Crocodilian Species Research Articles

Is there a mutation gradient along vertebrate mitochondrial genome mediated by genome replication?

There is a long-held belief that a mutation gradient exists along vertebrate mtDNA, mediated by mitochondrial replication that leaves different parts of the H-strand exposed in single-stranded state for different durations (DssH). However, the predicted mutation gradient and its tests suffer from both conceptual and empirical problems. I assembled representative mammalian, avian and crocodilian mtDNA to test this prediction. I measured substitution rates at codon positions 1 and 2 (S12) and at codon position 3 (S3), as well as synonymous and nonsynonymous substitution rates, and checked their change along the hypothetical gradient. Mammalian species do not support the predicted mutation gradient, although they should according to the model. Crocodilian species exhibit a pattern closest to the prediction, although they should not because their OL, if present, is not at a fixed position. Correlation between S3 and DssH is much weaker than that between S12 and DssH (contrary to the prediction). This is not due to substitution saturation but is instead due to differential gene conservation, e.g., COX1 is far more conserved than ND6 in all metazoans no matter where they are located along mtDNA. In vertebrates, conserved genes such as COX1 happen to have small DssH and variables genes such as ND6 happen to have large DssH. The observed “mutation gradient” is driven by nonsynonymous substitutions, with synonymous substitutions associated with a much weaker “mutation gradient” likely caused by differential codon re-adaptation after nonsynonymous substitutions. The mammalian and avian results are also confirmed by a much larger compilation and analysis of 691 mammalian and 462 avian mtDNAs. The results, however, does not reject paper is not a test of the strand-displacement model (SDM) of mtDNA replication because a mutation gradient is not a necessary consequence of SDM.

The Unlikely Triumph of Dinosaurs.

The article discusses research on the success of dinosaurs. Topics include a study of footprints from the species Prorotodactylus found in Poland, the evolution of dinosaur species, and the habitation of dinosaurs in deserts. The competition of dinosaurs with crocodile species is addressed in relation to the Triassic period. INSET: Family Feud.

Expression of the Sox9, Foxl2, Vasa, and TRPV4 genes in the ovaries and testes of the Morelet's crocodile, Crocodylus moreletii.

The Sox9 gene is important for determining sex in vertebrates, as well as for maintaining testis morphology and fertility during adult life. In the same way, Vasa is an important gene for the maintenance of the germinal lineage and has been highly conserved throughout evolution, as it is expressed in germ cells of both vertebrates and invertebrates. In the particular case of crocodiles, the expression of Sox9 during gonadal morphogenesis and in the testes of 3-month-old Alligator mississippiensis has been studied. However, it is interesting to carry out studies on other species of crocodiles in relation to their particular mechanism for sex determination influenced by temperature. In this work, we investigated the expression of the Sox9, Vasa, Foxl2, and TRPV4 genes in the ovaries and testes of 5-year-old juvenile crocodiles from Crocodylus moreletii. As expected, Sox9 expression was found in males, but surprisingly, it was also found in females. For the first time, the expression of Vasa was reported in spermatogonia, oogonia, and oocytes of 5-year-old crocodiles. Foxl2 is important for the development and maintenance of the ovary during adult life in vertebrates; moreover, Foxl2 protein and transcripts are both highly expressed in the ovaries compared to the testes. A possible upstream regulator of the Sox9 gene in reptiles has not yet been discovered; as such, the expression of the TRPV4 ion channel was evaluated. The TRPV4 ion channel was expressed in the cytoplasm of Sertoli and follicular cells and was therefore proposed as a possible regulator of SOX9.

