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Few aquatic animal negative-sense RNA viruses have been characterized, and their role in disease is poorly understood. Here, we describe a virus isolated from diseased freshwater turtles from a Florida farm in 2007 and from an ongoing epizootic among free-ranging populations of Florida softshell turtles (Apalone ferox), Florida red-bellied cooters (Pseudemys nelsoni), and peninsula cooters (Pseudemys peninsularis). Affected turtles presented with similar neurological signs, oral and genital ulceration, and secondary microbial infections. Microscopic lesions were most severe in the softshell turtles and included heterophilic/histiocytic meningoencephalitis, multi-organ vasculitis, and cytologic observation of leukocytic intracytoplasmic inclusions. The virus was isolated using Terrapene heart (TH-1) cells. Ultrastructurally, viral particles were round to pleomorphic and acquired an envelope with prominent surface projections by budding from the cell membrane. Viral genomes were sequenced from cDNA libraries of two nearly identical isolates and determined to be bi-segmented, with an ambisense coding arrangement. The larger segment encodes a predicted RNA-directed RNA polymerase (RdRP) and a putative zinc-binding matrix protein. The smaller segment encodes a putative nucleoprotein and an envelope glycoprotein precursor (GPC). Thus, the genome organization of this turtle virus resembles that of arenaviruses. Phylogenetic analysis shows that the RdRP of the turtle virus is highly diverged from the RdRPs of all known negative-sense RNA viruses and forms a deep branch within the phylum Negarnaviricota, that is not affiliated with any known group of viruses, even at the class level. In contrast, the GPC protein of the turtle virus is confidently affiliated with homologs from a distinct group of fish hantaviruses. Thus, the turtle virus is expected to become the founder of a new taxon of negative-sense RNA viruses, at least with a family rank, but likely, an order or even a class. These viruses probably evolved either by reassortment or by intrasegment recombination between a virus from a distinct branch of negarnaviruses distant from all known groups and a hanta-like aquatic virus. We suggest the provisional name Tosoviridae for the putative new family, with Turtle fraservirus 1 (TFV1) as the type species within the genus Fraservirus. A conventional RT-PCR assay, targeting the TFV1 RdRP, confirmed the presence of viral RNA in multiple tissues and exudates from diseased turtles. The systemic nature of the TFV1 infection was further supported by labeling of cells within lesions using in situ hybridization targeting the RNA of the TFV1 RdRP. Author summary: Freshwater turtles are an important part of aquatic ecosystems, but relatively little is known about diseases that affect them. We discovered a new virus associated with die-offs involving multiple turtle species in Florida. Affected turtles had inflammation involving the nervous system and other organs indicative of viral infection. We isolated a virus from diseased turtles and characterized its morphology using electron microscopy. We also sequenced its entire genome to explore its relatedness to other viruses and developed a PCR-based detection assay. The morphology of the virus particle and genome characteristics were reminiscent of an arenavirus. However, phylogenetic analyses of two viral proteins indicate that the turtle virus most likely evolved through genetic mixing between a distinct branch of RNA viruses distant from all known groups and an aquatic hantavirus. Thus, the turtle virus is expected to become a founding member of a new RNA virus family. We provide evidence linking this novel virus with disease in both wild and captive turtles. [ABSTRACT FROM AUTHOR]

Whether species coculture can overcome the shortcomings of crop monoculture requires additional study. Here, we show how aquatic animals (i.e. carp, crabs, and softshell turtles) benefit paddy ecosystems when cocultured with rice. Three separate field experiments and three separate mesocosm experiments were conducted. Each experiment included a rice monoculture (RM) treatment and a rice-aquatic animal (RA) coculture treatment; RA included feed addition for aquatic animals. In the field experiments, rice yield was higher with RA than with RM, and RA also produced aquatic animal yields that averaged 0.52-2.57 t ha-1. Compared to their corresponding RMs, the three RAs had significantly higher apparent nitrogen (N)-use efficiency and lower weed infestation, while soil N contents were stable over time. Dietary reconstruction analysis based on 13C and 15N showed that 16.0-50.2% of aquatic animal foods were from naturally occurring organisms in the rice fields. Stable-isotope-labeling (13C) in the field experiments indicated that the organic matter decomposition rate was greater with RA than with RM. Isotope 15N labeling in the mesocosm experiments indicated that rice used 13.0-35.1% of the aquatic animal feed-N. All these results suggest that rice-aquatic animal coculture increases food production, increases N-use efficiency, and maintains soil N content by reducing weeds and promoting decomposition and complementary N use. Our study supports the view that adding species to monocultures may enhance agroecosystem functions. [ABSTRACT FROM AUTHOR]

The black soldier fly (Hermetia illucens) larvae meal (BSFM) has been widely applied in aquaculture production due to its rich nutritional value, easy availability, and renewability. However, to our knowledge, it has not been used in the diets for turtles. Here, we evaluated the acceptability of partial substitution of fish meal (FM) with defatted BSFM in the diets of juvenile Chinese soft-shelled turtles (Pelodiscus sinensis). A total of 225 juvenile turtles were randomly divided into D1, D2, D3, D4, and D5 groups, which were fed with BSFM replacing 0, 5%, 10%, 15%, and 20% FM, respectively, for 70 days. The results showed that growth performances of groups D2 and D3 were not significantly different (P > 0.05) from that of group D1, whereas the growth performance compromised (P < 0.05) in groups D4 and D5. The activity of serum alkaline phosphatase was higher (P < 0.05) in group D3 relative to other groups, while alanine aminotransferase and aspartate aminotransferase activities were higher (P < 0.05) in group D4 when compared with groups D1, D2, and D3. Liver lysozyme activity and malonaldehyde level were significantly higher (P < 0.05) in group D1 relative to other groups, whereas total antioxidant capacity activity showed the opposite trend (P < 0.05). The activities of hepatic total superoxide dismutase and glutathione peroxidase displayed a linear elevation (P = 0.011 and P < 0.001 , respectively) from groups D1 to D3. Intestinal amylase and protease activities linearly increased (P < 0.001) with increasing BSFM levels, while intestinal lipase activity showed a quadratic increase and then a decrease with increasing BSFM levels (P = 0.011). Muscle crude protein and fat contents increased (P < 0.05) in group D3 when compared with groups D1 and D2. Muscle phenylalanine, glutamic, tryptophan, and arginine levels increased in group D1 in comparison with the other groups (P < 0.05), whereas the opposite was true for isoleucine and proline levels. The broken-line analysis based on specific growth rate estimated that the optimal level for replacing dietary FM with BSFM is 5.0%, which could elicit benefits on both the growth performance and physiological health condition of the juvenile Chinese soft-shelled turtles. [ABSTRACT FROM AUTHOR]

We describe a new subspecies of Amyda ornata (Gray, 1861) from the southeastern parts of Bangladesh (Chittagong hill tracts), adjacent regions of north-eastern India (Mizoram, Assam and Tripura) and Myanmar, based on morphological characteristics. The holotype comprises some skeletal elements held by the Natural History Museum of Vienna; the five paratypes consist of living specimens in the private collection of PP "Turtle Island" (a turtle conservation zoo in Graz, Austria). After the genetic identity had been determined in a previous investigation, the morphological description was carried out by examining living specimens and photo documents; and additionally, by evaluating the computed tomography of further living animals with exact location data. Amyda ornata jongli ssp. nov. differs from the other Amyda ornata subspecies mainly by its relatively compact and stout head in subadults and adults, its paler, greyish and less contrasting colouration missing distinct bright yellow pigments, ochre-coloured irises, and the existence of more distinct tubercles in the neck region and in the posterior section of the carapace than both other subspecies of Amyda ornata. The tubercles along the front of the carapace protrude clearly in subadult and adult specimens. This softshell turtle is heavily exploited within its limited distribution. Therefore, special comments on conservation aspects are given together with a final discussion on distribution and systematics. [ABSTRACT FROM AUTHOR]

