Miryam Venegas-Anaya

Sequencing three crocodilian genomes to illuminate the evolution of archosaurs and amniotes

The International Crocodilian Genomes Working Group (ICGWG) will sequence and assemble the American alligator (Alligator mississippiensis), saltwater crocodile (Crocodylus porosus) and Indian gharial (Gavialis gangeticus) genomes. The status of these projects and our planned analyses are described.

Genetic characterization of captive Cuban crocodiles (Crocodylus rhombifer) and evidence of hybridization with the American crocodile (Crocodylus acutus).

There is a surprising lack of genetic data for the Cuban crocodile (Crocodylus rhombifer), especially given its status as a critically endangered species. Samples from captive individuals were used to genetically characterize this species in comparison with other New World crocodilians. Partial mitochondrial sequence data were generated from cyt-b (843 bp) and the tRNA(Pro)- tRNA(Phe)-D-loop region (442 bp). Phylogenetic analyses were performed by generating maximum parsimony, maximum likelihood, and Bayesian-based topologies. In addition, in an effort to identify species-specific alleles, ten polymorphic microsatellite loci were genotyped. Distance and model-based clustering analyses were performed on microsatellite data, in addition to a model-based assignment of hybrid types. Both mitochondrial and nuclear markers identified two distinct C. rhombifer genetic sub-clades (alpha and beta); and microsatellite analyses revealed that most admixed individuals were F(2) hybrids between C. rhombifer-alpha and the American crocodile (C. acutus). All individuals in the C. rhombifer-beta group were morphologically identified as C. acutus and formed a distinct genetic assemblage. J. Exp. Zool. 309A:649-660, 2008. (c) 2008 Wiley-Liss, Inc.

Mitochondrial DNA Phylogeography of Caiman crocodilus in Mesoamerica and South America

The Neotropical crocodylian species, Caiman crocodilus, is widely distributed through Mesoamerica, northern South America, and the Amazon basin. Four subspecies are recognized within C. crocodilus, suggesting some geographic variation in morphology. In this study, we utilized mitochondrial DNA (mtDNA) sequence data from 45 individuals of C. crocodilus throughout its range to infer its evolutionary history and population structure, as well as to evaluate genealogical support for subspecies and their geographic distributions. Our molecular phylogenetic results identified five mtDNA haplotype clades with a mean sequence divergence of 3.4%, indicating considerable evolutionary independence among phylogeographic lineages. Our results were also broadly consistent with current subspecific taxonomy, with some important additional findings. First, we found substantial genetic structuring within C. c. fuscus from southern Mesoamerica. Second, though we confirmed the existence of a widespread Amazonian clade, we also discovered a cryptic and divergent mtDNA lineage that was indistinguishable from C. c. crocodilus based on external morphology. Third, we confirm the status of C. c. chiapasius as a distinct evolutionary lineage, and provide evidence that C. c. fuscus may be moving northward and hybridizing with C. c. chiapasius in northern Mesoamerica. Finally, our results parallel previous phylogeographic studies of other organisms that have demonstrated significant genetic structure over shorter geographic distances in Mesoamerica compared with Amazonia. We support conservation efforts for all five independent lineages within C. crocodilus, and highlight the subspecies C. c. chiapasius as a unit of particular conservation concern.

Evolutionary history of Cuban crocodiles Crocodylus rhombifer and Crocodylus acutus inferred from multilocus markers

Among crocodilians, Crocodylus rhombifer is one of the worlds most endangered species with the smallest natural distribution. In Cuba, this endemic species coexists with the American crocodile (Crocodylus acutus). Hybridization between these two species is well known in captivity and might occur in the wild, but has never been demonstrated genetically. Here, we combined molecular data with environmental, geographic, and fossil data to infer the evolutionary history of Crocodylus in the Cuban Archipelago, and to evaluate genealogical support for species boundaries. We analyzed seven microsatellite loci plus DNA sequence data from nuclear (RAG-1) and mitochondrial (cytochrome b and cytochrome oxidase I) genes from 89 wild-caught individuals in Cuba, Grand Cayman Island, Jamaica, and Central America, and two samples from zoo collections. Microsatellites showed evidence of introgression, suggesting potential hybridization among Cuban groups. In Cuba, C. acutus contained one mitochondrial DNA (mtDNA) haplotype, whereas C. rhombifer contained two haplotypes. MtDNA data showed that C. acutus is paraphyletic with respect to C. rhombifer, revealing 1% sequence divergence between species within Cuba vs. 8% divergence between Cuban forms and mainland C. acutus. We suggest that hybridization has been a historical as well as a current phenomenon between C. acutus and C. rhombifer. These findings suggest that long-term conservation of crocodiles in Cuba will require identification of genetically pure and hybrid individuals, and a decrease in anthropogenic activities. We also recommend more extensive morphological and genetic analyses of Cuban population to establish clear boundaries of the hybrid zone between C. acutus and C. rhombifer.

