Identification and expression analysis of dmc1 in Chinese soft-shell turtle (Pelodiscus sinensis) during gametogenesis.

DNA meiotic recombinase 1 (disrupted meiotic cDNA, Dmc1) protein is homologous to the Escherichia coli RecA protein, was first identified in Saccharomyces cerevisiae. This gene has been well studied as an essential role in meiosis in many species. However, studies on the dmc1 gene in reptiles are limited. In this study, a cDNA fragment of 1,111 bp was obtained from the gonadal tissues of the Chinese soft-shell turtle via RT-PCR, containing a 60 bp 3' UTR, a 22 bp 5' UTR, and an ORF of 1,029 bp encoding 342 amino acids, named Psdmc1. Multiple sequence alignments showed that the deduced protein has high similarity (>95 %) to tetrapod Dmc1 proteins, while being slightly lower (86-88 %) to fish species.Phylogenetic tree analysis showed that PsDmc1 was clustered with the other turtles' Dmc1 and close to the reptiles', but far away from the teleost's. RT-PCR and RT-qPCR analyses showed that the Psdmc1 gene was specifically expressed in the gonads, and much higher in testis than the ovary, especially highest in one year-old testis. In situ hybridization results showed that the Psdmc1 was mainly expressed in the perinuclear cytoplasm of primary and secondary spermatocytes, weakly in spermatogonia of the testes. These results indicated that dmc1 would be majorly involved in the developing testis, and play an essential role in the germ cells' meiosis. The findings of this study will provide a basis for further investigations on the mechanisms behind the germ cells' development and differentiation in Chinese soft-shell turtles, even in the reptiles.

Research on antibiotic resistance genes in wild and artificially bred green turtles (Chelonia mydas).

Sea turtles, vital to marine ecosystems, face population decline. Artificial breeding is a recovery strategy, yet it risks introducing antibiotic resistance genes (ARGs) to wild populations and ecosystems. This study employed metagenomic techniques to compare the distribution characteristics of ARGs in the guts of wild and artificially bred green turtles (Chelonia mydas). The findings revealed that the total abundance of ARGs in C. mydas that have been artificially bred was significantly higher than that in wild individuals. Additionally, the abundance of mobile genetic elements (MGEs) co-occurring with ARGs in artificially bred C. mydas was significantly higher than in wild C. mydas. In the analysis of bacteria carrying ARGs, wild C. mydas exhibited greater bacterial diversity. Furthermore, in artificially bred C. mydas, we discovered 23 potential human pathogenic bacteria (HPB) that contain antibiotic resistance genes. In contrast, in wild C. mydas, only one type of HPB carrying an antibiotic resistance gene was found. The findings of this study not only enhance our understanding of the distribution and dissemination of ARGs within the gut microbial communities of C. mydas, but also provide vital information for assessing the potential impact of releasing artificially bred C. mydas on the spread of antibiotic resistance.

Swimming through sand: using accelerometers to observe the cryptic, pre-emergence life-stage of sea turtle hatchlings.

Animals that hatch within a subterranean nest, such as turtle hatchlings, expend some of their limited energy reserves digging out through sand or soil to reach the surface. In sea turtles, this emergence process can take the hatchlings 3-7 days. However, we have a poor understanding of this process as it is difficult to observe what is occurring underground. Here, we utilize a novel method to characterize digging-out behaviour: affixing an accelerometer directly to newly hatched green turtles (Chelonia mydas) to record movement until nest emergence. Our data revealed that buried hatchlings maintained a head-up orientation but did not move in the expected left and right swaying motion associated with alternating limb crawling. Rather, they moved using dorsal-ventral heaving and pitching as if swimming vertically through the sand to the surface. Movement activity was irregular and brief, interspersed by many short periods of inactivity, mostly lasting less than 10 min. The first 24 h of head-up activity displayed no diel patterns, but the last 24 h prior to emergence involved more intense movement during night-time hours compared with daytime hours. Thus, our results add valuable new insight, and in some cases change previous assumptions, regarding the digging behaviours during the egg-to-emergence life stage in sea turtles.

