Range extension of
Ferguson’s Toad Duttaphrynus scaber(Schneider) (Amphibia: Anura:Bufonidae) up to the northern most limitof Western Ghats, with its advertisement call analysis
Anand Padhye1, Rohan Pandit2, Rajgopal Patil3, Swapnil Gaikwad4, Neelesh Dahanukar5 & Yogesh Shouche6
1 Department of Zoology, Abasahab Garware College, Karve Road, Pune, Maharashtra 411004, India
2 Department of Biodiversity, Abasahab Garware College, Karve Road, Pune, Maharashtra 411004, India
3 Ela Foundation,
C-9, Bhosale Park, Sahakarnagar-2, Pune, Maharashtra
411009, India
4,6 National Center for Cell
Sciences (NCCS), Ganeshkhind, Pune, Maharashtra
411007, India
5 Indian Institute of Science
Education and Research (IISER), Sai Trinity, Sus Road, Pashan, Pune, Maharahtra 411021, India and Zoo Outreach Organization, 96 Kumudham Nagar, VilankurichiRoad, Coimbatore, Tamil Nadu 641035, India
1 adpadhye@gmail.com
(corresponding author), 2 rohanpandit87@gmail.com, 3 rajnpatil@gmail.com,4 swapy28@gmail.com, 5 n.dahanukar@iiserpune.ac.in,6 yogesh@nccs.res.in
doi: http://dx.doi.org/10.11609/JoTT.o3345.4579-85 | ZooBank: urn:lsid:zoobank.org:pub:14242B29-CB85-461F-A8E4-81C59C225BE8
Editor: Anonymity requested. Date
of publication: 26 July 2013 (online & print)
Manuscript details: Ms #
o3345 | Received 13 September 2012 | Final received 24 June 2013 | Finally
accepted 29 June 2013
Citation: Padhye,
A., R. Pandit, R. Patil, S.Gaikwad, N. Dahanukar &
Y. Shouche (2013). Range extension of Ferguson’s ToadDuttaphrynus scaber (Schneider) (Amphibia: Anura:Bufonidae) up to the northern most limit of Western
Ghats, with its advertisement call analysis. Journal of Threatened Taxa5(11): 4579–4585; http://dx.doi.org/10.11609/JoTT.o3345.4579-85
Copyright: © Padhyeet al. 2013. Creative Commons Attribution 3.0 Unported License. JoTTallows unrestricted use of this article in any medium, reproduction and
distribution by providing adequate credit to the authors and the source of
publication.
Funding: None.
Competing Interest: None.
Acknowledgements: We
thank authorities of NCCS for providing the facility for molecular work. We
also thank Principal Abasaheb GarwareCollege as well as head of the Zoology Department for the infrastructural
facilities. Thanks to Ankur Padhye,Makarand Bhagwat, Vinayak Khade, Vitthal Bhoye and driver Anil Pujari who helped us in the field surveys. Special thanks to Dr. H.V. Ghate for his
valuable suggestions during the work and preparation of manuscript.
Abstract: Duttaphrynus scaber (Schneider) is known to
occur in the southern Western Ghats, Eastern Ghats and northeastern India. However, there is no report of its
occurrence from the northern Western Ghats from the states of Maharashtra and
Gujarat. Here we report the
occurrence of this species from the northernmost limit of the Western Ghats, a
substantial range extension (approx. 550km) from the nearest known
locality. We confirm our
identification on the basis of morphology by comparing our specimens with a previously collected Duttaphrynus scaber specimen from Thrissur in Kerala State as well as on the
molecular basis. Analysis based on
the advertisement call of Duttaphrynus scaber, albeit preliminary, is provided for the first
time for this species.
Keywords: Advertisement call analysis, Bufo fergusonii, Duttaphrynus scaber, range
extension.
The publication of this article is
supported by the Critical Ecosystem Partnership Fund (CEPF), a joint initiative
of l’Agence Française de Développement, Conservation International, the
European Commission, the Global Environment Facility, the Government of
Japan, the MacArthur Foundation and the World Bank.
