Barcoding fauna of India: An initiative by

Ministry of Environment Forest

& Climate Change
Zoological Survey of India
Vikas Kumar, Kailash Chandra, Kaomud Tyagi, Shantanu Kundu, John TD Caleb, Boni Amin Laskar,
Devkant Singha, Sumantika Chatterjee, Rajasree Chakraborty
Centre for DNA Taxonomy, Molecular Systematics Division, Zoological Survey of India, New Alipore-53, Kolkata, India
Corresponding author: Vikas Kumar (e-mail: vikaszsi77@gmail.com)
Abstract:
The Zoological Survey of India (ZSI), since its inception in 1916, has been maintaining several type materials in its National Zoological Collections at headquarter Kolkata as well as
in different regional museums. The survey is the premier institute on faunal research having over 5 million specimens including more than 17000 type specimens from protozoa
to mammal from India and from more than 60 other countries. Integration of molecular data with morphology is one of the major mandate for ZSI. As of now, over 2000 DNA
barcodes for a number of species falling in different groups like economically important Insects, indicator species of Lepidoptera, freshwater fishes, reptiles, and wildlife seizures
have been contributed to GenBank and BOLD. The study of barcode data from various groups resolved several problems of their identification, taxonomy and distribution. To
name a few, in lower groups of animal, we detected alien insect pest species (Thrips parvispinus), detected cryptic diversity in insect order Thysanoptera (for species complexes,
Frankliniella schultzei and Thrips palmi), detected host specific diversity in Hemiptera (Helopeltis theivora). In Araneae, we recorded for the first time from India the species
belonging to the genera Neriene and Psechrus. In higher animal, we first time recorded the distribution of a fish of the subfamily Gobionellinae, order Perciformes from northeast
India. We further detected three non-native species of turtles and tortoises from northeast India (Chitra chitra, Cyclemys fusca and Amyda ornata) along with the detection of
Nilssonia nigricans from wild habitat which is though categorized as Extinct in Wild in the IUCN Red List. DNA barcoding has to go a long way for covering all the extant species to
achieve its real advantage in biodiversity research and conservation. Our initial success from Indian region motivate towards future collaborative endeavour for barcoding and
generating mitogenomes from the archival specimens.

Introduction: In-silico analysis:

