Hindering the illegal trade in dog and cat furs through a DNA-based protocol for species identification
In Western countries dogs and cats are the most popular pets, and people are increasingly opposed to their rearing for the fur industry. In 2007, a Regulation of the European Union (EU) banned the use and trade of dog and cat furs, but an official analytical protocol to identify them as source speci...
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| Format: | Article |
| Language: | English |
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PeerJ Inc.
2018-06-01
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| Series: | PeerJ |
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| Online Access: | https://peerj.com/articles/4902.pdf |
| _version_ | 1797420123221393408 |
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| author | Luisa Garofalo Alessia Mariacher Rita Fanelli Rosario Fico Rita Lorenzini |
| author_facet | Luisa Garofalo Alessia Mariacher Rita Fanelli Rosario Fico Rita Lorenzini |
| author_sort | Luisa Garofalo |
| collection | DOAJ |
| description | In Western countries dogs and cats are the most popular pets, and people are increasingly opposed to their rearing for the fur industry. In 2007, a Regulation of the European Union (EU) banned the use and trade of dog and cat furs, but an official analytical protocol to identify them as source species was not provided, and violations of law are still frequent in all Member States. In this paper we report on the development and validation of a simple and affordable DNA method for species detection in furs to use as an effective tool to combat illegal trade in fur products. A set of mitochondrial primers was designed for amplification of partial cytochrome b, control region and ND1 gene in highly degraded samples, like furs and pelts. Our amplification workflow involved the use of a non-specific primer pair to perform a first test to identify the species through sequencing, then the application of species-specific primer pairs to use in singleplex end-point PCRs as confirmation tests. The advantage of this two-step procedure is twofold: on the one hand it minimises the possibility of negative test results from degraded samples, since failure of amplification with a first set of primers can be offset by successful amplification of the second, and on the other it adds confidence and reliability to final authentication of species. All designed primers were validated on a reference collection of tissue samples, obtaining solid results in terms of specificity, sensitivity, repeatability and reproducibility. Application of the protocol on real caseworks from seized furs yielded successful results also from old and dyed furs, suggesting that age and chemical staining do not necessarily affect positive amplifications. Major pros of this approach are: (1) sensitive and informative primer sets for detection of species; (2) short PCR amplicons for the analysis of poor quality DNA; (3) binding primers that avoid contamination from human DNA; (4) user-friendly protocol for any laboratory equipped for analysis of low-copy-number DNA. Our molecular procedure proved to be a good starting point for enforcing the EU Regulation against dog and cat fur trade in forensic contexts where source attribution is essential to the assignment of responsibilities. |
| first_indexed | 2024-03-09T06:58:08Z |
| format | Article |
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| institution | Directory Open Access Journal |
| issn | 2167-8359 |
| language | English |
| last_indexed | 2024-03-09T06:58:08Z |
| publishDate | 2018-06-01 |
| publisher | PeerJ Inc. |
| record_format | Article |
| series | PeerJ |
| spelling | doaj.art-615eb1c8018a46f7824cba589f6299802023-12-03T10:01:54ZengPeerJ Inc.PeerJ2167-83592018-06-016e490210.7717/peerj.4902Hindering the illegal trade in dog and cat furs through a DNA-based protocol for species identificationLuisa Garofalo0Alessia Mariacher1Rita Fanelli2Rosario Fico3Rita Lorenzini4Istituto Zooprofilattico Sperimentale delle Regioni Lazio e Toscana “M. Aleandri”, Centro di Referenza Nazionale per la Medicina Forense Veterinaria, Rieti, ItalyIstituto Zooprofilattico Sperimentale delle Regioni Lazio e Toscana “M. Aleandri”, Centro di Referenza Nazionale per la Medicina Forense Veterinaria, Grosseto, ItalyIstituto Zooprofilattico Sperimentale delle Regioni Lazio e Toscana “M. Aleandri”, Centro di Referenza Nazionale per la Medicina Forense Veterinaria, Rieti, ItalyIstituto Zooprofilattico Sperimentale delle Regioni Lazio e Toscana “M. Aleandri”, Centro di Referenza Nazionale per la Medicina Forense Veterinaria, Grosseto, ItalyIstituto Zooprofilattico Sperimentale delle Regioni Lazio e Toscana “M. Aleandri”, Centro di Referenza Nazionale per la Medicina Forense Veterinaria, Rieti, ItalyIn Western countries dogs and cats are the most popular pets, and people are increasingly opposed to their rearing for the fur industry. In 2007, a Regulation of the European Union (EU) banned the use and trade of dog and cat furs, but an official analytical protocol to identify them as source species was not provided, and violations of law are still frequent in all Member States. In this paper we report on the development and validation of a simple and affordable DNA method for species detection in furs to use as an effective tool to combat illegal trade in fur products. A set of mitochondrial primers was designed for amplification of partial cytochrome b, control region and ND1 gene in highly degraded samples, like furs and pelts. Our amplification workflow involved the use of a non-specific primer pair to perform a first test to identify the species through sequencing, then the application of species-specific primer pairs to use in singleplex end-point PCRs as confirmation tests. The advantage of this two-step procedure is twofold: on the one hand it minimises the possibility of negative test results from degraded samples, since failure of amplification with a first set of primers can be offset by successful amplification of the second, and on the other it adds confidence and reliability to final authentication of species. All designed primers were validated on a reference collection of tissue samples, obtaining solid results in terms of specificity, sensitivity, repeatability and reproducibility. Application of the protocol on real caseworks from seized furs yielded successful results also from old and dyed furs, suggesting that age and chemical staining do not necessarily affect positive amplifications. Major pros of this approach are: (1) sensitive and informative primer sets for detection of species; (2) short PCR amplicons for the analysis of poor quality DNA; (3) binding primers that avoid contamination from human DNA; (4) user-friendly protocol for any laboratory equipped for analysis of low-copy-number DNA. Our molecular procedure proved to be a good starting point for enforcing the EU Regulation against dog and cat fur trade in forensic contexts where source attribution is essential to the assignment of responsibilities.https://peerj.com/articles/4902.pdfDNA testingCytochrome bEU Regulation 1523/2007Control RegionMitochondrial ND1Fur industry |
| spellingShingle | Luisa Garofalo Alessia Mariacher Rita Fanelli Rosario Fico Rita Lorenzini Hindering the illegal trade in dog and cat furs through a DNA-based protocol for species identification PeerJ DNA testing Cytochrome b EU Regulation 1523/2007 Control Region Mitochondrial ND1 Fur industry |
| title | Hindering the illegal trade in dog and cat furs through a DNA-based protocol for species identification |
| title_full | Hindering the illegal trade in dog and cat furs through a DNA-based protocol for species identification |
| title_fullStr | Hindering the illegal trade in dog and cat furs through a DNA-based protocol for species identification |
| title_full_unstemmed | Hindering the illegal trade in dog and cat furs through a DNA-based protocol for species identification |
| title_short | Hindering the illegal trade in dog and cat furs through a DNA-based protocol for species identification |
| title_sort | hindering the illegal trade in dog and cat furs through a dna based protocol for species identification |
| topic | DNA testing Cytochrome b EU Regulation 1523/2007 Control Region Mitochondrial ND1 Fur industry |
| url | https://peerj.com/articles/4902.pdf |
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