For figures, images, tables -- click here
Duttaphrynus scaber (type locality ‘Orientali India’) was described by Schneider (1799). Boulenger(1892) described Bufo fergusonii (type locality ‘on the Cavalry Parade ground’ Thiruvananthapuram, Kerala)
as a different species. However,
after examining the lectotypes of D. scaber and comparing them with the holotypeof B. fergusonii, Dubois & Ohler (1999) found these two species to be conspecific and
hence they considered B. fergusonii as a
junior subjective synonym of D. scaber.
Duttaphrynus scaber has been reported in India
from Thriuvananthapuram in Kerala (Daniels 2005);
Mysore (Daniel 1963), Lakkavalli State Forest
(Krishnamurthy 1999) and Dharwad (Dutta1997) in Karnataka; Chennai in Tamil Nadu (Rao 1915; Ravichandran 1996; Daniels 2005); BanjaraHills, Hyderabad (Donahue & Daniel 1966) in Andhra Pradesh and Sambalpur District (Dutta 1988)
in Odisha. Recently the species has been recorded from the northeastern state of
Manipur by Mathew & Sen (2009, 2010). Outside of India, the species has also
been reported from Sri Lanka (Dutta & Manamendra-Arachchi 1996). Largely because of its wide
distribution, IUCN Red List status of this species is Least Concern (Dutta & Manamendra-Arachchi2004).
To date, Duttaphrynus scaber has not been reported from Maharashtra and
Gujarat states, despite a number of amphibian surveys having been carried out (Padhye & Ghate 2002 and
references therein; Naik & Vinod1992, 1993a,b; Bhatta et al. 1999; Vyas 2000a,b, 2002, 2004a,b, 2007, 2008, 2012; Dinesh et
al. 2009). There is a record of Bufo fergusonii (now Duttaphrynus scaber)
in the list of amphibians from ShoolpaneshwarWildlife Sanctuary, Gujarat (Sabnis & Amin 1992)
but Vyas (2008; 2012) has raised the question about
its validity. According to Vyas (2008), Naik & Vinod (1992, 1993a,b) never claimed any records of the
species from Shoolpaneshwar Wildlife Sanctuary, which
shows that the record of this toad species has been mistaken.
Therefore, we report this
species from both the states with molecular evidence for the first time and
describe its advertisement call.
Materials and Methods
We carried out amphibian
surveys of Surgana Talukain Nasik District, Maharashtra State and Ahwa Taluka, in Dang District, Gujarat State to assess the
amphibian diversity of the northern most regions of the Western Ghats. Surveys
were conducted from 1830-0230 hr on the 17 and 18 of
July 2010. Specimens were
identified in the field down to the species level and most were released. The presence of species was also
reported on the basis of calls. Two
specimens of Duttaphrynus scaber were collected from a road
side puddle and brought back to the laboratory for confirmation. One of the two specimens was preserved
in absolute ethanol and the other was preserved in 10% formalin and then transferred
to 70% ethanol. These specimens
were deposited at the museum of Zoology Research Laboratory, Abasaheb Garware College, Pune,
under the accession numbers AGCZRL Amphibia 41 and
AGCZRL Amphibia 42, respectively. Morphometrywas performed using digital Vernier caliper (Areospace® China, least count 0.01mm).
Thigh muscles of AGCZRL Amphibia 41 were used for the molecular analysis. The tissue was digested at 500C
for two hours using the extraction buffer (0.1M NaCl,
0.05M Tris-HCl, 0.01M EDTA, 1%SDS) with 15µl
Proteinase K (20mg/ml). DNA was
then extracted using the conventional phenol:chloroform method (Sambrooket al. 1989). The partial fragment
of each of the mitochondrial ribosomal RNA genes 12S (~400 bp)
and 16S (~520 bp) were amplified using PCR. The 12S segment of mitochondrial DNA was
amplified using universal primers L1091 (5’-AAACTGGGATTAGATACCCCACTA-3’) and
H1478 (5’-GAGGGTGACGGGCGGTGTGT-3’) as described by Kocher et al. (1989). While for 16S rDNAwe used primers 16SA (5’-CGCCTGTTTATCAAAAACAT-3’) and 16SB
(5’-CCGGTCTGAACTCAGATCACGT -3’) as described by Simon et al. (1994). PCR products were
purified by using PEG-NaCl protocol (Sambrook et al., 1989). Products were sequenced on ABI 3730 DNA
analyzer (Applied Biosystems, Hitachi), as per the
manufacturer’s instructions.
