Which frog's legs do froggies eat? The use of DNA barcoding for identification of deep frozen frog legs (Dicroglossidae, Amphibia) commercialized in France

  • Annemarie Ohler Institut de Systématique, Évolution, Biodiversité, ISYEB_UMR7205_CNRS, MNHN, UPMC, EPHE, Muséum national d’Histoire naturelle, Sorbonne Universités, Paris
  • Violaine Nicolas Institut de Systématique, Évolution, Biodiversité, ISYEB_UMR7205_CNRS, MNHN, UPMC, EPHE, Muséum national d’Histoire naturelle, Sorbonne Universités, Paris
Keywords: Fejervarya cancrivora, Limnonectes macrodon, frog leg trade, barcoding, conservation, Red List

Abstract

Several millions frogs captured in the wild in Indonesia are sold for food yearly in French supermarkets, as deep frozen frog legs. They are commercialized as Rana macrodon, but up to 15 look-alike species might also be concerned by this trade. From December 2012 to May 2013, we bought 209 specimens of deep frozen frog legs, and identified them through a barcoding approach based on the 16S gene. Our results show that 206 out of the 209 specimens belong to Fejervarya cancrivora, two to Limnonectes macrodon and one to F. moodiei. Thus only 0.96 % of the frogs were correctly identified. Unless misclassification was intentional, it seems that Indonesian frog leg exporters are not able to discriminate between the species. The quasi absence of L. macrodon in our samples might be an indication of its rarity, confirming that its natural populations are declining rapidly, in agreement with its “vulnerable” status according to the IUCN Red List. Our results show that the genetic and morphological diversity of the frogs in trade is much higher than the genetic and morphological diversity measured so far by scientific studies. These results underline the need for large scale studies to assess the status of wild populations.

References

Alcala A.C. 1962. Breeding behaviour and early development of frogs of Negros, Philippine Islands. Copeia 1962: 679–726.

Altherr S., Goyenechea A. & Schubert D. 2011. Canapés to Extinction—the International Trade in Frogs’ Legs and its Ecological Impact. Pro Wildlife, Defenders of Wildlife and Animal Welfare Institute, Munich and Washington DC.

Anonymous. 2001. Commission Regulation (EC) No. 2065/2001 of 22 October 2001 Laying Down Detailed Rules for the Application of Council Regulation (EC) No. 104/2000 as Regards Informing Consumers about Fishery and Aquaculture Products. Available from http://eur-lex.europa.eu/legal-content/EN/ALL/?uri=CELEX%3A32001R2065 [accessed 13 Jan. 2016].

Anonymous. 2007. Review of Non-CITES Amphibia Species that are Known or Likely to be in International Trade. Version edited for public release. Prepared for the European Commission Directorate General E –

Environment ENV.E.2. – Development and Environment. UNEP World Conservation Monitoring Centre, Cambridge.

Bandelt H. J., Forster P. & Rohl A. 1999. Median-joining networks for inferring intraspecific phylogenies. Molecular Biology and Evolution 16: 37–48. http://dx.doi.org/10.1093/oxfordjournals.molbev.a026036

Boulenger G.A. 1920. A monograph of the South Asian, Papuan, Melanesian and Australian frogs of the genus Rana. Records of the Indian Museum 20: 1–226.

Church G. 1960. The effects of seasonal and lunar changes on the breeding pattern of the edible Javanese frog, Rana cancrivora Gravenhorst. Treubia 25: 215–233.

Clement M., Posada D. & Crandall K.A. 2000. TCS: a computer program to estimate gene genealogies. Molecular Ecology 9: 1657–1659. http://dx.doi.org/10.1046/j.1365-294x.2000.01020.x

Dubois A. 1983. A propos de cuisses de grenouilles. Protection des amphibiens, arrêtés ministériels, projets d’élevage, gestion des populations naturelles, enquêtes de répartition, production, importations et consommation: une équation difficile à résoudre. Les propositions de la Société Batrachologique de France. Alytes 2 (3): 69–111.

Emerson S.B. 1978. Allometry and jumping in frogs – helping twain to meet. Evolution 32: 551–564.

