish. The new species represents the first record of Balitora inhabiting caves in China and increases the number of species in the genus Balitora in its present concept from 18 to 19. The study suggests that more evidence is needed to further clarify the taxonomic composition of the genus Balitora.
Nanpanjiang River, stone loach, taxonomy, phylogeny
The karst region of southwest China is well known as a distinctive center and hotspot of biodiversity (
The genus Balitora, Gray, 1830 was established with Balitora brucei as the type species, originally placed in the Cobitidae (Gray, 1830), and is now placed within the Balitoridae (
Few molecular markers have been used to assess the phylogeny of the genus Balitora. A phylogenetic tree reconstructed by
To date, the classification of the genus remains controversial, mainly because of the lack of clear phylogenetic relationships and stable morphological characters among Balitora, Hemimyzon, and Sinohomaloptera (Table
Taxonomic revision history of 19 species of the genus Balitora distributed in Asia. Species in bold are those which have a distribution in China.
ID # | Species |
|
|
|
|
Zhang and Zhao 2016 |
|
Zhang and Cao 2021 | This study |
---|---|---|---|---|---|---|---|---|---|
1 | Balitora annamitica Kottelat, 1988 | – | – | Balitora | – | – | Balitora | – | Balitora |
2 | Balitora brucei Gray, 1830 | – | – | Balitora | Balitora | – | Balitora | Balitora | Balitora |
3 | Balitora burmanica Hora, 1932 | – | – | Balitora | Balitora | – | Balitora | Balitora | Balitora |
4 | Balitora chipkali Kumkar, Katwate, Raghavan & Dahanukar, 2016 | – | – | – | – | – | Balitora | – | Balitora |
5 | Balitora eddsi Conway & Mayden, 2010 | – | – | Balitora | – | – | Balitora | – | Balitora |
6 | Balitora elongata Chen & Li, 1985 | Hemimyzon | Balitora | Hemimyzon | Balitora | Balitora | Hemimyzon | Balitora | Balitora |
7 | Balitora haithanhi Nguyen, 2005 | – | – | ? Balitora haithanhi | – | – | Balitora | – | Balitora |
8 | Balitora jalpalli Raghavan, Tharian, Ali, Jadhav & Dahanukar, 2013 | – | – | – | – | – | Balitora | – | Balitora |
9 | Balitora kwangsiensis (Fang, 1930) | Balitora | – | Balitora | Balitora | Balitora | Balitora | Balitora | Balitora |
10 | Balitora lancangjiangensis (Zheng, 1980) | Balitora | Balitora | Balitora | Balitora | Balitora | Balitora | Balitora | Balitora |
11 | Balitora laticauda Bhoite, Jadhav & Dahanukar, 2012 | – | – | Balitora | – | – | Balitora | – | Balitora |
12 | Balitora longibarbata (Chen, 1982) | Balitora | – | Balitora | Balitora | Balitora | Balitora | Balitora | Balitora |
13 | Balitora ludongensis Liu & Chen, 2012 | – | – | Balitora | Balitora | Balitora | Balitora | Balitora | Balitora |
14 | Balitora meridionalis Kottelat, 1988 | – | – | Balitora | Balitora | – | Balitora | Balitora | Balitora |
15 | Balitora mysorensis Hora, 1941 | – | – | Balitora | – | – | Balitora | – | Balitora |
16 | Balitora nantingensis Chen, Cui & Yang, 2005 | – | – | Balitora | Balitora | – | Balitora | Balitora | Balitora |
17 | Balitora vanlani Nguyen, 2005 | – | – | ? Balitora vanlani | – | – | Balitora | – | Balitora |
18 | Balitora vanlongi Nguyen, 2005 | – | – | ? Balitora vanlongi | – | – | Balitora | – | Balitora |
19 | Balitora tchangi Zheng, 1982 | Hemimyzon | Balitora | Hemimyzon | Balitora | Balitora | Hemimyzon | Balitora | Balitora |
In January 2023 during a survey of cavefishes in southwestern Guizhou Province, upper Pearl River, China, a number of specimens of the genus Balitora were collected, identified by two pairs of maxillary barbels. Morphological examination and molecular phylogenetic analysis suggested that these specimens can be distinguished from all other species presently assigned to the genus Balitora. Here, we describe these specimens as a new species, Balitora anlongensis sp. nov.
This work collected a total of 30 specimens from three species for morphological comparison and genetic analysis (Figs
Genomic DNA was extracted from muscle tissue using a DNA extraction kit from Tiangen Biotech Co., Ltd. (Beijing, China). Five tissue samples used for molecular analysis were amplified and sequenced for two mitochondrial gene and three nuclear genes: mitochondrial gene cytochrome b (Cyt b), cytochrome oxidase subunit 1 (COI), recombination activating gene 1 (RAG1), interphotoreceptor retinoid binding protein (IRBP), and early growth response protein 2B (EGR2B). Primer sequences are shown in Suppl. material
Localities, voucher information, and GenBank numbers for all samples used in this study. Bold sequences are new from this study.