Sequences were manually
checked for any ambiguity using BioEdit (Hall 1999)
and were deposited in GenBank(http://www.ncbi.nlm.nih.gov/) under the accession number JQ898085 (12S rDNA) and JQ898086 (16S rDNA).
Additional sequences of 12S and 16S rDNA for
comparison were downloaded from GenBank. We retrieved
12S and 16S rDNA sequence for Duttaphrynus scaber (FJ882785), Duttaphrynus melanostictus (DQ283333), Duttaphrynus stomaticus (FJ882787), Duttaphrynus parietalis (FJ882784), Duttaphrynus hololius (FJ882781), Duttaphrynus brevirostris (FJ882786), Pedostibes tuberculosus (FJ882793), Xanthophryne koynayensis (FJ882782) and Ansonia ornata (FJ882797) for comparision. Microhyla ornata from family Microhylidaewas used as an out-group and 12S (AB201176) and 16S (AY948728) sequence of the
same were retrieved from GenBank. Sequences were aligned independently for
each gene using MUSCLE (Edgar 2004). The individual gene sequences were trimmed using freeware DAMBE 5.2.77
(Xia & Xie 2001). Both the gene sequences were
concatenated before performing the phylogenetic analysis. Phylogenetic and
molecular evolutionary analyses were conducted using MEGA5 (Tamura et al.
2011). Best fitmodel for nucleotide substitution was selected based on minimum Akaike Information Criterion (AIC) value (Posada &
Crandall 2001). We constructed
phylogenetic trees based on maximum parsimony and maximum likelihood.
Reliability of the phylogenetic tree was estimated using bootstrap values run
for 1000 iterations.
Advertisement calls of two
frogs were recorded with a mobile phone in AMR (Adaptive Multi-Rate) format at
8kHz sampling rate. Because the
calls recorded on the mobile phone have limited sampling frequency we provide a
very preliminary account of the advertisement call. Nevertheless, our analysis is important
since there is no available information on the calls of this species. Calls were recorded at a distance of
approximately 0.5m from the calling frogs. The toads were sitting on the sides of a temporary rain-waterpool. The atmospheric temperature was 21.20C. Calls were analysedwith Raven Pro 1.4 software (Bioacoustics Research Program 2003) and Praat sound analysis program version 5.2 (Boersma & Weenink 2009). Audio spectrograms were calculated with
fast-Fourier transform (FFT) of 256 points, 50% overlap and 31.3 Hz grid-spacing, using Hanningwindows. We tested the hypothesis
that all different call parameters were same for both the individuals by
performing unpaired t test.
Results and Discussion
During our field surveys we
observed a total of three D. scaber. A single
male D. scaber (not collected) was observed on
17 July 2010 on the banks of Ahwa Lake (20045’9”N
& 73040’39”E, elevation 455m), Ahwa,
District Dang, Gujarat at 2340hr. This male was calling in the open on the
edge of the reservoir (Image 1). The two collected males were found calling in roadside puddles, which
were flanked by paddy fields (20037’18”N & 73038’56”E,
elevation 356m). These males were
collected on 18 July 2010 at Supdahad, Ahwa Taluka, Dang District,
Gujarat, at 0050hr. There was
sporadic rain on the same day. Further, calling individuals of this species
were observed near the Village Satkhamb, Taluka Surgana, District Nasik,
Maharashtra (20035’31”N & 73034’47”E, elevation
385m). Judging from the number of
calling males heard, the species appears to be locally abundant. The overall habitat was road surrounded
by paddy fields and fragmented patches of deciduous forest (Image 2). Our new records of D. scaber represent a substantial northwestern range
extension to the northernmost limits of the Western Ghats of India (Fig. 1).