The extant pig‐nosed turtle (Carettochelys insculpta), persisting in far northern Australia and southern New Guinea, is the last surviving member of Carettochelyidae and the only non‐Gondwanan freshwater turtle lineage in Australia. Despite having a global fossil record dating to the Cretaceous, the absence of carettochelyid fossils from Australia has implied a relatively recent colonization of this landmass. Here we report an upper Miocene to lower Pliocene carettochelyid fossil from Beaumaris, Victoria, in south‐eastern Australia. This record is the most southerly occurrence of the clade Carettochelyidae. The presence of carettochelyids in southern Australia, and a discontinuous record of trionychids (soft‐shell turtles) in the Cenozoic of Queensland, suggests at least two colonizations of Australia by Trionychia, pre‐dating the extant pig‐nosed turtle. This cryptic southern history of tropical soft‐shell and pig‐nosed turtles ended before the recent aridification of Australia, leaving the Gondwanan side‐necked turtles as the dominant turtle group in Australian freshwater ecosystems. Therefore, this singular fossil reveals a previously unknown shift in the diversity and evolutionary history of freshwater turtles in Australia. [ABSTRACT FROM AUTHOR]

Reptilian skull morphology is highly diverse and broadly categorized into three architectural patterns: anapsid, synapsid, and diapsid. The anapsid condition where the temporal region is completely roofed by bones is observed in both basal reptiles and turtles. According to recent phylogenetic studies, turtles are derived from diapsid reptiles, implying that the temporal fenestrae were secondarily closed in turtles. Although the repeated evolution of dermatocranial bone reductions in the skull's temporal region of the anapsid skull and the biomechanical factors driving the evolution of the anapsid skull in turtles have been actively discussed, its developmental basis for the anapsid skull remains poorly understood. To investigate the developmental mechanisms underlying the morphological evolution of turtle skulls, here we describe the cranial osteogenesis of three cryptodiran turtle species with distinct skull morphologies: Chelonia mydas (Cheloniidae), Mauremys reevesii (Geoemydidae), and Pelodiscus sinensis (Trionychidae). Our interspecific comparison of embryonic cranial osteogenesis suggests that the growth pattern of skull bone elements is presumed to be highly conserved during embryogenesis across cryptodiran turtles. We found that the growth pattern of the five dermatocranial elements that construct the temporal region of the skull (the parietal, postorbital, jugal, quadratojugal, and squamosal) is highly conserved among turtle species, and that the morphological diversity of turtle skulls arises from slight differences in growth patterns. Our study also suggests that interspecific morphological differences in the cheloniid adult skulls may become apparent in the post-hatching growth period rather than during embryogenesis. [ABSTRACT FROM AUTHOR]

The nesting activity of the Chinese softshell turtle (Pelodiscus sinensis) in its natural habitat is largely undocumented despite its broad geographical distribution across East Asia. Here, we report on female nesting behaviors, egg incubation, and hatchling emergence of P. sinensis on the Yellow River in northwestern China. In this region, the turtle is currently threatened by overharvesting, and successful conservation strategies should include strict monitoring of hunting and trade activities. [ABSTRACT FROM AUTHOR]

To conserve or restore riverine turtles, managers need baseline information on subpopulation structure and abundance in multiple rivers across large geographic areas. Assessing the demographics and morphological characteristics of different subpopulations can increase our understanding of how anthropogenic factors influence mortality and reproduction. We examined spiny softshell turtles (Apalone spinifera) in 5 rivers at the western edge of the species' range in southcentral Montana, where no commercial harvest is allowed. Over 4 yrs, we captured 637 spiny softshell turtles with fish-baited hoop traps. Our objective was to compare the subpopulation demographics in the Yellowstone River—considered one of the most intact rivers in the conterminous United States—to 3 Yellowstone River tributaries (Bighorn and Clarks Fork rivers and Pryor Creek) and the adjacent Musselshell River. Subpopulations differed significantly based on the demographic metrics we examined (e.g., mean sizes and sex ratios), and we documented limited numbers of males (4%–15%). Reproductive potential and mortality of adults among rivers appeared distinct based on juvenile and size class distribution of length-frequency histograms. This information from unharvested populations illustrates the variability in subpopulation demographics of riverine turtles. [ABSTRACT FROM AUTHOR]

This study was conducted to investigate the effects of single and mixed carotenoids dietary supplementation on the growth, immunity and skin colouration of Chinese soft‐shelled turtle. Six carotenoid‐supplemented diets (Diet 1/2/3/4/5/6) containing 0/50, 0/100, 0/120, 30/90, 60/60 and 90/30 mg/kg xanthophyll/canthaxanthin were prepared. A diet without carotenoid supplementation was used as the control. Turtles of the similar size (34.06 ± 0.56 g) were fed with these experimental diets for 72 days. The results showed that the turtles fed Diet 2 and Diet 4 had a better weight gain, specific growth and feed conversion ratio than those fed control diet (p <.05). Carotenoid supplementation improved antioxidant capacity and reduced lipid peroxidation in the serum and liver. The turtles in treatment diets showed higher skin redness and yellowness than those in control diet. Skin redness and yellowness were positively linearly related to canthaxanthin supplementation in feed, but the addition of xanthophyll achieved better skin yellowness. Moreover, dietary carotenoids increased the tissue carotenoid content and upregulated the expression of the enzyme β‐carotene oxygenase 2 (BCO2), catalase (CAT) and superoxide dismutase 2 (SOD2). According to the indexes of growth performance, antioxidant capacity and body colour, adding 100–120 mg/kg canthaxanthin or a 30/90–60/60 mg/kg ratio of xanthophyll/canthaxanthin in diets of Chinese soft‐shelled turtle was appropriate under the experimental conditions. [ABSTRACT FROM AUTHOR]

A 12‐week growth trial was conducted to investigate the impacts of dietary zinc on growth performance, whole‐body composition, haematological parameters, liver aspartate aminotransferase and alanine transaminase activity and tissue trace mineral contents of soft‐shelled turtle (Pelodiscus sinensis). Approximately 4 g of turtles were fed fishmeal‐based diets with 35.43, 46.23, 55.38, 66.74, 75.06 and 85.24 mg Zn/kg. Respiratory burst, red blood cells and intestine somatic index enhanced with escalating dietary Zn concentrations up to 55.38 mg/kg and then kept constant beyond this level. Escalating dietary Zn inclusion up to 66.74 mg/kg improved weight gain, survival rate, feed intake, albumin, haemoglobin, liver aspartate aminotransferase and alanine transaminase activity, carapace ratio and crude protein of turtles, beyond which they decreased. Osmolality, hepatosomatic index and crude lipid of turtles firstly declined with elevating dietary zinc levels from 35.43 to 55.38 mg/kg diet and then increased at a higher zinc level. Liver and muscle Zn concentrations enhanced with escalating dietary zinc inclusion, whereas Cu and Fe concentrations reduced. In summary, the optimal dietary zinc requirements for turtles were 61.27, 60.46 and 60.03 mg/kg diet, based on quadratic regression analysis derived from weight gain, aspartate aminotransferase and alanine transaminase activity, respectively. [ABSTRACT FROM AUTHOR]

