Anna B. Vassilieva

Two new species of Kalophrynus Tschudi, 1838 (Anura: Microhylidae) from the Annamite mountains in southern Vietnam.

We describe two new mountain-dwelling microhylid species of the genus Kalophrynus from the southern part of the Annamite Mountains in Vietnam. The two new species differ from all known congeners in morphological characters and mtDNA; phylogenetically, they form a sister clade to the large-bodied K. interlineatus (1009 bp, 16S rRNA gene, mtDNA). Both species share the following characteristics: snout pointed in dorsal and lateral views, slightly sloping in profile; tympanum distinct, smaller than eye in diameter; toe webbing moderate; outer metatarsal tubercle present; light dorsolateral line absent. Kalophrynus cryptophonus sp. nov. from Loc Bao, Lam Dong Province is a small-sized species distinguishing from its congeners by a combination of: SVL 27.9-30.4 mm in males, 23.4 mm in female; canthus rostralis indistinct; males with large sharp conical spines on the skin covering mandible margins and finely asperous nuptial pads on the dorsal surface of fingers I-III; dark ocelli in the inguinal region usually present, small, without a light border; anterior palatal dermal ridge short, restricted to medial part of palate. Kalophrynus honbaensis sp. nov. from Hon Ba, Khanh Hoa Province is a medium-sized Kalophrynus, distinguishing from its congeners by a combination of: SVL 26.7-36.8 mm in males; canthus rostralis distinct; males without distinguishable spines on the mandible margins nor the nuptial pads; dark ocelli in the inguinal region present, large, without a distinct light border, anterior palatal dermal ridge developed, parallel to posterior one. Kalophrynus cryptophonus sp. nov. reproduces in hollow bamboo stems; we describe larval morphology and bioacoustics of this species in relation to phytotelm breeding. A review of the distribution of the genus Kalophrynus in Indochina is provided.

Rhacophorus vampyrus (Anura: Rhacophoridae) Reproductive Biology: A New Type of Oophagous Tadpole in Asian Treefrogs

Abstract We report on the reproductive biology of Rhacophorus vampyrus (Rhacophoridae, Anura) from the Lang Bian Plateau in southern Vietnam. Unlike most other members of the genus Rhacophorus, R. vampyrus is a phytotelm breeder and reproduces in water-filled tree hollows. The species shows a unique tadpole mouthpart morphology not reported for any other anurans: presence of a specific serrated horny arch on the upper jaw and two large, fang-like horny teeth on the lower jaw. This, together with the presence of a large extensible stomach and relatively short digestive tract length, indicates an adaptation toward macrophagous feeding. We observed two clutch types in R. vampyrus: fertilized eggs included in a typical foam nest and unfertilized eggs, apparently having a trophic function, enveloped by dense mucous. The digestive tracts of all tadpoles examined contained up to 40 trophic eggs and showed no remains of any other food. Keratinous mouthparts undergo considerable remodeling during tadpole development presuming the change of feeding mode in ontogeny. We speculate that these findings indicate that the tadpole of R. vampyrus is an obligatorily and highly specialized oophagous type with unique mouthpart structures. The advanced form of parental care, including the maternal provisioning of unfertilized eggs, is reported for the first time for the genus Rhacophorus and is the second documented record for the Rhacophoridae.

Molecular, morphological and acoustic assessment of the genus Ophryophryne (Anura, Megophryidae) from Langbian Plateau, southern Vietnam, with description of a new species

Abstract Asian Mountain Toads (Ophryophryne) are a poorly known genus of mostly small-sized anurans from southeastern China and Indochina. To shed light on the systematics within this group, the most comprehensive mitochondrial DNA phylogeny for the genus to date is presented, and the taxonomy and biogeography of this group is discussed. Complimented with extensive morphological data (including associated statistical analyses), molecular data indicates that the Langbian Plateau, in the southern Annamite Mountains, Vietnam, is one of the diversity centres of this genus where three often sympatric species of Ophryophryne are found, O. gerti, O. synoria and an undescribed species. To help resolve outstanding taxonomic confusion evident in literature (reviewed herein), an expanded redescription of O. gerti is provided based on the examination of type material, and the distributions of both O. gerti and O. synoria are considerably revised based on new locality records. We provide the first descriptions of male mating calls for all three species, permitting a detailed bioacoustics comparison of the species. We describe the new species from highlands of the northern and eastern Langbian Plateau, and distinguish it from its congeners by a combination of morphological, molecular and acoustic characters. The new species represents one of the smallest known members of the genus Ophryophryne. At present, the new species is known from montane evergreen forest between 700–2200 m a.s.l. We suggest the species should be considered Data Deficient following IUCN’s Red List categories.

