Research Article |
Corresponding author: Alexander Balakirev ( alexbalakirev@mail.ru ) Academic editor: Kristofer M. Helgen
© 2014 Alexander Balakirev, Alexei Abramov, Viatcheslav Rozhnov.
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:
Balakirev A, Abramov A, Rozhnov V (2014) Phylogenetic relationships in the Niviventer-Chiromyscus complex (Rodentia, Muridae) inferred from molecular data, with description of a new species. ZooKeys 451: 109-136. https://doi.org/10.3897/zookeys.451.7210
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Based on molecular data for mitochondrial (Cyt b, COI) and nuclear (IRBP, GHR) genes, and morphological examinations of museum specimens, we examined diversity, species boundaries, and relationships within and between the murine genera Chiromyscus and Niviventer. Phylogenetic patterns recovered demonstrate that Niviventer sensu lato is not monophyletic but instead includes Chiromyscus chiropus, the only previously recognized species of Chiropus. To maintain the genera Niviventer and Chiropus as monophyletic lineages, the scope and definition of the genus Chiromyscus is revised to include at least three distinct species: Chiromyscus chiropus (the type species of Chiromyscus), C. langbianis (previously regarded as a species of Niviventer), and a new species, described in this paper under the name C. thomasi sp. n.
White-bellied rats, Fea’s tree rat, Southeast Asia, Vietnam, molecular phylogeny, taxonomy, new species
The genera Niviventer Marshall, 1976 and Chiromyscus Thomas, 1925 are members of the Dacnomys division of the tribe Rattini (
Taxonomic composition and preliminary views of relationships within the genus Niviventer were first established by
The monotypic genus Chiromyscus is most likely the closest relative to Niviventer. The only representative of this genus, Chiromyscus chiropus (Thomas, 1891), was first described as Mus chiropus from East Burma. This species is morphologically very similar to the Indochinese taxon Niviventer langbianis (
Newly collected museum specimens investigated here were obtained in Vietnam during a series of field expeditions of the Joint Russian-Vietnamese Tropical Research and Technological Centre between 2007 and 2013 and deposited at the Zoological Museum of Moscow State University (ZMMU, Moscow, Russia) and at the Zoological Institute of the Russian Academy of Sciences (ZIN, Saint Petersburg, Russia). Most specimens were collected by the authors (BAE, AAV). All animals were identified in the field based on external morphology according to field identification manuals (
Twenty eight specimens of N. langbianis and Chiromyscus from 6 localities in Vietnam were sampled for genetic analysis (See Suppl. material
The resulting double-stranded DNA products were directly sequenced in both directions using the Applied Biosystems 3130 Genetic Analyzer and the ABI PRISM BigDye Terminator Cycle Sequencing Ready Reaction Kit. All obtained sequences were deposited in GenBank (www.ncbi.nlm.nih.gov/genbank) under the accession numbers KF154023–KF154052 and KF154054–KF154085, and certain COI gene sequences were also uploaded into the BOLD database (www.barcodinglife.org project “Indochinese Muridae”, ICMBA).
We also analyzed 122 gene sequences of Niviventer (all “langbianis group” species sequences available, as well as some sequences from other species) and Chiromyscus that were available in the GenBank and BOLD databases as of 1 May 2013. Out of these 122 sequences, 35 were for Cyt b, 27 were for IRBP, 26 were for GHR, and 34 were for COI. The gene sequences from two outgroup species were used to root the phylogenetic tree [Mus musculus L., 1758 (V00711, complete mtDNA genome; AB033711, IRBP; NM001048147, GHR) and Rattus rattus L., 1758 (EU273707, complete mtDNA genome; AM408328, IRBP; DQ019074, GHR)].
