First record of Amolops truongi Pham, Pham, Ngo, Sung, Ziegler & Le, 2023 (Anura, Ranidae) from China

We report the first record of Amolops truongi Pham, Pham, Ngo, Sung, Ziegler & Le, 2023 from China, based on four specimens collected in Yunnan Huanglianshan National Nature Reserve. This species was previously known only from the type locality in north-western Vietnam. Morphologically, the specimens from China correspond to the original description of A. truongi with a few variations and, phylogenetically, they are clustered with the type specimens of A. truongi from Vietnam with strong support. In this study, we also provide an updated diagnosis of this species combining the original description and new data.


Introduction
Amolops Cope, 1865 is a genus with a rapid increase in the number of species recently, making it the most speciose genus within the family Ranidae and having a wide distribution range from southern and eastern Himalayas to the Peninsular Malaysia (Dever et al. 2012;Pham et al. 2019;Wu et al. 2020;Mahony et al. 2022;Tang et al. 2023;Frost 2024;Li et al. 2024).Currently, 85 species are included in the genus (Frost 2024) and 52 have been recorded in China (AmphibiaChina 2024).However, it is important to note that the taxonomic systems of Frost (2024) and AmphibiaChina (2024) have different views on the validity of some species, thus not allowing a direct comparison of the total number of spe-

Material and methods
The fieldwork was carried out under the permission of Yunnan Huanglianshan National Nature Reserve Management and Protection Bureau.Specimens were collected by hand at night.Liver tissues were stored in analytical pure ethanol and specimens were preserved in 75% ethanol.The newly-collected specimens were deposited at Kunming Natural History Museum of Zoology, Kunming Institute of Zoology, Chinese Academy of Sciences (KIZ).
Measurements were taken with a digital caliper to the nearest 0.1 mm.The methodology of measurements followed Pham et al. (2023) and the terminology describing the webbing formula followed Glaw and Vences (2007).The following morphological characteristics were recorded: snout-vent length (SVL), from the tip of the snout to the cloacal; head length (HL), from the back of the mandible to the tip of the snout; maximum head width (HW), across the angles of the jaws; snout length (SE), distance from the tip of the snout to the anterior corner of the eye; nostril to snout distance (SND), from the nostril to the tip of the snout; eye to nostril distance (END), from the anterior corner of the eye to the nostril; internarial distance (IND), the distance between nostrils; upper eyelid width (UEW), the maximum width of the upper eyelid; interorbital distance (IOD), the minimum distance between the upper eyelids; horizontal eye diameter (ED), from the anterior corner to the posterior corner of the eye; tympanum diameter (TD), the maximum diameter of the tympanum; tympanum-eye distance (TED), from the anterior margin of the tympanum to the posterior corner of the eye; fore-limb length (FLL), from the tip of the disc of the third finger to the axilla; finger III disc width (FTD), the maximum width of the disc of the third finger; hindlimb length (HLL), from the tip of the disc of the fourth toe to the groin; femur length (FL), from the vent to the knee; tibia length (TL), from the knee to the tarsus; foot and tarsus length (FOT), from the tarsus to the tip of the fourth toe; toe IV disc width (HTD), the maximum width of the disc of the fourth toe; inner metatarsal tubercle length (MTTi), the maximum length of the inner metatarsal tubercle.
The sequences were aligned using ClustalW (Thompson et al. 1994) integrated in MEGA X and the genetic divergences (uncorrected p-distance) were calculated in MEGA X (Kumar et al. 2018).The best substitution models GTR+F+I+G4 for Bayesian Inference and TIM+F+I+G4 for Maximum Likelihood phylogenetic analysis were selected using the Akaike Information Criterion (AIC) in ModelFinder (Kalyaanamoorthy et al. 2017).Maximum Likelihood phylogenetic analysis was performed in IQ-TREE 1.6.12(Nguyen et al. 2015) with the nodal support estimated by 1,000 ultrafast bootstrap replicates.Bayesian Inference was performed in MrBayes 3.2.7 (Ronquist et al. 2012) and the Markov chains run for 1,000,000 generations and sampled every 100 generations.The first 25% of the sampled trees were discarded as burn-in and the remaining trees were used to estimate Bayesian posterior probabilities.

Results
The morphological measurements of the specimens from China are presented in Table 2.There is no significant difference in morphological characteristics between the specimens from China and the type specimens of Amolops truongi from Vietnam, except for minor variations.Genetically, Bayesian Inference and Maximum Likelihood phylogeny obtained the same topology, with slightly different supports for some nodes.The sequences of the specimens from China clustered with the type specimens of A. truongi from Vietnam with strong support (Fig. 1).The genetic distance (uncorrected p-distance) between the specimens from China and the type specimens of A. truongi was only 0.4% (Table 3).Description of the specimens from China.Male body size relatively small, SVL 39.3-39.9mm in adult males; head moderate long (HL/SVL 0.36-0.38),longer than wide (HL/HW 1.09-1.15);snout relatively long (SE/SVL 0.16), projecting beyond lower jaw; canthus rostralis distinct; loreal region concave; distance from nostril to snout tip equal to or slightly greater than distance from eye to nostril (SND/END 1.00-1.03);internarial distance greater than interorbital distance (IND/IOD 1.22-1.39);upper eyelid width narrower than interorbital distance (UEW/ IOD 0.78-0.94);pupil oval, horizontal; tympanum distinct (TD/ED 0.35-0.40);tympanum-eye distance smaller than tympanum diameter (TED/TD 0.77-0.90);vomerine teeth present; choanae rounded; tongue cordiform, notched posteriorly; vocal sac opening on floor of mouth at corner, sac-like gular pouch, front margin positioned near to level of centre of orbit.
Dorsal and lateral surface of head and body smooth with few very small tubercles present on temporal sides of head, above tibiae and vent; supratympanic fold indistinct; dorsolateral fold distinct; ventral surface smooth with flat tubercles on basal ventral thigh.
Colouration in life.Dorsal sides of head and body green or olive brown with some black dots; lateral side of head and tympanum dark brown or black; a white stripe extending from tip of snout to shoulder on each side; iris pale gold; flanks light brown or brown; dorsal surface of fore-limbs and hind-limbs light brown or brown with dark bands; throat, chest and belly cream with some brown dots; vocal sac orange or light yellow; ventral surface of fore-limbs light red or flesh-coloured; ventral surface of hind-limbs red flesh-coloured or flesh-coloured with some dark brown dots; toe webbing dark brown.