Zinc (Zn) plays a role in the antioxidant capacity and immunity of aquatic animals. A twelve-week feeding experiment was performed to estimate the impact of dietary zinc on antioxidant enzyme-related gene expression, antioxidant enzyme activity and non-specific immune functions of soft-shelled turtles, Pelodiscus sinensis. Six fishmeal-based experimental diets with 32.45% protein were formulated, which contained 35.43, 46.23, 55.38, 66.74, 75.06 and 85.24 mg/kg Zn, respectively. Catalase (CAT), glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) levels improved with an elevation in dietary Zn from 35.43 to 55.38 mg/kg and then reduced when dietary Zn was further elevated. The expression levels of Nrf2 and antioxidant-related genes CuZnSOD, MnSOD, CAT, GPX1, GPX2, GPX3 and GPX4 escalated with elevating Zn concentration up to 55.38 mg/kg in diets and then reduced as dietary Zn elevated. The expression levels of Kelch-like ECH-associating protein 1 (keap1) showed a reverse trend with that of Nrf2. The contents of malondialdehyde (MDA) in the 55.38 and 66.74 mg/kg Zn diet-fed groups were the lowest. Alkaline phosphatase activity (AKP), superoxide anion (O 2 −), lysozyme activity and total antioxidant capacity (T-AOC) improved with an escalation in dietary Zn concentration up to 66.74 mg/kg. Optimal dietary Zn improved antioxidant capability, immunity, and antioxidant enzyme-related gene expression. The dietary Zn demand for soft-shelled turtles were 60.93 and 61.63 mg/kg, based on second regression analysis of SOD and T-AOC activity, respectively. • Optimal inclusions of zinc escalated antioxidant enzyme activity. • Zn deficiency or excess reduced non-specific immunity. • Zn deficiency or excess altered antioxidant enzyme expression via Nrf2 signaling. • Dietary Zn demand for soft-shelled turtles is 60.93 and 61.63 mg/kg, using SOD and T-AOC activity as indicators, respectively. [ABSTRACT FROM AUTHOR]

We mimicked daily fluctuations in temperature to examine whether the magnitude of the incubation temperature fluctuation affected the immunity of Chinese soft‐shelled turtle (Pelodiscus sinensis) hatchlings. The control incubation temperature was 29℃. Two of the temperatures used (29℃ ± 3℃ and 29℃ ± 5℃) were within the optimal range of the turtle's constant incubation temperature, and one (29℃ ± 9℃) was outside of this range. Data on hatching success, incubation period, and weight of the turtle hatchlings were collected. The incubation period and hatchling weight were significantly affected by temperature fluctuation, whereas the hatching success was not. After two months, the effect of the temperature fluctuation on hatchling weight was much more pronounced. After challenging hatchlings with pathogens, the hatchlings from the extremely fluctuating incubation temperature group were feeble. To explore the molecular changes of immunity associated with incubation temperature, serum lysozyme, acid and alkaline phosphatase activities, and seven immune‐related genes were examined in two‐month‐old turtles. Before pathogen infection, serum lysozyme activities were lower in turtles that experienced fluctuating temperature, and serum acid and alkaline phosphatase activities were higher in turtles exposed at 29℃ ± 3℃ and 29℃ ± 5℃. After pathogen challenge, lysozyme responded to the pathogen infection in 12 h, and acid and alkaline phosphatase activities were lower at 12 and 48 h in all four groups. Although the expression of the seven immune‐related genes was not affected before the bacterial challenge, they were upregulated after the bacterial challenge in all four groups. [ABSTRACT FROM AUTHOR]

The Euphrates softshell turtle (Rafetus euphraticus) is the most threatened chelonian species in the Middle East, where it is endemic to the Tigris and Euphrates basins. Since the ecology of this species is little known, it is difficult to plan any reliable action for its correct management and conservation.To enhance the scientific knowledge on R. euphraticus, and to establish a science‐based management strategy, a detailed investigation was undertaken on the distribution, habitat, activity and home range of R. euphraticus in Turkey. The previous distribution records in Turkey are for the 1990s; thus, this study is the first comprehensive field research on the species after nearly 3 decades.There was a significant sexual size dimorphism in the Turkish population, with males larger than females.Four individuals were radiotracked for 1 year. The mean home range was highly variable depending on how it was calculated, but that of males was considerably greater than that of females by all methods used.A quantitative evaluation was made of habitat change with time. The land cover classes potentially available to turtles in the Euphrates River basin increased by more than 100% between the 1990s and 2018: 92% of this enhancement was derived from water bodies and coastal lands.Fifteen dams have been built in the lower part of the basins in the last 30 years. In addition, continuing human interactions have caused the fragmentation or destruction of suitable habitats for R. euphraticus.The factors threatening R. euphraticus were assessed and a conservation action plan was developed. Some objectives such as monitoring and creating awareness of this action plan have been met while others are still waiting to be achieved. [ABSTRACT FROM AUTHOR]

Pan‐trionychids (soft‐shelled turtles) are known from the Neogene and Quaternary fossil record of Africa. They have so far been totally absent in Palaeogene localities of the continent although they are commonly found in Palaeogene localities across Europe, North America, and Australasia; as such, their absence from the Palaeogene of Africa has been envisaged to be genuine. Here, I describe a large costal from the Eocene of Mali, which documents for the first time the presence of pan‐trionychid turtles in the Palaeogene of the then isolated Afro‐Arabia. The affinities of the new Malian pan‐trionychid with Neogene and Quaternary forms from Africa are discussed, as well as its biogeographic origins and potential dispersal scenarios from other landmasses to Afro‐Arabia during the Palaeogene. [ABSTRACT FROM AUTHOR]

The nuclear-follower relationship is a specialized interspecific foraging association in which the nuclear species excavates the substrate while foraging, and follower species access food items flushed or uncovered by the nuclear species. We observed a nuclear-follower association between Apalone spinifera (Spiny Softshell Turtle) and fish (Micropterus salmoides [Largemouth Bass] and Lepomis spp.) in an urban drainage canal (Wards Creek) in Baton Rouge, LA, during June and July of 2020. Groups of Largemouth Bass and Lepomis spp. numbering 0-2 and 1-20, respectively, followed foraging Spiny Softshell Turtles. When foraging, Spiny Softshell Turtles were observed moving along the creek bottom, thrusting their probosces into and vigorously excavating the substrate. The accompanying fish then entered the cloud of suspended sediment to pursue and capture prey. Excavation of the substrate by Spiny Softshell Turtles appears to be the critical component of this nuclear-follower relationship, flushing or exposing prey that would otherwise be unavailable to fish. This nuclear-follower relationship is apparently one of commensalism, i.e., while fish probably benefit from increased foraging success, few if any benefits accr ue to foraging turtles. [ABSTRACT FROM AUTHOR]

Chinese softshell turtles (Pelodiscus sinensis) (CST) are susceptible to infections by bacteria belonging to the Bacilluscereus group (Bcg). Bcg includes several closely related species, two of which, B. cereus and B. thuringiensis, are pathogens of aquatic animals or insects. In the present study, we collected 57 Bcg isolates obtained from diseased CST from 2016 to 2019 in Kaohsiung and Pingtung, the areas with the most CST farms in Taiwan. All isolates were divided into four genotypes with two restriction enzymes, SmaI and NotI, by pulsed‐field gel electrophoresis and enterobacterial repetitive intergenic consensus–polymerase chain reaction (ERIC‐PCR). Representative isolates from each genotype were subjected to phylogenetic tree analysis using 16S rDNA and pyruvate carboxylase genes as phylogenetic markers, and these CST isolates appeared in different clades. PCR was performed targeting six selected virulence genes, four of which were detected in CST isolates, including cytotoxin K (1/57), hblC of the haemolysin BL complex (46/57), nheA of the non‐haemolytic enterotoxin complex (52/57) and enterotoxin FM (57/57), whereas cereulide synthetase and cereulide peptide synthase‐like genes were not detected in any isolates. [ABSTRACT FROM AUTHOR]

