Molecular Biology Reports

, Volume 39, Issue 1, pp 693–699 | Cite as

Forensic species identification of large macaws using DNA barcodes and microsatellite profiles

  • Hideaki Abe
  • Azusa Hayano
  • Miho Inoue-Murayama


Using mitochondrial and nuclear markers species identification was conducted in the case of seized feathers. Earlier, we had sequenced cytochrome c oxidase subunit I (COI) both from 10 seized specimens and 43 validation specimens from captive macaws belonging to 4 Ara species (A. macao, A. chloropterus, A. ararauna, and A. ambiguus) and identified 19 haplotypes based on COI sequences. Species-level identification using Barcode of Life Data Systems showed that seized feathers shared the highest similarity with scarlet macaws (A. macao), and this result was supported by the tree-base identification with high bootstrap values. Moreover, microsatellite profiles in AgGT17 locus showed that patterns of allelic distribution in the seized feathers were apparently distinct from those of red-and-green macaw (A. chloropterus), but were overlapped with those of A. macao, suggesting that all of seized feathers were derived from several individuals of A. macao. We also determined the parentage of hybrid macaws by the combination of COI barcodes and microsatellite profiles. The technique presented here will contribute to forensic identification and future conservation of large macaws that have been lost due to deforestation.


DNA barcoding Cytochrome c oxidase subunit I Microsatellite Macaw Seizure Forensic species identification 



The authors would like to thank the following zoos and aviaries in Japan for assisting us with sample collection: Adventure World, Asa Zoological Park, Companion Animals, Fuji Pet, Izu Animal Kingdom, Izu Shaboten Park, Kakegawa Kacho-en, Kobe Kacho-en, and Loros Bird Farm. Seized feathers were provided by the Aichi Prefectural Police. We also thank Eri Kawaguchi (Kyoto University) for technical support and Kara Gebhardt-Tessman (University of Idaho) for providing information on AgGT17 allele frequency in wild scarlet macaws. This study was supported financially by the Ministry of Education, Culture, Sports, Science and Technology (MEXT) with a Grant-in-aid for Science Research (#21310150 to MI-M) and Global Center of Excellence Program “Formation of a Strategic Base for Biodiversity and Evolutionary Research: from Genome to Ecosystem”.