Discussion
The morphological characteristics of the specimens of Amolops truongi from China mostly agree with the original description by Pham et al. (2023), but there are also a few differences between them.The distance from the nostril to the snout tip is equal to or slightly greater than the distance from the eye to the nostril in the specimens from China, while the distance from the nostril to the snout tip is smaller than the distance from the eye to the nostril in the original description.The upper eyelid width is narrower than the interorbital distance in the specimens from China.while the upper eyelid width is wider than the interorbital distance in the original description.In addition, according to the original description of this species, the colour of the dorsum is light grey, while some specimens from China have a green dorsum.Therefore, based on these variants, we revised the diagnosis of this species.
Yunnan Huanglianshan National Nature Reserve is located in the south of Yunnan Province, it has a variety of terrain and vegetation types, which has long established its rich species diversity.In recent years, many new species have been discovered in this region, such as frogs In addition, Yunnan Huanglianshan National Nature Reserve borders Vietnam in the south, so it is not surprising to find species that were previously considered to be distributed only in Vietnam.Previously, 52 species of the genus Amolops were recorded in China, 20 of which were distributed in Yunnan (Tang et al. 2023;Li et al. 2024).Our discovery of A. truongi in Yunnan increased the number of Amolops species in China to 53 and the number of Amolops species in Yunnan to 21.   17.2 13.1 15.1 13.0 15.4 14.2 16.5 16.5 9.5 15.0 10.3 13.9 15.0 14.5 14.5 14.8 16.9 16.8 20 Amolops vitreus 14.4 15.6 14.6 16.9 14.7 13.9 11.1 12.1 16.0 14.8 16.1 13.4 18.6 14.0 15.7 14.6 12.0 11.5 14.8 21 Amolops wenshanensis 15.0 15.5 14.3 16.6 15.1 13.6 6.8 6.7 15.8 15.7 15.3 14.0 17.1 13.9 14.8 14.7 7.1 6.8 14.5 10.2   Amolops truongi was previously known only from the type locality in north-western Vietnam (Pham et al. 2023).The new collection site in China extends the distribution range of this species to the northwest by approximately 290 km.The altitude of the type locality is 1360 m a.s.l., while the altitude of the new collection site in China is 1430 m a.s.l., the altitude difference between the two localities not being significant.According to Pham et al. (2023), the type specimens of this species were found on trees or limestone cliffs nearby a stream; however, the newly-collected specimens from China were all found on herbaceous plants nearby a river (Fig. 5), this being slightly different from those at the type locality.In addition, Pham et al. (2023) did not mention the courtship calls and breeding season of the species.During the surveys, we only collected males and did not find females or tadpoles and did not hear the courtship calls of males.Therefore, we are also unable to determine the breeding habits of this species.More field observations are needed to understand the habits and ecological information of A. truongi.the fieldwork.We would like to thank the editors and reviewers for their valuable comments on the manuscript.This work was supported by the Project of Huanglian Mountains National Nature Reserve Animal Diversity Expedition (Grant No. E2023HLS001), National Natural Science Foundation Project: Classificatory and Phylogenetic Studies on the Amolops frogs of China (Grant No. NSFC-31772424) and the project of the Ministry of Ecology and Environment of China: Investigation and assessment of amphibians and reptiles in southern Yunnan.

Figure 1 .
Figure 1.The Bayesian phylogenetic tree, based on the ND2 sequences.The numbers after and behind the "/" are the Bayesian posterior probabilities and the Maximum Likelihood ultrafast bootstrap values (> 0.90/90), respectively.

Figure 2 .
Figure 2. Dorsal view (top) and ventral view (bottom) of the specimens of Amolops truongi from China in preservative.

Figure 3 .
Figure 3. Dorsal, lateral and ventral views of the male specimen KIZ2023081 (A-C) and dorsal, lateral and ventral views of the male specimen KIZ2023082 (D-F) of Amolops truongi from China in life.

Figure 5 .
Figure 5. Habitat of the specimens of Amolops truongi collected in China.

Figure 4 .
Figure 4. Map showing the type locality of Amolops truongi in north-western Vietnam (black star) and the new collection site from Lvchun County, Yunnan Province, China (black dot).

Table 1 .
Samples used for the phylogenetic analyses in this study.

Table 2 .
Measurements (in mm) and proportions of the specimens of Amolops truongi from China (for abbreviations, see Material and methods).