The ZSI with its 16 regional centres and 5 DNA barcoding laboratory  The bi-directional chromatograms were checked through ABI
facilities, generated and contributed thousands of DNA barcode data Sequence Scanner, DNA Baser, Chromas, BLASTn, ORF Finder etc.
of various faunal groups in the global database to accelerate the  The bioinformatics softwares and online server were used:
taxonomic research. We have surveyed the faunal systems from Multiple and Pairwise alignment: MEGA, BioEdit, and ClustalX etc.
marine ecosystem to the alpine region including protected areas, Genetic divergence: MEGA, and PAUP etc.
captivity and commercialized markets. The current approaches Phylogeny: MEGA, PAUP, and MrBayes etc.
adjudicated to conserve the DNA of many highly threatened or Population genetics and Haplotyping: BEAST, DnaSP, and PopART etc.
endemic taxa from Indian zoogeography under ‘National Faunal Multiple species delimitation methods (BIN, ABGD, GMYC and bPTP).
Genome Resources’ program. Table 1: DNA barcoding data of various faunal systems from India
Materials and methods: Faunal system Order No. of DNA
Barcodes
Survey the samples with prior permission from wildlife authority Thrips Thysanoptera 604
where required. Molecular biology grade chemicals, Commercialized
Kits and instrumentation facilities. Butterflies & Moths Lepidoptera 499
Horse flies & deer flies Diptera 59
Classical taxonomy:
Spiders Araneae 664
 Process the specimens under Leica microscope and vouchered
with unique identification numbers. Turtles Testudines 40
 The morphological measurements were acquired by adopting Fishes Cypriniformes , Siluriformes, 441
appropriate methods specific to the studied groups. Perciformes, Mugiliformes etc.
 Morphological data was assembled and comparative studies were Confiscated Wildlife Carnivora, Perissodactyla, 25
perfomed with the type specimens or other sister species to samples (Mammals, Artiodactyla, Squamata, Testudines
validate the species identity Birds, Retiles etc.) etc.
Other invertebrates Hymenoptera, Coleotera, 120
Molecular taxonomy: Spongillida, Decapoda etc.
 DNA isolation from different biological samples (Blood, Tissue,
Saliva, Hair etc.) and checked in Agarose Gel Electrophoresis.
Results and discussion:
 The DNA barcode fragments (mtCOI~650bp) were amplified by
specific primer pairs through PCR.
 The PCR products were checked in Agarose Gel Electrophoresis
and cleaned by commercialized purification Kits.
 Bi-directional Cycle Sequencing were performed by Forward and
Reverse primer separately in Thermal Cycler.
 The DNA sequences were generated by using 48 capillary ABI-
3730 DNA Analyzer in the in-house facilities at ZSI, Kolkata.
Figure 1: Glimpse of Indian zoogeography and collection localities of
several faunal systems. Figure 2: The adopting DNA barcoding approaches for systematics
research by genetic divergence, phylogeny and estimated (molecular
operational taxonomic units (MOTUs).
 The study detected cryptic diversity, species complexes and alien
Alpine species of Thrips, an economically important species from India.
 The study demonstrated the presence of cryptic species in
Temperate Chrysops dispar and resolved the Tabanus striatus species
complex using DNA barcode.
 The DNA data clearly segregated the studied spider species and
identified many new species to science and new record to India.
Tropical  The aimed study detected three non-native and one ‘Extinct in the
Wild’ species of turtles and tortoises from northeast India.
 The DNA data clearly separated the studied fish species and first
Aquatic record of Gobionellinae in Indian waters, resolve the species
complexes in Amblyceps and Tor species.
References  The generated DNA data identified the confiscated, amorphous
Tyagi, K., Kumar, V., Singha, D., Chandra, K., Laskar, B.A., Kundu, S., Chakraborty, R., Chatterjee, wildlife materials seized from poachers hand by similarity search
S. (2017) DNA Barcoding studies on Thrips in India: Cryptic species, Species complexes.
Scientific Reports, 4898, 7:1-14. Doi:10.1038/s41598-017-05112-7
engine (BLASTn and BOLD-IDs) and helps to the court of law.
Laskar BA, Kumar V, Darshan A, Kundu S, Das DN. DNA barcoding of Amblyceps congeners  The DNA barcode data identified the freshwater sponge diversity
(Siluriformes: Amblycipitidae) from Brahmaputra drainage in northeast India. Mitochondrial from urban ponds, identified sea food (crab & prawn) and detected
DNA. DOI: 10.3109/24701394.2016.1174223.
Laskar BA, Kumar V, Kundu S, Tyagi K, Singha D, Chakraborty R, Chatterjee S, Saha S. DNA the contamination of Maxillopoda species.
barcoding of Gobiid fishes (Perciformes: Gobiidae) from eastern and northeastern India with
new record of a Gobionellinae species for the region. Mitochondrial DNA. doi: Conclusion: The success of DNA barcoding in systematics studies and
10.3109/24701394.2016.1143470. its countless applications in biodiversity research and conservation
Kundu S, Kumar V, Laskar BA, Chandra K, Tyagi K. Mitochondrial DNA effectively detects non-
native Testudines: Invisible wildlife trade in northeast India. GENE Reports.
activities, we are motivated to established long term collaboration with
doi:10.1016/j.genrep.2016.02.002. the global DNA barcoding networks to asses the Indian faunal
Tyagi, K., Kumar, V., Singha, D. and Chrakraborty, R. Morphological and DNA Barcoding biodiversity as well as the reexamine the oldest archives in National
Evidence for Invasive Pest thrips, Thrips parvispinus (Thripidae:Thysanoptera), newly
recorded from India. Journal of Insect Science, 15(1): 1-4. DOI: 10.1093/jisesa/iev087
Zoological collections at Zoological Survey of India.
Banerjee, D., Kumar, V., Maity, A., Ghosh, B., Tyagi, K., Singha, D., Kundu, S., Laskar, B. A., Funding Sources/Acknowledgements
Naskar, A. &Rath, S. 2015. Identification through DNA barcoding of Tabanidae (Diptera) The authors are thankful to the Director, Zoological Survey of India, Kolkata, for providing necessary
vectors of surra disease in India. Acta Tropica, 150: 52-58. facilities. The authors (VK,KT & BAL) acknowledge the core funding of ZSI, Ministry of Environment
Kundu, S., Laskar, B.A., Venkataraman, K., Banerjee, D. & Kumar, V. 2015. DNA barcoding of Forest and Climate Change (MoEF&CC), Govt. of India. The author (SK) acknowledge the DST-SERB
Nilsssonia congeners corroborates existence of wild N. nigricans in Northeast India. National Post-Doctoral fellowship (F. No. PDF/2015/000302). The author (JC & SC) acknowledge the
Mitochondrial DNA, 1-4. fellowship of National Mission on Himalayan Studies.