Chelonian Species Research Articles

Lack of detection of West Nile virus in an islander population of chelonians during a West Nile virus outbreak.

In 2011, several outbreaks of West Nile disease occurred in Sardinia (Italy). The region hosts several chelonian species. Because of the increasing concern on the potential role that ectotherms may play in the ecology of West Nile virus (WNV), in October 2011 blood samples were collected from 41 endemic Sardinian chelonians and tested for the presence of active WNV infection or neutralizing antibodies by real time polymerase chain reaction (RT-PCR) and serumneutralisation, respectively. Neither WNV neutralising antibodies (0%; 95% CI: 0‑8.4%) nor WNV RNA (0%; 95% CI: 0‑6.8%) were found in the tested samples. According to the results of this screening survey, it is unlikely that chelonians are involved in the epidemiology of the 2011 WNV outbreaks in Sardinia.

Ecological opportunities, habitat, and past climatic fluctuations influenced the diversification of modern turtles

Habitat may be viewed as an important life history component potentially related to diversification patterns. However, differences in diversification rates between aquatic and terrestrial realms are still poorly explored. Testudines is a group distributed worldwide that lives in aquatic and terrestrial environments, but until now no-one has evaluated the diversification history of the group as a whole. We aim here to investigate the diversification history of turtles and to test if habitat influenced speciation rate in these animals. We reconstructed the phylogeny of the modern species of chelonians and estimated node divergence dates using molecular markers and a Bayesian approach. Then, we used Bayesian Analyses of Macroevolutionary Mixtures to evaluate the diversification history of turtles and evaluate the effect of habitat on this pattern. Our reconstructed phylogeny covered 300 species (87% of the total diversity of the group). We found that the emydid subfamily Deirochelyinae, which forms the turtle hotspot in south-eastern United States, had an increase in its speciation rate, and that Galapagos tortoises had similar increases. Current speciation rates are lower in terrestrial turtles, contradicting studies supporting the idea terrestrial animals diversify more than aquatic species. Our results suggest that habitat, ecological opportunities, island invasions, and climatic factors are important drivers of diversification in modern turtles and reinforce the importance of habitat as a diversification driver.

Chelonian perivitelline membrane-bound sperm detection: A new breeding management tool

Perivitelline membrane (PVM)-bound sperm detection has recently been incorporated into avian breeding programs to assess egg fertility, confirm successful copulation, and to evaluate male reproductive status and pair compatibility. Due to the similarities between avian and chelonian egg structure and development, and because fertility determination in chelonian eggs lacking embryonic growth is equally challenging, PVM-bound sperm detection may also be a promising tool for the reproductive management of turtles and tortoises. This study is the first to successfully demonstrate the use of PVM-bound sperm detection in chelonian eggs. Recovered membranes were stained with Hoechst 33342 and examined for sperm presence using fluorescence microscopy. Sperm were positively identified for up to 206 days post-oviposition, following storage, diapause, and/or incubation, in 52 opportunistically collected eggs representing 12 species. However, advanced microbial infection frequently hindered the ability to detect membrane-bound sperm. Fertile Centrochelys sulcata, Manouria emys, and Stigmochelys pardalis eggs were used to evaluate the impact of incubation and storage on the ability to detect sperm. Storage at -20°C or in formalin were found to be the best methods for egg preservation prior to sperm detection. Additionally, sperm-derived mtDNA was isolated and PCR amplified from Astrochelys radiata, C. sulcata, and S. pardalis eggs. PVM-bound sperm detection has the potential to substantially improve studies of artificial incubation and sperm storage, and could be used to evaluate the success of artificial insemination in chelonian species. Mitochondrial DNA from PVM-bound sperm has applications for parentage analysis, the study of sperm competition, and potentially species identification.

An overview of same-sex mounting in turtles and tortoises

Same-sex mounting is an aspect of animal behavior that has received increased attention in recent years in an attempt to improve our limited understanding of the possible causal mechanisms. Here, to our knowledge, we review for the first time same-sex mounting in turtles and tortoises. To this end, we have compiled data on same-sex mounts in 13 chelonian species and discuss the data together with those hypotheses most commonly raised in turtle studies, namely, the intrasexual conflict, maladaptation, mistaken identify, and opposite-sex deprivation hypotheses. Compilation of the data revealed that in almost every species with reports of same-sex mounting there were dominance relationships mediated by aggressive/submissive interactions; most of these cases occurred in captivity, and the sex ratios were not skewed. We discuss future research directions, providing initial ideas of experimental testing on each hypothesis, in the hope of directing more research effort to this field.