  1. 1.
    Frézal L, Leblois R (2008) Four years of DNA barcoding: current advances and prospects. Infect Genet Evol 8:727–736. doi: 10.1016/j.meegid.2008.05.005 PubMedCrossRefGoogle Scholar
  2. 2.
    Waugh J (2007) DNA barcoding in animal species: progress, potential and pitfalls. BioEssays 29:188–197. doi: 10.1002/bies.20529 PubMedCrossRefGoogle Scholar
  3. 3.
    Ferri G, Alù M, Corradini B, Licata M, Beduschi G (2009) Species identification through DNA “barcodes”. Genet Test Mol Biomarkers 13:421–426. doi: 10.1089/gtmb.2008.0144 PubMedCrossRefGoogle Scholar
  4. 4.
    Johnson RN (2010) The use of DNA identification in prosecuting wildlife-traffickers in Australia: do the penalties fit the crimes? Forensic Sci Med Pathol 6:211–216. doi: 10.1007/s12024-010-9174-9 PubMedCrossRefGoogle Scholar
  5. 5.
    Dove CJ, Rotzel NC, Heacker M, Weigh LA (2008) Using DNA barcodes to identify bird species involved in birdstrikes. J Wildl Manage 72:1231–1236. doi: 10.2193/2007-272 CrossRefGoogle Scholar
  6. 6.
    Yang R, Wu X, Yan P, Li X (2009) Using DNA barcodes to identify a bird involved in a bird strike at a Chinese airport. Mol Biol Rep 37:3517–3523. doi: 10.1007/s11033-009-9945-0 PubMedCrossRefGoogle Scholar
  7. 7.
    Hebert PDN, Ratnasingham S, deWaard JR (2003) Barcoding animal life: cytochrome c oxidase subunit 1 divergences among closely related species. Proc R Soc Lond B 270:S96–S99. doi: 10.1098/rsbl.2003.0025 CrossRefGoogle Scholar
  8. 8.
    Ratnasingham S, Hebert PDN (2007) Bold: the barcode of life data system ( Mol Ecol Notes 7:355–364. doi: 10.1111/j.1471-8286.2007.01678.x Google Scholar
  9. 9.
    Barcode of Life website. Accessed 18 Jul 2010
  10. 10.
    Collar NJ (1997) Family Psittacidae (parrot). In: del Hoyo J, Elliott A, Sargatal J (eds) Handbook of the birds of the world. Sandgrouse to Cuckoos, vol 4. Lynx Edicions, Barcelona, pp 280–477Google Scholar
  11. 11.
    Bird Hybrid Database (2010) Bird hybrids database—search by name of birds. Accessed 18 Jul 2010
  12. 12.
    McCarthy EM (2006) Handbook of avian hybrids of the world. Oxford University Press, OxfordGoogle Scholar
  13. 13.
    Abramson J (1995) Captive breeding and conservation. In: Abramson J, Speer BL, Thomsen JB (eds) The large macaws. Their care, breeding and conservation, 1st edn. Raintree Publications, CA, pp 250–265Google Scholar
  14. 14.
    Rosen GE, Smith KF (2010) Summarizing the evidence on the international trade in illegal wildlife. EcoHealth 7:24–32. doi: 10.1007/s10393-010-0317-y PubMedCrossRefGoogle Scholar
  15. 15.
    Hebert PDN, Stoeckle MY, Zemlak TS, Francis CM (2004) Identification of birds through DNA barcodes. PloS Biol 2:1657–1663. doi: 10.1371/journal.pbio.0020312 CrossRefGoogle Scholar
  16. 16.
    Yoo HS, Eah JY, Kim JS, Kim YJ, Min MS, Peak WK, Lee H, Kim CB (2006) DNA barcoding Korean birds. Mol Cells 22:323–327PubMedGoogle Scholar
  17. 17.
    Hall TA (1999) Bioedit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symp Ser 41:95–98Google Scholar
  18. 18.
    Kimura M (1980) A simple method for estimating evolutionary rate of base substitutions through comparative studies of nucleotide sequences. J Mol Evol 16:111–120PubMedCrossRefGoogle Scholar
  19. 19.
    Tamura K, Dudley J, Nei M, Kumar S (2007) MEGA4: molecular evolutionary genetics analysis (MEGA) software version 4.0. Mol Biol Evol 24:1596–1599. doi: 10.1093/molbev/msm092 Google Scholar
  20. 20.
    Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425PubMedGoogle Scholar
  21. 21.
    Gebhardt KJ, Waits LP (2008) Cross-species amplification and optimization of microsatellite markers for use in six Neotropical parrots. Mol Ecol Resour 8:835–839. doi: 10.1111/j.1755-0998.2007.02083.x PubMedCrossRefGoogle Scholar
  22. 22.
    Russello M, Calcagnotto D, DeSalle R, Amato G (2001) Characterization of microsatellite loci in the endangered St. Vincent Parrot, Amazona guildingii. Mol Ecol Notes 1:162–164. doi: 10.1046/j.1471-8278.2001.00061.x CrossRefGoogle Scholar
  23. 23.
    Fridolfsson AK, Ellegren H (1999) A simple and universal method for molecular sexing of non-ratite birds. J Avian Biol 30:116–121CrossRefGoogle Scholar
  24. 24.
    Delhey K, Burger C, Fiedler W, Peters A (2010) Seasonal changes in colour: a comparison of structural, melanin- and carotenoid-based plumage colours. PLoS One 5:e11582. doi: 10.1371/journal.pone.0011582 PubMedCrossRefGoogle Scholar
  25. 25.
    Parrot Preservation Society (2010) Do you know the differences between Central American, South American and Mexican scarlet macaws? Accessed 15 Jun 2010
  26. 26.
    Gebhardt KJ, Brightsmith D, Powell G, Waits LP (2009) Molted feathers from clay licks in Peru provided DNA for three large macaws (Ara ararauna, A. chloropterus, and A. macao). J Field Ornithol 80:183–192. doi: 10.1111/j.1557-9263.2009.00221.x CrossRefGoogle Scholar
  27. 27.
    Proctor NS, Lynch PJ (1993) Manual of ornithology. Yale University Press, New Haven and LondonGoogle Scholar
  28. 28.
    Meyer CP, Paulay G (2005) DNA barcoding: error rates based on comprehensive sampling. PLoS Biology 3:e422. doi: 10.1371/journal.pbio.0030422 PubMedCrossRefGoogle Scholar
  29. 29.
    Lowenstein JH, Amato G, Kolokotronis S-O (2009) The real maccoyii: identifying tuna sushi with DNA barcodes—contrasting characteristic attributes and genetic distances PLoS ONE 4:e7866. doi: 10.1371/journal.pone.0007866
  30. 30.
    Moritz C, Cicero C (2004) DNA barcoding: promise and pitfalls. PLoS Biol 2:e354. doi: 10.1371/journal.pbio.0020354 PubMedCrossRefGoogle Scholar
  31. 31.
    Dubut V, Sinama M, Martin J, Meglécz E, Fernandez J, Chappaz R, Gilles A, Costedoat C (2010) Cross-species amplification of 41 microsatellites in European cyprinids: a tool for evolutionary, population genetics and hybridization studies. BMC Res Notes 3:135. doi: 10.1186/1756-0500-3-135 PubMedCrossRefGoogle Scholar
  32. 32.
    Abramson J (1995) Identification and profiles. In: Abramson J, Speer BL, Thomsen JB (eds) The large macaws. Their care, breeding and conservation, 1st edn. Raintree Publications, CA, pp 1–37Google Scholar
  33. 33.
    Alacs EA, Georges A, FitzSimmons NN, Robertson J (2010) DNA detective: a review of molecular approaches to wildlife forensics. Forensic Sci Med Pathol 6:180–194. doi: 10.1007/s12024-009-9131-7 PubMedCrossRefGoogle Scholar
  34. 34.
    Faria PJ, Guedes NMR, Yamashita C, Martuscelli P, Miyaki CY (2008) Genetic variation and population structure of the endangered hyacinth macaw (Anodorhynchus hyacinthinus): implications for conservation. Biodivers Conserv 17:765–779. doi: 10.1007/s10531-007-9312-1 CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Hideaki Abe
    • 1
  • Azusa Hayano
    • 1
  • Miho Inoue-Murayama
    • 1
  1. 1.Wildlife Research Center of Kyoto UniversityKyotoJapan

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