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Non-invasive genetic identification of two sympatric sister-species: ocelot (Leopardus pardalis) and margay (L. wiedii) in different biomes

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Abstract

The method presented in here allows for the unequivocal differentiation between non-invasive samples of ocelots and margays, when used in combination with the protocol by Roques et al. (Mol Ecol Resour 11:171–175, 2011). It reduced costs and laboratory processing, decreasing the chances of contamination, and therefore facilitating the identification of large numbers of samples by simple PCR amplification, without the need for sequencing. Our protocol was tested on 371 faeces collected in the field in 25 different sites across the distribution areas of both species, and compared to the mtDNA sequencing of 23 of the samples. The concatenation of a commonly used segment of Cytochrome b with other mitochondrial markers such as ATP-8 allows the identification of these two sister-species, but there was not enough resolution when used alone. With this new method, we could identify 98% of the field faeces, obtaining a global rate of 22.3 faeces of ocelot per margay sample in roads and trails located in potential areas of coexistence, suggesting generally lower densities of margay, although margays could be underestimated due to their arboreal habits. In Serra das Almas, located in the Brazilian Caatinga, the number of faeces of ocelot detected was especially elevated. Low densities of ocelots could be favoring higher densities of margays in areas like the Sinaloan dry forests and the Chimalapas montane forests, both in Mexico. Ocelots were found from open to forested habitats, including mosaic cropland and grassland, however margays were only found in forests where average canopy height exceeded 5 m.

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Acknowledgements

This study was carried out under project BIOCON 05–100/06 of Fundación BBVA, Brazil/Spain joint Project CNPq # 690085/02–8/CSIC # 2004BR0009, Project CGL2010-16902 of the Spanish Ministry of Research and Innovation, Project CGL2013-46026-P of MINECO, and the excellence Project RNM 2300 of Junta de Andalucía. MZ is supported by CNPq DCR fellowship Number 312627/2015-7. Samplings in Brazil were carried out under licenses # 131/2005 CGFAU/LIC of the Brazilian Institute for the Environment and Renewable Natural Resources—IBAMA, and# 11214-1, 13781-1, 13883-1 and 15664-1 SISBIO/Chico Mendes Institute for Biodiversity Conservation—ICMBio, in Mexico under licence # SGPA/DGVS/549 of Dirección General de Vida Silvestre (Semarnat) and in Bolivia under permission from the Sernap (Servicio Nacional de Areas Protegidas) and the DGBAP (Dirección General de Biodiversidad y Areas Protegidas). Faecal and blood samples were exported from Brazil to Spain for genetic analyses under IBAMA/CGEN authorization of access licence # 063/05 and IBAMA/CITES export licences # 0123242BR, 08BR002056/DF and 09BR003006/DF, from Mexico to Spain under export licences # MX33790 and MX42916 of Secretaría de Medio Ambiente/CITES and from Bolivia to Spain with an express permission from the MMayA (Ministerio de Medio Ambiente y Agua). We thank Cap. Ferreira and Lt. Carlos Palhari from the Veterinary Division of the Jungle Warfare Instruction Center of the Brazilian Armed Forces—CIGS, and Diogo Lagroteria and Carlos Abrahão, from IBAMA’s Wildlife Center—NUFAS, in Manaus (Amazonas, Brazil) for facilitating faeces and blood samples from captive felids in their facilities. Raphael de Almeida, Eduardo Ramos, Grasiela Porfírio, Tiago Boscarato, Javier Calzada, Miguel Delibes, Eloy Revilla, Julia Martínez, Gloria Clemencia Amaya, José da Silva Lópes, José Tavares, Carolina Jorge dos Santos, Xênya Garcia Bernardes da Luz, Denise Mello do Prado, Patricia Carignano Torres, Daniel Pererira Munari, Juan Carlos Faller, Meredic Calleja, Erik Joaquín Torres-Romero, Eduardo Espinoza-Medinilla, Ana Alicia Morales and Ricardo Barberí helped with the collection of samples, and Gerardo Ceballos and Heliot Zarza provided logistic support. We thank the managers of the Caiman Ecological Refuge, Serra da Capivara National Park, the Fundação Museu do Homem Americano, the Virua National Park (Bea Lisboa and Antonio Lisboa), Uatumã Biological Reserve, Maracá Ecological Station and the Serra das Almas Natural Reserve (Thiago Roberto Soares Vieira) in Brazil, the Edén Ecological Reserve (Marco Lazcano), El Zapotal Ecological Reserve (Pronatura Península de Yucatán, Juan Carlos Faller and María Andrade) and the Calakmul National Park in Mexico, and AMNI San Matías in Bolivia for their logistical support. Logistical support was provided by Laboratorio de Ecología Molecular, Estación Biológica de Doñana, CSIC (LEM-EBD). Séverine Roques, Noa González and Alberto García assisted in the analysis of samples. Two anonymous referees improved with their comments an earlier draft of the manuscript.

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Authors and Affiliations

  1. Department of Conservation Biology, Doñana Biological Station, CSIC, Avda. Américo Vespucio s/n, CP 41092, Seville, Spain

    Begoña Adrados, Pablo Villalva, Noa González-Borrajo & Francisco Palomares

  2. Departamento de Biologia, Centro de Ciências Humanas e Naturais, Universidade Federal do Espírito Santo, Vitória, Espírito Santo, Brazil

    Marina Zanin

  3. Jaguar Conservation Fund/Instituto Onça-Pintada, CP 193, Mineiros, GO, 75830-000, Brazil

    Leandro Silveira

  4. Universidad Autónoma Metropolitana-Unidad Lerma, Hidalgo Pte. 46, Col. La Estación, 52006, Lerma, Estado De México, Mexico

    Cuauhtemoc Chávez

  5. Department of Biodiversity, Instituto Nacional de Pesquisas da Amazônia – INPA, CP 2223, Manaus, Amazonas, 69060-000, Brazil

    Claudia Keller

  6. Panthera, 8 West 40th Street, 18th Floor, New York, NY, 10018, USA

    Bart J. Harmsen

  7. Environmental Research Institute, University of Belize, UB preschool Grounds, Price Centre Road, PO box 340, Belmopan, Belize

    Bart J. Harmsen

  8. Escuela de Biología, Universidad Autónoma de Sinaloa, Culiacán, Sinaloa, Mexico

    Yamel Rubio

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  1. Begoña Adrados
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Adrados, B., Zanin, M., Silveira, L. et al. Non-invasive genetic identification of two sympatric sister-species: ocelot (Leopardus pardalis) and margay (L. wiedii) in different biomes. Conservation Genet Resour 11, 203–217 (2019). https://doi.org/10.1007/s12686-018-0992-5

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