High Levels of Heavy Metals in Scutes and Eggs of Morelet's Crocodiles (Crocodylus moreletii) from Northeast Mexico

We evaluated the concentrations of three heavy metals (cadmium, chromium, lead) in eggs and caudal scutes of nesting female Crocodylus moreletii in El Carpintero Lagoon, Tamaulipas, Mexico, in the months of May to August 2013. Samples were processed and analyzed based on the method of flame atomic absorption spectrophotometry at the Centro de Investigacion and Tecnologia en Saneamiento Ambiental (CITSA). Our results showed the presence of cadmium, chromium, and lead in both caudal scutes and eggs. The three heavy metals showed values in nesting females of 20.3, 5.0, and 28.0 mg kg-1, respectively. We observed significant differences in levels of cadmium and lead among nests but not with levels of chromium. The variations in heavy metal concentrations in eggs suggest that extrinsic and intrinsic factors simultaneously affect crocodiles during the reproductive season. Our levels were high compared with other species of crocodilians and are above thresholds established by Mexican pollution laws.

Double gene targeting PCR assay for the detection of Crocodylus porosus in commercial products

ABSTRACTThe demand for crocodile meat is quickly growing because of its exotic and organoleptic appeal and also the low content of cholesterol and lipids. Moreover, crocodile oil and blood have been used in alternative medicines for treating asthma and several other ailments since ancient times. Furthermore, crocodile hides have great demand in leather industries. All of these have collectively contributed to the extensive hunting, illegal trading and consequent decline of crocodiles in most parts of the world. To keep space with the growing demands, some crocodile species such as Crocodylus porosus have been raised in farms and its commercial trades have been legalised. However, demand for wild crocodiles in foods and medicines has continued in high gear. Recently, several DNA-based methods have been proposed for crocodile detection, but those assays are based on single gene and longer-sized amplicon targets that break down during extensive processing. To address this gap, here we developed and validated a highly stable double gene targeted multiplex PCR assay for the identification of C. porosus materials in commercial products. The assay involved two short sites from C. porosus atp6 (77 bp) and cytb (127 bp) genes and a universal internal control (99 bp) for eukaryotes. The PCR primers were cross-tested against 18 species and validated under pure and mixed matrices under extensive boiling, autoclaving and microwave cooking conditions. Finally, it was used to identify five crocodile-based commercial products. The lower limits of detection for atp6 and cytb genes were 0.001 ng and 0.01 ng DNA, respectively, in pure meat and 1% under mixed matrices. Some inherent features, such as 77–127 bp amplicon sizes, exceptional stability and superior sensitivity, suggested the assay could be used for the identification of C. porosus in any forensic specimen.

Hematology and serum biochemistry parameters of captive Chinese alligators (Alligator sinensis) during the active and hibernating periods.

The Chinese alligator Alligator sinensis is an endangered freshwater crocodilian species endemic to China. Hematology and serum biochemistry reference range are useful in the assessment and management of animal health condition. In this study, a total of 74 Chinese Alligators (30 males and 44 females) were examined to establish reference range values of hematology and serum biochemistry parameters during the active and hibernating periods. We measured and analyzed 9 hematology and 21 serum biochemistry parameters including 4 serum electrolyte parameters, and described the morphology of different types of blood cells. No statistical differences between the sexes were found for hematology parameter, while significant differences were noted for some serum biochemistry parameters, with males having greater alkaline phosphatase activity level and lower globulin concentration value than females. There were some significant differences between the two different periods with alligators during the active period possessing lower values for mean corpuscular volume, mean corpuscular hemoglobin, total bilirubin and creatine kinase, but higher values for red blood cell and white blood cell counts, monocyte percentage, aspartate aminotransferase, a-amylase, blood urea nitrogen, creatinine, triglycerides, and cholesterol. These baseline data were essential for health condition evaluation and disease diagnosis of this endangered species.

Is There Evidence for Population Regulation in Amphibians and Reptiles?

Abstract From the 1950s to the present, many researchers have tested time series data for density dependence. All kinds of organisms have been studied, from microorganisms to insects and vertebrates to plants. A variety of techniques and population growth models were developed, and the conceptual framework to study populations has been improved. We searched for long time series data on amphibians and reptiles in the literature. From 102 population time series, and after filtering the dataset, we tested for density dependence in time series data for 69 populations (52 species) of amphibians (anurans and caudatans), serpents, lacertilians, chelonians, rhynchocephalians, and crocodilians. We used the exponential growth state-space model and the Ornstein-Uhlembeck state-space model as proxy models for density-independent and density-dependent population growth models, selecting between them with the parametric bootstrap likelihood ratio test. The hypothesis of density independence was rejected for 2 amphibian...

