Nanoplastic pollution changes the intestinal microbiome but not the morphology or behavior of a freshwater turtle.

Humans produce 350 million metric tons of plastic waste per year, leading to microplastic pollution and widespread environmental contamination, particularly in aquatic environments. This subsequently impacts aquatic organisms in myriad ways, yet the vast majority of research is conducted in marine, rather than freshwater systems. In this study, we exposed eggs and hatchlings of the Chinese soft-shelled turtle (Pelodiscus sinensis) to 80-nm polystyrene nanoplastics (PS-NPs) and monitored the impacts on development, behavior and the gut microbiome. We demonstrate that 80-nm PS-NPs can penetrate the eggshell and move into developing embryos. This led to metabolic impairments, as evidenced by bradycardia (a decreased heart rate), which persisted until hatching. We found no evidence that nanoplastic exposure affected hatchling morphology, growth rates, or levels of boldness and exploration, yet we discuss some potential caveats here. Exposure to nanoplastics reduced the diversity and homogeneity of gut microbiota in P. sinensis, with the level of disruption correlating to the length of environmental exposure (during incubation only or post-hatching also). Thirteen core genera (with an initial abundance >1 %) shifted after nanoplastic treatment: pathogenic bacteria increased, beneficial probiotic bacteria decreased, and there was an increase in the proportion of negative correlations between bacterial genera. These changes could have profound impacts on the viability of turtles throughout their lives. Our study highlights the toxicity of environmental NPs to the embryonic development and survival of freshwater turtles. We provide insights about population trends of P. sinensis in the wild, and future directions for research.

Microbial communities are thermally more sensitive in warm-climate lizards compared with their cold-climate counterparts.

Environmental temperature affects the composition, structure, and function of the gut microbial communities in host animals. To elucidate the role of gut microbiota in thermal adaptation, we designed a 2 species × 3 temperatures experiment, whereby we acclimated adult males of two agamid lizard species (warm-climate Leiolepis reevesii and cold-climate Phrynocephalus przewalskii) to 20, 28, and 36°C for 2 weeks and then collected their fecal and small-intestinal samples to analyze and compare the microbiota using 16S rRNA gene amplicon sequencing technology. The fecal microbiota displayed more pronounced interspecific differences in microbial community than the small-intestinal microbiota in the two species occurring in thermally different regions. The response of fecal and small-intestinal microbiota to temperature increase or decrease differed between the two species, with more bacterial taxa affected by acclimation temperature in L. reevesii than in P. przewalskii. Both species, the warm-climate species in particular, could cope with temperature change by adjusting the relative abundance of functional categories associated with metabolism and environmental information processing. Functional genes associated with carbohydrate metabolism were enhanced in P. przewalskii, suggesting the contribution of the fecal microbiota to cold-climate adaptation in P. przewalskii. Taken together, our results validate the two hypotheses tested, of which one suggests that the gut microbiota should help lizards adapt to thermal environments in which they live, and the other suggests that microbial communities should be thermally more sensitive in warm-climate lizards than in cold-climate lizards.

Normal table of embryonic development in the Anji salamander Hynobius amjiensis (Hynobiidae).

We established a normal embryonic development table for the Anji salamander Hynobius amjiensis, a critically endangered tailed amphibian of the family Hynobiidae with a very limited distribution in East China, following the standards set by the early developmental table of vertebrates. Put together 32 embryonic stages for the Anji salamander was defined. The total embryonic period from oviposition to hatching is approximately 30 days at 9 °C. Stages 1-16 represent early development from cleavage to neurulation. Stages 17-32 represent organogenesis documenting later developmental events such as tail, gill, and limb formation, and hatching (Stage 32). We provided a detailed description of the external morphology and color changes of the head, trunk, limbs, tail, external gills, and balancers at various stages from egg-laying to hatching. We also described several cases of abnormal embryonic development. The establishment of the embryonic development table in H. amjiensis contributes to better understanding of the ontogeny in tailed amphibians, distinguishing closely related species, and identifying abnormal embryonic amphibians.

Genomic insights into local adaptation in the Asiatic toad Bufo gargarizans, and its genomic offset to climate warming.

