Phylogenomics and historical biogeography of West Indian Rock Iguanas (genus Cyclura).

The genus Cyclura includes nine extant species and six subspecies of West Indian Rock Iguanas and is one of the most imperiled genera of squamate reptiles globally. An understanding of species diversity, evolutionary relationships, diversification, and historical biogeography in this group is crucial for implementing sound long-term conservation strategies. We collected DNA samples from 1 to 10 individuals per taxon from all Cyclura taxa (n = 70 ingroup individuals), focusing where possible on incorporating individuals from different populations of each species. We also collected 1-2 individuals from each of seven outgroup species of iguanas (Iguana delicatissima; five Ctenosaura species) and Anolis sagrei (n = 12 total outgroup individuals). We used targeted genomic sequence capture to isolate and to sequence 1,872 loci comprising of 687,308 base pairs (bp) from each of the 82 individuals from across the nuclear genome. We extracted mitochondrial reads and assembled and annotated mitogenomes for all Cyclura taxa plus outgroup species. We present well-supported phylogenomic gene tree/species tree analyses for all extant species of Cyclura using ASTRAL-III, SVDQuartets, and StarBEAST2 methods, and discuss the taxonomic, biogeographic, and conservation implications of these data. We find a most recent common ancestor of the genus 9.91 million years ago. The earliest divergence within Cyclura separates C. pinguis from a clade comprising all other Cyclura. Within the latter group, a clade comprising C. carinata from the southern Lucayan Islands and C. ricordii from Hispaniola is the sister taxon to a clade comprising the other Cyclura. Among the other Cyclura, the species C. cornuta and C. stejnegeri (from Hispaniola and Isla Mona) form the sister taxon to a clade of species from Jamaica (C. collei), Cuba and Cayman Islands (C. nubila and C. lewisi), and the eastern (C. rileyi) and western (C. cychlura) Lucayan Islands. Cyclura cychlura and C. rileyi form a clade whose sister taxa are C. nubila and C. lewisi. Cyclura collei is the sister taxon to these four species combined.

Heterozygosity-Fitness Correlations Reveal Inbreeding Depression in Neonatal Body Size in a Critically Endangered Rock Iguana.

Inbreeding depression, though challenging to identify in nature, may play an important role in regulating the dynamics of small and isolated populations. Conversely, greater expression of genetic load can enhance opportunities for natural selection. Conditional expression concentrates these opportunities for selection and may lead to failure of detection. This study investigates the possibility for age-dependent expression of inbreeding depression in a critically endangered population of rock iguanas, Cyclura nubila caymanensis. We employ heterozygote-fitness correlations to examine the contributions of individual genetic factors to body size, a fitness-related trait. Nonsignificant reductions in homozygosity (up to 7%) were detected between neonates and individuals surviving past their first year, which may reflect natural absorption of inbreeding effects by this small, fecund population. The majority of variation in neonate body size was attributed to maternal or environmental effects (i.e., clutch identity and incubation length); however, heterozygosity across 22 microsatellite loci also contributed significantly and positively to model predictions. Conversely, effects of heterozygosity on fitness were not detectable when adults were examined, suggesting that inbreeding depression in body size may be age dependent in this taxon. Overall, these findings emphasize the importance of taking holistic, cross-generational approaches to genetic monitoring of endangered populations.

Phylogeography of the endangered Lesser Antillean iguana, Iguana delicatissima: a recent diaspora in an archipelago known for ancient herpetological endemism.

Iguana delicatissima is an endangered endemic of the Lesser Antilles in the Caribbean. Phylogeographic analyses for many terrestrial vertebrate species in the Caribbean, particularly lizards, suggest ancient divergence times. Often, the closest relatives of species are found on the same island, indicating that colonization rates are so low that speciation on islands is often more likely to generate biodiversity than subsequent colonization events. Mitochondrial sequence analysis of the region spanning ND4 was performed on I. delicatissima individuals from islands across the species' range to estimate genetic divergence among geographically isolated populations. Five unique haplotypes were recovered from 46 individuals. The majority of animals carry a single common haplotype. Two of the haplotypes were only present in individuals classified as hybrids from Îles des Saintes. The final 2 haplotypes, single nucleotide substitutions, were present in animals from Îlet Chancel of Martinique and Saint Barthélemy, respectively. Despite the great distances between islands and habitat heterogeneity within islands, this species is characterized by low haplotype diversity. The low mtDNA variation of I. delicatissima suggests a single colonization coupled with rapid range expansion, potentially hastened by human-mediated dispersal.

Hand grab or noose pole? Evaluating the least stressful practice for capture of endangered Turks and Caicos Rock Iguanas Cyclura carinata.