Testing the efficiency of protected areas in the Amazon for conserving freshwater turtles

AbstractAimWe used chelonian distribution data to (1) predict suitable areas of the occurrence for freshwater turtle species using species distribution models and (2) evaluate whether these species are protected by the current network of protected areas (PAs).LocationThe Brazilian Amazon.MethodsWe generated predictions of suitable areas for chelonian occurrence based onBIOCLIM,SVM,GLMand maximum entropy modelling procedures. We used maximum entropy to run the gap analysis and compared the effectiveness of three kinds of protected areas with different levels of protection: (1) integral protection areas (IPA) only; (2) integral protection areas + sustainable use areas (IPA+SUA); and (3) integral protection areas + sustainable use areas + indigenous lands (IPA + SUA + IL).ResultsWe identified only one full gap species,Mesoclemmys nasuta, whose distribution is not included in anyPAs. Other chelonian species have at least a portion of their distribution included inPAs. Some protected species and partial gap species occur in areas with high rates of deforestation. ConsideringPAs with the highest level of protection (IPA), onlyRhinoclemmys punctulariaandKinosternon scorpioidesachieve their conservation targets. In theIPA + SUAscenario, conservation targets of some species with small range sizes are not achieved. When allPAtypes were considered (IPA + SUA + IL), only two species fail to achieve their conservation targets,Acanthochelys macrocephalaandM. nasuta.Main conclusionsDespite the large number ofPAs in the Brazilian Amazon,IPAs alone are not sufficient for capturing suitable areas for freshwater turtles. The inclusion ofSUAandILis crucial for achieving coverage targets for most species. However, chelonians may be overharvested inSUAs andILs, due to their importance as a food resource. Areas that have high turtle richness next to existingPAs and the needs of traditional cultures should be considered in management planning for freshwater turtles.

Threatening levels and extinction risks based on distributional, ecological and life-history datasets (DELH) versus IUCN criteria: example of Serbian reptiles

Recent studies on the world reptiles’ extinction risks pointed out that about 20 % of species are threatened, with another 20 % are data-deficient following IUCN criteria. IUCN set up criteria for assessment of extinction risk at global and regional scales, relying on distribution data, population parameters and quantitative analyses. Only 12 % of reptilian species were listed as globally threatened upon population data. Long-term population studies are time-consuming, thus it would be practical to use some other parameters for estimations of extinction risks. Threatening level (and extinction risks) of Serbian reptiles have never been investigated. We aimed to scrutinize species threatening level (thus set up conservation priorities) on the basis of distributional, ecological and life-history datasets (DELH). We estimated which traits are the most important for the conservation of species. Separately, we used IUCN criteria for regional level to assess extinction risk and compared them with the results obtained by analyses of DELH datasets. The two types of criteria gave similar estimates for the chelonian species. In snakes, two species exhibited higher conservation scores by DELH than by IUCN criteria, the opposite being true for two other species. In lizards, only two species had higher extinction risks by IUCN whereas five lizards were more threatened according to DELH. Our intention was not discredit the values of IUCN criteria for setting regional (or national) extinction risks, but to suggest that use of additional features other than IUCN could be valuable for estimation of threatening level and setting conservation priorities for cases where precise distribution and population data are not available.

Kinosternon scorpioides scorpioides Linnaeus, 1766: range extension and first records in the upper Paraguay River basin and Mato Grosso do Sul, Brazil

The Scorpion mud turtle (Kinosternon scorpioides) is the chelonian species with the largest distribution range in the Neotropics. The species is reportedly distributed as two disjunct populations in South America: the Amazonian and the Chacoan populations. We present new records of K. s. scorpioides which represent an expansion of the species distribution, with the first records for the Upper Paraguay River basin, the first records for the Mato Grosso do Sul state, Brazil, as well as an evidence that the two previously documented populations area not disjunct.