ID | Family/ species | Locality (* type localities) | Voucher | Cyt b | COI | RAG1 | IRBP | EGR2B |
---|---|---|---|---|---|---|---|---|
Balitoridae | ||||||||
1 | Hemimyzon yaotanensis | Mudong, Chongqing City, China | IHB0809019 | JN176994 | – | JN177053 | – | – |
2 | Sinogastromyzon szechuanensis | Neijiang, Sichuan Province, China | 20170920BB03 | MN241814 | MN241814 | – | – | – |
3 | Sinogastromyzon hsiashiensis | Taoyuan, Hunan Province, China | IHB1004171/IHB030105 | JN176997 | JN177109 | JN177054 | – | – |
4 | Lepturichthys dolichopterus | Jianou, Fujian Province, China | IHB0706070 | GU084245 | JN177091 | JN177029 | – | – |
5 | Lepturichthys fimbriata | Jinkou, Hubei Province, China | IHB0803128 | GU084229 | JN177102 | JN177182 | JN177272 | JN177251 |
6 | Sinogastromyzon nantaiensis | Taiwan Province, China | ASIZP0806662 | – | KU943003 | – | – | – |
7 | Sinogastromyzon puliensis | – | – | FJ605359 | FJ605359 | – | – | – |
8 | Metahomaloptera omeiensis | Mudong, Chongqing City, China | IHB0301071 | JN177000 | JN177080 | JN177041 | ||
9 | Sinogastromyzon sichangensis | Cishui, Guizhou Province, China | IHB0400184 | JN176998 | JN177078 | JN177040 | KP695068 | KP694496 |
10 | Sinogastromyzon sichangensis | – | P4 | OQ754146 | – | OQ754160 | OQ754156 | OQ754151 |
11 | Jinshaia sinensis | Chongqing City, China | IHB0301068 | JN176985 | JN177117 | JN177044 | – | – |
12 | Jinshaia abbreviata | Guizhou Province, China | IHB0709424 | JN176992 | JN177228 | JN177180 | JN177274 | JN177249 |
13 | Lepturichthys fimbriata | Mudong, Chongqing City, China | IHB0301070 | JN176942 | JN177139 | JN177020 | JN177273 | – |
14 | Hemimyzon formosanus | – | cyp903 | AY392484 | KU943001 | – | KP695096 | – |
15 | Hemimyzon taitungensis | Taiwan Province, China | – | KX056121 | – | – | – | – |
16 | Balitora ludongensis | Jingxi City, Guangxi Province, China | GZNU20230215023 | OQ754141 | – | – | – | – |
17 | Balitora ludongensis | Jingxi City, Guangxi Province, China | GZNU20230215024 | OQ754142 | – | – | – | – |
18 | Balitora ludongensis | Jingxi City, Guangxi Province, China | SCAU-20190805001 | MT157616 | MT157616 | – | – | – |
19 | Balitora anlongensis sp. nov. | Xinglong Town, Anlong County, Guzihou, China* | GZNU20230215018 | OQ754144 | OQ784688 | OQ754158 | OQ754154 | OQ754149 |
20 | Balitora anlongensis sp. nov. | Xinglong Town, Anlong County, Guzihou, China* | GZNU20230215019 | OQ754145 | OQ784690 | OQ754159 | OQ754155 | OQ754150 |
21 | Balitora anlongensis sp. nov. | Xinglong Town, Anlong County, Guzihou, China* | GZNU20230215020 | OQ754143 | OQ784689 | OQ754157 | OQ754152 | OQ754148 |
22 | Sinogastromyzon wui | Zhaoping, Guangxi Province, China | IHB0400321 | JN177001 | JN177076 | – | – | – |
23 | Hemimyzon nujiangensis | Nujiang, Yunnan Province, China | / | – | KM610757 | – | – | – |
24 | Hemimyzon nujiangensis | Nujiang, Yunnan Province, China | ihb201305588 | – | KM610756 | – | – | – |
25 | Sinogastromyzon tonkinensis | Yuanjiang, Yunnan Province, China | IHB0805543 | JN177002 | JN177074 | JN177056 | JN177277 | JN177247 |
26 | Balitora brucei | – | CIFEFGB-Bb-02 | MK732323 | MK388804 | – | – | – |
27 | Balitora brucei | – | HSLBB | – | KJ774109 | – | – | – |
28 | Balitora annamitica | – | – | – | – | EF056359 | – | – |
29 | Balitora mysorensis | India, Karnataka, Hattihole | WILD-15-PIS-231 | KU378018 | KU378005 | – | – | – |
30 | Balitora mysorensis | India, Karnataka, Hattihole | BNHS FWF 197 | KU378019 | KU378006 | – | – | – |
31 | Balitora chipkali | India, Karnataka, Ramnagar | BNHS FWF 193 | KU378016 | KU378003 | – | – | – |
32 | Balitora chipkali | India, Karnataka, Kamra, Joida | WILD-15-PIS-230 | KU378017 | KU378004 | – | – | – |
33 | Balitora jalpalli | / | KUFOS-19-AN-BA-34.1 | – | MT216524 | – | – | – |
34 | Balitora laticauda | India, Maharashtra, Venegaon | WILD-12-PIS-019 | KU378007 | KU377994 | – | – | – |
35 | Balitora laticauda | India, Maharashtra, Venegaon | ZSI-WRC P/2849 | KU378008 | KU377995 | – | – | – |
36 | Balitora elongata | Menglun, Yunnan Province, China | IHB0301053 | DQ105218 | – | – | – | – |
37 | Balitora elongata | Xishuangbanna, Yunnan Province, China | IHB0301030 | DQ105217 | – | – | – | – |
38 | Balitora meridionalis | – | – | – | – | KP322550 | ||
39 | Balitora kwangsiensis | Yuanjiang, Yunnan Province, China | IHB0805545 | JN177004 | – | JN177060 | – | – |