Genomic signatures of local adaptation have been identified in many species but remain sparsely studied in amphibians. Here, we explored genome-wide divergence within the Asiatic toad, Bufo gargarizans, to study local adaptation and genomic offset (i.e., the mismatch between current and future genotype-environment relationships) under climate warming scenarios. We obtained high-quality SNP data for 94 Asiatic toads from 21 populations in China to study spatial patterns of genomic variation, local adaptation, and genomic offset to warming in this wide-ranging species. Population structure and genetic diversity analysis based on high-quality SNPs revealed three clusters of B. gargarizans in the western, central-eastern, and northeastern portions of the species' range in China. Populations generally dispersed along two migration routes, one from the west to the central-east and one from the central-east to the northeast. Both genetic diversity and pairwise F ST were climatically correlated, and pairwise F ST was also correlated with geographic distance. Spatial genomic patterns in B. gargarizans were determined by the local environment and geographic distance. Global warming will increase the extirpation risk of B. gargarizans.

Dietary and Sexual Correlates of Gut Microbiota in the Japanese Gecko, Gekko japonicus (Schlegel, 1836).

Numerous studies have demonstrated that multiple intrinsic and extrinsic factors shape the structure and composition of gut microbiota in a host. The disorder of the gut microbiota may trigger various host diseases. Here, we collected fecal samples from wild-caught Japanese geckos (Gekko japonicus) and captive conspecifics fed with mealworms (mealworm-fed geckos) and fruit flies (fly-fed geckos), aiming to examine the dietary and sexual correlates of the gut microbiota. We used 16S rRNA gene sequencing technology to determine the composition of the gut microbiota. The dominant phyla with a mean relative abundance higher than 10% were Verrucomicrobiota, Bacteroidota, and Firmicutes. Gut microbial community richness and diversity were higher in mealworm-fed geckos than in wild geckos. Neither community evenness nor beta diversity of gut microbiota differed among wild, mealworm-fed, and fly-fed geckos. The beta rather than alpha diversity of gut microbiota was sex dependent. Based on the relative abundance of gut bacteria and their gene functions, we concluded that gut microbiota contributed more significantly to the host's metabolic and immune functions. A higher diversity of gut microbiota in mealworm-fed geckos could result from higher chitin content in insects of the order Coleoptera. This study not only provides basic information about the gut microbiota of G. japonicus but also shows that gut microbiota correlates with dietary habits and sex in the species.

Gut microbiota differs between two cold-climate lizards distributed in thermally different regions.

BACKGROUND: The metabolic cold-climate adaption hypothesis predicts that animals from cold environments have relatively high metabolic rates compared with their warm-climate counterparts. However, studies testing this hypothesis are sparse. Here, we compared gut microbes between two cold-climate lizard species of the genus Phrynocephalus to see if gut microbiota could help lizards adapt to cold environments by promoting metabolism. We conducted a 2 species (P. erythrurus and P. przewalskii) × 2 temperatures (24 and 30 °C) factorial design experiment, whereby we kept lizards of two Phrynocephalus species at 24 and 30 °C for 25 d and then collected their fecal samples to analyze and compare the microbiota based on 16S rRNA gene sequencing technology. RESULTS: The gut microbiota was mainly composed of bacteria of the phyla Proteobacteria, Firmicutes, Bacteroidetes, and Verrucomicrobia in both species (Proteobacteria > Firmicutes > Verrucomicrobiota in P. erythrurus, and Bacteroidetes > Proteobacteria > Firmicutes in P. przewalskii). Further analysis revealed that the gut microbiota promoted thermal adaptation in both lizard species, but with differences in the relative abundance of the contributory bacteria between the two species. An analysis based on the Kyoto Encyclopedia of Genes and Genomes revealed that the gut microbiota played important roles in metabolism, genetic information processing, cellular processes, and environmental information processing in both species. Furthermore, genes related to metabolism were more abundant in P. erythrurus at 24 °C than in other species ⋅ temperature combinations. CONCLUSION: Our study provides evidence that gut microbiota promotes thermal adaptation in both species but more evidently in P. erythrurus using colder habitats than P. przewalskii all year round, thus confirming the role of gut microbiota in cold-climate adaptation in lizards.

Single-cell transcriptomes from turtle livers reveal sensitivity of hepatic immune cells to bacteria-infection.