Distance to human populations influences epidemiology of respiratory disease in desert tortoises

We explored variables likely to affect health of Agassiz's desert tortoises (Gopherus agassizii) in a 1,183-km2 study area in the central Mojave Desert of California between 2005 and 2008. We evaluated 1,004 tortoises for prevalence and spatial distribution of 2 pathogens, Mycoplasma agassizii and M. testudineum, that cause upper respiratory tract disease. We defined tortoises as test-positive if they were positive by culture and/or DNA identification or positive or suspect for specific antibody for either of the two pathogens. We used covariates of habitat (vegetation, elevation, slope, and aspect), tortoise size and sex, distance from another test-positive tortoise, and anthropogenic variables (distances to roads, agricultural areas, playas, urban areas, and centroids of human-populated census blocks). We used both logistic regression models and regression trees to evaluate the 2 species of Mycoplasma separately. The prevalence of test-positive tortoises was low: 1.49% (15/1,004) for M. agassizii and 2.89% (29/1,004) for M. testudineum. The spatial distributions of test-positive tortoises for the 2 Mycoplasma species showed little overlap; only 2 tortoises were test-positive for both diseases. However, the spatial distributions did not differ statistically between the 2 species. We consistently found higher prevalence of test-positive tortoises with shorter distances to centroids of human-populated census blocks. The relationship between distance to human-populated census blocks and tortoises that are test-positive for M. agassizii and potentially M. testudineum may be related to release or escape of captive tortoises because the prevalence of M. agassizii in captive tortoises is high. Our findings have application to other species of chelonians where both domestic captive and wild populations exist. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.

Turtles and Tortoises of Togo: Historical Data, Distribution, Ecology, and Conservation

The chelonian fauna of Togo (West Africa) has been scarcely studied to date. In this article, we review and summarize all available data on the distribution, ecology, and conservation status of the chelonian species of Togo and present a short historical perspective on the development of studies on these reptiles. Overall, 13 chelonian species are found in Togo, 4 being marine, 3 terrestrial, and 6 freshwater. Among the marine species, only 2 of them nest on Togolese beaches (Lepidochelys olivacea and Dermochelys coriacea). Two species (Chelonia mydas and L. olivacea) are still locally common. Concerning the terrestrial and freshwater species, they differed remarkably in terms of their distribution by ecological zone. Cluster analyses revealed the existence of 3 clusters of species, 1 that contains the “forest” species, 1 accommodating the Sudanese savannah species, and a third cluster for the generalist savannah species. The conservation status of the various species and their dissimilarities in terms of threat score categories were assessed through a multivariate model following published work on African chelonian species in general. The results of this model indicate that the potentially most threatened species in Togo are Kinixys erosa, Kinixys homeana, and the 2 Cyclanorbis species (C. senegalensis and C. elegans), with Trionyx triunguis also being possibly seriously threatened. Overall, using the International Union for Conservation of Nature Red List criteria, the concurrent presence of 2 Critically Endangered (K. homeana, C. elegans), 1 Endangered (K. erosa), and several Vulnerable species shows that the chelonian fauna of Togo should be of priority relevance for the competent international agencies.

How to obtain a blood sample in reptiles via venepuncture

The use of venepuncture in reptiles is an important diagnostic step in determining illness and disease. There are several venepuncture sites that vary between species and knowledge of these will ensure a diagnostic sample. Knowing how to correctly handle the reptile will prevent any unnecessary injury to the handler or the animal. The correct gauge needle and smallest syringe need to be used so as not to damage the red blood cells, and the site needs to be cleaned appropriately. This article will look at the venepuncture sites and techniques used in the Chelonian and Squamata species.

Anterior segment morphology and morphometry in selected reptile species using optical coherence tomography