Trophic interactions between larger crocodylians and giant tortoises on Aldabra Atoll, Western Indian Ocean, during the Late Pleistocene.

Today, the UNESCO World Heritage Site of Aldabra Atoll is home to about 100 000 giant tortoises, Aldabrachelys gigantea, whose fossil record goes back to the Late Pleistocene. New Late Pleistocene fossils (age ca. 90–125 000 years) from the atoll revealed some appendicular bones and numerous shell fragments of giant tortoises and cranial and postcranial elements of crocodylians. Several tortoise bones show circular holes, pits and scratch marks that are interpreted as bite marks of crocodylians. The presence of a Late Pleistocene crocodylian species, Aldabrachampsus dilophus, has been known for some time, but the recently found crocodylian remains presented herein are distinctly larger than those previously described. This indicates the presence of at least some larger crocodylians, either of the same or of a different species, on the atoll. These larger crocodylians, likely the apex predators in the Aldabra ecosystem at the time, were well capable of inflicting damage on even very large giant tortoises. We thus propose an extinct predator–prey interaction between crocodylians and giant tortoises during the Late Pleistocene, when both groups were living sympatrically on Aldabra, and we discuss scenarios for the crocodylians directly attacking the tortoises or scavenging on recently deceased animals.

<b>Review and analysis of human and Mugger Crocodile conflict in Gujarat, India from 1960 to 2013</b>

Human-Crocodile conflict (HCC) occurs to varying degrees around the World, and with a number of crocodilian species (CrocBITE 2013). The Mugger or Marsh Crocodile Crocodylus palustris found in Gujarat State is the crocodilian species responsible for conflict with local people. This paper is a compilation of HCC occurring in various parts of Gujarat from 1960 to 2013. A total of 64 crocodile attacks were recorded: 44 (24 fatal & 20 non-fatal) on males, and 20 (9 fatal & 11 non-fatal) on females. By region 52 HCC were recorded in central Gujarat; five in Saurashtra, four in the northern region and three in Kutch; no crocodile attacks were recorded in southern Gujarat. Of the two major river systems in central Gujarat, 41 attacks occurred within the Vishwamitri-Dhadhar River System and 11 in the Narmada system. Most crocodile attacks happened between the months of April and September, peaking in May with 14 attacks. These months are the peak breeding season for the species in Gujarat. The most obvious contributors to HCC are lack of basic facilities in rural areas, poverty, illiteracy and the presence of adult animals close to human settlements and activities. Other contributing factors are lack of preventive measures by the forest department, absence of protocols for mugger crocodile rescue, and haphazard release of problematic animals.

Metallic elements in Nile Crocodile eggs from the Kruger National Park, South Africa

The Nile Crocodile is the largest predator on the African continent. Recent mass mortalities in the Kruger National Park (KNP) raised concerns about possible influence of pollution. We analysed eggs and their eggshells collected from nests inside the KNP and from a crocodile farm for metallic elements. We found that mercury, selenium, and copper occurred at levels of concern. Eggshells had very high concentrations of iron. Apart from toxicological implications associated with elevated concentrations in eggs, we found iron possibly contributing towards thicker eggshells. Thicker shells may act as a barrier to gas and water exchange, as well as possibly increasing the effort required for the hatchling to emerge from tightly packed shells under sand. Pollutants are transported into the KNP via rivers, and possibly via air. Mercury and copper pollution are waste-, industrial- and mining-related; ecotoxicological concern should therefore be extended to all areas where the four African crocodile species occur. Reptiles are under-represented in ecotoxicological literature in general, and especially from Africa. We know of only one previous report on metals and metalloids in crocodile eggs from Africa (Zimbabwe), published 30 years ago. Reduced fitness, endocrine disruption and effects on behaviour are other possible sub-lethal effects associated with metallic elements that may only become apparent decades later in a long-lived species such as the Nile Crocodile. In the face of habitat destruction, pollution, human population increases, and climate change, further research is needed regarding pollutant concentrations and effects in all African reptiles. The rivers that carry water from outside the park sustain its aquatic life, but also transport pollutants into the KNP. Therefore, improved source mitigation remains an important task and responsibility for all involved.