Plasma chemistry and hematology of eastern mediterranean sea green turtles undergoing rehabilitation.

The Eastern Mediterranean Sea (EMS) is oligotrophic, presenting low biological production and high temperature and salinity, while comprising a biodiversity hotspot, with a unique emblematic threatened species array, including sea turtles. The establishment of healthy captive green turtle (Chelonia mydas) baseline blood analyte reference intervals (RIs) will contribute to improving diagnosis, treatment, rehabilitation and conservation success rates for sea turtles, by offering information with therapeutic and prognostic implications. However, baseline plasma chemistry and hematological analytes of EMS C. mydas are still unavailable. Hence, this study aimed to establish RIs for several blood analytes in general and under different life and rehabilitation stages and compare results with congeneric and conspecifics of other regions: Mediterranean loggerhead turtles (Caretta caretta) and two Atlantic green turtle populations. The study engulfed 118 blood samples obtained from 72 turtles admitted to the Israel Sea Turtle Rescue Centre between 2008 and 2020. Blood analyses included plasma chemistry and packed cell volume (PCV), and their association with body size and health status, and enabled suggesting RIs for PCV, plasma glucose and potassium concentrations, and aspartate transaminase and alkaline phosphatase activities. The PCV were significantly associated with rehabilitation. Noted differences arising via population comparison are likely associated with interspecific dietary and metabolic traits. These novel physiological results form the basis for future studies, and provide caregivers a tool potentially increasing the success of recovery, rehabilitation and conservation efforts. Further research is warranted to fill knowledge gaps concerning physiology and potential ecological impacts on this species.

Thecal and Epithecal Ossifications of the Turtle Shell: Ontogenetic And Phylogenetic Aspects.

The problem of the origin of the bony shell in turtles has a two-century history and still has not lost its relevance. First, this concerns the issues of the homology, the sources of formation and the ratio of bones of different nature, that is, thecal and epithecal, in particular. This article analyzes various views on the nature of the shell elements, and proposes their typification, based on modern data on developmental biology. It is proposed that the defining characteristic of the types of shell ossifications is not the level of their anlage in the dermis (thecality or epithecality), but, first of all, the primary sources of their formation: (1) neural crest (nuchal and plastral plates); (2) vertebral and rib periosteum (neural and costal plates); and (3) dermal mesenchyme (peripheral, suprapygal and pygal plates, as well as epithecal elements). In addition, there is complete correspondence between these types of ossifications and the sequence of their appearance in the turtle ontogenesis. The data show fundamental coincidence of the modifications of the ontogenetic development and evolutionary formation of the shell ossifications and are in agreement with a stepwise model for the origin of the turtle body plan. Particular attention is paid to the origin of the epithecal elements of the turtle shell, which correspond to the additional or supernumerary ossifications and seem to have wider distribution among turtles, than previously thought.

Productive foraging grounds enhance maternal condition and offspring quality in a capital breeding species.

Feeding ecology is an essential component of an organism's life, but foraging comes with risks and energetic costs. Species in which populations exhibit more than one feeding strategy, such as sea turtles, are good systems for investigating how feeding ecology impacts life-history traits, reproduction and carried over effects across generations. Here, we investigated how the feeding ecology of loggerhead sea turtles (Caretta caretta) nesting at the Cabo Verde archipelago correlates with reproductive outputs and offspring quality. We determined the feeding ecology of female turtles before and during the breeding season from stable isotope analysis of carbon and nitrogen and correlated isotopic ratio with female and offspring traits. We found that female turtles feeding at higher trophic positions produced larger clutches. We also found that females with higher δ13C values, typical of productive foraging areas, had greater fat reserves, were less likely to be infected by leech parasites and produced heavier offspring. The offspring of infected mothers with higher δ13C values performed best in crawling and self-righting trials than those of non-infected mothers with higher δ13C values. This study shows adult female loggerheads that exploit productive areas build capital reserves that impact their reproductive success and multiple proxies for offspring quality. Overall, our findings provide valuable insights into the complex interplay between feeding ecology and reproductive success, and reveal the transgenerational carry-over effects of both feeding ecology and health on offspring quality in sea turtles.

