The new species belongs to the family Rhacophoridae, a group of shrub-dwelling frogs that ranges from sub-Saharan Africa and South Asia to Japan, the Philippines and Sulawesi. The family has expanded rapidly in the scientific literature, growing from roughly 300 recognized species in 2009 to about 485 today, distributed across 22 genera. Within this family, the genus Zhangixalus, established in 2019, currently contains 46 species and is one of the dominant treefrog lineages in Vietnam, where 88 treefrog species are known. Recent molecular work has repeatedly revealed that what were once considered single widespread species are actually complexes of distinct, often cryptic relatives, a pattern that has driven a steady stream of new descriptions across China, Laos and Vietnam over the past decade.

The road to the new species began with a field survey conducted from 25 April to 1 May 2019, when researchers working in Y Ty Commune recorded a Zhangixalus population that differed visibly from its congeners. Five adult male specimens were collected, photographed alive, euthanized humanely, fixed in 80 percent ethanol and transferred to 70 percent ethanol for permanent storage at the Institute of Biology in Hanoi. Liver tissue samples were preserved separately in 96 percent ethanol before fixation, allowing the team to extract and amplify fragments of the mitochondrial 16S rRNA gene, a standard barcode marker in amphibian taxonomy.

The molecular analysis was extensive. The researchers assembled 71 sequences representing 32 species of Zhangixalus, together with six outgroup sequences from related genera including Leptomantis, Rhacophorus and Theloderma. The aligned 16S dataset spanned 1,046 nucleotide sites, of which 364 were variable and 309 were parsimony-informative within the genus. Phylogenetic trees were built using both maximum likelihood, implemented in IQ-TREE with ultrafast bootstrap support evaluated over 1,000 replicates, and Bayesian inference in MrBayes, run for 10 million generations with two independent chains. The best-fit substitution model selected under the Bayesian information criterion was TIM2+F+I+G4 for the likelihood analysis, with the closely related GTR model applied in the Bayesian framework.

The results placed the Y Ty frogs firmly within the Zhangixalus dugritei species group, in a well-supported clade alongside eight relatives. Their closest relatives proved to be Z. duboisi and Z. puerensis, with genetic distances of 2.52 to 3.63 percent and 3.32 to 3.53 percent respectively on the 16S marker. Across the genus as a whole, the new species diverges from other Zhangixalus by between 2.5 and 12.9 percent, a range that comfortably exceeds the thresholds typically used to justify species-level separation in this group. Both analytical methods recovered the same topology, lending confidence to the conclusion that the Vietnamese population represents an independent evolutionary lineage.

Morphology told a matching story. The new species is medium-sized, with males measuring 35.5 to 37.6 millimeters in snout-vent length, and is diagnosed by a striking combination of traits: a smooth olive-green dorsum dotted with numerous small brown spots, grey axilla and groin marked with cream spots along the flanks, a cream throat flecked with grey, and a vivid red-orange belly and ventral surface of the hind limbs. The iris is pale golden-bronze with black pupils, and the finger and toe webbing follows distinctive formulas that separate the species from its relatives. As in other Zhangixalus, the fingertips expand into large adhesive disks ringed by circum-marginal grooves, an adaptation to life on branches and foliage.

Distinguishing the new species from its closest look-alikes required careful comparison. Z. matsuii is smaller than Z. duboisi and bears an unblotched orange belly rather than the cream venter with dark marbling seen in that species. Compared with Z. puerensis, its head is as long as it is wide rather than longer than wide, and its underside is orange rather than pale white. Against the many larger members of the genus, the size difference alone is decisive: species such as Z. dennysi, Z. feae, Z. franki and Z. smaragdinus all exceed 50 millimeters in males, in some cases reaching more than double the body length of the new frog. The formal description rests on a holotype collected at 1,883 meters elevation in May 2019, together with four male paratypes from the same locality.

The species name honors Professor Masafumi Matsui of Kyoto University, whose decades of work on amphibian systematics have profoundly shaped the understanding of Asian frog diversity, particularly within Rhacophoridae. The authors also dedicated the paper to the memory of Dr. Nikolai Orlov, a co-author of the survey who passed away recently. In English the animal is suggested as Matsui’s treefrog, while the Vietnamese common name is ech cay matsui. All known specimens were found between 19:00 and 24:00 hours, perched on tree branches about a meter above the ground near a small stream, within undisturbed evergreen forest on granite substrate dominated by large hardwood trees with an understory of ferns and shrubs.

Notably, only males have so far been found; females, eggs and tadpoles remain unknown to science, and the species is currently recorded only from its type locality at elevations between 1,880 and 2,000 meters. Because so little is known about its abundance and true range, the authors recommend classifying it as Data Deficient under the IUCN Red List criteria. They speculate that the species may also occur in the evergreen forests of Guangxi in southern China, which shares similar granite mountain terrain, unlike the limestone karst landscapes that dominate neighboring Yunnan, but confirming this will require further fieldwork on both sides of the border.

Beyond the description itself, the study highlights how much remains to be learned about Zhangixalus in the mountains of northern Vietnam and southern China. Four new species have been described in the Z. nigropunctatus group since 2016 and four more in the Z. dorsoviridis group since 2017, while the lineage around Z. puerensis has received comparatively little attention, and questions persist about whether species such as Z. dugritei and Z. hui are truly distinct. The authors also found that Vietnamese populations identified as Z. duboisi differ genetically from Z. omeimontis by roughly 4.8 to 5.4 percent, underscoring the need for integrative taxonomic studies in these homomorphic species groups. With seven Zhangixalus species now recorded in Lao Cai Province alone, the region’s complex topography appears to be a cradle of speciation, and researchers suspect that more undescribed frogs are still waiting among its mist-covered ridges.

Subject of Research: Description of a new treefrog species, Zhangixalus matsuii, from northwestern Vietnam based on molecular and morphological evidence

Article Title: A new species of Zhangixalus ( Anura , Rhacophoridae ) from Lao Cai Province, Vietnam

Article References: Ninh, H. T., Hoang, C. V., Orlov, N., Ichioka, Y., Ziegler, T., & Nguyen, T. T. (2026). A new species of Zhangixalus (Anura, Rhacophoridae) from Lao Cai Province, Vietnam. ZooKeys, 1294, 109-130. https://doi.org/10.3897/zookeys.1294.203393

Image Credits: AI Generated

DOI: 10.3897/zookeys.1294.203393

Keywords: Zhangixalus matsuii, treefrog, Vietnam, Lao Cai Province, Rhacophoridae, new species, 16S rRNA, phylogenetics, amphibian taxonomy, cryptic species, herpetology, biodiversity