Sex chromosome dosage compensation (SCDC) overcomes gene-dose imbalances that disturb transcriptional networks, as when ZW females or XY males are hemizygous for Z/X genes. Mounting data from non-model organisms reveal diverse SCDC mechanisms, yet their evolution remains obscure, because most informative lineages with variable sex chromosomes are unstudied. Here, we discovered SCDC in turtles and an unprecedented thermosensitive SCDC in eukaryotes. We contrasted RNA-seq expression of Z-genes, their autosomal orthologues, and control autosomal genes in Apalone spinifera (ZZ/ZW) and Chrysemys picta turtles with temperature-dependent sex determination (TSD) (proxy for ancestral expression). This approach disentangled chromosomal context effects on Z-linked and autosomal expression, from lineage effects owing to selection or drift. Embryonic Apalone SCDC is tissue- and age-dependent, regulated gene-by-gene, complete in females via Z-upregulation in both sexes (Type IV) but partial and environmentally plastic via Z-downregulation in males (accentuated at colder temperature), present in female hatchlings and a weakly suggestive in adult liver (Type I). Results indicate that embryonic SCDC evolved with/after sex chromosomes in Apalone's family Tryonichidae, while co-opting Z-gene upregulation present in the TSD ancestor. Notably, Apalone's SCDC resembles pygmy snake's, and differs from the full-SCDC of Anolis lizards who share homologous sex chromosomes (XY), advancing our understanding of how XX/XY and ZZ/ZW systems compensate gene-dose imbalance. [ABSTRACT FROM AUTHOR]

Turtle eggs containing embryos are exceedingly rare in the fossil record. Here, we provide the first description and taxonomic identification, to our knowledge, of a fossilized embryonic turtle preserved in an egg, a fossil recovered from the Upper Cretaceous Xiaguan Formation of Henan Province, China. The specimen is attributed to the Nanhsiungchelyidae (Pan-Trionychia), an extinct group of large terrestrial turtles (possibly the species Yuchelys nanyangensis). The egg is rigid, spherical, and is one of the largest and thickest shelled Mesozoic turtle eggs known. Importantly, this specimen allowed identification of other nanhsiungchelyid egg clutches and comparison to those of Adocidae, as Nanhsiungchelyidae and Adocidae form the basal extinct clade Adocusia of the Pan-Trionychia (includes living soft-shelled turtles). Despite the differences in habitat adaptations, nanhsiungchelyids (terrestrial) and adocids (aquatic) shared several reproductive traits, including relatively thick eggshells, medium size clutches and relatively large eggs, which may be primitive for trionychoids (including Adocusia and Carrettochelyidae). The unusually thick calcareous eggshell of nanhsiungchelyids compared to those of all other turtles (including adocids) may be related to a nesting style adaptation to an extremely harsh environment. [ABSTRACT FROM AUTHOR]

Objectives: Aeromonads cause severe diseases in farmed aquatic organisms. Herein, we examined 28 isolates causing disease in farmed aquatic organisms from India (n = 24) and Taiwan (n = 4) to gain insight of their genotypic and phenotypic properties. Results: API 20NE biochemical phenotyping showed ≥ 90% similarity classifying all isolates as Aeromonas hydrophila. 16S rRNA genotyping showed ≥ 98% homology among all isolates with A. sobria (NR119044.1ATCC), A. veronii (MK990549.1), A. caviae (NR029252.1) and A. hydrophila (MG984625.1ATCC) and other reference strains. In contrast, gyrB showed a higher intraspecies diversity (≥ 96%) than 16S rRNA delineating the 28 isolates into three groups. Group-I consisted of seven Indian isolates clustered with A. sobria (MK484163.1ATCC), group-II comprised of five Indian and two Taiwanese isolates clustered with A. veronii AF417626.1ATCC while group-III had 11 Indian and three Taiwanese isolates grouped with A. hydrophila (AY987520.1 and DQ519366.1) reference strains. None of our isolates clustered with A. caviae (AJ868400.1ATCC) reference strain. These findings suggest that A. sobria, A. veronii and A. hydrophila could be the etiological agents of diseases observed in farmed fish and soft-shelled turtles (Pelodiscus sinensis) examined in this study. Overall, our findings accentuate the importance of combining phenotyping with genotyping for correct taxonomic classification of Aeromonas spp. in Aquaculture. [ABSTRACT FROM AUTHOR]

The Chinese soft‐shelled turtle, Pelodiscus sinensis, is an important aquaculture species in China. Understanding the molecular mechanisms of growth regulation may contribute to its genetic improvement and lead to increases in the production. Growth rate in this species varies greatly among individuals, and the variation apparently has a genetic basis. To identify growth‐related genes and explore the molecular mechanism of its growth regulation, fast‐growing and slow‐growing turtles were selected from two growth stages, juveniles or adults, respectively, for transcriptome sequencing with liver tissue using the Illumina sequencing platform. Analyses showed that, in juveniles, 303 genes were differentially expressed, of which 217 genes were expressed at higher levels in the fast‐growing turtles. In adults, 1,093 genes were differentially expressed, of which 682 genes were expressed at higher levels in the fast‐growing turtles. Gene Ontology (GO) enrichment of differentially expressed genes (DEGs) identifies three functional groups being significantly enriched in juveniles, but 71 functional groups in adults. Kyoto Encyclopedia of Genes and Genomes enrichment analysis of DEGs in juveniles identified only one pathway being significantly enriched, the proxisome proliferator‐activated receptor (PPAR) signaling pathway, while eight pathways mostly related to fatty acid metabolism were identified in adults. These findings suggest that regulation of growth is more complicated and involves more genes and pathways in adults compared with juveniles, and metabolic or metabolic‐related genes may relate to the growth difference of P. sinensis. Some key genes related to growth were identified from another six growth‐related signaling pathways, and DEGs in GH‐IGF 1 axis genes and Jak–STAT signaling pathway might play important roles in the growth difference of P. sinensis. In addition, 73 single‐nucleotide polymorphisms (SNPs) from 23 growth‐related genes were successfully genotyped. Growth association analysis in 280 individuals revealed that 2 SNPs in 2 key genes (IGF2R and SLC27A2) were associated with growth of P. sinensis. This study provides information on key genes, SNPs, and biological processes that may be involved in growth and highlights the differentiation of growth regulation in different growth stages of P. sinensis. [ABSTRACT FROM AUTHOR]

In 1909, the famous paleontologist Louis Dollo identified two putative new species of giant soft-shelled turtles from the lowest Eocene record of Belgium, ' Trionyx erquelinnensis ' and ' Trionyx levalensis ', from Erquelinnes and Leval, respectively. However, these proposals did not meet the requirements of the International Code of Zoological Nomenclature, so they were considered as nomina nuda. The information on these specimens or about any other specimen of this lineage of giant turtles from the Belgian record is currently extremely limited. Relatively scarce material from giant trionychids has been described for the lower Eocene record of other European regions. Considering the available information, all the European material has recently been recognized as belonging to the genus Axestemys , which has a North American origin, and possibly attributable to a single species, Axestemys vittata , which currently lacks a diagnosis. Numerous and well-preserved Belgian specimens are deposited in the Royal Belgian Institute of Natural Sciences. In addition to the cited individuals from Erquelinnes and Leval, additional specimens from both localities, as well as others from Orp-le-Grand, are part of this collection. These specimens, found between the decades of 1910 and 1930, have been recently restored, and their study is carried out here. The presence of Axestemys vittata in Belgium (in Leval and Orp-le-Grand) is confirmed. Knowledge about this species is significantly improved, and a diagnosis is proposed. However, the hypothesis proposed by Dollo is here confirmed, this species being not the only identified in the Belgian record. So, Axestemys erquelinnensis nov. sp. is defined based on the carapace from Erquelinnes known by Dollo, suggesting that the genus probably reached Europe during the Paleocene Eocene Thermal Maximum. [ABSTRACT FROM AUTHOR]