As the analysis of blood metabolites has become more readily accessible thanks to the use of point-of-care analyzers, it is now possible to evaluate stress level of wild animals directly in the field. Lactate is receiving much attention as a good stress level proxy in individuals subjected to capture, manual restraint, and data sampling in the wild, and appropriate protocols to maintain lactate values low should be preferred. In this study we compare how two different capture methodologies, hand grab vs. noose pole, affect the variation of blood lactate values in Cyclura carinata iguanas when captured for sampling. We used blood lactate concentration, measured immediately upon- and 15 min after-capture, as a proxy for stress level. While the primary goal of this work is to determine the least stressful capture methodology to be favored when sampling this and other wild iguanas, we also evaluated additional baseline physiological parameters relevant to the health and disease monitoring for this species. Our results show that while initial lactate values level-out in sampled individuals after 15 min in captivity, regardless of the capture methodology, rock iguanas captured by noose pole showed significantly higher lactate concentration and increased heartbeat rate immediately after capture. While the overall health evaluation determined that all analyzed individuals were in good health, based on our results we recommend that, when possible, hand capture should be preferred over noose pole when sampling wild individuals.

Locating Nesting Sites for Critically Endangered Galápagos Pink Land Iguanas (Conolophus marthae).

Invasive alien species control is recognized worldwide as a priority action to preserve global biodiversity. However, a lack of general life history knowledge for threatened species can impede the effectiveness of conservation actions. Galápagos pink land iguanas (Conolophus marthae) are endemic to Wolf Volcano, Galápagos, Ecuador. These iguanas are threatened by invasive alien species, particularly feral cats, that may affect their small population size. To guarantee the long-term survival of C. marthae, the Galápagos National Park Directorate is considering, along with an ongoing campaign of feral cat control, the implementation of a head-start program. However, the success of this management strategy necessarily relies on the identification of pink iguana nesting grounds, which were still unknown at the onset of this study. We modeled the movement patterns of male and female iguanas during the reproductive season, using location data collected from custom-made remote tracking devices installed on adult pink iguanas in April 2021. We first calculated for each individual the vector of distances from its starting location, which was defined as net displacement. We then used net displacement as the response variable in a generalized additive mixed model with day of the year as the predictor. Based on the hypothesis that males and females may behaviorally differ after mating, we looked for female-specific migratory behavior suggesting females were moving toward nesting areas. The results obtained confirmed our hypothesis, as females exhibited a distinct migratory behavior, reaching a small plateau area inside of Wolf Volcano's caldera and ca. 400 m below the volcano's northern rim. Moreover, once inside the caldera, females displayed a more aggregated distribution pattern. The movement data obtained allowed Galápagos National Park rangers to locate individual pink iguana nests and subsequently to sight and collect the first observed hatchlings of the species. This work constitutes a necessary baseline to perform dedicated studies of pink iguana nests and emerging hatchling iguanas, which is an essential step toward the development of an effective head-start program.

Conservation of a flagship species: Health assessment of the pink land iguana, Conolophus marthae.

The pink land iguana, Conolophus marthae, is one of four species of iguanas (three terrestrial and one marine) in the Galápagos Islands, and the only one listed as critically endangered by the IUCN. The species can only be found on the north-west slopes of the highest volcano on Isabela Island and was first described to science in 2009. As part of a population telemetry study, a health assessment was authorized by the Galápagos National Park. Wild adult iguanas were captured on Wolf Volcano in September 2019 and April 2021 to record morphological and physiological parameters including body temperature, heart rate, intraocular pressures, tear formation, and infrared iris images. Blood samples were also collected and analyzed. An i-STAT portable blood analyzer was used to obtain values for base excess in the extracellular fluid compartment (BEecf), glucose (Glu), hematocrit (HctPCV), hemoglobin (Hb), ionized calcium (iCa), partial pressure of carbon dioxide (pCO2), partial pressure of oxygen (pO2), percent oxygen saturation (sO2%), pH, potassium (K), and sodium (Na). When possible, data were compared to previously published and available data for the other Galápagos iguanas. The results reported here provide baseline values that will be useful in detecting changes in health status among pink land iguanas affected by climate change, invasive species, anthropogenic threats, or natural disturbances. The collected data also provide an invaluable resource for conservation scientists planning to implement conservation strategies, like translocations, that may temporarily alter these baseline values.

Conditional female strategies influence hatching success in a communally nesting iguana.