Sequences were aligned using BIOEDIT 3.0 (
A phylogeny was first estimated for each gene independently, and subsequently for the concatenated dataset once the four genes were manually combined into a single data set in BIOEDIT 3.0 to produce combined samples. This restricted subset (12 variables/taxa in total, see Suppl. material
Bayesian analysis for the combined data set was performed using four independent runs of 2 × 106 generations each. The most complex substitution model, GTR+G, was used for the combined data set to avoid multi-partition calculation procedures and relax computing process (even though the mitochondrial genes appeared to evolve under the more simple TN93+G+I substitution model). We used a flat Dirichlet prior distribution for the relative nucleotide frequencies and for the relative rate parameters, a discrete uniform prior distribution for the topologies and an exponential distribution for the gamma shape parameter and all branch lengths. A burn-in period of 500,000 generations was determined graphically using TRACER v1.4 (
Single gene phylogenies revealed that relationships across the overall taxon sampling could not be reliably resolved for most basal nodes, irrespective of the phylogenetic approach (results not shown). The trees obtained from the different genes and methods differed mostly in the topology of the branches of species within the niviventer/fulvescens/langbianis groups of species and in the level of nodal supports. In Fig.
The ultrametric ML phylogenetic tree constructed based on complete Cyt b gene sequences (TN93+G+I; 1-2-3 pos. inc.) of the Niviventer-Chiromyscus complex. The scale bars at the bottom represent the level of divergence (d, T3P). The bars at the nodes represent the level of confidence of branch lengths.
The combined analysis using all four genes on a reduced dataset resulted in a well-supported phylogeny (Fig.
Given that Niviventer is shown not to be monophyletic, two taxonomic approaches to generic nomenclature could be undertaken. Either Niviventer could be regarded as a junior synonym of Chiromyscus based on taxonomic priority (
Three morphologically distinct groups can be traced from the N. langbianis/C. chiropus complex (i.e., the redefined content of the genus Chiromyscus) that correspond to species-level phylogenetic clades revealed within the “langbianis-chiropus” cluster obtained from analyses of mitochondrial and nuclear genes (Figs
Thirteen adult specimens identified here as Chiromyscus langbianis (Robinson & Kloss, 1922) were collected in the highlands of the Dalat Plateau, Lam Dong Province, southern Vietnam, close to the type locality of this taxon, and in the Huu Lien Nature Reserve, Lang Son Province, northern Vietnam. The corresponding samples formed two independent but closely related clusters, labeled as clade 1 with subclades A and B in Fig.
Another thirteen adult animals we identified as C. chiropus (Fig.
Chiromyscus chiropus, southern Vietnam. A Dorsal view B Ventral view C Lateral view D General appearance (photo by Alexander E. Balakirev) E Head, face-lateral view F Head, lateral view G Head, dorsal view H Hind foot, dorsal view I Hind foot, plantar surface. A–C, E–I specimen from the Binh Chau Nature Reserve, Ba Ria – Vung Tau Province, southern Vietnam, ZMMU S-191972, genetic voucher BT10-2 D specimen from the Bao Loc Forestry, Lam Dong Province, southern Vietnam, ZIN 100966, genetic voucher 12-068.
Chiromyscus chiropus was first described as Mus chiropus by
The holotype of Chiromyscus chiropus, stuffed skin (MSNG 18396) and skull (MSNG 18397). A Stuffed skin, dorsal view B Skull, ventral view; hind foot, dorsal view C Stuffed skin, ventral view D Head, lateral view E Head, face view F Hind foot, plantar surface. Images were kindly provided by the Museo Civico di Storia Naturale “Giacomo Doria”, Genoa, Italy.
The holotype of Niviventer cremoriventer, stuffed skin and skull, USNM 86770. A Stuffed skin, dorsal view B Stuffed skin, ventral view; the natural coloration of front legs and feet is changed due to chemical treatment C Skull. Images were kindly provided by the National Museum of Natural History, Smithsonian Institution, Washington, USA.