International overview of sea turtle fibropapillomatosis: a survey of expert opinions and trends.

Introduction: Marine environments offer a wealth of opportunities to improve understanding and treatment options for cancers, through insights into a range of fields from drug discovery to mechanistic insights. By applying One Health principles the knowledge obtained can benefit both human and animal populations, including marine species suffering from cancer. One such species is green sea turtles (Chelonia mydas), which are under threat from fibropapillomatosis (FP), an epizootic tumor disease (animal epidemic) that continues to spread and increase in prevalence globally. In order to effectively address this epizootic, a more thorough understanding is required of the prevalence of the disease and the approaches to treating afflicted turtles. Methods: To identify knowledge gaps and assess future needs, we conducted a survey of sea turtle FP experts. The survey consisted of 47 questions designed to assess general perceptions of FP, the areas where more information is needed, local FP trends, the disease status, and mitigation needs, and was voluntarily completed by 44 experts across a broad geographic range. Results: Over 70% of respondents both recognized FP as a cancerous panzootic disease, and reported that FP is increasing in prevalence. They report several factors contributing to this increase. Nearly all of the respondents reported that FP research, patient treatment and rehabilitation required more funding in their area, and reported inadequate facilities and capacity for dealing with FP patients. Treatment approaches varied: just over 70% of the medical experts that responded surgically remove FP tumors, either using laser or scalpel. Just under half of respondents use anti-cancer drugs in their treatment of FP. Internal tumors were reported as justification for euthanasia by 61.5% of respondents, and 30.8% reported severe external tumors to be sufficient grounds for euthanasia. Most medical respondents (93.3%) routinely perform necropsy on deceased or euthanized FP-afflicted turtles. Over 80% of respondents considered large-scale multidisciplinary collaboration 'extremely important' for advancing the field of FP research. Discussion: The survey responses provide a valuable insight into the current status of FP in sea turtles, FP treatment, rehabilitation and research, and help to identify critical FP-related areas most in need of attention.

Status of Hawksbill Turtle and Green Turtle in Negeri Sembilan, Malaysia.

This paper highlighted the annual distribution, seasonality and reproduction status of two species of sea turtles in Negeri Sembilan, Peninsular Malaysia between January 2016 and July 2020 (55 months). These data were officially provided by the Department of Fisheries Malaysia (Negeri Sembilan's state), as a part of a conservation effort made by them to protect the endangered species, as the turtle's population residing in Negeri Sembilan is one of the smallest in Malaysia. Current status shows that the hawksbill turtles, Eretmochelys Imbricata population in Negeri Sembilan is in stable decline with an average of <30 nests per year, and an average of <10 nests per year for the green turtle, Chelonia mydas. The overall sum was 122 nests collected for both species from 2016-2020. This paper updated the status of the hawksbill and green turtle after 30 years from the first study recorded by Mortimer et al. in 1993.

Development and validation of a quantitative PCR assay for detection of Sulawesi tortoise adenovirus.

In 2007, a mortality event involving over 100 Sulawesi tortoises (Indotestudo forsteni), two Impressed tortoises (Manouria impress) and a critically endangered Burmese star tortoise (Geochelone platynota) was attributed to Sulawesi tortoise adenovirus (STADV; genus Siadenovirus). We developed a TaqMan quantitative PCR assay targeting the DNA polymerase gene of STADV for use in clinical diagnosis and epidemiologic surveillance. This assay failed to amplify five closely-related chelonian adenoviruses, indicating high analytical specificity. The assay performed with high efficiency (slope = -3.337; R2 = 0.999) and high inter- and intra-assay repeatability (coefficient of variation <1.36 % at all standard curve dilutions). Dynamic range included 1.00 × 107 to 1.00 × 101 target copies per reaction and limit of detection was 101 target copies per reaction, though 100 target copies per reaction were intermittently detected. This qPCR assay provides a valuable diagnostic tool for characterization of STADV epidemiology, including potential identification of the North American reservoir host.

