Identificación molecular mediante código de barras de DNA de moscas Lucilia (Diptera: Calliphoridae) recolectadas en Costa Rica
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Publicado:
feb. 27, 2020
Palabras clave:
Código de barras ADN, COI, 28S, Lucilia, Costa Rica
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Resumen
La identificación del género Lucilia es importante cuando se recolectan adultos o larvas en casos forenses porque el intervalo post-mortem puede ser estimado. En este trabajo se analizó morfológica y molecularmente un panel de especímenes de Lucilia (n = 42) recolectados en Costa Rica, de los cuales el 21% de las muestras corresponden a moscas criadas a partir de larvas tomadas de cadáveres humanos. Morfológicamente, los ejemplares fueron identificados como Lucilia cuprina [1] (33,3% de los ejemplares), Lucilia eximia [1] (33,3%), Lucilia purpurascens [2] (21,4%) o miembros del complejo Lucilia (11,9%). La identificación molecular, con las regiones de la citocromo oxidasa I (COI) o de la subunidad ribosomal 28S, sólo fue posible para el 43% de las muestras. La mayoría de las secuencias obtenidas con COI fueron concordantes con los análisis morfológicos, pero la región 28S carecía de suficiente resolución para identificar muestras al nivel de especies, excepto para L. cuprina. Juntos, los datos genéticos y morfológicos indican que L. cuprina y L. eximia fueron las especies más comúnmente encontradas; las moscas criadas a partir de larvas tomadas de cadáveres humanos correspondieron sólo a estas especies. Hasta donde sabemos, este trabajo es el primero en Costa Rica y en la región centroamericana en describir la entomofauna cadavérica del género Lucilia, lo cual es valioso para el desarrollo de potenciales aplicaciones forenses.
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Cómo citar
Identificación molecular mediante código de barras de DNA de moscas Lucilia (Diptera: Calliphoridae) recolectadas en Costa Rica. (2020). Revista Tecnología En Marcha, 33(1), Pág. 99–110. https://doi.org/10.18845/tm.v33i1.5025
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Citas
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[2] F. Walker, “Descriptions, & c. of the Diptera”. In: Curtis, J., Haliday, A.H. & Walker, F. (Eds.), Descriptions, & c. of the insects collected by Captain P.P. King, R.N., F.R.S., in the survey of the Straits of Magellan. Transactions of the Linnean Society of London, 17(3): 331–359, 1836.
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[5] S. Y. Cho et al., “DNA barcode library and its efficacy for identifying food-associated insect pests in Korea”. Bull. Entomol. Res. vol. 43, pp. 253–261, 2013. https://doi.org/10.1111/1748-5967.12034
[6] T. Withworth, “A revision of the neotropical species of Lucilia Robineau-Desvoidy”, Zootaxa, vol. 3810, pp. 1–76, 2014. http://dx.doi.org/10.11646/zootaxa.3810.1.1
[7] S. N. Kutty et al., “The Muscoidea (Diptera: Calyptratae) are paraphyletic: Evidence from four mitochondrial and four nuclear genes”, Mol Phylogenet Evol, vol. 49, pp. 639–652, 2008. https://doi.org/10.1016/j.ympev.2008.08.012
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[11] F.A.H. Sperling et al., “A DNA-Based Approach to the Identification of Insect Species Used for Postmorten Interval Estimation”, J Forensic Sci, vol. 39, pp. 418–427, 1994. https://doi.org/10.1520/JFS13613J
[12] P. D. N. Hebert et al., “Biological identifications through DNA barcodes”, Proc R Soc Lond [Biol],vol. 270, pp. 313–21, 2003. https://doi.org/10.1098/rspb.2002.2218
[13] J. F. Wallman and S.C. Donnellan, “The utility of mitochondrial DNA sequences for the identification of forensically important blowflies (Diptera: Calliphoridae) in southeastern Australia”, Forensic Sci Int, vol. 120, pp. 60–67, 2001. https://doi.org/10.1016/S0379-0738(01)00426-1
[14] J. D. Wells and F. A. H. Sperling, “DNA-based identification of forensically important Chrysomyinae (Diptera: Calliphoridae)”, Forensic Sci Int, vol. 120, pp. 110–115, 2001. https://doi.org/10.1016/S0379-0738(01)00414-5