Unfortunately, we did not have an opportunity to include the holotype of C. chiropus in our genetic comparisons. Nevertheless, based on apparent morphological similarity we attributed our southern Vietnamese specimens to Chiromyscus chiropus proper. Because of the scarcity of museum specimens and DNA-confirmed records for this species, it is difficult to estimate the true distributional range for this species. However, there are no substantial geographic barriers over the vast area stretching from the lowlands of southern Vietnam through Cambodia and central Thailand and west up to the hilly country of Peninsular Thailand and the eastern regions of Myanmar. Thus, there is every reason to believe the species may be distributed over substantial areas in Thailand and Cambodia, most likely scattered over patches of forested areas.
The third distinct species-level genetic lineage within Chiromyscus is labeled here as C. chiropus clade 3 (Fig.
Vietnam, Son La Province, Muong Thai Village, near Lung Lo pass, 21°18'31"N, 104°41'34"E, elevation ~ 450 m above sea level.
BMNH 25.1.1.110, skin and skull, male, Bao Ha, Lao Cai Province, Vietnam; BMNH 26.10.4.167, skin and skull, female, Dak To, Kon Tum Province, Vietnam; BMNH 26.10.4.166, skin, male, Xieng Kuang, Laos.
This species is set apart from all other described species within the genus Chiromyscus by the following combination of morphological traits: (1) Appreciably larger size. This species is the largest in size of any species of Chiromyscus. Head and body length is 145–180 mm, tail length 200–231 mm, length of hind foot 27–29 mm, ear length 18–20 mm, greatest skull length 41.0–43.0 mm, upper molar lengths 7.0–8.0 mm; the supraorbital ridges are more developed than in other species, forming a distinct pointed triangle shelf at the point where the frontal and palatal bones come into contact. This shelf is very perceptible in the frontal view of the skull. (2) The upper parts are orange-brown. From the face to behind the ears, the pelage is bright orange, with a prominent darker ring around the eye forming a “mask” on the face. The under parts are pure white and sharply demarcated from the upper parts. The feet and toes are generally white with orange hairs on top. The tail is bicolored, dark on top and appreciably lighter below, where there is a pinkish hue. The hallux is shortened with rounded nails instead of pointed claws. The species is well differentiated genetically from other Chiromyscus. The DNA sequences that are deposited in GenBank under IDs JQ755933–JQ755934, JQ755964–JQ755965, KF154025 and KF154068–KF154069 may be used as genetic vouchers for this species.
The fur is dense, smooth and downy. The coloration of the upper side is a bright fulvous with a perceptible orange hue, which is most prominent in the humeral area. On the underside, the belly is pure white without patches or creamy hues. The sides are more brightly colored than the back. The cheek, lateral surface of the neck and the front legs are a bright yellowish-orange. The rump, hips, and base of tail are also, like the cheek, a rich ochraceous color. A very prominent black strip passes over the eye, forming a very characteristic “mask” on the face. The vibrissae are long (over 60 mm), both black- and white-colored, and the ears are small (18–20 mm), pale-brown colored and rounded. The dorsal sides of both the front and hind feet are completely buffy-orange. The pads both in the front and hind feet are well developed. The claws are large (4.2–5.0 mm in length), curved and appreciably sharp. The hallux bears a plain nail instead of a claw. The tail is very long, slender and hairy; it is much longer than the body (128–132% of body length). It is rather thick and almost uniformly tinged pale-brown from the proximal part to the tip.
