Microbial diversity in mountain-dwelling amphibians: The combined effects of host and climatic factors.

Comprehending the determinants of host-associated microbiota is pivotal in microbial ecology. Yet, the links between climatic factors and variations in host-associated microbiota necessitate further clarification. Mountain-dwelling amphibians, with limited dispersal abilities, serve as valuable models for addressing these questions. Our study, using 126 amphibian-associated microbial samples (64 gut and 62 skin) and 101 environmental microbial samples (51 soil and 50 water) from the eastern Tibetan Plateau, revealed host factors as primary drivers of the variations in host-associated microbiota. However, climatic factors contributed to additional variations in gut microbial beta-diversity and skin microbial function. Water microbiota were identified as a significant contributor to the amphibian-associated microbiomes, with their climate-driven variations mediating an indirect association between the variations in climatic factors and host-associated microbiota. These findings extend our understanding of the assembly of host-associated microbiota in amphibians, emphasizing the significance of microbiota in evaluating the impact of climate change on animals.

Single cell RNA analysis uncovers the cell differentiation and functionalization for air breathing of frog lung.

The evolution and development of vertebrate lungs have been widely studied due to their significance in terrestrial adaptation. Amphibians possess the most primitive lungs among tetrapods, underscoring their evolutionary importance in bridging the transition from aquatic to terrestrial life. However, the intricate process of cell differentiation during amphibian lung development remains poorly understood. Using single-cell RNA sequencing, we identify 13 cell types in the developing lungs of a land-dwelling frog (Microhyla fissipes). We elucidate the differentiation trajectories and mechanisms of mesenchymal cells, identifying five cell fates and their respective driver genes. Using temporal dynamics analyses, we reveal the gene expression switches of epithelial cells, which facilitate air breathing during metamorphosis. Furthermore, by integrating the published data from another amphibian and two terrestrial mammals, we illuminate both conserved and divergent cellular repertoires during the evolution of tetrapod lungs. These findings uncover the frog lung cell differentiation trajectories and functionalization for breathing in air and provide valuable insights into the cell-type evolution of vertebrate lungs.

What frog gill resorption brings: loss of function, cell death, and metabolic reorganization.

BACKGROUND: Anuran metamorphosis, which is driven by thyroid hormone (TH)-mediated processes, orchestrates intricate morphological and functional transformations for the transition from aquatic tadpoles to terrestrial life, providing a valuable model for studying organ functionalization, remodeling, and regression. Larva-specific organ regression is one of the most striking phenomena observed during the anuran metamorphic climax. While previous studies extensively analyzed the regression mechanisms of the tail, the molecular processes governing gill resorption remain elusive. RESULTS: We employed Microhyla fissipes as a model, and utilized a comprehensive approach involving histological analysis, transmission electron microscopy, and transcriptomics to unravel gill development and resorption. The pro-metamorphic stages revealed highly developed gill structures, emphasizing their crucial role as the primary respiratory organ for tadpoles. The transcriptomic analysis highlighted the upregulation of genes associated with enhanced respiratory efficiency, such as hemoglobin and mucins. However, as metamorphosis progressed, gill filaments underwent shrinkage, decreases in blood vessel density, and structural changes that signified a decline in respiratory function. The molecular mechanisms driving gill resorption involved the TH pathway-in particular, the upregulation of thyroid hormone receptor (TR) ß, genes associated with the tumor necrosis factor pathway and matrix metalloproteinases. Two distinct pathways orchestrate gill resorption, involving apoptosis directly induced by TH and cell death through the degradation of the extracellular matrix. In addition, metabolic reorganization during metamorphosis is a complex process, with tadpoles adapting their feeding behavior and mobilizing energy storage organs. The gills, which were previously overlooked, have been unveiled as potential energy storage organs that undergo metabolic reorganization. The transcriptomic analysis revealed dynamic changes in metabolism-related genes, indicating decreased protein synthesis and energy production and enhanced substrate transport and metabolism during metamorphic climax. CONCLUSION: This study sheds light on the structural, molecular, and metabolic dynamics during gill development and resorption in M. fissipes. The findings deepen our understanding of the intricate mechanisms governing organ regression and underscore the pivotal role of the gills in facilitating the transition from aquatic to terrestrial habitats.

Gut microbiota reflect adaptation of cave-dwelling tadpoles to resource scarcity.