The liver is important in the synthesis, metabolism and storage of nutrients, detoxification and immune response of the body, and the liver immune response against exogenous pathogens from the intestinal tract plays a key role in the immune activities. However, the cellular composition of the liver immune atlas remains sparsely studied in reptiles. We used single-cell RNA sequencing to identify the cellular profile of the liver of the Chinese soft-shelled turtle (Pelodiscus sinensis). We obtained the transcriptional landscape based on 9938 cells from the fractionation of fresh hepatic tissues from two individuals, uninfected and infected with bacteria (Aeromonas hydrophila). We identified seven hepatic immune cell subsets, including plasma, erythroid, T/NK, B, endothelial, dendritic and Kupffer cells. Bacteria-infection altered the number of liver immune cells, as revealed by the fact that the infected turtle had more plasma, endothelial and Kupffer cells and fewer T/NK, dendritic and erythroid cells than did the uninfected turtle. Our study is the first to provide a comprehensive view of the hepatic immune landscape of P. sinensis at the single-cell resolution that outlines the characteristics of immune cells in the turtle liver and provides a liver transcriptome baseline for turtle immunology.

Shell-Isolated Nanoparticle-Enhanced Electrochemiluminescence.

Enhanced electrochemiluminescence (ECL) aims to promote higher sensitivity and obtain better detection limit. The core-shell nanostructures, owing to unique surface plasmon resonance (SPR) enabling distance-dependent strong localized electromagnetic field, have attracted rising attention in enhanced ECL research and application. However, the present structures usually with porous shell involve electrocatalytic activity from the metal core and adsorption effect from the shell, which interfere with practical SPR enhancement contribution to ECL signal. Herein, to exclude the interference and unveil exact SPR-enhanced effect, shell-isolated nanoparticles (SHINs) whose shell gets thicker and becomes pinhole-free are developed by modifying pH value and particles concentration. Furthermore, allowing for the distribution of hotspots and stronger enhancement, excitation intensity and ECL reaction layer thickness are mainly investigated, and several types of SHINs-enhanced ECL platforms are prepared to fabricate distinct hotspot distribution via electrostatic attraction (submonolayer) and a layer-by-layer deposition method (monolayer). Consequently, the strongest enhancement up to ≈250-fold is achieved by monolayer SHINs with 10 nm shell, and the platform is applied in a "turn-off" mode sensing for dopamine. The platform provides new guidelines to shell preparation, interface engineering and hotspots fabrication for superior ECL enhancement and analytical application with high performance.

Identification of a quasi-liquid phase at solid-liquid interface.

An understanding of solid-liquid interfaces is of great importance for fundamental research as well as industrial applications. However, it has been very challenging to directly image solid-liquid interfaces with high resolution, thus their structure and properties are often unknown. Here, we report a quasi-liquid phase between metal (In, Sn) nanoparticle surfaces and an aqueous solution observed using liquid cell transmission electron microscopy. Our real-time high-resolution imaging reveals a thin layer of liquid-like materials at the interfaces with the frequent appearance of small In nanoclusters. Such a quasi-liquid phase serves as an intermediate for the mass transport from the metal nanoparticle to the liquid. Density functional theory-molecular dynamics simulations demonstrate that the positive charges of In ions greatly contribute to the stabilization of the quasi-liquid phase on the metal surface.

Novel Flexible Vacuum-Assisted Ureteral Access Sheath Can Actively Control Intrarenal Pressure and Obtain a Complete Stone-Free Status.

Objectives: To compare the safety and effectiveness of a novel flexible vacuum-assisted ureteral access sheath (FV-UAS) and traditional ureteral access sheath (UAS) in simulating retrograde intrarenal surgery (RIRS). Materials and Methods: A manometric model was established in porcine kidneys to observe the change in intrarenal pressure (IRP) in the FV-UAS and traditional UAS groups at different irrigation fluid velocities of 30, 50, 80, and 100 mL/min. Establish a kidney stone model (with 0.2 g, dry, ≤5 mm stones) to simulate RIRS. A total of 20 porcine kidneys were randomly numbered from 1 to 20 (FV-UAS group, 1 - 10; traditional UAS group, 11 - 20). The stone volume clearance rate and operation time were compared between the two groups. ("Stonevolumeclearancerate=1-ResidualstonevolumePreoperativestonevolume×100%"). Stone volume was obtained by CT pre- and postoperatively. Results: FV-UAS can follow flexible ureteroscopy (f-URS) to cross the ureteropelvic junction (UPJ) and into the renal pelvis and calices. FV-UAS can actively make IRP <10 cmH2O by adjusting the negative values at different irrigation fluid velocities. The mean residual stone volume of the FV-UAS vs traditional UAS groups was 33.7 vs 92.5 mm3 (p = 0.017). The mean stone volume clearance rates of the FV-UAS vs traditional UAS groups were 98.5% and 95.9%, respectively (p = 0.017). Seven cases achieved complete stone-free status in the FV-UAS group. All patients had residual fragments postoperatively in the traditional UAS group. Conclusions: FV-UAS can follow f-URS to cross the UPJ and into the renal pelvis and calices, avoiding the interference of UPJ in controlling IRP. FV-UAS can actively control the IRP to be reduced to the desired range by adjusting the negative value under any irrigation fluid velocity. FV-UAS close to the stone can achieve complete stone-free status in RIRS.

