Research Article |
Corresponding author: Sylvia Hofmann ( s.hofmann@leibniz-zfmk.de ) Academic editor: Angelica Crottini
© 2021 Sylvia Hofmann, Rafaqat Masroor, Daniel Jablonski.
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:
Hofmann S, Masroor R, Jablonski D (2021) Morphological and molecular data on tadpoles of the westernmost Himalayan spiny frog Allopaa hazarensis (Dubois & Khan, 1979). ZooKeys 1049: 67-77. https://doi.org/10.3897/zookeys.1049.66645
|
Little is known about the life history, ecology, and distribution of the genus Allopaa (Dicroglossidae) and far less recent data are available about the larvae of this taxon. Here, we provide data on the larval stage of Allopaa hazarensis (Dubois & Khan, 1979) from northern Pakistan based on the examination of three tadpoles. Specimens were obtained from two sites in Buner, Khyber Pakhtunkhwa province, Pakistan. Morphological and genetic analysis (mtDNA and nDNA) confirmed the identity of the tadpoles as A. hazarensis. Tadpole characterizations were illustrated by detailed imagery. Basic measurements and details on oral apparatus provide relevant taxonomic characteristics to distinguish the tadpoles of this species from other spiny frogs. The illustration and description of the tadpole of A. hazarensis should facilitate the identification of this species in the field.
Chaparana, Dicroglossidae, DNA barcoding, larva, Paa, Pakistan, species identification
The Hazara (Torrent) Frog, Allopaa hazarensis (Dubois & Khan, 1979), belongs to the tribe Paini (Dicroglossidae), which are found across the Himalayan mountain arc from northern Afghanistan, Pakistan, and northern India, through Nepal, Sikkim, and Bhutan, and in the valleys of southern and eastern Tibet, eastwards to eastern China, and southwards to the mountains of Indochina (Myanmar, Thailand, Laos, northern Vietnam;
Allopaa hazarensis was described from near Datta (~34.30°N, 73.26°E), northern Pakistan (Manshera District, Hazera Division, about 1,200 m a.s.l.) and is known to occur in Khyber Pakhtunkhwa Province, Pakistan, and in adjacent Kashmir, India (see
In the present study, we provide detailed photographs and a brief description of A. hazarensis tadpoles from northern Pakistan. We used mitochondrial and nuclear DNA sequence data to validate the identity of our specimens by assigning them to existing Allopaa sequences. These recent data may support future research on this taxon in the Kashmir Himalaya and adjacent regions.
Two larvae (ZFMK 103353, ZFMK 103354) were collected in September 2020 during night time in Buner, Khyber Pakhtunkhwa province, Pakistan (34.66°N, 72.50°E, 1,520 m a.s.l.; Fig.
Sampling was conducted according to the regulations for the protection of terrestrial wild animals under the permits of the Pakistan Museum of Natural History, Islamabad, Pakistan [no. PMNH/EST-1(89)/05]. A small piece of the tail was taken from two of the tadpoles (ZFMK 103354, ZFMK 103351), transferred into absolute ethanol, and stored at −20 °C. All investigated specimens are deposited in the Zoological Research Museum Alexander Koenig, Bonn, Germany.
Tadpoles were staged according to
Genomic DNA was extracted from tissues using the DNeasy Blood & Tissue Kit (Qiagen, Venlo, The Netherlands) following the manufacturer’s protocol. Approximately 546 bp of the 16S ribosomal RNA (rRNA), 499 bp of the Cytochrome c oxidase I (COI), and a fragment of 1,207 bp of the Recombination activating gene 1 (Rag1) gene were amplified via the polymerase chain reaction (PCR) using primers and PCR conditions as previously described (
We aligned the new sequences (accession numbers 16S: MW723172–MW723173, COI: MW723177–MW723178, Rag1: MW728951–MW728952) to data available from our previous studies (
We confirmed the identity of the two tadpoles as Allopaa hazarensis. Our tadpole sequences nested in the clade of A. hazarensis that includes sequences from the type locality; the placement within this clade was highly supported (Suppl. material
Tadpoles identified as A. hazarensis have been described by Khan and Dubois (1979) and
Here we report on larvae of the dicroglossid frog Allopaa hazarensis. Our work is based on morphological and DNA data of A. hazarensis tadpoles and previously compiled molecular data sets (
Our new data mainly agree with previous descriptions (
Based on the original description, A. hazarensis has been assumed to be most closely related to Nanorana minica (Dubois, 1995), which has been reported to occur in Indian Uttar Pradesh and Himachal Pradesh, and in western Nepal (
Our tadpoles differ significantly from those of Nanorana vicina (Stoliczka, 1872), which is endemic to uplands in northern Pakistan and India. Compared to A. hazarensis, the number of tooth rows on the upper labia of the N. vicina tadpoles is fewer (5 vs. 7 or 8; no difference on lower labia), and the submarginal papillae are not cumulated laterally (
The illustration and description of the tadpole of A. hazarensis should facilitate the identification of this species, for example, during tadpole surveys in Pakistan. As the tadpoles of this species need several weeks to even months to complete development, they can be often more easily detected than their adult conspecifics, both at night and during the day. The sparse knowledge about the genus Allopaa, and particularly of their larval stages, requires more research to utilize the valuable tadpole data, especially for monitoring and conservation efforts.
We thank Morris Fleck for assistance with tadpole photographs, Sandra Kukowka, Anja Bodenheim and Jana Poláková for their technical support in the lab, and a number of students and colleagues of RM in Pakistan for their support during the fieldwork. This work was funded by the German Research Foundation (DFG, grant no. HO 3792/8-1 to SH), and by the Slovak Research and Development Agency (contract no. APVV-19-0076 to DJ).
Tables S1, S2, Figures S1, S2
Data type: occurences, morphological, and phylogenetic
Explanation note: List of Allopaa hazarensis specimens used in the present study; Measurements (in mm) and counts of voucher specimens (V-ID) of tadpole series of Allopaa hazarensis; Maximum-likelihood tree based on concatenated mtDNA and nuDNA sequence data; Tadpole (103351; Gosner stage 26) of Allopaa hazarensis from Buner, Pakistan.