C. thomasi is a brightly colored species, a feature that obviously distinguishes it from C. langbianis, which is generally dull in coloration. With its bright fulvous or orange coloration C. thomasi is similar to C. chiropus but may be distinguished from it by its dorso-ventral coloration demarcation line. In C. thomasi, the white-colored belly replaces the bright orange ventral side coloration abruptly, without any intermediate zone, whereas a lighter-colored fulvous intermediate zone (0.5–1.0 cm in width) is perceptible on the back sides of C. chiropus. However, the most apparent distinguishing feature of C. thomasi is a dark “mask” on the face around the eyes, which may be used to visually separate it from any another Chiromyscus or Niviventer species. Chiromyscus thomasi is the largest species in the genus, appreciably bigger than C. chiropus and C. langbianis. Its skull well exceeds the known range of size variation for other Chiromyscus as well as for the majority of Niviventer species, with the exception of N. tenaster and the “andersoni” group, both of which are roughly equal in size to, or larger than, C. thomasi. In comparison with other Chiromyscus species, the skull of C. thomasi is also the most “heavily-built”, with supraorbital ridges that are more developed, forming prominent wide shelves. The skull of C. langbianis is much smaller and gracile, and the shelves are not so apparent, whereas in C. chiropus the skull has an obviously convex profile (when viewed from the side), in contrast with C. thomasi, which appears rather flattened when viewed from the side.
The new species is named in honor of Oldfield Thomas (1858–1929), the British zoologist who named and described the genus Chiromyscus and the species chiropus.
Thomas' masked tree rat.
Confirmed specimens of Chiromyscus thomasi have been recorded from the provinces of Son La and Lao Cai in northern Vietnam, the provinces of Kon Tum and Nhge An in central Vietnam, and the provinces of Xieng Khouang and Luang Prabang in northern Laos, based on published data and our (BAE) most recent and unpublished data. This species may have a wider distribution in central Vietnam (
In spite of the close phylogenetic relationships evident within the Niviventer-Chiromyscus complex, the taxonomic composition within genera can be reliably resolved by a combination of mitochondrial and nuclear gene analyses, which provide support to the traditional morphological segregation initially suggested for a langbianis-Chiromyscus cluster by
This study was realized with the support of the Joint Russian-Vietnamese Tropical Research and Technological Centre, Southern Division, Ho Chi Minh City, Vietnam. We thank Dr. Victor V. Suntsov, Dr. Nina I. Suntsova (A.N. Severtsov Institute of Ecology and Evolution, Russian Academy of Sciences, Moscow, Russia), Dr. Sergey V. Kruskop (Zoological Museum of Moscow State University, Moscow, Russia), and Mr. Anton V. Shchinov, Dr. Tran Cong Huan, Dr. Nguyen Dang Hoi, Dr. Bui Xuan Phuong, Tran Quang Tien, and Pham Mai Phuong (all from the Joint Russian-Vietnamese Tropical Research and Technological Centre, Hanoi, Vietnam), who made considerable efforts in preparing for a number of expeditions and who supplied us with a significant number of specimens. We also want to express our warmest gratitude to Dr. Giuliano Doria (Museo Civico di Storia Naturale “Giacomo Doria”, Genoa, Italy), Dr. Michael D. Carleton, Dr. Kristofer M. Helgen, Mr. Craig Ludwig and Ms. Renee Regan (all from the Smithsonian Institution; National Museum of Natural History, Division of Mammals, Washington, USA) for their kind assistance in the investigation of holotype specimens. Finally, we thank the administrations of Hoang Lien, Nam Cat Tien, Bi Dup-Nui Ba, Bu Gia Map, Lo Go Xa Mat, Dong Nai and Binh Chau National Parks and Nature Reserves for their aid in the management of our studies. We are also very grateful to Darrin Lunde, Dr. Kristofer M. Helgen, Dr. Guy G. Musser and several anonymous reviewers for their helpful and constructive comments on earlier versions of this manuscript. This study was supported in part by the Research Program “Living Nature: Modern State and Problems of Development” of the Presidium of the Russian Academy of Sciences.
Complete list of samples used for phylogenetic reconstructions
Data type: species data
The list of samples used for combined Cyt b+COI+IRBP+GHR analysis
Data type: species data
Cranial measurements, range, and standard deviation
Data type: species measurements
Explanation note: Cranial measurements, including range and standard deviation (SD), for Chiromyscus species from Vietnam (intact, adult skulls of both sexes).