40 | Balitora kwangsiensis | GZNU20230215022 | OQ754147 | OQ754161 | OQ754153 | – | ||
41 | Balitora elongata | / | cyp74 | – | – | KP695617 | KP695065 | – |
42 | Balitora kwangsiensis | Yuanjiang, Yunnan Province, China | IHB0805546 | JN177006 | JN177071 | JN177058 | ||
43 | Balitora kwangsiensis | Yuanjiang, Yunnan Province, China | IHB0805547 | JN177005 | JN177072 | JN177059 | JN177276 | JN177248 |
44 | Balitora kwangsiensis | / | / | DQ105216 | – | – | – | – |
45 | Homaloptera bilineata | – | – | – | – | KP322549 | – | – |
46 | Homaloptera confuzona | – | CBM:ZF 11705 | NC_033955 | NC_033955 | KP322543 | – | – |
47 | Homaloptera parclitella | – | – | NC_031634 | NC_031634 | EU409610 | EU409668 | EU409732 |
48 | Homaloptera ocellata | CBM:ZF 12287 | NC_033953 | NC_033953 | KP322539 | – | – | |
49 | Homaloptera ogilviei | – | – | NC_031635 | NC_031635 | – | – | – |
50 | Pseudohomaloptera leonardi | – | – | AB242165 | AB242165 | EU711130 | FJ197076 | AB531164 |
51 | Pseudohomaloptera sexmaculata | – | – | – | ON903153 | KP322545 | – | – |
52 | Balitoropsis ophiolepis | – | Vial 2006-0588 | – | KR052868 | KP322540 | – | – |
53 | Balitoropsis zollingeri | – | Vial 2005-0948 | – | KR052865 | KP322535 | – | – |
54 | Bhavania australis | – | KUFOS.2019.12.54 | MT002547 | MT002512 | MT002571 | – | – |
55 | Homaloptera montana | – | NBFGR:8118E | – | HQ219123 | – | – | – |
56 | Travancoria elongata | – | – | – | MT216535 | – | – | – |
57 | Travancoria jonesi | – | KUFOS-19-AN-TR-49.1 | – | MT216539 | – | – | – |
58 | Ghatsa montana | – | KUFOS-19-RR-GA-39.1 | – | MT216529 | – | – | – |
59 | Ghatsa santhamparaiensis | – | KUFOS-19-AN-GA-42.1 | – | MT216532 | – | – | – |
60 | Ghatsa pillaii | – | KUFOS-19-AN-GA-40.1 | – | MT216530 | – | – | – |
61 | Homalopteroides smithi | – | CBM:ZF 12281 | NC_033957 | NC_033957 | KP322546 | – | – |
62 | Homalopterula gymnogaster | – | Vial SN25 | – | – | KP322554 | – | – |
63 | Neohomaloptera johorensis | – | CBM:ZF 12286 | NC_033952 | NC_033952 | – | – | – |
Gastromyzontidae | ||||||||
64 | Beaufortia liui | Panzhihua, Sichuan Province, China | IHB1004172 | JN177009 | JN177069 | – | – | – |
65 | Beaufortia szechuanensis | Zhaotong, Yunna Province, China | IHB0709025 | JN177007 | JN177067 | JN177061 | JN177281 | JN177253 |
66 | Formosania chenyiyui | Changting, Fujia Province, China | IHB0301051 | MK135435 | MK135435 | – | – | – |
67 | Vanmanenia caldwelli | Wuyishan, Fujian Province, China | IHB0706028 | JN177011 | JN177232 | JN177178 | JN177280 | JN177252 |
Catostomidae | ||||||||
68 | Myxocyprinus asiaticus | Mudong, Chongqing City, China | IHB0809033 | JN176936 | – | JN177063 | – | JN177263 |
A total of 168 sequences (48 Cyt b, 55 COI, 36 RAG1, 18 IRBP, and 17 EGR2B sequences) was used for molecular analysis, including 30 newly sequenced sequences and 143 sequences downloaded from GenBank. This work followed the phylogenetic study reported by
All sequences were assembled and aligned using the MUSCLE (
The analysis suggested the best partition scheme for each codon position of Cyt b, COI, RAG1, IRBP, and EGR2B. As a result, SYM+G was selected as the best model for the first codon of Cyt b; HKY+I was selected as the best model for the second codon of Cyt b, COI, EGR2B, IRBP, and RAG1, and the first codon of EGR2B; TRN+I+G was selected as the best model for the third codon of Cyt b; TRNEF+G and TIM+G were selected as the best models for the first and third codons of COI; TVM was selected as the best model for the third codon of EGR2B; TVM+I was selected as the best model for the first codon of IRBP and RAG1; and K80+G was selected as the best model for the third codon of IRBP and RAG1. Two independent runs were conducted in BI analysis, each of which was performed for 2 × 107 generations and sampled every 1,000 generations. The first 25% of the samples were discarded as burn-in, resulting in a potential scale reduction factor of < 0.01. Nodes in the trees were considered well supported when the Bayesian posterior probabilities (BPP) were ≥ 0.95 and the ML UFB value was ≥95%. Uncorrected p-distances (1000 replicates) based on Cyt b and COI were estimated using MEGA 7.0.
Morphometric data were collected from 11 well-preserved specimens of the new species (Table
Morphological characters and measurement data (mm) of the new species Balitora anlongensis sp. nov. described in this study.