To investigate the effects of sunlamp-based lighting mode on the growth performance, survival rate, stress response, and oxidative stress of juvenile soft-shelled turtles in a greenhouse, four lighting modes with three replicates were used in this study: lighting the basking area and water simultaneously (LBW), lighting the water only (LW), lighting the basking area only (LB), and no light (NL). Results showed that the final weight, weight gain rate, and specific growth rate had no significant differences among treatments. The survival rate was significantly lower in LBW than in NL, while no significant differences were observed among NL, LB, and LW. On the physiological level, serum cortisol level was significantly lower in LB compared to the other two treatments with light, and glucose level was significantly lower in LB than in LW and NL. With regard to oxidative stress, indexes indicated that the turtles suffered less oxidative stress in LB. Glutathione peroxidase (GSH-PX) was significantly lower in LB compared to the other three treatments. Total antioxidant capacity (T-AOC) was significantly lower in LB, as well as in LW and NL, than in LBW, and no significant differences were found among LB, LW, and NL. Additionally, complement 3 (C3) was significantly higher in LB than in NL, which indicated that the turtles had higher immune-related protein in LB. On the whole, the results demonstrated that lighting mode had significant impacts on survival rate, stress response, and oxidative stress. It can be concluded that lighting the basking area only is the optimal lighting mode for soft-shelled turtles in a greenhouse. From the perspective of breeding welfare, soft-shelled turtles are suggested to be provided with a separate area for basking when cultured in a greenhouse. [ABSTRACT FROM AUTHOR]

Sarju river flowing through the Himalayan foothills is recognized as a key turtle conservation habitat holding remarkable freshwater turtle diversity. This study is a strenuous attempt to document the diversity, distribution and relative abundance of freshwater chelonians in the lower Sarju River along with their bathymetric preferences in order to prioritize certain river sections as critical habitats of various threatened species including the elusive, Crowned River Turtle (Hardella thurjii). The entire length of the river (ca. 205 km) from source to confluence was trekked and canoed. A total of 289 turtles, representing nine species were encountered via 50 observation points. Six of the nine species recorded in this short unprotected river stretch are endangered in IUCN Redlist. Indian Roofed Turtle (Pangshura tecta) was the most abundant and widely distributed species observed at 28 sites followed by Indian Flapshell turtle (Lissemys punctata) and H. thurjii recorded at 16 and 8 sites, respectively. Nine important habitats were identified and ranked considering the species diversity, abundance, extent of habitat, and its potential to support conservation intervention. Interesting observations were made with the species such as P. tecta, H. thurjii, Indian Peacock Softshell Turtle (Nilssonia hurum) and Spotted pond turtle (Geoclemys hamiltonii), which utilized a variety of depth levels from bottom to top of the water column as compared to the other species. Though utilizing varied depth range, H. thurjii was largely observed when river formed deeper pools with heavily vegetated foraging areas. The information will help in the future to refine conservation strategies as well as the supplementation and recovery program of one of the most enigmatic species across North Indian subcontinent. [ABSTRACT FROM AUTHOR]

Sheikh Jamal Inani National Park (Inani) is a wildlife habitat in Bangladesh located under the Cox’s Bazar South Forest Division, Cox’s Bazar. It constitutes significant habitat for the charismatic and globally ‘Endangered’ Western Hoolock Gibbon Hoolock hoolock in Bangladesh. Here, we show that Inani is a poorly-known gibbon habitat with a population of seven groups, comprising a total of 18 individuals. Among them, 77.8% were adults (males and females), and 11.1%, 5.6%, and 5.6% were sub-adults, juveniles, and infants, respectively, indicating low reproductive output. Five of seven groups had no off spring present in the group, and the mean group size of 2.57 individuals/group is low compared to other habitats in Bangladesh. Beside Western Hoolock Gibbon, Inani is home to many threatened wildlife species. The first record of the Slaty-backed Flycatcher Ficedula erithacus in Bangladesh occurred in Inani, adding this new species to the national bird checklist of Bangladesh. The presence of the globally ‘Endangered’ Asian Elephant Elephas maximus, Phayre’s Langur Trachypithecus phayrei, & Elongated Tortoise Indotestudo elongata and the globally ‘Vulnerable’ Northern Pig-tailed Macaque Macaca leonina, Capped Langur Trachypithecus pileatus, Indian Leopard Panthera pardus, & Asiatic Softshell Turtle Amyda cartilaginea highlight the importance of Inani as a conservation area in Bangladesh. The Western Hoolock Gibbon and other threatened wildlife of Inani are now on the verge of local extinction due to a sharp increase in forest resource extraction resulting from the recent influx of large numbers of Rohingya refugees from Myanmar, many of whom have settled around Inani. Through stakeholder interviews in the area, we have identified feasible and measurable conservation actions at Inani that are urgently needed to prevent further loss of wildlife and to protect this important gibbon habitat. [ABSTRACT FROM AUTHOR]

We review the historical and current distribution for Cantor's giant softshell turtle (Pelochelys cantorii) in India. We report 13 new records from Kerala State. Based on our intensive survey in Kasaragod District, we report a potential breeding population from this region. [ABSTRACT FROM AUTHOR]

A detailed understanding of the spatial ecology and habitat requirements of endangered species is critical for population restoration and conservation. Home ranges and movements of the endangered Chinese softshell turtle (Pelodiscus sinensis) were investigated in the Yellow River, northwestern China, from October 2016 to July 2018. We monitored 9 adult turtles (5 females and 4 males) with radio transmitters. Mean linear range size was 440 ± 161 m SD, mean river channel area was 1.98 ± 0.72 ha SD, the average minimum convex polygon was 1.36 ± 0.65 ha SD, average 95% kernel density estimator measured 0.84 ± 0.51 ha SD, with a core area (50% kernel density estimator) of 0.30 ± 0.20 ha SD. Home range values were not significantly different between the sexes nor were they related to straight-line carapace length or mass. Daily movements of P. sinensis averaged 35 ± 18 m SD for males and 43 ± 18 m SD for females, and there was no significant difference between sexes or significant correlation between movement patterns and body size. However, during the nesting season, there was a significant difference between sexes. Turtle activity was highest in May (average daily movement 59 ± 6 m SD) and lowest in January (0.4 ± 0.6 m SD). Given that P. sinensis individuals have small home ranges and are capable of existing in small rivers, management efforts should protect smaller rivers that may be easily overlooked. Results from this study provide the first assessment of home range requirements for P. sinensis, which may be used in future population modeling efforts and are important for establishing conservation strategies for this vulnerable species. [ABSTRACT FROM AUTHOR]