French Customs. 2012. Données d’importations CAF et d’exportation FAB de cuisses de grenouilles, fraîches, réfrigéres ou congélées. Available from http://www.insee.fr/ [accessed 16 Jan. 2012].

Grosjean S., Ohler A., Chuaynkern Y., Cruaud C. & Hassanin A. 2015. Improving biodiversity assessment of anuran amphibians using DNA barcoding of tadpoles. Case studies from Southeast Asia. Comptes Rendus Biologies 338: 351–361. http://dx.doi.org/10.1016/j.crvi.2015.03.015

Hasan M., Islam M.M., Khan M.R., Alam M.S., Kurabayashi A., Igawa T., Kuramoto M. & Sumida M. 2012. Cryptic anuran biodiversity in Bangladesh revealed by mitochondrial 16S rRNA gene sequences. Zoological Science 29: 162–172. http://dx.doi.org/10.2108/zsj.29.162

Hebert P.D., Cywinska A., Ball S.L. & deWaard J.R. 2003. Biological identifications through DNA barcodes. Proceedings of the Royal Society London B 270: 313–321. http://dx.doi.org/10.1098/rspb.2002.2218

Huelsenbeck J.P. & Ronquist F. 2001. MrBayes: Bayesian inference of phylogenetic trees. Bioinformatics 17: 754–755. http://dx.doi.org/10.1093/bioinformatics/17.8.754

Inger R.F. 1966. The systematics and zoogeography of the Amphibia of Borneo. Fieldiana, Zoology 52: 1–402.

Inger R.F. & Stuebing R.B. 1997. A Field Guide to the Frogs of Borneo. Natural History Publications (Borneo) Limited, Kota Kinabalu, Malaysia.

Iskandar D.T. 1998. The Amphibians of Java and Bali. Research and Development Centre for Biology –

LIPI, Bogor, Indonesia.

Iskandar D., Mumpuni, Das I., Shrestha T.K. & Ohler A. 2004. Limnonectes macrodon. The IUCN Red List of Threatened Species 2004: e.T58351A11770829. Available from http://dx.doi.org/10.2305/IUCN.UK.2004.RLTS.T58351A11770829.en [accessed 22 Jan. 2016].

Kotaki M., Kurabayashi A., Matsui M., Khonsue W., Djong T.H., Tandon M. & Sumida M. 2008. Genetic divergences and phylogenetic relationships among the Fejervarya limnocharis complex in Thailand and neighboring countries revealed by mitochondrial and nuclear genes. Zoological Science 25: 381–390. http://dx.doi.org/10.2108/zsj.25.381

Kurabayashi A., Kuramoto M., Joshy H. & Sumida M. 2005. Molecular phylogeny of the ranid frogs from southwest India based on the mitochondrial ribosomal RNA gene sequences. Zoological Science 22: 525–534. http://dx.doi.org/10.2108/zsj.22.525

Kurniawan N., Islam M.M., Djong T.H., Igawa T., Daicus M.B., Yong H.S., Wanichanon R., Khan M.M., Iskandar D.T., Nishioka M. & Sumida M. 2010. Genetic divergence and evolutionary relationship in Fejervarya cancrivora inferred from allozyme and mtDNA sequence analyses. Zoological Science 27: 222–233. http://dx.doi.org/10.2108/zsj.27.222

Kurniawan N., Djong T.H., Islam M.M., Nishizawa T., Belabut D.M., Sen Y.H., Wanichanon R., Yasir I. & Sumida M. 2011. Taxonomic status of three types of Fejervarya cancrivora from Indonesia and other Asian countries based on morphological observations and crossing experiments. Zoological Science 28: 12–24. http://dx.doi.org/10.2108/zsj.28.12

Kurniawan N., Djong T.H., Maideliza T., Hamidy A., Hasan M., Igawa T. & Sumida M. 2014. Genetic divergence and geographic distribution of frogs in the genus Fejervarya from Indonesia inferred from mitochondrial 16S rRNA gene analysis. Treubia 41: 1–16.