Characters | GZNU2023 0215007 | GZNU2023 0215008 | GZNU2023 0215009 | GZNU2023 0215010 | GZNU2023 0215011 | GZNU2023 0215012 | GZNU2023 02150113 | GZNU2023 0215014 | GZNU2023 0215015 | GZNU2023 0215016 | GZNU2023 0215017 | Range | Mean ± SD |
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Dorsal-fin rays | iii, 8½ | iii, 8½ | iii, 8½ | iii, 8½ | iii, 8½ | iii, 8½ | iii, 8½ | iii, 8½ | iii, 8½ | iii, 8½ | iii, 8½ | / | / |
Pectoral-fin rays | viii, 11 | viii, 11 | viii, 11 | viii, 11 | viii, 11 | viii, 11 | viii, 11 | viii, 11 | viii, 11 | viii, 11 | viii, 11 | / | / |
Pelvic-fin rays | ii, 9 | ii, 9 | ii, 9 | ii, 9 | ii, 9 | ii, 9 | ii, 9 | ii, 9 | ii, 9 | ii, 9 | ii, 9 | / | / |
Anal-fin rays | iii, 5½ | iii, 5½ | iii, 5½ | iii, 5½ | iii, 5½ | iii, 5½ | iii, 5½ | iii, 5½ | iii, 5½ | iii, 5½ | iii, 5½ | / | / |
Caudal-fin rays | 17 | 17 | 17 | 17 | 17 | 17 | 17 | 17 | 17 | 17 | 17 | / | / |
Lateral-line pores/scales | 68 | 68 | 68 | 68 | 67 | 68 | 68 | 67 | 66 | 68 | 68 | 66–68 | / |
Total length | 60.4 | 57.7 | 51.7 | 56.9 | 52.2 | 59.6 | 56.7 | 53.6 | 49.7 | 60.4 | 55.8 | 49.7–60.4 | 55.9 ± 3.7 |
Standard length | 47.9 | 44.6 | 42.4 | 44.7 | 42.3 | 47.7 | 45.9 | 42.5 | 39.8 | 48.2 | 44.3 | 39.8–48.2 | 44.6 ± 2.7 |
Body depth | 5.2 | 5.3 | 5.6 | 5.5 | 4.7 | 5.8 | 5.6 | 5.0 | 4.5 | 5.8 | 5.1 | 4.5–5.8 | 5.3 ± 0.4 |
Body width | 6.5 | 5.4 | 5.8 | 5.2 | 5.3 | 6.0 | 5.9 | 5.4 | 5.1 | 6.4 | 5.3 | 5.1–6.5 | 5.7 ± 0.5 |
Head length | 10.3 | 10.0 | 9.3 | 10.6 | 9.4 | 10.5 | 10.4 | 9.9 | 8.4 | 9.9 | 9.9 | 8.4–10.6 | 9.9 ± 0.6 |
Head depth | 4.9 | 4.2 | 4.3 | 4.3 | 4.2 | 4.1 | 4.0 | 3.8 | 3.6 | 4.5 | 4.1 | 3.6–4.9 | 4.2 ± 0.3 |
Head width | 8.7 | 6.8 | 7.0 | 6.7 | 6.6 | 7.2 | 7.2 | 6.1 | 6.3 | 7.0 | 6.9 | 6.1–8.7 | 7.0 ± 0.7 |
Pre-anterior distance | 3.5 | 4.3 | 3.4 | 3.8 | 3.7 | 4.6 | 3.6 | 3.9 | 3.8 | 3.9 | 4.4 | 3.4–4.6 | 3.9 ± 0.4 |
Distance between anterior nostrils | 2.8 | 2.9 | 2.8 | 3.1 | 2.6 | 3.0 | 2.9 | 2.3 | 2.1 | 2.4 | 2.1 | 2.1–3.1 | 2.6 ± 0.4 |
Distance between posterior nostrils | 3.8 | 3.7 | 3.6 | 3.6 | 3.8 | 3.4 | 3.6 | 3.2 | 2.9 | 3.4 | 3.1 | 2.9–3.8 | 3.5 ± 0.3 |
Distance between anterior and posterior nostrils | 2.0 | 1.7 | 1.1 | 1.6 | 1.5 | 1.6 | 1.3 | 1.3 | 1.4 | 1.5 | 1.3 | 1.1–2.0 | 1.5 ± 0.2 |
Snout length | 5.9 | 5.3 | 5.2 | 5.6 | 5.2 | 6.1 | 5.4 | 5.1 | 4.9 | 5.6 | 5.6 | 4.9–6.1 | 5.4 ± 0.4 |
Upper jaw length | 2.8 | 2.5 | 2.6 | 2.5 | 2.5 | 2.8 | 2.1 | 2.7 | 2.3 | 2.8 | 2.6 | 2.1–2.8 | 2.6 ± 0.2 |
Lower jaw length | 2.0 | 1.8 | 1.6 | 1.8 | 2.0 | 1.9 | 1.4 | 1.9 | 1.4 | 2.1 | 2.0 | 1.4–2.1 | 1.8 ± 0.2 |
Mouth width | 3.6 | 3.8 | 3.5 | 3.8 | 4.1 | 4.2 | 4.0 | 3.6 | 3.4 | 4.0 | 3.7 | 3.4–4.2 | 3.8 ± 0.3 |
Eye diameter | 1.3 | 1.1 | 1.0 | 0.9 | 1.0 | 0.9 | 0.8 | 0.9 | 0.8 | 1.1 | 0.9 | 0.8–1.3 | 1.0 ± 0.1 |