Development of the bony skeleton in two salamander species (Mertensiella caucasica and Chioglossa lusitanica: Salamandridae) with partial embryonization

168 The Caucasian salamander (Mertensiella cauca sica) and golden striped salamander (Chioglossa lusi tanica) are the most closely related taxa in the group of true salamanders belonging to the family Salaman dridae [1, 2]. In addition, they stand out among the other salamanders by a considerable similarity in their morphological constitution [3] and life style [3–5]. A study on the reproduction biology of these species has also demonstrated that they are very similar in a num ber of reproductive parameters, namely, both species reproduce in brooks and lay large (5–7 mm in diame ter) eggs in small clutches (on average, 17–18 eggs) [4–6]. These traits distinguish M. caucasica and C. lusitanica from the majority of the other true sala manders, characteristic of which is ovoviviparity or viviparity, and the remaining members of the family Salamandridae, newts, which lay considerably smaller eggs. On the other hand, the large size of the eggs and hatching of larvae at advanced development stages (as was demonstrated for M. caucasica [7]) suggest a pro nounced embryonization of development, i.e., transi tion of the initial stages in larval development to the embryonic period. Among amphibians, this evolutionary process leads to establishment of a direct development (lacking the larval stage), which is present only in members of the family Plethodontidae within the order Urodela. The effect of the degree of embryonization on the specific morphoge netic traits and, in particular, craniogenesis, has been also demonstrated for these amphibians [8, 9]. As we see it, a study of the skeletal development in the amphibian spe cies with partial embryonization of development will help to shed light onto the initial stages in the evolution path way that had led to direct development. For this purpose, we have studied the skeletal development of M. caucasica and C. lusitanica in the ontogenesis. Clutches of both salamander species were incubated under laboratory conditions; the embryos at late developmental stages (staring from limb differen tiation), larvae, and metamorphosing animals were regularly fixed with 10% formaldehyde and stained with alizarin red and alcian blue to detect bones and cartilages, respectively. The developmental stages were determined according to the table of M. caucasica normal development [7], which, except for some col oration traits, equally well described the C. lusitanica developmental stages. GENERAL BIOLOGY

The phytotelm tadpoles of Microhyla arboricola (Anura: Microhylidae) from Vietnam, with comments on reproductive biology and development

The reproductive biology of Microhyla arboricola (Microhylidae) was studied in two regions of the southern Annamite Mountains in Vietnam. M. arboricola is an obligate phytotelm-breeder that reproduces in water-filled tree hollows in montane evergreen forests. Clutches are attached above the water level in the hollows and contain 5-37 pigmented, relatively large eggs. Larvae hatch at markedly advanced stages and develop in water until metamorphosis is completed. The developing tadpoles are obligately oophagous and feed on conspecific eggs and embryos. M. arboricola tadpoles differ from tadpoles of pond-dwelling Microhyla species in their external morphology (extremely long tails, dorsolateral position of the eyes, dark pigmentation), digestive tract morphology (large, extensible larval stomach and short intestine), and oral morphology. The larval chondrocranium and hyobranchiumof M. arboricola is described. M. arboricola shares its habitat with other hollow-breeding species of anurans. To date, M. arboricola is the only known arboreal species of the genus Microhyla that has a unique reproductive mode. The ecological niche of this species differs greatly from those occupied by other microhylids of Indochinese Peninsula.

Role of heterochronies in the morphogenesis of amphibian skull bones: An experimental study

Heterochronies, i.e., changes in the relative timingof the development of different morphological struc-tures, are widespread in amphibians, which makesthese animals a suitable object for studying both naturaland experimental heterochronies. Temporal parametersof skull morphogenesis, including the timing and rateof the development of cranial elements, are affected bymany factors, including endocrine ones, the main ofwhich are hormones synthesized by the thyroid glandor thyroid hormones (THs). The use of exogenous THsand goitrogens (substances suppressing the thyroidsecretion, thereby decreasing the plasma content ofendogenous THs) allows artificial simulation of hetero-chronies.Experimental heterochronies are expressed in alter-ations of the rate and order of the formation of skullbones depending on the hormonal background of theanimal development. For example, exogenous THsaccelerate bone growth, whereas goitrogens, con-versely, retard it. In addition, high doses of THscause premature development of the skull ossifica-tions that normally occur at the ontogenetic stagesclose to metamorphosis, and goitrogens retard thedevelopment of these bones or prevent their appear-ance altogether [1–3].The study of the role of THs in the control of cran-iogenesis in various species of Urodela and Anura asdescribed in [1–3] demonstrated that THs and goitro-gens could alter not only the rate and timing of thedevelopment of skull bones, but also the morphogeneticpattern of some bones.For example, a dermal bone that is located in thenasal region of the skull of the Siberian salamanderSalamandrella keyserlingi (Urodela, Hynobiidae) andis traditionally termed the nasale develops from twoossification centers, one of which appears in the tectumnasale, and the other, on the tectum internasale cartilagemedial to former. According to Lebedkina [4], they arehomologous to the nasale and postrostrale of Crossop-terygii, respectively. Both ossification centers soon fusebehind the apex of the ascending process of the prae-maxillare to form a single bone, the naso-postrostrale.However, the naso-postrostrale of animals growingunder the conditions of an enhanced hormonal back-ground (treated with exogenous THs) [1] is morpholog-ically typical but develops from a single ossificationcenter.The frontoparietale of the common frog Rana tem-poraria (Anura, Ranidae) normally develops from oneossification center. Thiourea (TU, a goitrogen) retardsits appearance, the retardation being longer, the higherthe goitrogen concentration. In some of animals kept ina 0.02% TU solution, two ossification centers appear atthe site of the future frontoparietale. In animals kept in0.04% TU, the frontoparietale always develops fromtwo separate centers, which partly fuse later in ontog-eny [3].The dermal bone angulospleniale, part of the mandi-ble of