[15] R. Zehner B. K. Zajac et al., “Genetic identification of forensically important flesh flies (Diptera: Sarcophagidae)” Int J Legal Med., vol. 118, pp. 245–247, 2004. https://doi.org/10.1007/s00414-004-0445-4
[16] W. Y. Chen et al., “Molecular identification of forensically important blow fly species (Diptera: Calliphoridae) in Taiwan”, J Med Entomol, vol. 41, pp. 47–57, 2004. https://doi.org/10.1603/0022-2585-41.1.47
[17] L. Cainé et al., “Genetic identification of forensically important Calliphoridae in Portugal”, Int Congr Ser, vol. 1288, pp. 846–848, 2006. https://doi.org/10.1016/j.ics.2005.09.153
[18] J. D. Wells and D. W. Williams, “Validation of a DNA-based method for identifying Chrysomyinae (Diptera: Calliphoridae) used in a death investigation”, Int J Legal Med., vol. 121, pp. 1–8, 2007. https://doi.org/10.1007/s00414-005-0056-8
[19] M. L. Harvey et al., “A global study of forensically significant calliphorids: Implications for identification”, Forensic Sci Int, vol. 177, pp. 66–76, 2008. https://doi.org/10.1016/j.forsciint.2007.10.009
[20] S. Reibe et al., “Molecular identification of forensically important blowfly species (Diptera: Calliphoridae) from Germany”, Parasitol Res, vol. 106, pp. 257–261, 2009. https://doi.org/10.1007/s00436-009-1657-9
[21] P. Boehme et al., “Molecular identification of carrion-breeding scuttle flies (Diptera: Phoridae) using COI barcodes”, Int J Legal Med., vol. 124, pp. 577–581, 2010. https://doi.org/10.1007/s00414-010-0429-5
[22] P. Boehme et al., “The use of COI barcodes for molecular identification of forensically important fly species in Germany”, Parasitol Res, vol. 110, pp. 2325–2332, 2012. https://doi.org/10.1007/s00436-011-2767-8
[23] K.A. Meiklejohn et al., “DNA-based identification of forensically important Australian Sarcophagidae (Diptera)”, Int J Legal Med., vol. 125, pp. 27–32, 2011. https://doi.org/10.1007/s00414-009-0395-y
[24] K. Jordaens et al., “Identification of forensically important Sarcophaga species (Diptera: Sarcophagidae) using the mitochondrial COI gene”, Int J Legal Med., vol. 127, pp. 491–504, 2013. https://doi.org/10.1007/s00414-012-0767-6
[25] M. GilArriortua et al., “Molecular differentiation of Central European blowfly species (Diptera, Calliphoridae) using mitochondrial and nuclear genetic markers”, Forensic Sci Int, vol. 242, pp. 274–282, 2014. https://doi.org/10.1016/j.forsciint.2014.07.018
[26] Y. Guo et al., “Identification of forensically important sarcophagid flies (Diptera: Sarcophagidae) in China based on COI and period gene”, Int J Legal Med., vol. 128, pp. 221–228, 2014. https://doi.org/10.1007/s00414-013-0923-7
[27] B. K. Zajac et al., “Application of DNA barcoding for identifying forensically relevant Diptera from northern Thailand”, Parasitol Res, vol. 115, pp. 2307–2320, 2016. https://doi.org/10.1007/s00436-016-4977-6
[28] J. D. Wells et al., “Phylogenetic analysis of forensically important Lucilia flies based on cytochrome oxidase I sequence: A cautionary tale for forensic species determination” Int J Legal Med., vol. 121, pp. 229–233, 2007. https://doi.org/10.1007/s00414-006-0147-1
[29] G. Sonet et al., “Why is the molecular identification of the forensically important blowfly species Lucilia caesar and L. illustris (family Calliphoridae) so problematic?”, Forensic Sci Int, vol. 223, pp. 153–159, 2012. https://doi.org/10.1016/j.forsciint.2012.08.020
[30] R. W. Debry et al., “DNA-Based Identification of Forensically Important Lucilia (Diptera: Calliphoridae) in the Continental United States”, J Forensic Sci, vol. 58, pp. 73–78, 2013. https://doi.org/10.1111/j.1556-4029.2012.02176.x
[31] M. GilArriortua et al., “Technical Note: “Mitochondrial and nuclear DNA approaches for reliable identification of Lucilia (Diptera, Calliphoridae) species of forensic interest from Southern Europe”, Forensic Sci Int, vol. 257, pp. 393–397, 2015. https://doi.org/10.1016/j.forsciint.2015.10.010
[32] J. Vargas “Distribución y morfología de adultos e inmaduros de moscas califóridas (Diptera: Calliphoridae) de importancia forense en Costa Rica”, Lic. Universidad de Costa Rica, 1999.