Population Ecology of American Crocodile (Crocodylus acutus) in Coiba National Park, Panama

Abstract We conducted nocturnal surveys in the insular and coastal areas of Coiba National Park (CNP) and its mainland buffer zone in Panama (Chiriquí conservation site) from 2009–2012 to determine the conservation status of Crocodylus acutus. In 99 nights, we surveyed 147.2 km and captured 185 animals during nocturnal transects inspection with headlamps. Overall, sex ratio was 1.00:1.01 female/male with significant differences by size/age class and year. Females were slightly larger in total length than males (115.1 ± 56.9 cm-females, 105.4 ± 71.8 cm-males). The encounter rate was calculated based on number of animals captured per km of surveyed transect. The C. acutus encounter rate per year was 1.8 ind/km (60 ind/33.5 km/12 places visited) in 2009, 1.0 ind/km (90 ind/87.4 km/18 places visited) in 2010, and 1.3 ind/km (35 ind/26.3 km/8 places visited) in 2012. Based on our spatial analysis, the animals showed a dispersed pattern in most sites on CNP. Captured C. acutus were found in 581.1 km2 total area within 78% natural habitat, including mangroves and beaches, and 22% disturbed habitat on both the mainland and the islands. In addition, the spatial analysis showed reduction in natural land cover; crocodile habitat showed limited conversion to agricultural land use; and we found correlation between crocodile population size and protected areas. The differences between mainland and island populations regarding ecology suggest that a long-term monitoring program for American Crocodiles is necessary to distinguish between natural fluctuations and anthropogenic changes on population dynamics and conservation status. Resumen Entre 2009 al 2012 hicimos varias expediciones en la parte insular y en las áreas costeras del Parque Nacional Coiba y su zona de influencia en Panamá (Sitio de conservación para la diversidad), para determinar el estado de conservación de Crocodylus acutus y sus hábitats asociados. En 99 noches recorrimos 147.2 km y capturamos 185 animales usando linternas de cabeza. La proporción de sexos fue 1.00:1.01 hembra/macho, la cual fue significativamente diferente cuando los animales fueron clasificados por grupo etario y año. Las hembras fueron un poco más grandes que los machos (115.1 ± 56.9 cm-hembras, 105.4 ± 71.8 cm-machos). La tasa de encuentro fue calculada con base en el número de animales capturados por kilómetro. La tasa de encuentro para C. acutus por año fue de 1.8 ind/km (n = 60; 33.5 km; lugares visitados: 12) en 2009, 1.0 ind/km (n = 90; 87.4 km; lugares visitados: 18) in 2010 y 1.3 ind/km (n = 35; 26.3 km; 26.3; lugares visitados: 8) en 2012. El análisis espacial mostró a C. acutus distribuido en la isla de Coiba en un patrón disperso, excepto en El María y el Playa Blanca donde el patrón de dispersión fue de agrupamiento. Los animales capturados se encontraban en un área total de 581.12 km2 cubierta por un 78% de manglares y playas y un 22% por hábitats modificados. Comparando la cobertura vegetal de 1992 y 2000, nuestros resultados mostraron baja densidad poblacional asociada a la reducción de hábitat. Las diferencias en la ecología poblacional entre los sitios continentales y los insulares sugieren que un programa de monitoreo a largo plazo del Cocodrilo Americano permite distinguir entre las fluctuaciones naturales y los cambios antropogénicos sobre la dinámica poblacional y el estado de conservación.

Reproductive Ecology and Hatchling Growth Rates of the American Crocodile (Crocodylus acutus) on Coiba Island, Panama