Duttaphrynus scaber individuals were identified
by the presence of distinct parietal ridges on head, numerous spiny warts on the
entire body, a fawn coloured underside, the 1stand 2nd fingers equal in length, toes scarcely webbed, parotid
glands rounded and tympanum less than half the diameter of eye (Daniel 1963;
Dubois & Ohler 1999; Daniels 2005; Mathews & Sen 2010) (Image 3a–c). The specimens were also compared with a
previously collected museum specimen from Thrissur, Kerala(Voucher No. AGCZRL Amphibia 98),
which was identified by Dr. Annemarie Ohler (Museum
National d’Histoire Naturelle,
Paris). Morphometricsof the two specimens collected during the current survey and the specimen from Thrissur are given in Table 1.
Further identification of the
specimens was confirmed by comparing the 12S and 16S mitochondrial rDNA partial sequences with the available Gene Bank
sequence of Duttaphrynus scaber and other allied species. Percent divergence in
partial sequences of 12S and 16S genes between the freshly collected specimens
of D. scaber and GenBanksequences was 0.4%. As compared to other Duttaphrynusspecies considered in the study, D. scaber in
our collection showed a minimum divergence of 5.3% and maximum of 8%. Modeltestperformed in MEGA 5 revealed that the nucleotide substitution rates could be
best described by General Time Reversal model with Gamma Distribution (AIC = 5247.40,lnL = -2595.61). Phylogenetic analysis based on maximum likelihood method suggests
further conspecificity with a known Duttaphrynus scaberspecimen from Western Ghats for which the sequences are available in GenBank (Fig. 2).
The advertisement call of Duttaphrynus scaberis a sustained call that is mildly harsh and scratchy in tone (Audio
1). It has a regular beat pattern giving the
feel of a small saw quickly going back and forth with a brief interlude at each
end. The call file is provided as supplementary
data. The frequency of the call is
spread over a band mostly in the region of 2.5–4.5 kHz (Fig. 3a). The
dominant frequency was 3.6 kHz (Table 2, Fig. 3c). Though the sampling freqwas 8 kHz the software used for analysis could render the upper bound unto 4
kHz that is evident from the fig. 6a. Due to the limitations of the sampling frequency the energy levels were
cut off at 4 kHz however it can be predicted that the frequency range could
reach 4.5 kHz which is evident from the Fig. 3a. Each note thus had an average duration
of 155.8 (sd = 24.50) ms,
the note interval was about 85.6 (sd = 10.60) ms and number of notes per minute was 251 (Table 2). The
waveform (Fig. 3b) shows the beat pattern of the call and the rapid amplitude
modulation within each note highlights the scratchy sound each note makes. Comparison of the two calls from two
different males revealed that there was no significant difference in higher
frequency (t = -0.562, df = 40, P= 0.577), dominant frequency (t = -1.959, df= 40, P = 0.057) and call duration (t = 1.295, df = 40, P = 0.203) of the two calls but
there was significant difference in the lower frequency (t = -3.683, df = 40, P = 0.001) and inter-note duration (t= 2.495, df = 38, P = 0.017). This
indicates that except for the lower frequency and inter-note duration the call
pattern for two individuals was not significantly different. Even though the recorded calls had
limitations in terms of sampling frequency, the preliminary analysis provided
here is important as, to our knowledge, there is no
other information about the calls of this species.
Our report of Duttaophrynus scaberbased on both morphological and genetic data extends the geographical
distribution of the species up to the northern most limit of the Western Ghats
of India. From the previous
reports, this species was known from southern Western Ghats up to Karnataka,
Eastern Ghats up to Odisha and Manipur from
northeastern India. However its
presence in northern Western Ghats indicates that the species may be
distributed throughout most of peninsular India. ShoolpaneshwarWildlife Sanctuary, the place from where Sabnis &
Amin (1992) reported the species, is far away from the location indicated by
the current locality records. Further, the specimens collected by them (if any) should be confirmed by
DNA analysis. This emphasizes the
need for detailed surveys for the presence of this species in Satpura ranges as well as plains of Maharashtra State.
References
Bhatt, K., R. Vyas & M. Singh (1999). Herpetofauna of GirProtected Area. Zoos’ Print Journal 14(5): 27–30; http://dx.doi.org/10.11609/JoTT.ZPJ.14.5.27-30
Bioacoustics
Research Program (2003). Raven Lite: Interactive Sound Analysis Software Version 1.0.