Notes on the Courtship and Captive Reproduction of the Southern Vietnam Box Turtle, Cuora picturata, at the Turtle Conservation Center in Cuc Phuong National Park, Vietnam.

Endemic to only three known provinces in South Central Vietnam, Cuora picturata have faced extensive collection pressures for the food, medicinal, and pet trades. Further exacerbating their decline is the lack of protected areas where wild populations exist, with only one known population occurring within a protected area. With threats to wild populations persisting, the development of an assurance colony has been prioritized for C. picturata. The Asian Turtle Program of Indo-Myanmar Conservation has been operating the Turtle Conservation Center (TCC) in Cuc Phuong National Park since 1998. From 2020 to 2022, 24 C. picturata were acquired by the TCC from the illegal trade. With no ideal repatriation plan, these animals provided an opportunity for developing a captive assurance colony at the TCC to safeguard the species from extinction. As semi-aquatic species, box turtles in the genus Cuora are a unique group that shows variations in habitat use and behavior between species. Herein, we documented some specific courtship behaviors not yet described for C. picturata, providing insight into the unique reproductive behaviors of this species.

Moving Toward Evidence-Based Care: Measuring Preferences in Zoo-Housed Aquatic Turtles Using 24-h Remote Monitoring.

Providing evidence-based care to animals is a common goal in zoos and aquariums. However, there are many understudied species with a lack of information available, which may result in suboptimal care for individuals of these species. Using preference tests to ask individual animals what they want can be an effective means to enhance their care, and subsequently, their welfare. Using 24-h remote monitoring, we assessed substrate, enclosure feature, and lighting preferences of five aquatic turtles (two wood turtles, one Blanding's turtle, and two spotted turtles) that share an indoor enclosure at Lincoln Park Zoo. In addition, we compared enclosure feature and lighting preferences between winter and summer to determine whether the turtles had changing seasonal preferences despite being housed indoors. We found that the wood turtles and Blanding's turtle had substrate preferences, whereas the spotted turtles did not. In addition, we found that the turtles' enclosure feature preferences were consistent between winter and summer, but the amount of time spent on land, logs, and in water varied by individual. Finally, we found that the turtles utilized areas with ambient lighting more than areas with a heat lamp or UV lamp; this was consistent between winter and summer. The turtle preferences observed in this study can contribute to the development of care strategies that support their welfare and, in turn, may inspire similar preference studies at other institutions as we collectively advance toward providing animals of all taxonomic groups with evidence-based care.

Potential drivers and implications of a balanced breeding sex ratio in a small population of an imperiled species with environmental sex determination.

Small populations of imperiled species are susceptible to the negative consequences of skewed sex-ratios. In imperiled species with environmental sex determination such as sea turtles, examining sex ratios across a range of environments and population abundance levels can provide insight into factors that influence population resilience, which can then be the foci of management plans for these species. Breeding sex ratios (the ratio of actively breeding males to females during a reproductive season; BSRs) extrapolated from genetic parentage analyses are a common approach for enumerating sex ratios in sea turtles. Such analyses also allow for the characterization of multiple paternity within sea turtle clutches, which should reflect BSRs and breeding behaviors. We characterized the first BSR for a breeding assemblage of loggerhead sea turtles (Caretta caretta) belonging to the temperate, low-abundance Northern Gulf of Mexico Recovery Unit using genotypes of 16 microsatellite loci from nesting females and hatchlings. Unlike prior studies at both more-tropical and more-temperate, and higher-abundance, Recovery Units in this region, we found a balanced BSR of 1.3:1 males:female and a low incidence (~17%) of multiple paternity. This suggests that there are relatively few males breeding at this assemblage and within this Recovery Unit. Beaches in this region are expected to produce substantial numbers of male hatchlings based on sand temperature data. The relative dearth of mature males may then be due to hydrologic disturbances that disproportionately affect the fitness and survival of male hatchlings, or due to demographic stochasticity. More work is needed to study the factors that might influence male hatchling production and fitness in this region, particularly as climate change is predicted to lead to feminization in global sea turtle populations. Our work demonstrates the broad utility of characterizing BSRs and other sex ratios across a range of populations in imperiled, environmentally sensitive species.