Gut microbiota are significant to the host's nutrition and provide a flexible way for the host to adapt to extreme environments. However, whether gut microbiota help the host to colonize caves, a resource-limited environment, remains unknown. The nonobligate cave frog Oreolalax rhodostigmatus completes its metamorphosis within caves for 3-5 years before foraging outside. Their tadpoles are occasionally removed from the caves by floods and utilize outside resources, providing a contrast to the cave-dwelling population. For both cave and outside tadpoles, the development-related reduction in their growth rate and gut length during prometamorphosis coincided with a shift in their gut microbiota, which was characterized by decreased Lactobacillus and Cellulosilyticum and Proteocatella in the cave and outside individuals, respectively. The proportion of these three genera was significantly higher in the gut microbiota of cave-dwelling individuals compared with those outside. The cave-dwellers' gut microbiota harbored more abundant fibrolytic, glycolytic, and fermentative enzymes and yielded more short-chain fatty acids, potentially benefitting the host's nutrition. Experimentally depriving the animals of food resulted in gut atrophy for the individuals collected outside the cave, but not for those from inside the cave. Imitating food scarcity reproduced some major microbial features (e.g. abundant Proteocatella and fermentative genes) of the field-collected cave individuals, indicating an association between the cave-associated gut microbiota and resource scarcity. Overall, the gut microbiota may reflect the adaptation of O. rhodostigmatus tadpoles to resource-limited environments. This extends our understanding of the role of gut microbiota in the adaptation of animals to extreme environments.

Predicting abiotic stress-responsive miRNA in plants based on multi-source features fusion and graph neural network.

BACKGROUND: More and more studies show that miRNA plays a crucial role in plants' response to different abiotic stresses. However, traditional experimental methods are often expensive and inefficient, so it is important to develop efficient and economical computational methods. Although researchers have developed machine learning-based method, the information of miRNAs and abiotic stresses has not been fully exploited. Therefore, we propose a novel approach based on graph neural networks for predicting potential miRNA-abiotic stress associations. RESULTS: In this study, we fully considered the multi-source feature information from miRNAs and abiotic stresses, and calculated and integrated the similarity network of miRNA and abiotic stress from different feature perspectives using multiple similarity measures. Then, the above multi-source similarity network and association information between miRNAs and abiotic stresses are effectively fused through heterogeneous networks. Subsequently, the Restart Random Walk (RWR) algorithm is employed to extract global structural information from heterogeneous networks, providing feature vectors for miRNA and abiotic stress. After that, we utilized the graph autoencoder based on GIN (Graph Isomorphism Networks) to learn and reconstruct a miRNA-abiotic stress association matrix to obtain potential miRNA-abiotic stress associations. The experimental results show that our model is superior to all known methods in predicting potential miRNA-abiotic stress associations, and the AUPR and AUC metrics of our model achieve 98.24% and 97.43%, respectively, under five-fold cross-validation. CONCLUSIONS: The robustness and effectiveness of our proposed model position it as a valuable approach for advancing the field of miRNA-abiotic stress association prediction.

The Significance of Insulinoma-Associated Protein 1 in the Pathological Diagnosis of Small-Cell Lung Cancer in Biopsy Specimens.

Objective: Our purpose was to investigate the clinicopathological diagnostic value of immunohistochemical antibody for insulinoma-associated protein 1 (INSM1) in biopsy specimens of SCLC. Methods: Biopsy specimens of SCLC diagnosed at the pathology department of Tangshan Gongren Hospital from January 2022 to June 2023 were selected. INSM1 expression was detected and compared with conventional neuroendocrine markers synaptophysin (SYP), chromogranin A (CHGA), and CD56 regarding expression sensitivity and specificity. Results: The sensitivity of INSM1 expression was significantly higher than that of CHGA (95% vs 50%, P = .000), but there was no statistically significant difference in the specificity of INSM1, SYP, CHGA, and CD56 expression (100% vs 94% vs 98% vs 92%, respectively, P = .241, 1.000, .126). Conclusions: INSM1 antibody shows high sensitivity and specificity in the expression of SCLC and serves as a reliable immunohistochemical marker in the clinicopathological diagnosis of SCLC in biopsy specimens.

Why Bufo gargarizans tadpoles grow bigger in Pb-contaminated environments? The gut microbiota matter.