Delimitation of evolutionary units in Cuvier’s dwarf caiman, Paleosuchus palpebrosus (Cuvier, 1807): insights from conservation of a broadly distributed species

An important goal of evolutionary and conservation biology is the identification of units below the species level, such as Evolutionarily Significant Units (ESUs), providing objectively delimited units for species conservation and management. In this study we tested the hypothesis that Cuvier’s dwarf caiman (Paleosuchus palpebrosus)—a species broadly distributed across several biomes and watersheds of South America—is comprised of different ESUs. We analyzed mitochondrial cytochrome b sequences of 206 individuals and 532 unlinked ddRAD loci of 20 individuals chosen from amongst the mitochondrial haplogroups. Analysis of the cytochrome b sequences revealed four mitochondrial clusters, while STRUCTURE analysis of ddRAD loci detected three genomic clusters with different levels of mixture between them. Using the Adaptive Evolutionary Conservation (AEC) framework we identified three ESUs: “Amazon”, “Madeira-Bolivia” and “Pantanal”; one of them composed of two different Management Units (MUs), “Madeira” and “Bolivia”. In general, based on the comparisons with other crocodilian species, genetic diversity of each lineage was moderate however, the Madeira MU showed fivefold lower genetic diversity than other geographic groups. Considering the particularities of each Paleosuchus palpebrosus conservation unit, we recommend that the conservation status of each is evaluated separately. Tropical biodiversity is largely underestimated and in this context the broadly distributed species are the most likely candidates to harbor distinct evolutionary lineages. Thus, we suggest that conservation research should not neglect species that are generally considered of Least Concern by IUCN due to the taxon’s broad geographic distribution.

Variation in crocodilian dorsal scute organization and geometry with a discussion of possible functional implications.

Dermal ossifications, including osteoderms, are present in many vertebrates and are frequently interpreted as a defense against predators. Nevertheless, osteoderms remain ubiquitous in adult crocodilians while being absent in hatchlings, even though adults rarely experience predation. In other biological systems, increased variation, particularly fluctuating asymmetry, have proven useful for identifying biological structures likely to have evolved under relaxed selection, which in turn may inform their function. Therefore, using the keratinous scutes as proxies for the underlying osteoderm morphology, I investigated the average intraspecific variability of geometry and fluctuating asymmetry in dorsal scutes in five species of crocodilians. I first tested for differences in variability of scute length and width, then for differences in bilateral fluctuating asymmetry of scute number, before finally investigating scute distribution patterns for each species compared to hypothetical rectangular and hexagonal scute arrangements. The American crocodile, Crocodylus acutus, shows significantly more asymmetry than other species, which is consistent with relaxed selection on osteoderms in this species. A suspected decrease in intraspecific aggression within Crocodylus acutus, in conjunction with the inferred relaxed selection, suggests that, in general, crocodilian osteoderms function primarily as defensive armor in aggressive encounters with conspecifics. The smooth-fronted caiman, Paleosuchus trigonatus, exhibits increased variation in scute dimensions linked to the mediolateral offset of osteoderms in adjacent rows, possibly resulting in a more rigid carapace. Unfortunately, comparative data on crocodilian behavior, physiology, and development is extremely limited and restricts the ability to explore other potential explanations for the patterns observed, highlighting the need for more research on rare and cryptic crocodylians.