[33] R. Morales, Elaboración de una clave taxonómica para adultos y larvas de las especies de los géneros Lucilia y Hemilucilia (Diptera: Calliphoridae) de Costa Rica. Lic. Universidad de Costa Rica, 2014. http://hdl.handle.net/10669/29686
[34] T. Hall, “BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT”, Nucleic Acids Symp Ser, vol. 41, pp. 95–98, 1999.
[35] K. Tamura et al., “MEGA6: Molecular evolutionary genetics analysis version 6.0”, Mol Biol Evol, vol. 30, pp. 2725–2729, 2013. https://doi.org/10.1093/molbev/mst197
[36] R. C. Edgar “MUSCLE: Multiple sequence alignment with high accuracy and high throughput”, Nucleic Acids Res, vol. 32, pp. 1792–1797, 2004.
[37] J. Amendt et al., “Best practice in forensic entomology - Standards and guidelines”, Int J Legal Med., vol. 121, pp. 90–104, 2007. https://doi.org/10.1007/s00414-006-0086-x
[38] J. Macquart, “New or little known exotic diptera”, Volume two, 3rd part. Roret, Paris, 304 pp, 1844.
[39] J.W. Meigen, “Systematic description of the known European two-winged insects”, Fifth part. Schulz, Hamm, 412 pp, 1826.
[40] R.C. Shannon, “Synopsis of the American Calliphoridae (Diptera)”. Proceedings of the Entomological Society of Washington, 28, 115–139, 1926.
[41] R. W. Debry et al., “MtDNA-based identification of Lucilia cuprina (Wiedemann) and Lucilia sericata (Meigen) (Diptera: Calliphoridae) in the continental United States”, Forensic Sci Int, vol. 202, pp. 102–109, 2010. https://doi.org/10.1016/j.forsciint.2010.04.038
[42] M. GilArriortua et al., “Cytochrome b as a useful tool for the identification of blowflies of forensic interest (Diptera, Calliphoridae)”, Forensic Sci Int, vol. 228, pp. 132–136, 2013. https://doi.org/10.1016/j.forsciint.2013.02.037
[43] J. F. Wallman and S.C. Donnellan, “The utility of mitochondrial DNA sequences for the identification of forensically important blowflies (Diptera: Calliphoridae) in southeastern Australia”, Forensic Sci Int, vol. 120, pp. 60–67, 2001. https://doi.org/10.1016/S0379-0738(01)00426-1
[2] F. Walker, “Descriptions, & c. of the Diptera”. In: Curtis, J., Haliday, A.H. & Walker, F. (Eds.), Descriptions, & c. of the insects collected by Captain P.P. King, R.N., F.R.S., in the survey of the Straits of Magellan. Transactions of the Linnean Society of London, 17(3): 331–359, 1836.
[3] J. Amendt et al., “Forensic entomology”, Naturwissenschaften, vol. 91, pp. 51–65, 2004. https://doi.org/10.1007/s00114-003-0493-5
[4] M.A.T. Marinho et al., “Molecular phylogenetics of Oestroidea (Diptera: Calyptratae) with emphasis on Calliphoridae: Insights into the inter-familial relationships and additional evidence for paraphyly among blowflies”, Mol Phylogenet Evol, vol. 65, pp. 840–854, 2012. https://doi.org/10.1016/j.ympev.2012.08.007
[5] S. Y. Cho et al., “DNA barcode library and its efficacy for identifying food-associated insect pests in Korea”. Bull. Entomol. Res. vol. 43, pp. 253–261, 2013. https://doi.org/10.1111/1748-5967.12034
[6] T. Withworth, “A revision of the neotropical species of Lucilia Robineau-Desvoidy”, Zootaxa, vol. 3810, pp. 1–76, 2014. http://dx.doi.org/10.11646/zootaxa.3810.1.1
[7] S. N. Kutty et al., “The Muscoidea (Diptera: Calyptratae) are paraphyletic: Evidence from four mitochondrial and four nuclear genes”, Mol Phylogenet Evol, vol. 49, pp. 639–652, 2008. https://doi.org/10.1016/j.ympev.2008.08.012
[8] J. Vargas and D.M. Wood, “Calliphoridae (blow flies)”, in Manual of Centro-American Diptera, B. Brown et al., M. (eds), Ottawa: NRC Research Press, pp. 1297–1304, 2010.