Interorbital distance | 4.7 | 4.4 | 4.4 | 4.4 | 4.1 | 4.7 | 3.5 | 3.7 | 3.9 | 4.3 | 3.7 | 3.5–4.7 | 4.2 ± 0.4 |
Predorsal length | 22.3 | 21.2 | 19.2 | 20.9 | 19.5 | 21.9 | 21.0 | 20.1 | 18.2 | 22.0 | 20.5 | 18.2–22.3 | 20.6 ± 1.3 |
Dorsal-fin base length | 7.4 | 6.0 | 6.2 | 6.3 | 5.3 | 7.0 | 6.7 | 6.7 | 6.1 | 7.7 | 6.4 | 5.3–7.7 | 6.5 ± 0.7 |
Dorsal-fin length | 10.9 | 10.5 | 9.6 | 10.3 | 10.2 | 10.4 | 10.1 | 10.0 | 8.9 | 11.0 | 10.1 | 8.9–11.0 | 10.2 ± 0.6 |
Pectoral-fin length | 11.7 | 10.4 | 10.0 | 10.9 | 9.8 | 11.1 | 11.6 | 9.9 | 10.2 | 11.8 | 11.2 | 9.8–11.8 | 10.8 ± 0.8 |
Pectoral-fin base length | 4.9 | 4.0 | 3.8 | 3.4 | 3.8 | 4.2 | 4.1 | 3.3 | 4.0 | 4.7 | 4.3 | 3.3–4.9 | 4.0 ± 0.5 |
Pre-pectoral length | 9.3 | 8.7 | 7.3 | 8.2 | 7.2 | 8.4 | 7.8 | 8.2 | 7.4 | 8.1 | 7.8 | 7.2–9.3 | 8.0 ± 0.6 |
Pelvic-fin length | 10.0 | 10.1 | 8.5 | 9.4 | 8.8 | 10.4 | 9.8 | 9.2 | 8.5 | 10.3 | 9.2 | 8.5–10.4 | 9.5 ± 0.7 |
Pelvic-fin base length | 3.4 | 2.6 | 3.2 | 2.6 | 2.6 | 2.8 | 2.9 | 2.9 | 2.7 | 2.9 | 2.6 | 2.6–3.4 | 2.8 ± 0.3 |
Pre-pelvic length | 22.6 | 21.7 | 19.7 | 20.1 | 20.2 | 22.7 | 21.5 | 20.3 | 18.8 | 21.8 | 20.1 | 18.8–22.7 | 20.9 ± 1.3 |
Anal-fin length | 8.7 | 8.5 | 7.3 | 8.4 | 7.6 | 8.5 | 8.0 | 7.9 | 6.9 | 8.6 | 7.8 | 6.9–8.7 | 8.0 ± 0.6 |
Anal-fin base length | 3.9 | 3.4 | 3.6 | 3.8 | 3.4 | 3.3 | 4.0 | 3.8 | 3.3 | 3.6 | 3.4 | 3.3–4.0 | 3.6 ± 0.3 |
Pre-anal length | 34.8 | 33.0 | 30.4 | 32.5 | 31.6 | 35.3 | 33.5 | 31.2 | 29.8 | 35.2 | 31.9 | 29.8–35.3 | 32.7 ± 1.9 |
Distance between origin of pectoral fin and origin of ventral fin | 10.0 | 9.4 | 9.0 | 9.8 | 9.3 | 10.8 | 10.2 | 9.1 | 8.2 | 10.7 | 9.7 | 8.2–10.8 | 9.7 ± 0.8 |
Distance between origin of ventral fin and origin of Anal fin | 10.6 | 8.9 | 8.8 | 9.2 | 10.0 | 10.0 | 10.0 | 8.3 | 8.4 | 10.3 | 9.5 | 8.3–10.6 | 9.5 ± 0.8 |
Distance between end of Anal fin and anus | 1.8 | 2.4 | 2.4 | 2.2 | 2.9 | 3.0 | 2.3 | 1.9 | 1.7 | 2.4 | 2.1 | 1.7–3.0 | 2.3 ± 0.4 |
Caudal peduncle length | 8.6 | 9.6 | 8.5 | 8.7 | 7.5 | 9.6 | 9.1 | 8.1 | 7.8 | 9.9 | 9.2 | 7.5–9.9 | 8.8 ± 0.8 |
Caudal peduncle depth | 3.1 | 2.9 | 3.1 | 2.7 | 2.7 | 3.1 | 2.8 | 3.0 | 2.3 | 3.2 | 3.0 | 2.3–3.2 | 2.9 ± 0.3 |
Inner maxillary barbel length | 1.4 | 2.0 | 1.4 | 2.1 | 1.7 | 2.5 | 1.5 | 2.4 | 1.7 | 2.0 | 1.5 | 1.4–2.5 | 1.8 ± 0.4 |
Outer maxillary barbel length | 1.1 | 1.0 | 1.2 | 0.9 | 0.9 | 1.5 | 1.0 | 1.2 | 0.9 | 1.1 | 0.9 | 0.9–1.5 | 1.1 ± 0.2 |
Inner rostral barbel length | 1.4 | 1.1 | 1.0 | 1.2 | 1.1 | 1.6 | 1.3 | 1.4 | 1.1 | 1.6 | 1.1 | 1.0–1.6 | 1.3 ± 0.2 |
Outer rostral barbel length | 1.9 | 2.0 | 1.7 | 2.0 | 1.9 | 1.8 | 1.9 | 1.8 | 1.8 | 2.2 | 1.7 | 1.7–2.2 | 1.9 ± 0.1 |
Comparison of the diagnostic characters of the new species described here with those selected for the 16 species of the genus Balitora. Gray shading indicates clear differences in characters compared to Balitora anlongensis sp. nov.