Trismus in cold-stunned Kemp's ridley (Lepidochelys kempii) and loggerhead (Caretta caretta) sea turtles.

OBJECTIVE: To describe the presentation, clinical findings, diagnosis, treatment, and outcome of cases of trismus (lockjaw) in cold-stunned sea turtles. ANIMALS: 4 Kemp's ridley (Lepidochelys kempii) and 1 loggerhead (Caretta caretta) sea turtle. METHODS: Cold-stunned sea turtles that presented with difficulty or inability to open their jaw between 2009 and 2023 were included. Information retrieved from medical records included signalment, physical exam findings, diagnostic information, definitive diagnosis via either advanced imaging or histopathology, treatment, and clinical outcome. RESULTS: Turtles presented between 4 and 48 days into rehabilitation. Three were diagnosed by advanced imaging (CT or MRI), and 2 were diagnosed based on clinical signs and postmortem histopathology. Treatment was multimodal and consisted of antibiotics (5/5), nonsteroidal anti-inflammatories (5/5), vitamin E (3/5), intralesional steroid therapy (3/5), acupuncture (3/5), antifungals (2/5), anti-inflammatory parenteral steroids (2/5), physical therapy (2/5), therapeutic laser (2/5), and supportive feeding via either total parenteral nutrition (1/5), or tube feedings (2/5). Two animals were released, 2 died naturally, and 1 was euthanized. CLINICAL RELEVANCE: Trismus (lockjaw) is an uncommon finding in stranded cold-stunned sea turtles that can have a significant impact on animal welfare if not diagnosed and treated. This report describes the condition to aid clinical case management and resource allocation in rehabilitation facilities.

Variability in thermal tolerance of clutches from different mothers indicates adaptation potential to climate warming in sea turtles.

The current climate warming is a challenge to biodiversity that could surpass the adaptation capacity of some species. Hence, understanding the means by which populations undergo an increase in their thermal tolerance is critical to assess how they could adapt to climate warming. Specifically, sea turtle populations could respond to increasing temperatures by (1) colonizing new nesting areas, (2) nesting during cooler times of the year, and/or (3) by increasing their thermal tolerance. Differences in thermal tolerance of clutches laid by different females would indicate that populations have the potential to adapt by natural selection. Here, we used exhaustive information on nest temperatures and hatching success of leatherback turtle (Dermochelys coriacea) clutches over 14 years to assess the occurrence of individual variability in thermal tolerance among females. We found an effect of temperature, year, and the interaction between female identity and nest temperature on hatching success, indicating that clutches laid by different females exhibited different levels of vulnerability to high temperatures. If thermal tolerance is a heritable trait, individuals with higher thermal tolerances could have greater chances of passing their genes to following generations, increasing their frequency in the population. However, the high rate of failure of clutches at temperatures above 32°C suggests that leatherback turtles are already experiencing extreme heat stress. A proper understanding of mechanisms of adaptation in populations to counteract changes in climate could greatly contribute to future conservation of endangered populations in a rapidly changing world.

Correlations between local geoclimatic variables and hatchling body size in the sea turtles Caretta caretta and Chelonia mydas.

It has been widely demonstrated that air and sand temperatures influence the anatomy of sea turtle hatchlings. We examined the impact of precipitation during the nesting season on the hatchling body size of loggerhead and green turtles from 37 beaches worldwide. Longitudinal data collected between 2012 and 2018 from Florida (US) and from a sample on Bõa Vista Island (Cabo Verde) carried out in 2019 showed that loggerhead body size at hatching was negatively correlated with precipitation, while precipitation was not correlated with hatchling body size in green turtles. A meta-analysis revealed that precipitation is positively correlated with hatchling mass in loggerhead turtles, while it is positively correlated with straight carapace length and width in green turtle hatchlings. The strongest influence of precipitation was found in the middle of the incubation period of loggerhead turtles in Cabo Verde, and we posit that this is due to an increase in the uptake of water for embryonic growth. These findings highlight the great importance of understanding the correlated effects of regional environmental variables, such as precipitation, on the development of sea turtle hatchlings and will have an impact on the evaluation of ongoing conservation and climate change discussions.