Ithaca, NY: The Cornell Lab of Ornithology. Available from
http://www.birds.cornell.edu/raven.
Boersma P. & D. Weenink (2009). Praat: doing
phonetics by computer (Version 5.2). Accessed on 1 May 2011
http://www.praat.org/.
Boulenger, G.A. (1892). Description
of a new toad from Travancore. Journal of the
Bombay Natural History Society 7: 317–318.
Daniel, J.C. (1963). Field
guide to the amphibians of western India. Part I. Journal of the
Bombay Natural History Society 60: 415–438.
Daniels, R.J.R. (2005). Amphibians
of Peninsular India. Universities Press, Hyderabad, 268pp.
Dinesh, K.P., C. Radhakrishnan, K.V. Gururaja& G.K. Bhatta (2009). An
annotated checklist of amphibia of India with some
insight into the pattern of species discoveries, distribution and endemism. Records of the Zoological Survey of India (Occasional
Paper) 302: 1–153.
Donahue, J.P.
& J.C. Daniel (1966). Occurrence of the toad Bufo fergusonii Boulenger in Hyderabad, Andhra Pradesh, India (Anura: Bufonidae). Journal of the Bombay
Natural History Society 63: 447.
Dubois, A. & A. Ohler (1999). Asian and oriental toads of the Bufo melanostictus, Bufo scaber and Bufo stejnegeri groups (Amphibia,Anura): a list of available and valid names and redescription of some name-bearing types. Journal of the South Asian Natural History 4: 133–180.
Dutta, S. & K. Manamendra-Arachchi (2004). Duttaphrynus scaber. In: IUCN 2010. IUCN Red
List of Threatened Species. Version 2010.4. <www.iucnredlist.org>.
Downloaded on 14 June 2011.
Dutta, S.K. (1988). First
records of Bufo stomaticusand Bufo fergusonii(Anura: Bufonidae) from
Orissa, with comments on their distribution. Journal
of the Bombay Natural History Society 63: 439–441.
Dutta, S.K. (1997). Amphibians
of India and Sri Lanka (Checklist and Bibliography). Odyssey
Publishing House, Bubaneswar, Orissa.
Dutta, S.K. & K.N. Manamendra-Arachchi (1996). Amphibian
Fauna of Sri Lanka. Wildlife Heritage Trust of Sri Lanka, Colombo.
Edgar, R.C. (2004). MUSCLE: multiple sequence
alignment with high accuracy and high throughput. Nucleic Acids Research32: 1792–1797; http://dx.doi.org/10.1093/nar/gkh340
Hall, T.A. (1999). BioEdit: a user-friendly biological
sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium Series 41: 95–98.
Kocher, T.D., W.K. Thomas, A.
Meyer, S.V. Edwards, S.F. Paabo, S.F. Villablanca & A. Wilson (1989). Dynamics of mitochondrial
DNA evolution in animals: amplification and sequencing with conserved primers. Proceedings of the National Academy of Science USA 86:
6196–6200.
Krishnamurthy, S.V. (1999). Amphibian
diversity in a few selected environs of Western Ghats, pp. 107–117.In: Hussain, S.A. & K.P. Achar(eds.). Biodiversity of the Western Ghats Complex of
Karnataka. Biodiversity Initiative Trusts.
Mathew,
R. & N. Sen, (2009). Studies on
little known amphibians of Northeast India. Records of the Zoological
Survey of India, Occasional Papers 293: 1–64 + 23 plates.
Mathew,
R. & N. Sen (2010). Pictorial
Guide to the Amphibians of North East India. Zoological Survey of India, Kolkata, 144pp.
Naik, Y.M. & K.R. Vinod (1992). Amphibia of Shoolpaneshwar Sanctuary. Cobra 8: 7–10.
Naik, Y.M. & K.R. Vinod (1993a). Record of the verrucosefrog Rana keralensis(Dubois) in Shoolpaneshwar Wildlife Sanctuary
(Bharuch Dist., Gujarat). Journal of the Bombay
Natural History Society 90: 521–522.