ID | Species | Maxillary barbels | Dorsal fin rays | Anal fin rays | Pectoral fin rays | Pelvic fin rays | Lateral-line scales | Dorsal black spot | Pectoral fin tip | Position of origin of dorsal-fin and origin of pelvic-fin | Tip of pelvic fin reaching anus |
---|---|---|---|---|---|---|---|---|---|---|---|
1 | Balitora anlongensis sp. nov. | 2 | iii, 8½ | iii, 5½ | viii, 11 | iii, 9 | 66–68 | 6–7 | Not reaching to pelvic fin origin | Anterior to the pelvic-fin origin | Yes |
2 | B. annamitica | 1 | – | – | viii–x, ? | – | 61–62 | – | – | – | – |
3 | B. brucei | 1 | iii, 8 | i, 5 | ix, 11 | ii, 9 | 65–69 | – | – | – | – |
4 | B. burmanica | 1 | iii, 8 | i, 5 | ix, 11 | ii, 8 | 70–72 | – | – | – | – |
5 | B. chipkali | 1 | iii, 8 | iii, 5 | iii–ix, 11–12 | ii, 9 | 66–68 | 7 | Not reaching to pelvic fin origin | Opposite to pelvic-fin origin | Yes |
6 | B. eddsi | 1 | iii, 9 | iii, 5\7 | vi, 10–12 | ii, 8–9 | 66–67 | Without | Not reaching to pelvic fin origin | Slightly anterior to pelvic-fin origin | NO |
7 | B. jalpalli | 1 | iii, 8 | ii, 5 | ix, 10–11 | ii, 8–9 | 64–66 | 9 | Not reaching to pelvic fin origin | Opposite to pelvic-fin origin | NO |
8 | B. kwangsiensis | 2 | iii, 8 | ii, 5 | vi–viii, 10–13 | ii, 8 | 61–65 | 6–8 | Not reaching to pelvic fin origin | Anterior to the pelvic-fin origin | Yes |
9 | B. lancangjiangensis | 1 | iii, 8 | ii, 5 | viii, 10–12 | ii, 8–9 | 68–70 | 7–8 | Not reaching to pelvic fin origin | Opposite to pelvic-fin origin | NO |
10 | B. laticauda | 1 | iii, 8 | iii, 5 | viii–ix, 10–11 | ii, 8–9 | 66–68 | 10 | Not reaching to pelvic fin origin | Opposite to pelvic fin origin | NO |
11 | B. longibarbata | 2 | iii, 8 | ii, 5 | viii–x, 11–14 | ii, 9–11 | 74–76 | 8–9 | Not reaching to pelvic fin origin | Opposite to pelvic-fin origin | NO |
12 | B. ludongensis | 2 | iii, 8 | ii, 5 | vi–vii, 11–12 | ii, 6–7 | 69–74 | 6–9 | Not reaching to pelvic fin origin | Slightly anterior to pelvic-fin origin | Yes |
13 | B. meridionalis | 1 | – | – | – | – | – | – | – | – | – |
14 | B. mysorensis | 1 | iii, 8–9 | ii, 5 | viii-ix 10–12 | ii, 8–9 | 68–69 | – | – | – | – |
15 | B. nantingensis | 1 | iii, 8 | ii, 5 | viii–x, 9–12 | ii, 9 | 59–64 | – | Not reaching to pelvic fin origin | Slightly anterior to pelvic-fin origin | – |
16 | B. elongata | 1 | iii, 8 | ii, 5 | x, 10–12 | iii, 8 | 67 | 7 | Not reaching to pelvic fin origin | Opposite to pelvic-fin origin | NO |
17 | B. tchangi | 1 | iii, 7 | ii, 5 | xii, 13 | v, 13 | 74 | 8 | Beyond the origin of pelvic fin | Posterior to the pelvic-fin origin | NO |
Both ML and BI phylogenies were constructed based on two mitochondrial and three nuclear genes, with a sequence length of 5382 base pairs. The BI and ML phylogenetic trees show a highly consistent topology that strongly supports the monophyly of the family Balitoridae and can be divided into five major clades (Fig.
A Distributions of the different clades of the genus Balitora in Asia B phylogenetic tree reconstructed based on combined two mitochondrial (Cyt b and COI) and three nuclear gene (RAG1, IRBP, and EGR2B) fragments. In this phylogenetic tree, ultrafast bootstrap (UFB) supports from maximum likelihood (ML) analyses/Bayesian posterior probabilities (BPP) from Bayesian inference (BI) analyses are noted beside nodes. The scale bar represents 0.04 nucleotide substitutions per site. The numbers at the tips of branches correspond to the ID numbers in Table
The phylogenetic tree shows that Balitora is not a monophyletic and contains three distant clades: Clade I, including B. meridionalis Kottelat, 1988, B. elongata, B. laticauda Bhoite, Jadhav & Dahanukar, 2012, B. jalpalli Raghavan, Tharian, Ali, Jadhav & Dahanukar, 2013, B. chipkali Kumar, Katwate, Raghavan & Dahanukar, 2016, B. mysorensis Hora, 1941, B. annamitica Kottelat, 1988, and B. brucei Gray, 1830 (type species of the genus Balitora); Clade II, containing only B. kwangsiensis; and Clade III, including B. ludongensis and the new species described in this study (Fig.
Uncorrected p-distance (%) between eight species of the genus Balitora based on mitochondrial Cyt b.
ID | Species | 1 | 2 | 3 | 4 | 5 | 6 | 7 |
---|---|---|---|---|---|---|---|---|
1 | Balitora anlongensis sp. nov. | |||||||
2 | Balitora ludongensis | 7.3 | ||||||
3 | Balitora brucei | 10.4 | 9.5 | |||||
4 | Balitora chipkali | 12.0 | 11.9 | 9.9 | ||||
5 | Balitora elongata | 9.2 | 9.5 | 10.7 | 11.1 | |||
6 | Balitora kwangsiensis | 9.5 | 9.0 | 9.6 | 12.8 | 10.5 | ||
7 | Balitora laticauda | 10.9 | 10.6 | 8.3 | 4.5 | 10.3 | 11.4 | |
8 | Balitora mysorensis | 10.6 | 10.3 | 9.0 | 11.7 | 10.3 | 11.3 | 10.4 |
Uncorrected p-distance (%) between nine species of the genus Balitora based on mitochondrial COI.