Impact of poxvirus lesions on saltwater crocodile (Crocodylus porosus) skins

Cutaneous poxvirus infections are common in several crocodilian species and are of importance in crocodile farming due to their potential impact on the tanned hide. To confirm poxvirus infection and understand the impact on saltwater crocodile (Crocodylus porosus) skin, fourteen animals from different age groups (five hatchlings, five yearlings and four grow-outs) were selected based on a criterion of ten poxvirus-like lesions per animal. One lesion on each animal was extruded for genetic analysis and transmission electron microscopy. Both methods confirmed poxvirus so the remainder of lesions were re-examined every six weeks over a 24 week study period. Each lesion went through four distinct phases: early active, active, expulsion and healing. To understand how these lesions impact on the final skin product, one crocodile from each age group was euthanised and the lesions examined. Using standard skin grading techniques (light-table), the early phase (early active – expulsion) lesions were all translucent and would lead to downgrading of the skin or, at worst, rendering them unsaleable. At the later stages of healing, the translucency reduces. Histological examination of the phases confirm that the basement membrane is not breached by the infection further indicating that poxvirus lesions, given enough time, will eventually have no detrimental effect on skin quality. This is obviously dependent upon no more lesions developing in the interim.

Differential response to disturbance factors for the population of sympatric crocodilians (Gavialis gangeticus and Crocodylus palustris) in Katarniaghat Wildlife Sanctuary, India

Abstract Katarniaghat Wildlife Sanctuary along the Indo‐Nepal border in India harbours a well‐known breeding gharial (Gavialis gangeticus) population in its global distribution range together with a substantial population of breeding muggers (Crocodylus palustris). However, no systematic information on size and structure is available for either of the species' populations in the protected area. This study was undertaken in winter and the ensuing summer of 2010–2011 to estimate the relative population density and structure of the two crocodilian species and to examine the effect of various disturbance factors, with the main focus on the effects of tourism and illegal fishing on the behavioural attributes of the two species. Five daytime surveys were conducted from December 2010 to April 2011. Relative density based on encounter rate (number per 20 km) was highest for gharial juveniles followed by gharial adults and gharial sub‐adults in all the five surveys. Muggers, on the other hand, showed the opposite trend, with the lowest encounter rate being for juveniles, followed by sub‐adults and adults. The encounter rate of both species declined with increase in the mean ambient temperature from December to April. The river habitat was divided into 2 km segments and disturbance factors were recorded at intervals of 100 m on both river banks. Wariness was taken as an indicator of response to disturbance caused by human beings and was measured from mechanized boats used for tourism and non‐mechanized boats used for illegal fishing. Segments with sandbars, in spite of moderate to high disturbance rate, were preferred for basking by both species. In conclusion, with suitable habitat for basking, gharials and muggers were observed to tolerate moderate levels of disturbance. Wariness resulting from disturbances from the non‐mechanized boats was higher than that from the mechanized boats. Gharials tend to avoid humans, possibly with age/size acquired experience and knowledge, whereas muggers become more tolerant to human presence with increased age/size. Despite many disturbance factors, the crocodilian population in Katarniaghat Wildlife Sanctuary is doing relatively well compared with populations in other habitats in India. With the involvement of local stakeholders and strict implementation of forest laws, the habitat can be further improved and a healthy gharial population can be ensured.

Trackways of the American Crocodile (Crocodylus acutus) in Northwestern Costa Rica: Implications for Crocodylian Ichnology

ABSTRACTWe documented trackways of free-living Crocodylus acutus on beaches at the mouths of Tamarindo and Ventanas estuaries, Costa Rica. Our crocodiles had estimated total lengths of 1–3 meters or more. Manus prints have five digits, with digits I–III bearing claw marks. Pes prints have four digits, with claw marks on digits I–III. The pes is plantigrade. Claws generally dig into the substrate. Apart from claw marks, digit I and the heel of the pes are usually the most deeply impressed parts of footprints. Trackways are wide-gauge. Pes prints are usually positioned just behind ipsilateral manus prints of the same set and may overlap them. Manus and pes prints angle slightly outward with respect to the crocodile's direction of movement. Claw-bearing digits of both the manus and pes may create curved, concave-toward-the-midline drag marks as the autopodium is protracted. The tail mark varies in depth and clarity, and in shape from nearly linear to markedly sinuous. Sometimes the tail mark hugs the trackway midline, but sometimes it is closer to, or even cuts across, prints of one side. American crocodile footprints and trackways are similar to those observed in other extant crocodylian species, indicating substantial trackway conservatism across the group.