Recreational activities can be detrimental to biodiversity; for example, off-road vehicle traffic (e.g., ATV riding), which has become increasingly popular in recent decades, can threaten wildlife. Although ATV riding around wetlands may threaten the shallow nests of turtles, there are no data on the effect of ATVs on turtle nests. We studied nest site choice and nest survival in two species of softshell turtles (Apalone mutica and A. spinifera) along a river in Louisiana before (1993–1994) and after (2015–2016) ATV riding became popular at the site to determine whether ATVs were an important source of nest mortality, and whether there was an effect of nest site choice on nest survival. ATVs were the most common source of nest mortality (one-third of nests destroyed); nest mortality was significantly positively related to increased ATV traffic but was not influenced by species or nest site choice. Experiments with surrogate eggs and an ATV revealed that the most vulnerable nests to ATV mortality were those that were shallower, were driven over more slowly, and were turned upon. We recommend restricting the access of riding clubs to the river; enforcement of regulations on isolated riders from adjacent residential areas will be logistically and financially challenging. [ABSTRACT FROM AUTHOR]

The impacts of dietary protein on growth, body proximate composition, feed utilization, amino acid profile, digestive enzyme activity and amino acid metabolizing enzymes in soft‐shelled turtles (Pelodiscus sinensis) (initial weight: 4.02 ± 0.06 g) were measured during an 8‐week feeding trail. Six diets containing 8%, 18%, 28%, 38%, 48% and 60% (control group) fishmeal were formulated, which contained 14.38%, 20.41%, 26.19%, 32.23%, 37.63% and 45.23% protein respectively. Weight gain, mean feed intake, skirt ratio, pepsin activity of stomach, total amino acid of the muscle and crude protein levels of the whole body improved as dietary protein content increased from 14.38% to 26.19% but remained stable thereafter. The hepatosomatic index, feed conversion ratio, intestinal alpha‐amylase activity, moisture content of the whole body and serum alanine transaminase and aspartate aminotransferase activity decreased with increasing protein content up to 26.19% and then plateaued. The protein productive value and protein efficiency ratio decreased as protein content increased. Intestinal lipase activity and crude lipid levels of the whole body were unaffected by dietary protein level. Hepatic alanine transaminase and aspartate aminotransferase activity first significantly increased with increasing protein but remained stable at protein content ≥26.19%. The optimum dietary protein level for soft‐shelled turtles was in the range of 27.36%–28.46% based on weight gain, ALT activity and AST activity results. [ABSTRACT FROM AUTHOR]

Extracellular vesicles (EVs) are a heterogeneous group of cell-derived membranous structures comprising exosomes and microvesicles that originate from the endosomal system or are shed from the plasma membrane respectively. As mediators of cell communication, EVs are present in biological fluids and are involved in many physiological and pathological processes. The role of EVs has been extensively investigated in the mammalian male reproductive tract, but the characteristics and identification of EVs in reptiles are still largely unknown. In this review we focus our attention on EVs and their distribution in the male reproductive tract of the Chinese softshell turtle Pelodiscus sinensis, mainly discussing the potential roles of EVs in intercellular communication during different phases of the reproductive process. In softshell turtles, Sertoli-germ cell communication via multivesicular bodies can serve as a source of EVs during spermatogenesis, and these EVs interact with epithelia of the ductuli efferentes and the principal cells of the epididymal epithelium. These EVs are involved in sperm maturation, transport and storage. EVs are also shed by telocytes, which contact and exchange information with other, as well as distant interstitial cells. Overall, EVs play an indispensable role in the normal reproductive function of P. sinensis and can be used as an excellent biomarker for understanding male fertility. [ABSTRACT FROM AUTHOR]

In this study, waste fat from the Chinese soft-shelled turtle (Pelodiscus sinensis) was used as the raw material, and soft-shelled turtle oil (SSTO) was extracted by water heating. Analysis of the fatty acid composition of SSTO revealed that unsaturated fatty acids (UFAs) comprised more than 70% of the oil, of which more than 20% were omega-3 poly-UFAs. DPPH radical scavenging and cellular ROS assays confirmed the reduction of oxidative stress by SSTO. In D-galactose-induced aging rats, SSTO feeding alone or in combination with swimming training resulted in improved memory and physical strength. In addition, SSTO feeding with swimming intervention significantly increased the SOD level and maintained better blood pressure in the aged rats. The serum DHEAS and soleus muscle glycogen level were also highly correlated with SSTO feeding and swimming training. In conclusion, the results of this study demonstrated that SSTO has the potential to be developed into a health food that exerts anti-aging effects, and those effects are stronger when combined with daily swimming exercise. [ABSTRACT FROM AUTHOR]

Organisms have evolved various mechanisms to cope with the differences in the gene copy numbers between sexes caused by degeneration of Y and W sex chromosomes. Complete dosage compensation or at least expression balance between sexes has been reported predominantly in XX/XY systems, but rarely in ZZ/ZW systems. However, this often‐reported pattern is based on comparisons of lineages where sex chromosomes evolved from nonhomologous genomic regions, potentially differing in sensitivity to differences in gene copy numbers. Here we document that two reptilian lineages (XX/XY iguanas and ZZ/ZW softshell turtles), which independently co‐opted the same ancestral genomic region for the function of sex chromosomes, evolved different gene dose regulatory mechanisms. The independent co‐option of the same genomic region for the role of sex chromosomes as in the iguanas and the softshell turtles offers great opportunity for testing evolutionary scenarios on sex chromosome evolution under the explicit control of the genomic background and gene identity. We show that the parallel loss of functional genes from the Y chromosome of the green anole and the W chromosome of the Florida softshell turtle led to different dosage compensation mechanisms. Our approach controlling for genetic background thus does not support that the variability in the regulation of gene dose differences is a consequence of ancestral autosomal gene content. [ABSTRACT FROM AUTHOR]

Antioxidant defense is essential for animals to cope with homeostasis disruption during hibernation. The present study aimed to investigate the antioxidant defense response of juvenile soft-shelled turtle Pelodiscus sinensis during hibernation and following arousal. Turtle brain, liver, and kidney samples were collected at pre-hibernation (17 °C mud temperature; MT), during hibernation (5.8 °C MT) and after arousal (20.1 °C MT) in the field. Transcript levels of NF-E2-related factor 2 (Nrf2) decreased significantly during hibernation and recovered after arousal in all tissues. Cerebral and nephric copper-zinc superoxide dismutase (Cu/Zn SOD), catalase (CAT), glutathione peroxidase 3 (GPx3) and nephric GPx4 mRNA showed similar changing patterns as Nrf2. Cerebral Mn SOD , GPx1 and nephric GPx1 up-regulated after arousal. Hepatic Cu/Zn SOD , GPx1 and GPx3 mRNA kept stable, except hepatic GPx4 increased during hibernation. Hepatic Mn SOD and CAT increased after arousal. In the GSH system, mRNA levels of glutathione synthetases (GS s) kept stable during hibernation and up-regulated after arousal in most tissues except nephric GS2 mRNA remained unchanged. Gene expressions of glutathione reductase (GR) exhibited a tissue specific changing pattern, while those of glutathione-S-Transferase (GST) shared a similar pattern among tissues: remained stable or down-regulated during hibernation then recovered in arousal. In contrast to these diverse responses in gene expressions, most of the antioxidant enzyme activities maintained high and stable. Overall, no preparation for oxidative stress (POS) strategy was found in enzymatic antioxidant system in P. sinensis juveniles during hibernation, the Chinese soft-shelled turtles were able to stay safe from potential oxidative stress during hibernation by maintaining high level activities/concentrations of the antioxidant enzymes/antioxidants. [ABSTRACT FROM AUTHOR]

