Morphometric and DNA-based identification of horseshoe crab Carcinoscorpius rotundicauda (Latreille, 1802) and Tachypleus gigas (Müller, 1785) from Bangladesh coast

Wahida  Haque; Sanjana  Enam; Md. Tarikul  Islam; Sujan Kumar  Datta; Md. Sagir  Ahmed

doi:10.3329/brc.v10i2.74568

Authors

Wahida Haque Department of Fisheries, University of Dhaka, Dhaka 1000, Bangladesh
Sanjana Enam Department of Zoology, University of Dhaka, Dhaka 1000, Bangladesh
Md. Tarikul Islam Bangladesh Oceanographic Research Institute, Cox’s Bazar 4730, Bangladesh
Sujan Kumar Datta Department of Zoology, University of Dhaka, Dhaka 1000, Bangladesh
Md. Sagir Ahmed Department of Zoology, University of Dhaka, Dhaka 1000, Bangladesh

DOI:

https://doi.org/10.3329/brc.v10i2.74568

Keywords:

Horseshoe crab, Carcinoscorpius rotundicauda, Tachypleus gigas, DNA Barcoding, COI

Abstract

Horseshoe crabs are the living fossils of xiphosurans that have survived for over 200 million years without morphological change and records fossil dating back 500 million years. A study was conducted on morphometric and molecular characterization of horseshoe crab Carcinoscorpius rotundicauda (Latreille, 1802) and Tachypleus gigas (Müller, 1785) from Bangladesh waters. A total of ten samples of C. rotundicauda and one sample of T. gigas were collected from Dublar Char, Sundarbans and Cox’s bazar from June 2016 to May 2017 and February 2024, respectively. Among them, four specimens were male and seven female. Morphometric measurements were taken from all the collected samples and differentiated male and female based on the second pair of legs, the pedipalps. The species were genetically identified using mitochondrial cytochrome c oxidase subunit I (COI) gene. An average length of about 670.5 bp nucleotide sequences were obtained. Average percentage of nucleotide frequencies were T (30.3), C (16.3), A (34.4) and G (19.0) for C. rotundicauda and T (34.45), C (22.01), A (27.91) and C (15.63) for T. gigas. GC content analysis showed that average GC 36.20% and AT 63.80%. The average GC content at the 1^st, 2^nd and 3^rd codon position were 38.82%, 37.87% and 31.92% respectively. This is the first report of DNA barcoding of a living fossil C. rotundicauda and T. gigas from Bangladesh and will serve as baseline information for future research, conservation and management of this invaluable primitive creature.

References

Ahmed, A. T. A., Kabir, S. M. H., Ahmad, M., Rahman, A. K. A., Haque, E. U., Ahmad, Z. U., Begum, Z. N. T., Hassan, M.A. and Khondokar, M. (eds)., 2008. Encyclopedia of Flora and Fauna of Bangladesh, Vol. 18. Part I. Arthropoda: Arachnida. Asiatic Society of Bangladesh, Dhaka. pp.3-10.

Ahmed, M.S., Afrin, T. and Barua, A., 2021b. New distributional record of Charybdis japonica, Coenobita violascens, Galene bispinosa, and Portunus reticulatus (Crustacea: Decapoda) from Bangladesh waters of the Bay of Bengal, Regional Studies in Marine Science. 44: 101785

Ahmed, M.S., Barua, A., Datta, S.K., Saha, T., Antu, D.R. and Ahmed, S., 2022. Characterization of spiny lobsters from Bangladesh waters using morphology, COI and 16S rRNA sequences. Heliyon. 8(2):1-9

Ahmed, M.S., Datta, S.K., Saha, T. and Hossain, Z., 2021a. Molecular characterization of marine and coastal fishes of Bangladesh through DNA barcodes, Ecology and Evolution. 11(9): 3696-3709

Alcock, A., 1900. Materials for a carcinological fauna of India. No. 5. Brachyura Primigenia or Dromiacea. Journal of the Asiatic Society of Bengal, Calcutta. 68(2):1-104.

Arnold, C., 2020. Horseshoe crab blood is key to making a COVID-19 vaccined but the ecosystem may suffer. https://www.nationalgeographic.com/animals/ 2020/07/covid-vaccine-needs-horseshoe-crab-blood/.

Chowdhury, M.M., Rahman, A.S.M.S., Nahar, L., Rahman, M., Reza, H.A. and Ahmed, M.S., 2016. Efficiency of Different DNA Extraction Methods for Fish Tissues: A Comparative Analysis. IOSR Journal of Pharmacy and Biological Sciences 11(3):11-15.

Chowdhury, S.H., and Harizuddin, A.K.H., 1980. Horse-shoe crabs (Chelicerata: Merostomata) occurring along the south-east coasts of Bangladesh. Bangladesh J. Zool. 8: 5-13.