Kusrini M.D. 2005. Edible Frog Harvesting in Indonesia: Evaluating its Impact and Ecological Context. PhD Thesis in Zoology and Tropical Ecology. James Cook University, School of Tropical Biology, Townsville, Australia.

Kusrini M.D. & Alford R.A. 2006. Indonesia’s exports of frog’s legs. Traffic Bulletin 21: 13–20.

Le Serrec G. 1988. France’s frog consumption. Traffic Bulletin 10: 17.

Manthey U. & Grossmann W. 1997. Die Amphibien und Reptilien Südostasiens. Natur und Tier-Verlag, Münster, Germany.

Meier R., Shiyang K., Vaidya G. & Ng P.K. 2006. DNA barcoding and taxonomy in Diptera: a tale of high intraspecific variability and low identification success. Systematic Biology 55: 715–728. http://dx.doi.org/10.1080/10635150600969864

Neveu A. 2004. La raniculture est-elle une alternative à la récolte? Etat actuel en France. INRA Production Animale 17: 167–175.

Nijman V. 2010. An overview of international wildlife trade from Southeast Asia. Biodiversity and Conservation 19: 1101–1114. http://dx.doi.org/10.1007/s10531-009-9758-4

Nylander J.A. 2004. MrModeltest v. 2. Program distributed by the author. Evolutionary Biology Centre, Uppsala University, Uppsala, Sweden.

Palumbi S.R., Martin A., Romano S., McMillan W.O., Stice L. & Grabowski G. 1991. The Simple Fool’s Guide to PCR. University of Hawaii Press, Honolulu.

Phelps J., Webb E.L., Bickford D., Nijman V. & Sodhi N.S. 2010. Boosting CITES. Science 330: 1752–1753.

Ronquist F., Huelsenbeck J.P. & Van der Mark P. 2005. MrBayes v. 3.1 Manual. Available from http://cs.mcgill.ca/~birch/doc/mrbayes/mb3.1_manual.pdf [accessed 1 Jan. 2016].

Thompson J.D., Higgins D.G. & Gibson T.J. 1994. Clustal W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Research 22: 4673–4680. http://dx.doi.org/10.1093/nar/22.22.4673

Veith M., Kosuch J., Feldmann R. Martens H. & Seitz A. 2000. A test for correct species declaration of frog legs imports from Indonesia into the European Union. Biodiversity and Conservation 9: 333–341. http://dx.doi.org/10.1023/A:1008906918236

Vences M., Thomas M., van der Meijden A., Chiari Y. & Vieites D.R. 2005. Comparative performance of the 16S rRNA gene in DNA barcoding of amphibians. Frontiers in Zoology 2 (5): 1–12. http://dx.doi.org/10.1186/1742-9994-2-5

Vidal-Garcia M., Byrne P.G., Roberts J.D. & Keogh J.S. 2014. The role of phylogeny and ecology in shaping morphology in 21 genera and 127 species of Australo-Papuan myobatrachid frogs. Journal of Evolutionary Biology 27: 181–192. http://dx.doi.org/10.1111/jeb.12292

Warkentin I.G., Bickford D., Sodhi N.S. & Bradshaw C.J.A. 2009. Eating frogs to extinction. Conservation Biology 23: 1056–1059. http://dx.doi.org/10.1111/j.1523-1739.2008.01165.x

Yuan Z., Zhao E., Shi H., Diesmos A., Alcala A., Brown R., Afuang L., Gee G., Sukumaran J., Yaakob N.,

Leong T.M., Chuaynkern Y., Thirakhupt K., Das I., Iskandar D., Mumpuni & Inger R. 2004. Fejervarya cancrivora. The IUCN Red List of Threatened Species 2004: e.T58269A11759436. http://dx.doi.org/10.2305/IUCN.UK.2004.RLTS.T58269A11759436.en

Published
2017-02-06
How to Cite
Ohler, A., & Nicolas, V. (2017). Which frog’s legs do froggies eat? The use of DNA barcoding for identification of deep frozen frog legs (Dicroglossidae, Amphibia) commercialized in France. European Journal of Taxonomy, (271). https://doi.org/10.5852/ejt.2017.271