The biochronology and palaeobiogeography of Baru (Crocodylia: Mekosuchinae) based on new specimens from the Northern Territory and Queensland, Australia.

New records of the Oligo–Miocene mekosuchine crocodylian, Baru, from Queensland and the Northern Territory are described. Baru wickeni and Baru darrowi are accepted as valid species in the genus and their diagnoses are revised. Both species are present in Queensland and the Northern Territory but are restricted in time, with B. wickeni known from the late Oligocene and B. darrowi from the middle Miocene. The broad geographic distributions and restricted time spans of these species indicate that this genus is useful for biochronology. The record of B. wickeni from the Pwerte Marnte Marnte Local Fauna in the Northern Territory establishes that the species inhabited the north-western margin of the Lake Eyre Basin (LEB) drainage system. More southerly Oligo–Miocene sites in the LEB contain only one crocodylian species, Australosuchus clarkae. The Pwerte Marnte Marnte occurrence of B. wickeni indicates that the separation of Baru and Australosuchus did not correspond with the boundaries of drainage basins and that palaeolatitude was a more likely segregating factor.

The metabolic cost of nesting: body condition and blood parameters of Caiman crocodilus and Melanosuchus niger in Central Amazonia

Although nesting ecology is well studied in several crocodilian species, it is not known how nest attendance influences physiology and body condition of nesting females. In this study, we describe body condition and serum biochemical values of nesting female, non-nesting female and male spectacled caiman (Caiman crocodilus) and black caiman (Melanosuchus niger) in two areas of Central Amazonia. We also evaluated the effect of nest age and nest distance to water on body condition and blood parameters of nesting females. Body condition and plasmatic concentrations of glucose, triglycerides, lactate and uric acid of nesting females were significantly different from those of non-nesting females and males in C. crocodilus, but not in M. niger. Our study also demonstrated that nest age and distance to water had a negative effect on female body condition in C. crocodilus, but not in M. niger. Female C. crocodilus attending older nests or nests built further away from permanent water bodies tended to have lower body condition. Our results demonstrate that the nesting strategy of C. crocodilus has a metabolic cost associated with nest attendance for nesting females, which appear to depend on accumulated energetic reserves during nest attendance. In contrast, nest attendance had little effect on the physiology of female M. niger.

Opportunistic top predators partition food resources in a tropical freshwater ecosystem

Abstract The structure of food webs may be strongly influenced by the distribution of top predators in space and time. The Amazon biome is the only region in the world where four alligatorid species are known to occur in sympatry, and they attain high densities in some regions. As top predators with a diverse range of prey species occupying different trophic levels, their impact upon food webs should be substantial, but the degree to which crocodilians differ in their food sources, and potentially avoid competitive exclusion where they occur syntopically is not well understood. Although most crocodilians are considered generalist opportunistic predators that feed on any source of protein available in the environment, Amazonian crocodilians show broad differences in the proportions of prey items they consume. It is believed that these differences may in part reflect habitat use, but it is unknown to what extent they represent interspecific differences in prey preferences or are a direct function of habitat selection. Stable carbon isotope data (δ13C) of crocodilians and their potential prey were used to assess differences in reliance on terrestrial versus aquatic resources. These data were then placed in a spatial context using classified maps that reflect habitat types (headwater streams, mid‐order flooded‐forest streams and várzea floodplains) to elucidate whether dietary differences are explained by habitat selection or are more likely a reflection of prey preferences. We found evidence for differences in types of basal resources supporting these crocodilians. Mean δ13C values were highest in Paleosuchus trigonatus (Schneider's dwarf caiman, −25.7 ± 1.2‰), intermediate in Caiman crocodilus (spectacled caiman, −27.4 ± 1.2‰) and Paleosuchus palpebrosus (Cuvier's dwarf caiman, −27.7 ± 1.1‰) and lowest in Melanosuchus niger (black caiman, −29.9 ± 1.3‰). A progressive decrease in δ13C values of crocodilian tissues occurred from headwaters to floodplains, which most likely reflects a progressive increase in autochthonous over allochthonous inputs in lower reaches of streams. The shift from terrestrial to aquatic resources sustaining these sympatric predators mirrors their spatial distribution along this ecotone. However, after taking into account the habitat in which pairs of syntopic individuals of distinct species occurred, significant differences in δ13C values suggest that P. trigonatus and P. palpebrosus have different prey bases. Thus, despite being opportunistic predators, our results show that differences in crocodilian diets likely result from prey preferences and not only from habitat selection. These findings suggest that some species of crocodilians may be less generalist than traditionally thought and their influence on terrestrial or aquatic food webs might be species specific.