To provide new and original images of the anterior segment (AS) of the eye of selected Ophidian, Chelonian, and Saurian species and to compare the AS architecture among and within these three groups. 17 Saurians, 14 Ophidians, and 11 Chelonians with no concurrent systemic or eye disease were included in the study. Age, weight, nose-cloaca distance (NCD), and pupil shape were collected for each animal. The AS was examined by optical coherence tomography (OCT). After gross description of the appearance of the AS, the central and peripheral corneal thickness (CCT, PCT) and anterior chamber depth (ACD) were measured using the software provided with the OCT device. The ratio CCT/ACD was then calculated for each animal. Pupil shape was a vertical slit in all the crepuscular or nocturnal animals (except for 1 chelonian and 1 ophidian). Each group had its own particular AS architecture. Saurians had a regularly thin cornea with a flat anterior lens capsule and a deep anterior chamber. Ophidians had a thick cornea with a narrow anterior chamber due to a very anteriorly anchored spherical lens. The spectacle was difficult to identify in all ophidians except in Python molurus bivitattus in which it was more obvious. Chelonians displayed an intermediate architecture which more closely resembled the Saurian type than the Ophidian type. Despite grossly similar AS architecture, the three groups of reptiles in the study demonstrated differences that are suggestive of a link between anatomical disparities and variations in environment and lifestyle.

Blood gases, biochemistry, and hematology of Galapagos green turtles (Chelonia mydas).

The green turtle, Chelonia mydas, is an endangered marine chelonian with a circum-global distribution. Reference blood parameter intervals have been published for some chelonian species, but baseline hematology, biochemical, and blood gas values are lacking from the Galapagos sea turtles. Analyses were done on blood samples drawn from 28 green turtles captured in two foraging locations on San Cristóbal Island (14 from each site). Of these turtles, 20 were immature and of unknown sex; the other eight were males (five mature, three immature). A portable blood analyzer (iSTAT) was used to obtain near immediate field results for pH, lactate, pO2, pCO2, HCO3 −, Hct, Hb, Na, K, iCa, and Glu. Parameter values affected by temperature were corrected in two ways: (1) with standard formulas; and (2) with auto-corrections made by the iSTAT. The two methods yielded clinically equivalent results. Standard laboratory hematology techniques were employed for the red and white blood cell counts and the hematocrit determination, which was also compared to the hematocrit values generated by the iSTAT. Of all blood analytes, only lactate concentrations were positively correlated with body size. All other values showed no significant difference between the two sample locations nor were they correlated with body size or internal temperature. For hematocrit count, the iSTAT blood analyzer yielded results indistinguishable from those obtained with high-speed centrifugation. The values reported in this study provide baseline data that may be useful in comparisons among populations and in detecting changes in health status among Galapagos sea turtles. The findings might also be helpful in future efforts to demonstrate associations between specific biochemical parameters and disease.

Population pharmacokinetics of a single intramuscular administration of tulathromycin in adult desert tortoises (Gopherus agassizii)

Tulathromycin, a long acting macrolide antibiotic, has demonstrated efficacy against respiratory pathogens including Mycoplasma bovis and M.hyopneumoniae. A pharmacokinetic study was performed to evaluate the clinical applicability of tulathromycin in desert tortoises following a single intramuscular dose of 5mg/kg. A single blood sample was collected from 110 different desert tortoises at 0.25, 0.5, 1, 4, 8, 24, 48, 72, 120, and 240h following drug administration. Plasma concentrations of the parent form of tulathromycin were measured using liquid chromatography/mass spectrometry. As each tortoise was only bled once, pharmacokinetic parameters were initially estimated using a naïve pooled data approach. Given the variability in the data, population-based compartmental modeling was also performed. Using nonparametric population compartmental modeling, a two-compartment model with first-order absorption and elimination best fit the data. An observed Cmax of 36.2±29.7μg/mL was detected at 0.25h (observed Tmax ). The elimination half-life (T½el ) was long (77.1h) resulting in detectable plasma concentrations 240h postadministration. This study represents a preliminary step in evaluating the utility of tulathromycin in chelonian species and demonstrates that population data modeling offers advantages for estimating pharmacokinetic parameters where sparse data sampling occurs and there is substantial variability in the data.