[9] B. K. Zajac et al., “Molecular identification and phylogenetic analysis of the forensically important family Piophilidae (Diptera) from different European locations”, Forensic Sci Int, vol. 259, pp. 77–84, 2016. https://doi.org/10.1016/j.forsciint.2015.12.024
[10] U. Jinbo et al., “Current progress in DNA barcoding and future implications for entomology”, Entomol Sci, vol. 14, pp. 107–124, 2011. https://doi.org/10.1111/j.1479-8298.2011.00449.x
[11] F.A.H. Sperling et al., “A DNA-Based Approach to the Identification of Insect Species Used for Postmorten Interval Estimation”, J Forensic Sci, vol. 39, pp. 418–427, 1994. https://doi.org/10.1520/JFS13613J
[12] P. D. N. Hebert et al., “Biological identifications through DNA barcodes”, Proc R Soc Lond [Biol],vol. 270, pp. 313–21, 2003. https://doi.org/10.1098/rspb.2002.2218
[13] J. F. Wallman and S.C. Donnellan, “The utility of mitochondrial DNA sequences for the identification of forensically important blowflies (Diptera: Calliphoridae) in southeastern Australia”, Forensic Sci Int, vol. 120, pp. 60–67, 2001. https://doi.org/10.1016/S0379-0738(01)00426-1
[14] J. D. Wells and F. A. H. Sperling, “DNA-based identification of forensically important Chrysomyinae (Diptera: Calliphoridae)”, Forensic Sci Int, vol. 120, pp. 110–115, 2001. https://doi.org/10.1016/S0379-0738(01)00414-5
[15] R. Zehner B. K. Zajac et al., “Genetic identification of forensically important flesh flies (Diptera: Sarcophagidae)” Int J Legal Med., vol. 118, pp. 245–247, 2004. https://doi.org/10.1007/s00414-004-0445-4
[16] W. Y. Chen et al., “Molecular identification of forensically important blow fly species (Diptera: Calliphoridae) in Taiwan”, J Med Entomol, vol. 41, pp. 47–57, 2004. https://doi.org/10.1603/0022-2585-41.1.47
[17] L. Cainé et al., “Genetic identification of forensically important Calliphoridae in Portugal”, Int Congr Ser, vol. 1288, pp. 846–848, 2006. https://doi.org/10.1016/j.ics.2005.09.153
[18] J. D. Wells and D. W. Williams, “Validation of a DNA-based method for identifying Chrysomyinae (Diptera: Calliphoridae) used in a death investigation”, Int J Legal Med., vol. 121, pp. 1–8, 2007. https://doi.org/10.1007/s00414-005-0056-8
[19] M. L. Harvey et al., “A global study of forensically significant calliphorids: Implications for identification”, Forensic Sci Int, vol. 177, pp. 66–76, 2008. https://doi.org/10.1016/j.forsciint.2007.10.009
[20] S. Reibe et al., “Molecular identification of forensically important blowfly species (Diptera: Calliphoridae) from Germany”, Parasitol Res, vol. 106, pp. 257–261, 2009. https://doi.org/10.1007/s00436-009-1657-9
[21] P. Boehme et al., “Molecular identification of carrion-breeding scuttle flies (Diptera: Phoridae) using COI barcodes”, Int J Legal Med., vol. 124, pp. 577–581, 2010. https://doi.org/10.1007/s00414-010-0429-5
[22] P. Boehme et al., “The use of COI barcodes for molecular identification of forensically important fly species in Germany”, Parasitol Res, vol. 110, pp. 2325–2332, 2012. https://doi.org/10.1007/s00436-011-2767-8
[23] K.A. Meiklejohn et al., “DNA-based identification of forensically important Australian Sarcophagidae (Diptera)”, Int J Legal Med., vol. 125, pp. 27–32, 2011. https://doi.org/10.1007/s00414-009-0395-y
[24] K. Jordaens et al., “Identification of forensically important Sarcophaga species (Diptera: Sarcophagidae) using the mitochondrial COI gene”, Int J Legal Med., vol. 127, pp. 491–504, 2013. https://doi.org/10.1007/s00414-012-0767-6
[25] M. GilArriortua et al., “Molecular differentiation of Central European blowfly species (Diptera, Calliphoridae) using mitochondrial and nuclear genetic markers”, Forensic Sci Int, vol. 242, pp. 274–282, 2014. https://doi.org/10.1016/j.forsciint.2014.07.018
