Noninvasive tracking of jaguars (Panthera onca) and co-occurring Neotropical felids in Belize, Central America by genotyping feces and remote camera trapping

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Virginia Tech


The elusive jaguar (Panthera onca) is extremely difficult to study due to its wide-ranging behavior, crepuscular activity peaks and its occurrence in low population densities in often dense forest habitats. Jaguars are also a species of concern, but our ability to provide for their survival is hampered by our inability to obtain reliable information on the status of their wild populations.

This study combines innovative noninvasive research techniques such as scat detector dogs and molecular scatology to conduct the first genetic study on wild populations of Neotropical felids coexisting across fragmented forest habitats in Belize, Central America. Specifically, we analyzed multi-locus data in jaguars (Panthera onca), pumas (Puma concolor) and ocelots (Leopardus pardalis) collected from 1053 scat samples across their range in the country. First, we optimized 14 polymorphic microsatellite loci for jaguars (Panthera onca), pumas (Puma concolor), and ocelots (Leopardus pardalis), and assessed their utility for cross-species amplification. Additionally, we tested their reliability for species and individual identification using fecal DNA as he primary DNA source. All microsatellite loci examined successfully cross-amplified in the three target species, and were polymorphic. Second, to maximize PCR amplification success and genotyping accuracy rates, and to minimize genotyping error rates for fecal DNA samples, we evaluated the performance of two fecal DNA storage techniques (dimethyl sulfoxide saline solution/DET buffer, 95% EtOH) suitable for long-term preservation at remote tropical sites. Additionally, we tested fecal DNA samples collected from four different scat locations (top, side, bottom, inside). DET buffer was the superior fecal DNA preservation method and collecting fecal DNA from side and top locations of the scat resulted in the highest PCR success rates.

For the main genetic study, we assessed the genetic conservation status of all three target species across the country of Belize. We examined levels of genetic diversity within different sites, (2) defined potential genetic clusters/populations, (3) and examined levels of gene flow and population structure for all three target species on a countrywide scale. Furthermore, we compared genetic diversity and gene flow levels among the three target species. Wild felids in Belize showed moderate levels of heterozygosity (HE = 0.60 - 0.70) with jaguars having the lowest genetic diversity with average expected heterozygosities of HE = 0.60 ± 0.05 and allelic richness (AR) of 4.94 ± 0.44 followed by pumas with HE = 0.65 ± 0.06 and AR of 7.52 ± 0.86 and ocelots with HE = 0.70 ± 0.05 and AR of 3.89 ± 0.23. We observed low to moderate levels of differentiation (FST = 0.00 - 0.15) and weak population structure using spatial Bayesian clustering techniques for all three target species. Although levels of genetic diversity and gene flow across the country are still fairly high, we did detect evidence of fragmentation indicating the risk of further habitat loss and fragmentation for wild felids.

Felids were simultaneously monitored across all study sites by remote sensing camera traps, which allows for a comparison of density estimates obtained from two different noninvasive survey approaches. Furthermore, analytical methods for density estimation are advancing rapidly, making it difficult to choose the optimal technique. Thus, we compared a variety of density estimators including the conventional approach of estimating abundance ( ) in programs CAPTURE and MARK and dividing abundance by the effective trapping area (ETA), the recently developed spatially explicit capture-recapture (SECR) models, both the likelihood-based approach (ML-SECR) in program DENSITY and the Bayesian approach (B-SECR) in program SPACECAP, and finally the genetic-based mark-recapture one sampling occasion estimator in program CAPWIRE. Although different survey methods using various density estimators produced similar density estimates, confidence levels and coefficients of variation varied, with SECR methods resulting in the least precise estimates. Detection probabilities were generally higher for noninvasive genetic sampling than for camera trapping. Both techniques were shown to be reliable and highly efficient survey methods for density estimation of low-density Neotropical felids living in challenging environments such as the tropics. While less precise, SECR CMR models are probably a more realistic reflection of our uncertainty. They hold great promise for density estimation studies for wide-ranging and territorial carnivore species, especially if precision can be improved through study design or analysis advancements in the future.

In conclusion, our results demonstrated that noninvasive sampling techniques such as molecular scatology and remote camera trapping are efficient research approaches to study multiple Neotropical felids in a multifaceted way and on a countrywide scale. We believe that the techniques and analyses developed in this study are widely applicable and relevant to the conservation and management of other elusive and difficult to study wild felids worldwide.



Panthera onca, Puma concolor, Leopardus pardalis, genetic diversity, gene flow, dispersal, microsatellites, molecular scatology