Influence of male courtship intensity and male-male competition on paternity distribution in Hermann's tortoise,Testudo hermanni hermanni(Chelonia: Testudinidae)

Honest-advertisement models of sexual selection suggest that condition-dependent male secondary sexual characters could function as reliable signals of male quality, enabling females to discriminate among potential partners, both in the pre- and post-copulatory phases. In this context, many studies have revealed the importance of promiscuous mating systems and female sperm storage in determining the occurrence of such a model of sexual selection. By contrast, few studies have investigated the presence and extent of post-copulatory female choice in chelonian species. The present study aimed to investigate the effect of male size, male–male competition, and courtship intensity on paternity distribution in Testudo hermanni hermanni, combining behavioural and genetic data. We created experimental groups composed of two males of different sizes and three or four randomly selected females. Observations conducted during social interactions between males revealed that a hierarchy, unrelated to male size, was soon established: Alpha males were more aggressive towards competitors and courted females more intensively. Alpha males also achieved a higher mounting success than Beta males. Paternity analysis performed on hatchlings produced from experimental females revealed that male reproductive success was not correlated with male–female size ratio. Finally, despite the higher mounting success of Alpha males, paternity analysis revealed that male reproductive success did not differ between Alpha and Beta males. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 111, 656–667. ADDITIONAL KEYWORDS: assignment – female choice – intrasexual competition – reproductive success – sperm storage.

Dietary variation and overlap in D’Orbigny’s slider turtles Trachemys dorbigni (Duméril and Bibron 1835) (Testudines: Emydidae)

Although D’Orbigny’s slider (Trachemys dorbigni) is the most abundant freshwater chelonian species in southern Brazil, little is known about its feeding habits. Our goal was therefore to evaluate this species’ dietary composition and niche variation there. For this, we collected road-killed animals (n = 73) on a federal highway (BR 392) between 2002 and 2003, and analysed their gut contents. We identified 26 different dietary items, and our results indicated that D’Orbigny’s slider is omnivorous in this area. Total food volume, as well as the degree of herbivory and carnivory, were similar between males and females. However dietary composition of plants was different: although both males and females fed on underwater plant matter, only females consumed surface macrophytes. This finding suggested differential microhabitat usage between males and females throughout the swamps.

Molecular epidemiology and genetic diversity of Entamoeba species in a chelonian collection

Veterinary medicine has focused recently on reptiles, due to the existence of captive collections in zoos and an increase in the acquisition of reptiles as pets. The protozoan parasite, Entamoeba can cause amoebiasis in various animal species and humans. Although amoebiasis disease is remarkably rare in most species of chelonians and crocodiles, these species may serve as Entamoeba species carriers that transmit parasites to susceptible reptile species, such as snakes and lizards, which can become sick and die. In this study, we identified the Entamoeba species in a population of healthy (disease-free) chelonians, and evaluated their diversity through the amplification and sequencing of a small subunit rDNA region. Using this procedure, three Entamoeba species were identified: Entamoeba invadens in 4.76 % of chelonians, Entamoeba moshkovskii in 3.96 % and Entamoeba terrapinae in 50 %. We did not detect mixed Entamoeba infections. Comparative analysis of the amplified region allowed us to determine the intra-species variations. The E. invadens and E. moshkovskii strains isolated in this study did not exhibit marked differences with respect to the sequences reported in GenBank. The analysis of the E. terrapinae isolates revealed three different subgroups (A, B and C). Although subgroups A and C were very similar, subgroup B showed a relatively marked difference with respect to subgroups A and C (Fst = 0.984 and Fst = 1.000, respectively; 10-14 % nucleotide variation, as determined by blast) and with respect to the sequences reported in GenBank. These results suggested that E. terrapinae subgroup B may be either in a process of speciation or belong to a different lineage. However, additional research is necessary to support this statement conclusively.

Socio-economic and spatial determinants of anthropogenic predation on Yellow-spotted River Turtle, Podocnemis unifilis (Testudines: Pelomedusidae), nests in the Brazilian Amazon: Implications for sustainable conservation and management

Human expansion has drastically affected wildlife species across Amazonian waterways and the continued increase in rural populations across Amazonia is likely to increase pressure on widely exploited chelonian species. The lack of information evaluating determinants of human consumption patterns limits the effective implementation of conservation strategies. Our objective was to determine the relative importance of social, economic, and spatial variables on human exploitation of Podocnemis unifilis Troschel, 1848 around a sustainable-use protected area in the eastern Brazilian Amazon. We conducted interviews with 51 riverine residents to evaluate attitudes towards and exploitation of P. unifilis. We used multimodel inference to evaluate 12 working hypothesis predicting social, economic, and spatial influences on the occurrence and intensity of P. unifilis egg consumption. None of the respondents reported catching and/or eating adult P. unifilis and none sold adults or eggs during the previous year. Although the majority (58.8%) of respondents had eaten P. unifilis eggs during the previous year and the species was occasionally caught in fishing nets, P. unifilis was rarely cited as a liked or disliked species. Our information theoretic analysis showed that spatial hypotheses were the most strongly supported whereas social and economic hypotheses were only weakly supported in explaining if and how many P. unifilis eggs had been consumed by riverine residents during the previous year. Our findings suggest that current Amazonian development patterns may be associated with reduced consumption of adult P. unifilis, but consumption of eggs together with other indirect anthropogenic perturbations continue to threaten remaining P. unifilis populations.