[26] Y. Guo et al., “Identification of forensically important sarcophagid flies (Diptera: Sarcophagidae) in China based on COI and period gene”, Int J Legal Med., vol. 128, pp. 221–228, 2014. https://doi.org/10.1007/s00414-013-0923-7
[27] B. K. Zajac et al., “Application of DNA barcoding for identifying forensically relevant Diptera from northern Thailand”, Parasitol Res, vol. 115, pp. 2307–2320, 2016. https://doi.org/10.1007/s00436-016-4977-6
[28] J. D. Wells et al., “Phylogenetic analysis of forensically important Lucilia flies based on cytochrome oxidase I sequence: A cautionary tale for forensic species determination” Int J Legal Med., vol. 121, pp. 229–233, 2007. https://doi.org/10.1007/s00414-006-0147-1
[29] G. Sonet et al., “Why is the molecular identification of the forensically important blowfly species Lucilia caesar and L. illustris (family Calliphoridae) so problematic?”, Forensic Sci Int, vol. 223, pp. 153–159, 2012. https://doi.org/10.1016/j.forsciint.2012.08.020
[30] R. W. Debry et al., “DNA-Based Identification of Forensically Important Lucilia (Diptera: Calliphoridae) in the Continental United States”, J Forensic Sci, vol. 58, pp. 73–78, 2013. https://doi.org/10.1111/j.1556-4029.2012.02176.x
[31] M. GilArriortua et al., “Technical Note: “Mitochondrial and nuclear DNA approaches for reliable identification of Lucilia (Diptera, Calliphoridae) species of forensic interest from Southern Europe”, Forensic Sci Int, vol. 257, pp. 393–397, 2015. https://doi.org/10.1016/j.forsciint.2015.10.010
[32] J. Vargas “Distribución y morfología de adultos e inmaduros de moscas califóridas (Diptera: Calliphoridae) de importancia forense en Costa Rica”, Lic. Universidad de Costa Rica, 1999.
[33] R. Morales, Elaboración de una clave taxonómica para adultos y larvas de las especies de los géneros Lucilia y Hemilucilia (Diptera: Calliphoridae) de Costa Rica. Lic. Universidad de Costa Rica, 2014. http://hdl.handle.net/10669/29686
[34] T. Hall, “BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT”, Nucleic Acids Symp Ser, vol. 41, pp. 95–98, 1999.
[35] K. Tamura et al., “MEGA6: Molecular evolutionary genetics analysis version 6.0”, Mol Biol Evol, vol. 30, pp. 2725–2729, 2013. https://doi.org/10.1093/molbev/mst197
[36] R. C. Edgar “MUSCLE: Multiple sequence alignment with high accuracy and high throughput”, Nucleic Acids Res, vol. 32, pp. 1792–1797, 2004.
[37] J. Amendt et al., “Best practice in forensic entomology - Standards and guidelines”, Int J Legal Med., vol. 121, pp. 90–104, 2007. https://doi.org/10.1007/s00414-006-0086-x
[38] J. Macquart, “New or little known exotic diptera”, Volume two, 3rd part. Roret, Paris, 304 pp, 1844.
[39] J.W. Meigen, “Systematic description of the known European two-winged insects”, Fifth part. Schulz, Hamm, 412 pp, 1826.
[40] R.C. Shannon, “Synopsis of the American Calliphoridae (Diptera)”. Proceedings of the Entomological Society of Washington, 28, 115–139, 1926.
[41] R. W. Debry et al., “MtDNA-based identification of Lucilia cuprina (Wiedemann) and Lucilia sericata (Meigen) (Diptera: Calliphoridae) in the continental United States”, Forensic Sci Int, vol. 202, pp. 102–109, 2010. https://doi.org/10.1016/j.forsciint.2010.04.038
[42] M. GilArriortua et al., “Cytochrome b as a useful tool for the identification of blowflies of forensic interest (Diptera, Calliphoridae)”, Forensic Sci Int, vol. 228, pp. 132–136, 2013. https://doi.org/10.1016/j.forsciint.2013.02.037
[43] J. F. Wallman and S.C. Donnellan, “The utility of mitochondrial DNA sequences for the identification of forensically important blowflies (Diptera: Calliphoridae) in southeastern Australia”, Forensic Sci Int, vol. 120, pp. 60–67, 2001. https://doi.org/10.1016/S0379-0738(01)00426-1