Chemokines are small, secreted proteins with chemoattractive properties, which play an important role in the recruitment and activation of immune cells. CXCL11 is a CXC chemokine specific for the CXCR3 receptors, which has been shown to mediate the generation of Th1-type immune responses and have bactericidal effects similar to defensins. Herein, we cloned the full-length cDNA of Chinese soft-shelled turtle (Pelodiscus sinensis) CXCL11, designated as PsCXCL11, which consist of an open reading frame (ORF) of 282 bp encoding 93 amino acids, with estimated molecular weight of 10.055 kDa and isoelectric point of 10.37. The deduced PsCXCL11 sequence had a signal peptide, a highly conserved family-specific small cytokine (SCY) domain, one putative N-glycosylation site and ten potential phosphorylation sites. Phylogenetic analysis showed a close relationship between P. sinensis and Chelydra Serpentina CXCL11. P. sinensis CXCL 11 basal expression levels were higher in heart, kidney and spleen than in other organs of health turtles. Infections of Aeromonas hydrophila and Staphylococcus aureus led to significant upregulation of P. sinensis CXCL11 in the blood, while significant upregulation of PsCXCL11 were observed in liver and spleen after infection of A. hydrophila , but not S. aureus. PsCXCL11 recombinant protein with His-tag was successfully expressed by an auto-inducible expression system, and purified by Ni-NTA affinity chromatography. These findings laid a solid foundation for further research towards development of the Chinese soft-shelled turtle as a model for the role of CXCL11 in regulating inflammatory responses to stimulation by invading pathogens. • The full-length cDNA of Chemokine CXCL11 was identified and characterized in Chinese soft-shelled turtle Pelodiscus sinensis. • Infections of Aeromonas hydrophila and Staphylococcus aureus led to varying degrees of upregulation of CXCL11 in blood, spleen and liver. [ABSTRACT FROM AUTHOR]

• Chromatin remodeler genes might be involved in gonadal differentiation of the turtle. • Cyp19a1 and Foxl2 exhibited female-specific expression at different embryonic stages. • Dmrt1 , Amh , and Sox9 displayed male-specific expression at different embryonic stages. • miRNA might play an essential role in early gonadal differentiation in Pelodiscus sinensis. The Chinese soft-shelled turtle (Pelodiscus sinensis) is extensively cultured in Asia for its nutritional and medical value. Gonadal differentiation is fantastic in turtles, whereas morphologic, mRNA, and miRNA expressions were insufficient in the turtle. In this study, ovaries and testes histomorphology analysis of 14–23 stage embryos were performed, and mRNA and miRNA expression profiles were analyzed. Histomorphology analysis revealed that gonads were undifferentiated at embryonic stage 14. Ovarian morphological differentiation became evident from stage 15, which was characterized by the development of the cortical region and degeneration of the medullary region. Concurrently, testicular morphological differentiation was apparent from stage 15, marked by the development of the medullary region and degeneration of the cortical region. qRT-PCR results showed that Cyp19a1 and Foxl2 exhibited female-specific expression at stage 15 and the expression increased throughout most of the embryonic development. Dmrt1 , Amh , and Sox9 displayed male-specific expression at stage 15 and tended to increase substantially at later developmental stages. The expression of miR-8356 and miR-3299 in ZZ gonads were significantly higher than that in ZW gonads at stage 15, 17 and 19, and they had the highest expression at stage 15. While the expression of miR-8085 and miR-7982 had the highest expression at stage 19. Furthermore, chromatin remodeler genes showed differential expression in female and male P. sinensis gonads. These results of master sex-differentiation genes and morphological characteristics would provide a reference for the research of sex differentiation and sex reversal in turtles. Additionally, the expression of chromatin remodeler genes indicated they might be involved in gonadal differentiation of P. sinensis. [ABSTRACT FROM AUTHOR]

The Chinese Soft-Shelled Turtle (CSST) is a significant commercial aquatic animal in China. However, the increased frequency of diseases in the breeding process has hindered industrial development. A gram-negative bacterium, Z617 strain, was isolated from diseased CSST on a farm in Hubei Province, China. It was identified as Morganella morganii by 16S rDNA. The median lethal dose of Z617 strain was (8.32 ± 0.09) × 104 colony-forming units/g CSST weight. The histopathological changes of infected CSSTs, including hepatic sinus congestion, vacuolization of hepatocytes, ulceration of the intestinal mucosa, destruction of the intestinal villi, an unclear distinction between red and white pulp of the spleen, and swelling and necrosis of the glomerular of the kidney, were examined. The structure of the intestinal community of CSSTs was analyzed, and the distinction between richness and diversity was found. The intestinal microbiota community structure differed between the control and infected groups. The relative abundance of Cetobacterium , Lawsonia , and Bacteroides was increased in the infected group compared with the control group at the genus level. Z617 strain is susceptible to antibiotics such as Ceftazidime and Piperacillin. Moreover, it showed various degrees of drug resistance to Enrofloxacin, Doxycycline, Cefuroxim, Streptomycin, Midecamy Cinum, Minocycline, Gentamicin, Neomycin, Florfenicol, and Carbenicillin. This research indicated that M. morganii handled numerous CSSTs' death and clarified the biological profile and potential mechanisms of the Z617 strain. It throws some light on the disease diagnosis and may provide the referenced value for disease treatment. • Morganella morganii was isolated from diseased CSST, named Z617. • The LD 50 of Z617 strain for CSST is (8.32 ± 0.09) × 104 CFU/g. • Liver, spleen, intestine and kidney of disease CSST showed pathological changes after infection. • Z617 changed the intestinal flora of diseased CSST. • Z617 strain was sensitive to Ceftazidime, Ceftriaxone, Cefoperazone, Piperacillin. [ABSTRACT FROM AUTHOR]

A feature of the Chinese soft-shelled turtle (Pelodiscus sinensis) is seasonal spermatogenesis; however, the underlying molecular mechanism is not well clarified. Here, we firstly cloned and characterized P. sinensis DKKL1 , and then performed comparative genomic studies, expression analysis, and functional validation. P. sinensis DKKL1 had 2 putative N-glycosylation sites and 16 phosphorylation sites. DKKL1 also had classic transmembrane structures that were extracellularly localized. DKKL1 's genetic distance was close to turtles, followed by amphibians and mammals, but its genetic distance was far from fishes. DKKL1 genes from different species shared distinct genomic characteristics. Meanwhile, they were also relatively conserved among themselves, at least from the perspective of classes. Notably, the transcription factors associated with spermatogenesis were also identified, containing CTCF , EWSR1 , and FOXL2. DKKL1 exhibited sexually dimorphic expression only in adult gonads, which was significantly higher than that in other somatic tissues (P < 0.001), and was barely expressed in embryonic gonads. DKKL1 transcripts showed a strong signal in sperm, while faint signals were detected in other male germ cells. DKKL1 in adult testes progressively increased per month (P < 0.05), displaying a seasonal expression trait. DKKL1 was significantly downregulated in testes cells after the sex hormones (17β-estradiol and 17α-methyltestosterone) and Wnt/β-catenin inhibitor treatment (P < 0.05). Likewise, the Wnt/β-catenin inhibitor treatment dramatically repressed CTCF, EWSR1, and FOXL2 expression. Conversely, they were markedly upregulated after the 17β-estradiol and 17α-methyltestosterone treatment, suggesting that the three transcription factors might bind to different promoter regions, thereby negatively regulating DKKL1 transcription in response to the changes in the estrogen and androgen pathways, and positively controlling DKKL1 transcription in answer to the alterations in the Wnt/β-catenin pathway. Knockdown of DKKL1 significantly reduced the relative expression of HMGB2 and SPATS1 (P < 0.01), suggesting that it may be involved in seasonal spermatogenesis of P. sinensis through a positive regulatory interaction with these two genes. Overall, our findings provide novel insights into the genome evolution and potential functions of seasonal spermatogenesis of P. sinensis DKKL1. • DKKL1 had a relatively conserved function across species, but also underwent functional evolution. • Expression of DKKL1 exhibited testis-biased, age variation, and seasonal variation characteristics. • DKKL1 predominantly was localized in sperm and could be used as a molecular marker for male germ cells in P. sinensis. • CTCF , EWSR1 , and FOXL2 most likely bind to different promoter regions regulating DKKL1 transcription in different pathways. • DKKL1 may be involved in seasonal spermatogenesis of P. sinensis through a positive regulation with HMGB2 and SPATS1. [ABSTRACT FROM AUTHOR]