Lessons from the diet: Captivity and sex shape the gut microbiota in an oviparous lizard (Calotes versicolor).

Studies have indicated that the abundance and community structure of gut microbiota are altered by diet. In this study, next-generation sequencing of the 16S rRNA gene amplicon was performed to evaluate variations in the gut microbiota of wild and captive individuals of both sexes of Calotes versicolor. The results showed that there was a significant sex difference in microbial community structure for wild C. versicolor, Bacteroide was the dominant genus in wild females (WF), whereas Ochrobactrum was the dominant genus in wild males (WM). Acinetobacter and Hymenobacter were the dominant genera in WF, while Clostridium was the dominant genus in captive females (CF). The results indicated that differences in diet between wild and captive C. versicolor also resulted in variations in gut microbiota. Thus, it was not surprising that captivity and sex shape the gut microbiota in C. versicolor. In summary, the fundamental information presented about the gut microbiota of both sexes of wild (and captive females) C. versicolor, indicates that the artificial environments are not suitable for the wild C. versicolor.

Ultrasensitive Temperature Sensing Based on Ligand-Free Alloyed CsPbClx Br3-x Perovskite Nanocrystals Confined in Hollow Mesoporous Silica with High Density of Halide Vacancies.

Temperature sensing based on fluorescent semiconductor nanocrystals has recently received immense attention. Enhancing the trap-facilitated thermal quenching of the fluorescence should be an effective approach to achieve high sensitivity for temperature sensing. Compared with conventional semiconductor nanocrystals, the defect-tolerant feature of lead halide perovskite nanocrystals (LHP NCs) endows them with high density of defects. Here, hollow mesoporous silica (h-SiO2 ) template-assisted ligand-free synthesis and halogen manipulation (chloride-importing) are proposed to fabricate highly defective yet fluorescent CsPbCl1.2 Br1.8 NCs confined in h-SiO2 (CsPbCl1.2 Br1.8 NCs@h-SiO2 ) for ultrasensitive temperature sensing. The trap barrier heights, exciton-phonon scattering, and trap state filling process in the CsPbCl1.2 Br1.8 NCs@h-SiO2 and CsPbBr3 NCs@h-SiO2 are studied to illustrate the higher temperature sensitivity of CsPbCl1.2 Br1.8 NCs@h-SiO2 at physiological temperature range. By integrating the thermal-sensitive CsPbCl1.2 Br1.8 NCs@h-SiO2 and thermal-insensitive K2 SiF6 :Mn4+ phosphor into the flexible ethylene-vinyl acetate polymer matrix, ratiometric temperature sensing from 30.0 °C to 45.0 °C is demonstrated with a relative temperature sensitivity up to 13.44% °C-1 at 37.0 °C. The composite film shows high potential as a thermometer for monitoring the body temperature. This work demonstrates the unparalleled temperature sensing performance of LHP NCs and provides new inspiration on switching the defects into advantages in sensing applications.

Venom of the Annulated Sea Snake Hydrophis cyanocinctus: A Biochemically Simple but Genetically Complex Weapon.

Given that the venom system in sea snakes has a role in enhancing their secondary adaption to the marine environment, it follows that elucidating the diversity and function of venom toxins will help to understand the adaptive radiation of sea snakes. We performed proteomic and de novo NGS analyses to explore the diversity of venom toxins in the annulated sea snake (Hydrophis cyanocinctus) and estimated the adaptive molecular evolution of the toxin-coding unigenes and the toxicity of the major components. We found three-finger toxins (3-FTxs), phospholipase A2 (PLA2) and cysteine-rich secretory protein (CRISP) in the venom proteome and 59 toxin-coding unigenes belonging to 24 protein families in the venom-gland transcriptome; 3-FTx and PLA2 were the most abundant families. Nearly half of the toxin-coding unigenes had undergone positive selection. The short- (i.p. 0.09 µg/g) and long-chain neurotoxin (i.p. 0.14 µg/g) presented fairly high toxicity, whereas both basic and acidic PLA2s expressed low toxicity. The toxicity of H. cyanocinctus venom was largely determined by the 3-FTxs. Our data show the venom is used by H. cyanocinctus as a biochemically simple but genetically complex weapon and venom evolution in H. cyanocinctus is presumably driven by natural selection to deal with fast-moving prey and enemies in the marine environment.