Congenital malformation in green turtle embryos and hatchlings.

Successful embryonic development depends on the interaction between genetic factors and environmental variables. Congenital malformations in sea turtles can result from extreme conditions during the incubation period, reducing hatching success and potentially impeding population recovery. We aimed to characterize the congenital malformations found in green turtle nests, determine their prevalence and severity, and understand their drivers during the 2022 nesting season on Samandag beach on northern Mediterranean nesting beaches. A total of 2986 examples of congenital malformations were observed in 362 out of 907 green turtle nests. The prevalence of congenital malformations per nest was 39%, and the severity (the number of malformed individuals per nest) was 3.8%. Nests with congenital malformations exhibited a lower mean distance from the sea, a shorter incubation duration (a proxy for incubation temperature), lower hatching success, a larger clutch size, and higher mortality at late embryonic and hatchling stages than nests without congenital malformations. There was no significant difference in total mortality between these two nest types. A total of 52 different congenital malformations were recorded, 2 of which were observed for the first time in sea turtles and 28 for the first time in green turtles. The results suggest that congenital malformations may be related to nest temperature and clutch size, while overall mortality may be independent of malformations. Pigmentation disorders and craniofacial malformations typically coexist in cases of multiple malformations. Long-term monitoring of congenital malformations is crucial, as it can provide clues about the health status of the nesting beach and nesting colony.

Emydomyces testavorans Surveillance in Multiple Free-Ranging Terrestrial and Aquatic Chelonian Species in Illinois, USA.

The order Onygenales contains multiple fungal pathogens that affect free-ranging and zoo-housed reptilian species. Emydomyces testavorans, an onygenalean fungus associated with skin and shell disease, has been sporadically detected in aquatic chelonians. Because of the recent discovery of this organism, little is known about its prevalence in free-ranging chelonians. The objective of this study was to perform surveillance for E. testavorans in six free-ranging aquatic and terrestrial chelonian species in Illinois, USA: Blanding's turtles (n=437; Emydoidea blandingii), painted turtles (n=199; Chrysemys picta), common snapping turtles (n=35; Chelydra serpentina), red-eared sliders (n=62; RES; Trachemys scripta elegans), eastern box turtles (n=73; Terrapene carolina carolina) and ornate box turtles (n=29; Terrapene ornata). Combined cloacal-oral swabs (COSs) or shell (carapace and plastron surfaces) swabs were collected from 2019 to 2021 and tested for E. testavorans using quantitative PCR. The PCR detected E. testavorans in COSs of an adult male, subadult female, and juvenile male Blanding's turtle (0.6%; 95% confidence interval [CI], 0.2-1.9%) and a shell swab from an adult female RES (1.6%; 95% CI, 0-8.7%). Shell lesions consistent with E. testavorans infection were present in two of the positive Blanding's turtles. These results document the rarity of this pathogen on the landscape in Illinois. Additional studies should determine this pathogen's impact on individuals and clarify its significance for conservation efforts of Blanding's turtle, in which E. testavorans has not been reported previously.

Unusual Patterns of Lateral Scutes in Two Olive Ridley Turtles and Their Genetic Assignment to the Thai Andaman Sea Populations of Lepidochelys olivacea Eschscholtz, 1829.

Two stranded Lepidochelys-like sea turtles were rescued from the Thai Andaman Sea coastline by veterinarians of the Phuket Marine Biological Center (PMBC), one in May of 2019 and another in July of 2021. They were first identified as olive ridley turtles (Lepidochelys olivacea), as the external appearance of both turtles was closer to that species than the other four species found in the Thai Andaman Sea. In fact, when carefully examined, an unusual pattern of the lateral scutes on each turtle was observed, specifically symmetric 5/5 and asymmetric 5/6, both of which are considered rare for L. olivacea and had never been reported in the Thai Andaman Sea. In contrast, this characteristic was more common for the closely related species, Kemp's ridley (L. kempii), although this species is not distributed in the Indo-Pacific Ocean. Thus, we further investigated their genetic information to confirm species identification using two molecular markers, namely the mtDNA control region and nDNA RAG2. The results from the mtDNA control region sequences using the Basic Local Alignment Search Tool (BLAST) indicated that both individuals exhibited a higher percent identity with L. olivacea (99.81-100.00%) rather than L. kempii (94.29-95.41%) or any other species. A phylogenetic tree confirmed that these two turtles belonged to the L. olivacea clade. Moreover, the results of RAG2 also supported the mtDNA result, as both individuals shared the same RAG2 haplotype with L. olivacea. Thus, we have concluded that the two turtles with unusual lateral scute patterns exhibited genetic consistency with their original species, L. olivacea, which has brought attention to the importance of exploring rare phenotypes in sea turtle populations residing in Thai Seas.