ID | Species | 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 |
---|---|---|---|---|---|---|---|---|---|
1 | Balitora anlongensis sp. nov. | ||||||||
2 | Balitora ludongensis | 10.5 | |||||||
3 | Balitora brucei | 10.2 | 11.1 | ||||||
4 | Balitora chipkali | 10.5 | 11.1 | 9.4 | |||||
5 | Balitora jalpalli | 10.3 | 11.3 | 9.7 | 1.1 | ||||
6 | Balitora kwangsiensis | 12.1 | 11.7 | 12.2 | 12.3 | 12.8 | |||
7 | Balitora laticauda | 11.1 | 11.7 | 9.3 | 2.5 | 2.1 | 12.5 | ||
8 | Balitora mysorensis | 9.2 | 10.0 | 6.8 | 9.0 | 8.6 | 11.5 | 8.4 | |
9 | Balitora nujiangensis | 9.6 | 10.0 | 9.4 | 9.6 | 9.8 | 10.0 | 9.8 | 8.4 |
Thus, the population at this locality represents an independently evolved lineage and is described as a new species, Balitora anlongensis sp. nov., below.
Holotype. GZNU20230215007 (Fig.
Morphological characters of holotype GZNU20230215007 of Balitora anlongensis sp. nov. in preservative (10% formalin) A lateral view B dorsal view C ventral view D ventral side view of head, and E dorsal side view of head. Photos from Tao Luo. Abbreviations: M, maxillary barbels; AN, anterior nostril.
Paratypes. Ten specimens from the same locality as the holotype: GZNU20230215008–215017, collected by Tao Luo, Xin-Rui Zhao, Wei-Feng Wang, Jing Yu, and Chang-Ting Lan on 15 February 2023.
The new species is assigned to the genus Balitora based on the combination of the following diagnostic characters (
Balitora anlongensis sp. nov. can be distinguished from other congeners by the following combination of characters: (1) two pairs of maxillary barbels; (2) dorsal fin rays iii, 8½; (3) pectoral fin viii, 11; (4) pelvic fin rays ii, 9; (5) anal fin rays iii, 5½; (6) lateral-line scales 66–68; (7) tip of pectoral fin not reaching to the pelvic fin origin; (8) dorsal fin origin anterior to the pelvic fin origin; (9) tip of the pelvic fin reaching to the anus; (10) eyes small, eye diameter equal to outer maxillary barbel length; (11) six to seven indistinctly separated transversely oval blotches on the dorsal side; and (12) each fin transparent and unpigmented in life.
Morphological data of the 11 specimens of the Balitora anlongensis sp. nov. are provided in Table
Dorsal fin rays iii, 8½, pectoral fin rays viii,11, pelvic fin rays ii, 9, anal fin rays iii, 5½, and 17 branched caudal fin rays. Dorsal fin long, 22–24% of SL, nearly equal to head length, distal margin truncated, origin anterior to pelvic fin insertion, situated slightly anterior to midpoint between snout tip and the caudal fin base, first branched ray longest, shorter than HL, tip of the dorsal fin extending to the vertical of the anus. Pectoral fin elongated and developed, distal margin rounded, pectoral fin length slightly greater than HL, 23–25% of SL, tip of the pectoral fin extends backward beyond 3/4 of the distance between the origin of the pectoral fin and the origin of the pelvic fin, without reaching to the pelvic fin-origin. Pelvic fin moderately developed, distal margin rounded, pectoral fin length approximately equal to HL, 23–26% of SL, vertically aligned with the third branched ray of the dorsal fin, pelvic fin origin closer to the snout tip than the caudal fin base and closer to the anal fin origin than the snout tip, tips of the pelvic fin reaching to the anus. Anus ~ 4/5 distance from posterior end of the pelvic fin base to the anal fin origin. Anal fin short, 17–19% of SL, distal margin truncated, origin close to the anus and far from the caudal fin base, spacing ~ 2.9 mm, tips of the anal fin extending backwards and not reaching caudal fin base, distance between the end of the anal fin and the anus 5.4-times the eye diameter. Caudal fin deeply forked, upper lobe equal in length to the lower one, tips pointed, caudal peduncle length 8.8 mm, caudal peduncle depth 2.9 mm, without adipose crests along both dorsal and ventral sides. Vertebrae 36: nine pre-dorsal abdominal, 19 abdominal (including four Weberian and 15 pre-dorsal ones), and 17 caudal (including three pre-anal caudal and the hypural complex) (Fig.
Body smooth, covered with thin scales all over except for on the ventral side, head, and fins. Lateral line complete and straight, with 66–68 lateral line scales, exceeding the tip of the pectoral fin and reaching the base of the caudal fin. Two air bladder chambers, posterior chamber of the air bladder slightly developed and closed (Fig.
Dwelling in the water bodies of the cave (Fig.
No sexual dimorphism was observed based on the present specimens of Balitora anlongensis sp. nov.