Abstract. We assessed the reproductive ecology of the American crocodile (Crocodylus acutus) on Coiba Island, Panama from January–December 2013. We examined nest site characteristics from January–April and hatchling survivorship from April–December. Ten nests were examined at three nesting localities where 30% of the nests were found under forest canopies and 70% were exposed to sunlight (distance to nearest tree = 280 ± 110 cm). Half of the nests were built closer to the sea and the other half closer to bodies of freshwater (700 ± 360 cm). The nest dimensions were 17.5 ± 7.8 cm from the top of the clutch to the surface, 42.9 ± 9.9 cm from the bottom of the clutch to the surface, and 35.9 ± 3.6 cm wide at the top of the nest cavity. The average soil conditions in the nests consistently had high concentrations of potassium (69.3 mL/L) and manganese (9.2 mg/L), moderate concentrations of phosphorus (6.6 mg/L) and iron (3.7 mg/L), and low concentrations of zinc (0.5 mg/L) and copper (0.0 mL/L). Cation exchange capacity showed consistently high concentrations of calcium (2.2 cmol/kg), moderate of magnesium (1.1 cmol/kg), and low in aluminum (0.1 cmol/L). Volumetric water content was about 25.0 ± 2.6% at the bottom and 22.8 ± .3% in the middle of the clutches. Hatching success was 88.9%, of which 68.3% hatched by themselves or with the mothers aid and 20.6% hatched with our aid. Mean size of the mother was 219 ± 6.2 cm total length (TL) and 115.9 ± 3.0 cm snout-vent length (SVL). The incubation period was estimated to be 85–88 days. TL and SVL growth rate of those individuals were 0.03–0.16 cm/day and 0.00–0.09 cm/day, respectively. Population size was estimated to be 218.6 hatchlings in 22.4 km2; the hatchling population declined 65.7% after the first 2 months (May and June) and 95.9% by July, leaving only 0.5% remaining by December. This is the first study to assess nest-site characteristics and estimate hatchling survival in a Pacific population of American crocodiles.

Spatial Ecology of the American Crocodile in a Tropical Pacific Island in Central America

Conservation of large predators has long been a challenge for biologists due to the limited information we have about their ecology, generally low numbers in the wild, large home ranges and the continuous expansion of human settlements. The American crocodile (Crocodylus acutus) is a typical apex predator, that has suffered from all of these characteristic problems, especially the latter one. Humans have had a major impact on the recovery of this species throughout its range, even though most of the countries it inhabits have banned hunting. The last decade has made it clear that in order to implement sound conservation and management programs, we must increase our understanding of crocodile spatial ecology. However, in only two countries where American crocodiles have telemetry studies even been published. Herein we have characterized the spatial ecology of C. acutus on Coiba Island, Panama, by radio-tracking (VHF transmitters) 24 individuals between 2010 and 2013, to determine movement patterns, home range, and habitat use. We have then compared our findings with those of previous studies to develop the most comprehensive assessment of American crocodile spatial ecology to date. Females showed a higher average movement distance (AMD) than males; similarly, adults showed a higher AMD than sub-adults and juveniles. However, males exhibited larger home ranges than females, and concomitantly sub-adults had larger home ranges than juveniles, hatchlings, and adults. There was an obvious relationship between seasonal precipitation and AMD, with increased AMD in the dry and “low-wet” seasons, and reduced AMD during the “true” wet season. We found disaggregate distributions according to age groups throughout the 9 habitat types in the study area; adults and hatchlings inhabited fewer habitat types than juveniles and sub-adults. These sex- and age-group discrepancies in movement and habitat choice are likely due to the influences of reproductive biology and Coiba’s precipitation cycle. Juveniles also showed distinct movement patterns and home ranges; however, with sexual maturation and development, these behaviors became more characteristic of adults and sub-adults. Ours is one of a very small number of studies that will allow future management and conservation planning to be based on the comprehensive integration of the spatial ecology of a Neotropical crocodylian apex predator.

The Biology and Conservation Status of the American Crocodile in Colombia

Abstract Along with the Orinoco and Cuban Crocodiles (Crocodylus intermedius and Crocodylus rhombifer), the American Crocodile (Crocodylus acutus) is among the most threatened crocodylian species in the Americas. Although this species is distributed widely, its IUCN “threat category” in several countries is “of concern.” We review the literature dealing with the biology and conservation status of C. acutus in Colombia and assess its Extent of Occurrence (EoO) and its EoO-Habitat Status. We propose a way to estimate the effective area of optimal habitable occurrence on the basis of the EoO (EoO-Effective Habitable Area). We found that only 37% of the national territories have been sampled in ecological studies during the last decade. The relatively few reports of abundance values and the lack of long-term projects that can produce accurate estimates of population sizes make it difficult to establish the conservation status of this species in Colombia. Our analyses indicate that the EoO-Habitat Status has a higher proportion of disturbed landscapes, as well as agricultural areas, than natural forests and protected areas. We considered that EoO-Effective Habitable Area is a good parameter to estimate the amount of suitable habitat and status of optimal habitat because it does not overestimate the actual areas that crocodiles inhabit. Spatial information gaps, combined with the absence of knowledge about the population ecology and habitat reduction are indeed the principal threats to the American Crocodile in Colombia.