BACKGROUND There are abundant resources of Carapax Trionycis from soft‐shelled turtle processing wastes each year in China. Our preliminary work showed that Carapax Trionycis ultrafine powder (CTUP) obtained using ball‐milling with a particle size of 2.24 μm (D0.025) contained more active ingredients. The CTUP D0.025 has a good bioaccessibility, but there has been no report about the immunomodulatory function of CTUP. Therefore, using a cyclophosphamide‐induced immunosuppression mice model, we investigated the immunomodulatory effects of CTUP D0.025. RESULTS: The results indicated that CTUP D0.025 administration significantly improved the immune organ (bone marrow, thymus and spleen) indices, ameliorated spleen tissue morphology and increased the capacity of splenocyte proliferation and the activity of macrophage phagocytosis. CTUP D0.025 also significantly promoted the secretion of cytokines (IL‐2, IL‐4, IL‐10, IFN‐γ and TNF‐α), improved the related mRNA expression levels of IL2, IFN‐γ, T‐bet and GATA3 in immunosuppressed mice and increased the production of serum hemolysin and the levels of IgG, IgM as well as complement C3. Moreover, CTUP D0.025 administration enhanced the antioxidant capacity of mice, exhibited a moderating effect on the damage of bone and skeletal muscle and improved the recovery of bone mineral density and calcium metabolism. CONCLUSIONS: These findings demonstrated that CTUP D0.025 had an effective immune‐enhancing function in immunosuppressive Balb/c mice and also exhibited anti‐osteoporosis properties. © 2020 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

The research on hatching ecology of the Chinese softshell turtle Trionyx sinensis has essential guiding roles to clarify the physiological and ecological mechanism of reptile evolution. The aim of this study is to describe the histological changes, differentiation, and maturation of some functional cells during the genesis and development of the liver and pancreas of the Chinese softshell turtle T. sinensis. Softshell turtle eggs were incubated under artificial conditions and hatched within 41–45 days. Hematoxylin and eosin-stained embryonic pancreas and liver were examined at various time points from 2 to 31 days and compared with that of other reptiles, amphibians, fishes, and birds in the literature. Immunohistochemical assay for glucagon and insulin was performed on paraformaldehyde-fixed embryos to identify functional cells in the pancreas. Pancreatic endocrine cells of T. sinensis have secretory ability at day 26 of embryonic development, and the dispersed pancreatic endocrine cells may be the result of the incomplete pancreatic development. [ABSTRACT FROM AUTHOR]

North American soft-shelled turtles, including trionychines and plastomenids, are incredibly abundant in latest Cretaceous through earliest Paleocene sediments. Here we describe a new species of plastomenid turtle, Plastomenus joycei, based on a nearly complete early Danian skeleton from the Denver Formation in the Denver Basin, Colorado. Plastomenus joycei is differentiated from all other plastomenid turtles based on the presence of large eighth costals that are much longer than wide, sinusoidal raised ridges on the carapace, flat posterior edge of the carapace, spike-like epiplastra that lack callosities, a broad entoplastron that lacks a callosity, hyoplastra with two lateral processes, presence of metaplastically ossified hyoplastral shoulders (i.e., anteriorly protruding lappets), and a broad midline contact between the hypo- and xiphiplastra. Plastomenus joycei broadly resembles the Cretaceous Gilmoremys spp. and the Eocene Plastomenus thomasii and exhibits intermediate morphology between the two, most notably in the degree of ossification of the plastron. The increase in plastral ossification, as well as a decrease in overall size and an increase in the doming of the shell, co-occurs with the Cretaceous–Paleogene mass extinction and these morphological changes may be in response to an increase in mammalian predators during the early Paleogene. Plastomenus joycei is most commonly found in riverine sandstone sediments and is hypothesized to be a riverine turtle. [ABSTRACT FROM AUTHOR]

In this study, we use a specimen from wild-caught individual to determine the complete mitochondrial genome of the Amur soft-shelled turtle (Pelodiscus maackii). The complete mitogenome of P. maackii has 16,258 bp in length and consists of 13 protein-coding genes (PCGs), 22 tRNAs, two rRNAs, and one control region. The arrangement of genes of P. maackii is identical with previously reported mitogenomes in the family Trionychoidea. According to our result, the ML tree for the phylogenetic reconstruction revealed that the individuals used in present study is closely related with the previously reported sequences of P. sinensis (AY962573 and MG431983) in p-distance 0.7% and 2.5%. [ABSTRACT FROM AUTHOR]

Polystomes (Monogenea: Polystomatidae) of freshwater turtles are currently represented by five genera, namely Neopolystoma, Polystomoides, Polystomoidella, Uropolystomoides and Uteropolystomoides. These parasites can infect the urinary, oral and/or the conjunctival sac systems of their hosts, showing strict site specificity. A recent phylogenetic study showed that the two most diverse genera within chelonian polystomes, i.e. Neopolystoma and Polystomoides, are not monophyletic. Furthermore, polystomes infecting the conjunctival sacs of their host, except for one species, formed a robust lineage. A fusiform egg shape has been reported for conjunctival sac polystomes and it was assumed that this characteristic could be a good character for the systematics of polystomes. Our objective in the present work was, therefore, to study more in depth the morphology of polystomes collected from the conjunctival sacs of chelonians to find characters defining a putative new genus. To achieve this objective, more specimens were collected in 2018 and 2019 from turtles sampled in North Carolina and Florida (USA) to extend taxon sampling for the phylogenetic analysis. Morphological characters of relevant polystome specimens were re-examined from several collections from Asia, Australia, Europe, South Africa, South America and North America. Based on a Bayesian tree inferred from the analysis of four concatenated genes, namely 12S, 18S, 28S and COI, polystomes found in the conjunctival sacs were grouped in three distinct lineages, the first one including a single species infecting an Australian pleurodire turtle; the second one including eleven species infecting cryptodire turtles of South America, North America and Asia; and the last one including a single species infecting a softshell cryptodire turtle of North America. Based on observations of live specimens by Dr. Sylvie Pichelin and our morphological analysis, the conjunctival sac polystomes from Australian turtles are small, cannot extend their body significantly, have a spherical ovary and egg, have a large genital bulb and possess latero-ventral vaginae at the level of the testis. Based on observations of live specimens and morphological analysis of whole mounted specimens, polystomes of the second lineage share the following morphological characteristics: the ability to stretch out and double their length, a long oval ovary, a separate egg-cell-maturation-chamber, fusiform to diamond-shaped eggs with acute tips, small genital bulb and vaginae peripheral on the side of the body at the level of the testis. The polystome species of the third lineage occupies a basal position, has the ability to stretch out and possess an elongated ovary, a large fusiform egg with rounded tips, a small genital bulb and small latero-ventral vaginae at the level of the ovary. These three distinct conjunctival sac polystome lineages are herein described as separate new genera, Aussietrema, Fornixtrema and Apaloneotrema, respectively. [ABSTRACT FROM AUTHOR]