Comparative data of Balitora anlongensis sp. nov. with 19 species within the genus Balitora are given in Table
Balitora anlongensis sp. nov. differs from B. annamitica, B. brucei, B. burmanica Hora, 1932, B. chipkali, B. eddsi Conway & Mayden, 2010, B. jalpalli, B. lancangjiangensis, B. laticauda, B. meridionalis, B. mysorensis, B. nantingensis Chen, Cui & Yang, 2005, B. elongata, and B. tchangi based on the presence of two maxillary barbels at each corner of the mouth (vs one maxillary barbel at each corner of the mouth). Balitora anlongensis sp. nov. can be further distinguished from B. eddsi and B. tchangi based on dorsal fin rays (iii, 8½ vs iii, 9 in B. eddsi and iii, 7 in B. tchangi); from B. jalpalli, B. lancangjiangensis, B. laticauda, B. nantingensis, B. elongata, and B. tchangi by the tip of pelvic fin reaching to the anus (vs not reaching to the anus); from B. brucei by pectoral fin rays (viii, 11 vs ix, 11); and from B. jalpalli, B. lancangjiangensis, B. laticauda, B. elongata, and B. tchangi by the dorsal fin origin being anterior to the pelvic fin origin (vs opposite/posterior to pelvic fin origin; see Table
Balitora anlongensis sp. nov. and B. kwangsiensis, B. longibarbata, and B. ludongensis are distributed in the Pearl River and share two maxillary barbels at each corner of the mouth, but can be distinguished by the combination of serial characters. Balitora anlongensis sp. nov. can be distinguished from B. kwangsiensis by anal fin rays (iii, 5½ vs ii, 5), pelvic fin rays (ii, 9 vs ii, 8), lateral-line scales (66–68 vs 61–65), and each fin being transparent and unpigmented in life (vs each fin having black spots); from B. longibarbata by the pelvic fin rays (ii, 8 vs ii, 9–11), anal fin rays (iii, 5½ vs ii, 5), lateral-line scales (66–68 vs 74–76), the dorsal fin origin being anterior to the pelvic fin origin (vs opposite to the pelvic fin origin), and the tip of the pelvic fin reaching to the anus (vs not reaching to the anus).
Phylogeny constructed based on combined mitochondrial and nuclear genes shows that Balitora anlongensis sp. nov. is close to B. ludongensis and can be distinguished by the combination of the following morphological characters: eight unbranched pectoral fin rays (vs six to seven), anal fin rays (iii, 5½ vs ii, 5), pelvic fin rays (ii, 9 vs ii, 6–7), lateral-line scales (66–68 vs 69–74), dorsal fin origin far anterior to the pelvic fin origin (vs slightly anterior to the pelvic fin origin), body depth 13–14% of the SL (vs 15–20%), body width 12–14% of SL (vs 18–21%), head depth 38–48% of HL (vs 51–67%), head width 109–134% of body width (vs 87–107%), transversely oval blotches on dorsal side indistinctly separated (vs distinctly separated), and each fin transparent and unpigmented in life (vs each fin having black spots).
Balitora anlongensis sp. nov. is only known from the type locality, a vertical cave some distance from NaNao Village, Xinglong Town, Anlong County, Guizhou Province, China at an elevation of 1387 m (Figs
The specific epithet “anlongensis” is in reference to the type locality of the new species: NaNao Village, Xinglong Town, Anlong County, Guizhou Province, China. We propose the common English name “Anlong stone loach” and the Chinese name “ān lóng Pá Qīu (安龙爬鳅)”.
This work described a new cavefish species from Xinglong Town, Anlong County, Guizhou, named Balitora anlongensis sp. nov., based on morphological comparisons (see the comparison above) and genetic differences (Fig.
The genus Balitora may need to be revised in the future by combining evidence from phylogenetic and morphological evidence. The taxonomy of the genus Balitora based on morphological characters has undergone two major revisions, with the main controversy being over its key diagnostic characteristics, i.e., one pair of maxillary barbel (
Balitora ludongensis (n = 2): China: Guangxi: Jingxi County: Ludong Town (type locality): GZNU 20230207001–0207002.
Balitora kwangsiensis (n = 2): China: Yunnan: Amojiang River basin: GZNU20230226001–0226002.
We thank Wei-Feng Wang, Xing-Liang Wang, Li Wu, Cui Fan, and other for help with specimen collection. We thank LetPub (www.letpub.com) for its linguistic assistance during the preparation of this manuscript.
The authors have declared that no competing interests exist.
No ethical statement was reported.
This research was supported by the programs of the Diversity and Distribution Survey of Chiroptera species in China (2021FY100302), the Guizhou Province Top Discipline Construction Program Project (Qianjiao Keyan Fa [2019] 125), and the Guizhou Normal University Academic Emerging Talent Fund Project (Qianshi Xin Miao [2021] 20).
Jiang Zhou and Tao Luo conceived and designed the research; Tao Luo, Zhi-Xia Chen, Xin-Rui Zhao, Jing Yu, and Chang-Ting Lan conducted field surveys and collected samples; Tao Luo, Jing Yu, Ning Xiao and Chang-Ting Lan performed molecular work; Zhi-Xia Chen, Jia-Jun Zhou, and Xin-Rui Zhao collected distribution data and mapped the distribution; and Jiang Zhou, Ning Xiao and Tao Luo wrote and discussed and revised the manuscript. All authors read and approved the final version of the manuscript.
Tao Luo https://orcid.org/0000-0003-4186-1192
Zhi-Xia Chen https://orcid.org/0009-0002-2066-494X
Xin-Rui Zhao https://orcid.org/0000-0002-9125-6276
Jing Yu https://orcid.org/0009-0004-3629-3826
Chang-Ting Lan https://orcid.org/0009-0007-2381-3601
Jia-Jun Zhou https://orcid.org/0000-0003-1038-1540
Ning Xiao https://orcid.org/0000-0002-7240-6726
Jiang Zhou https://orcid.org/0000-0003-1560-8759
All of the data that support the findings of this study are available in the main text or Supplementary Information.
List of cavefishes in China
Data type: xlsx
Specific primers used to amplify two mitochondrial and three nuclear genes
Data type: xlsx