Calcium-binding proteins and cytochrome oxidase activity in the turtle optic tectum with special reference to the tectofugal visual pathway

Using immunohistochemistry and a tracer technique we investigated the distribution in the optic tectum of turtles (Emys orbicularis and Testudo horsfieldi) of the calcium-binding proteins (CaBPr) parvalbumin (PV), calbindin (CB) and calretinin (CR) before and after labeling of the nucleus rotundus (Rot) with horseradish peroxidase. The optic tectum activity of the cytochrome oxidase (CO) was studied in parallel. In the principal link of the tectofugal visual pathway (central gray layer, SGC) in both chelonian species, the sparse PV-ir as well as CB- and CR-ir neurons were found significantly varying both in number and the intensity of immunoreactivity of their bodies and dendrites. In contrast, the superficial (SGFS) and deeper periventricular (SGP) tectal layers comprised numerous cells immunoreactive to all three CaBPr in different proportions. Only few retrogradely labeled tectorotundal SGC neurons expressed PV, CB or CR. The very large PV-ir neurons in SGC and SAC were not retrogradely labeled; morphologically they matched the efferent neurons with descending projections. SGC neurons of two chelonian species differed in the level of CO activity. Intense immunoreactivity to all three CaBPr and high CO activity were detected in both species in SGFS neuropil with some differences in sublaminar distribution patterns. The peculiarities of the CaBPr and CO activity distribution patterns in different segments of SGC neurons are discussed as related to the laminar organization of the turtle tectum and its retinal innervation. It is suggested that in the projection tectorotundal SGC neurons the CaBPr are concentrated mainly in their distal dendrites that contact retinal afferents in the superficial retinorecipient tectal layer.

Determination of Glomerular Filtration Rate in Juvenile Kemp's Ridley Turtles (Lepidochelys kempii) Using Iohexol Clearance, with Preliminary Comparison of Clinically Healthy Turtles vs. Those with Renal Disease

Abstract Glomerular filtration rates (GFR) were determined with the use of iohexol clearance testing for 17 clinically healthy, captive, juvenile Kemp's ridley turtles (Lepidochelys kempii) and three Kemp's ridley turtles with histopathologically confirmed renal disease. Iohexol was administered intravenously at 75 mg/kg, and blood was collected 4, 8, and 24 h after injection. Plasma iohexol concentrations were determined by high-performance liquid chromatography, and GFR was calculated. Iohexol clearance rates for healthy turtles were 9–17.4 ml/kg/h (mean: 12.6, median: 13.2, standard deviation: 2.17), and those for renal disease turtles were 10.2, 3.0, and 9.2 ml/kg/h, respectively. Administration of intravenous iohexol to Kemp's ridley turtles appeared to be safe, as no adverse effects were detected. This study provides the first data for iohexol clearance in a chelonian species, and the first GFR data for Kemp's ridley turtles. Although the data are limited, this study suggests that iohexol clearance ...

One Extinct Turtle Species Less: Pelusios seychellensis Is Not Extinct, It Never Existed

Pelusios seychellensis is thought to be a freshwater turtle species endemic to the island of Mahé, Seychelles. There are only three museum specimens from the late 19th century known. The species has been never found again, despite intensive searches on Mahé. Therefore, P. seychellensis has been declared as “Extinct” by the IUCN and is the sole putatively extinct freshwater turtle species. Using DNA sequences of three mitochondrial genes of the historical type specimen and phylogenetic analyses including all other species of the genus, we provide evidence that the description of P. seychellensis was erroneously based on a widely distributed West African species, P. castaneus. Consequently, we synonymize the two species and delete P. seychellensis from the list of extinct chelonian species and from the faunal list of the Seychelles.