Venom-gland transcriptomic, venomic, and antivenomic profiles of the spine-bellied sea snake (Hydrophis curtus) from the South China Sea.

BACKGROUND: A comprehensive evaluation of the -omic profiles of venom is important for understanding the potential function and evolution of snake venom. Here, we conducted an integrated multi-omics-analysis to unveil the venom-transcriptomic and venomic profiles in a same group of spine-bellied sea snakes (Hydrophis curtus) from the South China Sea, where the snake is a widespread species and might generate regionally-specific venom potentially harmful to human activities. The capacity of two heterologous antivenoms to immunocapture the H. curtus venom was determined for an in-depth evaluation of their rationality in treatment of H. curtus envenomation. In addition, a phylogenetic analysis by maximum likelihood was used to detect the adaptive molecular evolution of full-length toxin-coding unigenes. RESULTS: A total of 90,909,384 pairs of clean reads were generated via Illumina sequencing from a pooled cDNA library of six specimens, and yielding 148,121 unigenes through de novo assembly. Sequence similarity searching harvested 63,845 valid annotations, including 63,789 non-toxin-coding and 56 toxin-coding unigenes belonging to 22 protein families. Three protein families, three-finger toxins (3-FTx), phospholipase A2 (PLA2), and cysteine-rich secretory protein, were detected in the venom proteome. 3-FTx (27.15% in the transcriptome/41.94% in the proteome) and PLA2 (59.71%/49.36%) were identified as the most abundant families in the venom-gland transcriptome and venom proteome. In addition, 24 unigenes from 11 protein families were shown to have experienced positive selection in their evolutionary history, whereas four were relatively conserved throughout evolution. Commercial Naja atra antivenom exhibited a stronger capacity than Bungarus multicinctus antivenom to immunocapture H. curtus venom components, especially short neurotoxins, with the capacity of both antivenoms to immunocapture short neurotoxins being weaker than that for PLA2s. CONCLUSIONS: Our study clarified the venom-gland transcriptomic and venomic profiles along with the within-group divergence of a H. curtus population from the South China Sea. Adaptive evolution of most venom components driven by natural selection appeared to occur rapidly during evolutionary history. Notably, the utility of commercial N. atra and B. multicinctus antivenoms against H. curtus toxins was not comprehensive; thus, the development of species-specific antivenom is urgently needed.

Complete mitochondrial genome of the Bingzhi's stout newt (Pachytriton granulosus Chang, 1933) and its phylogenetic placement.

The Bingzhi's stout newt (Pachytriton granulosus Chang, 1933) is distributed in mountainous areas of Zhejiang, China. The first complete mitochondrial genome (mitogenome) of P. granulosus was determined by next-generation sequencing. The size of the assembled mitogenome for P. granulosus was 16,293 bp, which included 13 protein coding genes, 22 tRNAs, 2 rRNAs, a non-coding region, and a control region (D-loop). The phylogenetic analysis using Bayesian Inference validated the taxonomic status of P. granulosus, showing the close relationship with the other two species from the genus Pachytriton.

Dietary Correlates of Oral and Gut Microbiota in the Water Monitor Lizard, Varanus salvator (Laurenti, 1768).

Numerous studies have demonstrated that food shapes the structure and composition of the host's oral and gut microbiota. The disorder of oral and gut microbiota may trigger various host diseases. Here, we collected oral and gut samples from wild water monitor lizards (Varanus salvator) and their captive conspecifics fed with bullfrogs, eggs, and depilated chicken, aiming to examine dietary correlates of oral and gut microbiota. We used the 16S rRNA gene sequencing technology to analyze the composition of the microbiota. Proteobacteria and Bacteroidota were the dominant phyla in the oral microbiota, and so were in the gut microbiota. The alpha diversity of microbiota was significantly higher in the gut than in the oral cavity, and the alpha diversity of oral microbiota was higher in captive lizards than in wild conspecifics. Comparing the relative abundance of oral and gut bacteria and their gene functions, differences among different animal groups presumably resulted from human contact in artificial breeding environments and complex food processing. Differences in gene function might be related to the absolute number and/or the taxonomic abundance of oral and gut microorganisms in the wild and the water environment. This study provides not only basic information about the oral and gut microbiota of captive and wild water monitor lizards, but also an inference that feeding on frogs and aquatic products and reducing human exposure help water monitor lizards maintain a microbiota similar to that in the wild environment.