Tadpole of Microhyla picta Shenkel (Anura: Microhylidae), an endemic narrow-mouthed frog from Vietnam

The Painted Rice Frog, Microhyla picta Schenkel, is one of the least studied species among narrow-mouthed frogs of the genus Microhyla. To date it is known only from Vietnam, where it is distributed mostly in eastern coastal areas in central and southern parts of the country (Nguyen et al. 2009; Nguyen Hoang 2013). The species was also found in Con Dao archipelago (Poyarkov Vassilieva 2011). No data are available on the ecology and reproductive biology of the species, and its larval morphology is also not described. Microhyla frogs are widespread and numerous in terrestrial ecosystems and microhylid tadpoles are known to play an important role in the aquatic communities of temporary waterbodies of monsoon ecosystems in Southeast Asia (Heyer 1973; Vassilieva et al. 2017). The ability to identify microhylid tadpoles in the field is pertinent for biodiversity and ecological studies. We provide a description of larvae of M. picta from various areas in southern Vietnam.

Heterochronies in the cranial development of Asian tree frogs (Amphibia: Anura: Rhacophoridae) with different life histories

The development of bony skull was studied in four species of Asian tree frogs (Rhacophoridae) with different life histories: biphasic development with free larval stage and direct development. In biphasic rhacophorids the sequence of the appearance of cranial bones generally followed the generalized pattern of craniogenesis, which was described for most studied anurans. In contrast, direct-developing species displayed some heterochronies in the formation of skull bones, namely, the accelerated formation of the anlagen of jaw and suspensorium bones. The obtained results support that the embryonization in amphibians is regularly accompanied by a heterochronic repatterning of craniogenesis, rather similar in different phyletic groups.

Larval morphology and complex vocal repertoire of Rhacophorus helenae (Anura: Rhacophoridae), a rare flying frog from Vietnam

We present new data on the distribution, reproduction, larval morphology and vocalization of Rhacophorus helenae (Rhacophoridae), a narrowly distributed frog from southern Vietnam. Two new populations of R. helenae were discovered during field surveys in the lowland monsoon forests in Dong Nai and Ba Ria-Vung Tau provinces in 2010-2013. Spawning was observed in May 2013. Egg clutches containing small (2.3±0.1 mm) unpigmented eggs were embedded in a foam nest and suspended high on trees above temporary ponds. The tadpoles of R. helenae have a morphology typical of pond-dwelling Rhacophorus larvae with a moderate tail length and a labial tooth row formula of 5(2-5)/3. Postmetamorphic juveniles differed from adult frogs in the features of their coloration and less developed webbing. The complex vocal repertoire of R. helenae included five types of tonal, wideband and pulsed calls and several transitional signal types differentiated by frequency and amplitude parameters. Calls were uttered as singular signals (pulsed calls) or within non-stereotyped series of variable duration (other call types). The complex structure of the advertisement call markedly distinguishes R. helenae from other members of the Rhacophorus reinwardtii species complex.

A new species of the genus Oligodon Fitzinger, 1826 (Squamata: Colubridae) from coastal southern Vietnam

A new species of the genus Oligodon from the coastal area of Binh Chau-Phuoc Buu Nature Reserve, Ba Ria-Vung Tau Province, southern Vietnam, is described. Oligodon arenarius sp. nov. is distinguishable from all other species by the unique combination of the following characters: medium size; 17 dorsal scale rows; 6-8 maxillary teeth, the posterior three being enlarged; head scalation lacking a loreal but usually including a presubocular; divided nasal; two postoculars; 131-144 ventrals; 36-60 subcaudals; unforked hemipenis, without spines or obvious papillae; sexual dimorphism displayed in the number of subcaudals (36-40 in females, 58-60 in males) and a relative tail length, tails being quite long in males (TaL/TL = 0.26-0.28) and moderate in females (TaL/TL = 0.13-0.17); head coloration pattern including ocular band, temporal bands and chevron-shaped mark on nape; dorsal coloration without distinct pattern, uniform or with dark speckling; ventrals pinkish in life, immaculate. To date, this species appear to be endemic to Vietnam.