Comparison of Two Intravenous Propofol Doses after Jugular Administration for Short Non-Surgical Procedures in Red-Eared Sliders (Trachemys scripta elegans).

This study compares the effects of two different doses of propofol administered intravenously (IV), in the jugular vein, to red-eared sliders (Trachemys scripta elegans). In this crossover study, 5 or 10 mg/kg propofol was administered to six Trachemys scripta elegans after cannulation of the jugular vein. Each turtle received each dose, G1 (5 mg/kg IV) and G2 (10 mg/kg IV), after a 7-day washout period. The parameters evaluated were heart rate, palpebral reflex, cloacal reflex, muscle relaxation, ease of handling, sensitivity to anterior and posterior pinch stimuli, and possibility of intubation. Additionally, respiratory rate was measured when possible, and the times from propofol administration to full recovery and from intubation to extubation were recorded. None of the turtles in G1 could be intubated, and this dose provided little relaxation and ease of handling, with a duration of effect until full recovery of 12.16 ± 8.32 (SD) min for this group. In G2, five out of the six turtles could be intubated, and the duration of effect was 32.33 ± 5.85 (SD) min. Heart rates were influenced by manipulation for catheter placement. There were statistically significant differences (p value ≤ 0.05) between the two groups in muscle relaxation degree, handling, cloacal reflex, and possibility of intubation. The 5 mg/kg propofol dose was not sufficient to induce anesthesia, even when administered in the jugular vein, in red-eared sliders. A dose of 10 mg/kg IV or higher should be used.

New insights into the early morphological evolution of sea turtles by re-investigation of Nichollsemys baieri, a three-dimensionally preserved fossil stem chelonioid from the Campanian of Alberta, Canada.

The early evolution of Pan-Chelonioidea (sea turtles) is poorly understood. This is in part due to the rarity of undeformed skulls of definitive early stem chelonioids. In this work, we redescribe the holotype of Nichollsemys baieri using µCT scans and segmentations of the skull. This fossil is the best 3D preserved skull of any Campanian sea turtle, and includes partial "soft tissue" preservation. Nichollsemys is morphologically similar but clearly distinct from Toxochelys spp., and both show a mosaic of plesiomorphic and derived chelonioid features. The internal cranial anatomy documents the presence of derived characters in Nichollsemys baieri that are absent in Toxochelys spp., such as the loss of the epipterygoids and the rod-like shape of the rostrum basisphenoidale. Among the numerous plesiomorphic characters is the presence of a splenial bone, which was unnoticed before. An updated phylogenetic analysis retrieves Nichollsemys baieri as a non-protostegid early stem chelonioid in a slightly more crownward position than Toxochelys latiremis. Our phylogeny includes macrobaenids and protostegids as pan-chelonioids, and we find unorthodox results for dermochelyids. Thus, although Nichollsemys baieri provides important new insights into the early morphological evolution of sea turtles, much work remains to be done. As a completely 3D preserved specimen, we included Nichollsemys baieri into a recent landmark-based skull shape dataset of turtles. Morphospace analysis reveals an intermediate position between cryptodires and crown chelonioids. Based on these data, we also predict that Nichollsemys baieri was still capable of neck retraction, constraining the loss of this trait to more crownward pan-chelonioids. Supplementary Information: The online version contains supplementary material available at 10.1186/s13358-024-00323-8.