The decision of females to nest communally has important consequences for reproductive success. While often associated with reduced energetic expenditure, conspecific aggregations also expose females and offspring to conspecific aggression, exploitation, and infanticide. Intrasexual competition pressures are expected to favor the evolution of conditional strategies, which could be based on simple decision rules (i.e., availability of nesting sites and synchronicity with conspecifics) or on a focal individual's condition or status (i.e., body size). Oviparous reptiles that reproduce seasonally and provide limited to no postnatal care provide ideal systems for disentangling social factors that influence different female reproductive tactics from those present in offspring-rearing environments. In this study, we investigated whether nesting strategies in a West Indian rock iguana, Cyclura nubila caymanensis, vary conditionally with reproductive timing or body size, and evaluated consequences for nesting success. Nesting surveys were conducted on Little Cayman, Cayman Islands, British West Indies for four consecutive years. Use of high-density nesting sites was increasingly favored up to seasonal nesting activity peaks, after which nesting was generally restricted to low-density nesting areas. Although larger females were not more likely than smaller females to nest in high-density areas, larger females nested earlier and gained access to priority oviposition sites. Smaller females constructed nests later in the season, apparently foregoing investment in extended nest defense. Late-season nests were also constructed at shallower depths and exhibited shorter incubation periods. While nest depth and incubation length had significant effects on reproductive outcomes, so did local nest densities. Higher densities were associated with significant declines in hatching success, with up to 20% of egg-filled nests experiencing later intrusion by a conspecific. Despite these risks, nests in high-density areas were significantly more successful than elsewhere due to the benefits of greater chamber depths and longer incubation times. These results imply that communal nest sites convey honest signals of habitat quality, but that gaining and defending priority oviposition sites requires competitive ability.

Chemical signatures of femoral pore secretions in two syntopic but reproductively isolated species of Galápagos land iguanas (Conolophus marthae and C. subcristatus).

The only known population of Conolophus marthae (Reptilia, Iguanidae) and a population of C. subcristatus are syntopic on Wolf Volcano (Isabela Island, Galápagos). No gene flow occurs suggesting that effective reproductive isolating mechanisms exist between these two species. Chemical signature of femoral pore secretions is important for intra- and inter-specific chemical communication in squamates. As a first step towards testing the hypothesis that chemical signals could mediate reproductive isolation between C. marthae and C. subcristatus, we compared the chemical profiles of femoral gland exudate from adults caught on Wolf Volcano. We compared data from three different years and focused on two years in particular when femoral gland exudate was collected from adults during the reproductive season. Samples were processed using Gas Chromatography coupled with Mass Spectrometry (GC-MS). We identified over 100 different chemical compounds. Non-Metric Multidimensional Scaling (nMDS) was used to graphically represent the similarity among individuals based on their chemical profiles. Results from non-parametric statistical tests indicate that the separation between the two species is significant, suggesting that the chemical profile signatures of the two species may help prevent hybridization between C. marthae and C. subcristatus. Further investigation is needed to better resolve environmental influence and temporal reproductive patterns in determining the variation of biochemical profiles in both species.

Genetic structure at three spatial scales is consistent with limited philopatry in Ricord's Rock Iguanas (Cyclura ricordii).

Cyclura ricordii is an endemic iguana from Hispaniola Island and is threatened on the IUCN Red List. The main threats are predation by introduced mammals, habitat destruction, and hunting pressure. The present study focused on two nesting sites from Pedernales Province in the Dominican Republic. The hypothesis that natal philopatry influences dispersal and nest-site selection was tested. Monitoring and sampling took place in 2012 and 2013. Polymorphic markers were used to evaluate whether natal philopatry limits dispersal at multiple spatial scales. Ripley's K revealed that nests were significantly clustered at multiple scales, when both nesting sites were considered and within each nesting site. This suggests a patchy, nonrandom distribution of nests within nest sites. Hierarchical AMOVA revealed that nest-site aggregations did not explain a significant portion of genetic variation within nesting sites. However, a small but positive correlation between geographic and genetic distance was detected using a Mantel's test. Hence, the relationship between geographic distance and genetic distance among hatchlings within nest sites, while detectable, was not strong enough to have a marked effect on fine-scale genetic structure. Spatial and genetic data combined determined that the nesting sites included nesting females from multiple locations, and the hypothesis of "natal philopatry" was not supported because females nesting in the same cluster were no more closely related to each other than to other females from the same nesting site. These findings imply that nesting aggregations are more likely associated with cryptic habitat variables contributing to optimal nesting conditions.

Isolation and characterization of 23 polymorphic microsatellite loci for a West Indian iguana (Cyclura pinguis) from the British Virgin Islands.

Twenty-three polymorphic microsatellite markers were identified and characterized for Cyclura pinguis, a critically endangered species of lizard (Sauria: Iguanidae) native to Anegada Island in the British Virgin Islands. We examined variation at these loci for 39 C. pinguis, finding up to five alleles per locus and an average expected heterozygosity of 0.55. Allele frequency estimates for these microsatellite loci will be used to characterize genetic diversity of captive and wild C. pinguis populations and to estimate relatedness among adult iguanas at the San Diego Zoo that form the nucleus of a captive breeding programme for this critically endangered species.