Naik, Y.M. & K.R. Vinod (1993b). The distribution of amphibians in Gujarat
State, India. Hamadryad 18: 28–34.
Padhye, A.D. & H.V. Ghate (2002). An overview of amphibian
fauna of Maharashtra state. Zoos’ Print Journal 17:
735–740; http://dx.doi.org/10.11609/JoTT.ZPJ.17.3.735-40
Posada,
D. & K.A. Crandall (2001). Selecting the best-fit model of nucleotide substitution. SystemsBiolology 50: 580–601; http://dx.doi.org/10.1080/10635150118469
Rao, C.R.N. (1915). Notes on
some south Indian Batrachia. Records of the Indian Museum 11: 31–38.
Ravichandran, M.S. (1996). Amphibia of KalakadWildlife Sanctuary, Tamil Nadu, India. Cobra23: 15–31.
Sabnis, S. D.
& J. V. Amin (1992). Eco-environmental studies of Sardar Sarovar Environs, a Report Published by M.S.
University, Baroda.
Sambrook, J., E.F. Fritsch & T. Maniatis (1989). Molecular Cloning: A Laboratory Manual. Second Edition.Cold Spring Harbor Laboratory Press, New York.
Schneider, J.G. (1799). Historia Amphibiorum Naturalis et Literarariae. Fasciculus
Primus. Continens Ranas, Calamitas, Bufones, Salamandras et Hydrosin Genera et Species Descriptos Notisque suis Distinctos. Jena: Friederici Frommanni.
Simon, C., F. Frati, A. Beckenbach, B. Crespi, H. Liu & P. Flook(1994).Evolution, weighting, and phylogenetic utility of mitochondrial gene sequences
and a compilation of conserved polymerase chain reaction primers. Annals of the Entomological Society of America 87:
651–701.
Tamura,
K., D. Peterson, N. Peterson, G. Stecher, M. Nei & S. Kumar (2011). MEGA5: Molecular evolutionary
genetics analysis using maximum likelihood, evolutionary distance, and maximum
parsimony methods. Molecular Biology and Evolution 28: 2731–2739; http://dx.doi.org/10.1093/molbev/msr121
Vyas, R. (2000a). Supplimentary note on herpetofauna of Gir Forests. Zoos’ Print Journal 15(5): 263–264; http://dx.doi.org/10.11609/JoTT.ZPJ.15.5.263-4
Vyas, R. (2000b). Herpetofauna of HingolgadhNature Education Sanctuary, Gujarat. Zoos’ Print Journal 15(6):
285–286; http://dx.doi.org/10.11609/JoTT.ZPJ.15.6.285-6
Vyas, R. (2002). Priliminary survey of herpetofauna of Narayan SarovarSanctuary, Gujarat. Zoos’ Print Journal 17(6): 812–814; http://dx.doi.org/10.11609/JoTT.ZPJ.17.6.812-4
Vyas, R. (2004a). Herpetofauna of VansdaNational Park, Gujarat. Zoos’ Print Journal 19(6):
1512–1514; http://dx.doi.org/10.11609/JoTT.ZPJ.1036.1512-4
Vyas, R. (2004b). Note on amphibians of Barda Wildlife Sanctuary, Gujarat. Zoos’ Print Journal19(7): 1545; http://dx.doi.org/10.11609/JoTT.ZPJ.1034.1545
Vyas, R. (2007). A field
guide to amphibians of Gujarat. Nature Club, Surat, 30pp. Online access:
http://www.natureclubsurat.org/Products/ambiphianbook.pdf, accessed on 3 April
2012.
Vyas, R. (2008). Review of
the current diversity and richness of amphibians of Gujarat, India. The Indian Forester 134 (10): 1381–1392.
Vyas, R. (2012). Frogs of Shoolpaneswr Wildlife Sanctuary, Gujarat, India. Froglog 101: 54–56.
Xia, X. & Z. Xie (2001). DAMBE: Software package for
data analysis in molecular biology and evolution. Journal of Heredity92: 371─373; http://dx.doi.org/10.1093/jhered/92.4.371