Assessment of trophic ecomorphology in non-alligatoroid crocodylians and its adaptive and taxonomic implications.

Although the establishment of trophic ecomorphology in living crocodylians can contribute to estimating feeding habits of extinct large aquatic reptiles, assessment of ecomorphological traits other than the snout shape has scarcely been conducted in crocodylians. Here, I tested the validity of the proposed trophic ecomorphological traits in crocodylians by examining the correlation between those traits and the snout shape (an established trophic ecomorphology), using 10 non-alligatoroid crocodylian species with a wide range of snout shape. I then compared the ontogenetic scaling of trophic ecomorphology to discuss its adaptive and taxonomic significance. The results demonstrated that degree of heterodonty, tooth spacing, size of supratemporal fenestra (STF), ventral extension of pterygoid flange and length of lower jaw symphysis are significantly correlated with snout shape by both non-phylogenetic and phylogenetic regression analyses. Gavialis gangeticus falls outside of 95% prediction intervals for the relationships of some traits and the snout shape, suggesting that piscivorous specialization involves the deviation from the typical transformation axis of skull characters. The comparative snout shape ontogeny revealed a universal trend of snout widening through growth in the sampled crocodylians, implying the existence of a shared size-dependent biomechanical constraint in non-alligatoroid crocodylians. Growth patterns of other traits indicated that G.gangeticus shows atypical trends for degree of heterodonty, size of STF, and symphysis length, whereas the same trends are shared for tooth spacing and ventral extension of pterygoid flange among non-alligatoroid crocodylians. These suggest that some characters are ontogenetically labile in response to prey preference shifts through growth, but other characters are in keeping with the conserved biomechanics among non-alligatoroid crocodylians. Some important taxonomic characters such as the occlusal pattern are likely correlated with ontogeny and trophic ecomorphology rather than are constrained by phylogenetic relationships, and careful reassessment of such characters might be necessary for better reconstructing the morphological phylogeny of crocodylians.

Crocodilian perivitelline membrane-bound sperm detection.

Advanced reproductive technologies (ART's) are often employed with various taxa to enhance captive breeding programs and maintain genetic diversity. Perivitelline membrane-bound (PVM-bound) sperm detection has previously been demonstrated in avian and chelonian species as a useful technique for breeding management. In the absence of embryotic development within an egg, this technique can detect the presence of sperm trapped on the oocyte membrane confirming breeding, male reproductive status, and pair compatibility. PVM-bound sperm were successfully detected in three clutches of Cuban crocodile (Crocodylus rhombifer) eggs at the Smithsonian's National Zoological Park (NZP) for the first time in any crocodilian species. PVM-bound sperm were detected in fresh and incubated C. rhombifer eggs, as well as eggs that were developing (banded) and those that were not (not banded). The results of this study showed significant differences in average sperm densities per egg between clutches (p = 0.001). Additionally, there was not a significant difference within clutches between eggs that banded and those that did not band (Clutch A, p = 0.505; Clutch B, p = 0.665; Clutch C, p = 0.266). The results of this study demonstrate the necessity to microscopically examine eggs that do not develop (do not band), to determine if sperm is present, which can help animal managers problem solve reproductive shortcomings. PVM-bound sperm detection could be a useful technique in assessing crocodilian breeding programs, as well as have potential uses in studies assessing sperm storage, artificial insemination, and artificial incubation.