Costa, F.O., Dewaard, J.R., Boutillier, J., Ratnasingham, S., Dooh, R.T., Hajibabaei, M. and Hebert, P.D.N., 2007. Biological identifications through DNA barcodes: the case of the Crustacea. Can. J. Fifh. Aquat. Sci. 64: 272-295.

Cumberlidge, N. and Sachs, R., 1989. Zeitschrift Fur Angewandte Zoologie. German Journal for Applied Zoology. pp. 220-229.

Dybas, C. L., 2019. New Lifeblood for Atlantic Horseshoe Crabs. Oceanography, 32(2): 12- 14.

Edgar, R. 2004., MUSCLE: A multiple sequence alignment method with reduced time and space complexity. BMC bioinformatics. 5: 113. https://doi.org/10.1186/1471-2105-5-113

Folmer, O., Black, M., Hoeh, W., Lutz, R. and Vrijenhoek, R., 1994. DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates. Mol. Mar. Biol. Biotech. 3(5):294-299.

Gibson, R. N., Atkinson, R. J. A., & Gordon, J. D. M., 2010. Historical reconstruction of human- induced changes in US estuaries. Oceanography and marine biology: an annual review, 48, 267-338.

Hebert, P.N.D., Penton, E.H., Burns, J.M., Janzen, D.H. and Hallwachs, W., 2004. Ten species in one: DNA barcoding reveals cryptic species in the neo tropical skipper butterfly Astraptes fulgerator. Proc. Natl. Acad. Sci. U.S.A. 101: 14812–14817.

Hebert, P.N.D., Ratnasingham, S. and Dewaard, J.R., 2003. Barcoding animal life: cytochrome c oxidase subunit 1 divergences among closely related species. Proc. R. Soc. Lond. B 270: S96–S99.

Itow, T., Misra, J.K. and Ahmed, A.T.A., 2004. Horseshoe crabs, (King crabs) in the Bay of Bengal, South Asia, Shizuoka University. Bull. Fac. Educ., Nat. Sci. Seri. 54:13-30.

IUCN (International Union for Conservation of Nature)., 2015. Red Book of Threatened Species of Bangladesh, Crustacean. Vol.6.

Nandi, N.C. and Pramanik, S.K., 1994. Crabs and Crab Fisheries of Sundarban. Hindustan Publishing Corporation, Delhi. pp. 34-54.

Pocock, R. I., 1902. The taxonomy of recent species of Limulus. Ann. Mag. Nat. Hist. Ser. 7, 9:256- 266.

Remigio, E.A. and Hebert, P.D.N., 2003. Testing the utility of partial COI sequences for phylogenetic estimates of Gastropod relationships. Mol. Phylogenet. Evol. 29: 641–647.

Sambrook, J. and Russell, R.W., 2001. Molecular Cloning: A laboratory manual, 3rd ed. Cold spring harbor laboratory press, cold spring harbor, N.Y.

Sekiguchi, K., Nakamura, K., and Scshimo, H., 1978. Morphological variation of a horseshoe crab, Carcinoscorpius rotundicauda, from the bay of Bengal and gulf of Siam. Proc. Jap. Soc. Syst. Zool. 15:24-30.

Siddique, M.Z.H. and Zafor, M., 2002. Crabs in the Chakaria Sundarban area of Bangladesh. The Journal of NOAMI. 19(2): 61-75.

Tamura, K., Stecher, G. and Kumar, S., 2021. MEGA11: molecular evolutionary genetics analysis version 11. Molecular biology and evolution, 38(7): 3022- 3027.https://doi.org/10.1093/molbev/msab120

Vestbo S, Obst M, Quevedo Fernandez FJ, Intanai I, Funch P. 2018. Present and poten-tial future distributions of Asian horseshoe crabs determine areas for conservation.Frontier Marine Science 5:164 DOI 10.3389/fmars.2018.00164.

Ward, R.D., Zemlak, T.S., Innes, B.H., Last, P.R. and Hebert, P.D.N., 2005. DNA barcoding Australia’s fish species. Philos. Trans. R. Soc. Lond. B 360: 1847–1857.

Waycott, B., 2020. Can farming horseshoe crabs help the COVID-19 cause? https://www.aquaculturealliance.org/advocate/can-farming-horseshoe-crabs-help-the-covid-19-cause/. (Accessed 30 August 2020).

Wong, L. L., Peatman, E., Lu, J., Kucuktas, H., He, S., Zhou, C., Na-nakorn, U. and Liu, Z., 2011. DNA barcoding of Catfish: species authentication and phylogenetic assessment. Plos One 6 (3): e17812.