The impacts of lead/Pb2+ on ecosystems have received widespread attention. Growth suppression is a major toxic effect of Pb compounds on aquatic animals, however, some studies have also reported their growth-promoting effects. These complex outcomes may be explained by anions that accompany Pb2+ or by the multiple toxic mechanisms/pathways of Pb2+. To examine these hypotheses, we tested how Bufo gargarizans tadpoles responded to Pb(NO3)2 (100 and 200 µg/L Pb2+) using transcriptomics and microbiomics, with NaNO3 and blank groups as controls. Tadpoles exposed to Pb(NO3)2 showed delayed development while increased somatic growth in a dose-dependent manner, which can be attributed to the effects of NO3- and Pb2+, respectively. Tadpole transcriptomics revealed that exposure to NO3- downregulated the MAPK pathway at transcriptional level, explaining the development-suppressing effect of NO3-; while Pb2+ upregulated the transcription of detoxification pathways (e.g., xenobiotics metabolism by cytochrome P450 and glutathione metabolism), indicating cellular stress and thus contradicting the growth advantage of Pb2+-exposed tadpoles. Pb2+ exposure changed the tadpole gut microbiota drastically, characterized by increased polysaccharides and carbohydrate utilization while decreased fatty acid and amino acid consumption according to microbial functional analysis. Similar gut microbial variations were observed in field-collected tadpoles from different Pb2+ environments. This metabolic shift in gut microbiota likely improved the overall food utilization efficiency and increased the allocation of fatty acids and amino acids to the host, explaining the growth advantage of Pb2+-exposed tadpoles. In summary, our results suggest multiple toxic pathways of Pb2+, and the gut microbiota may affect the pollution outcomes on animals.

Resection for malignant tumors in the elbow and individualized reconstruction under assistance of 3D printing technology: A case report.

RATIONALE: With a high failure rate and multiple postoperative complications, the resection for tumors in the elbow and reconstruction present a formidable challenge to orthopedic surgeons. The maturation of 3-dimension (3D) printing technology has facilitated the preoperative design, intraoperative navigation, and reconstruction of bone defects in patients with complex malignant tumors of the elbow joint. In order to improve prognosis, we explored a method of tumor resection and elbow reconstruction aided by 3D printing technology in this research. PATIENT CONCERNS: The patient underwent nephrectomy for clear cell carcinoma of the left kidney 3 years ago. Six months ago, the patient presented with limited movement and lateral tenderness in the right elbow joint. The tumor puncture biopsy demonstrated renal clear cell carcinoma metastasis. DIAGNOSES: Renal clear cell carcinoma with distal humerus bone metastasis. INTERVENTIONS: Thin-layer CT scan data of the patient was acquired, and a 3D reconstruction of both upper limb bones and joints was conducted, followed by a simulation of diseased tissue excision. According to the model, individualized osteotomy guidelines and elbow prostheses were designed and manufactured. Then, prior to the completion of the actual operation, a simulation of the preoperative phase was performed. OUTCOMES: The operation was completed without incident. At the 1-, 3-, and 6-month postoperative examinations, both the position and mobility of the prosthesis were found to be satisfactory, and no complications were observed. The hospital for special surgery score and mayo elbow performance score scores increased in comparison to the preoperative period. LESSONS: For patients with complex tumors in the elbow joint, 3D printing technology may assist in the precise excision of the tumor and provide an individualized elbow joint prosthesis that is more precise and effective than traditional surgery. It can accomplish a satisfactory treatment effect for patients when combined with early postoperative scientific rehabilitation training, so it is a method worth promoting.

Analysis, Occurrence and Exposure Evaluation of Antibiotic and Anthelmintic Residues in Whole Cow Milk from China.

An optimized QuEChERS method for the simultaneous extraction of 26 antibiotics and 19 anthelmintics in whole cow milk was established, followed by UHPLC-MS/MS analysis. Briefly, 20 mL acetonitrile with 1 g disodium hydrogen citrate, 2 g sodium citrate, 4 g anhydrous MgSO4, and 1 g sodium chloride were added to 10 g milk for target chemical extraction, followed by 50 mg anhydrous MgSO4 for purification. Satisfactory recoveries were obtained using the modified QuEChERS method, with recoveries of the antibiotics ranging from 79.7 to 117.2%, with the exception of norfloxacin, which was at 53.4%, while those for anthelmintics were in the range of 73.1-105.1%. The optimized QuEChERS method presented good precision, with relative standard deviations ranging from 7.2 to 18.6% for both antibiotics and anthelmintics. The method was successfully applied to analyze the antibiotics and anthelmintics in 56 whole cow milk samples from China. Briefly, the detection frequencies and concentrations of most of the antibiotics and anthelmintics were low in the whole cow milk samples, with concentrations ranging from below LOD to 4296.8 ng/kg. Fenbendazole, febantel, enrofloxacin, levofloxacin, sulfadiazine, and sulfamethoxazole were the predominant drug residues in the whole cow milk samples. Spatial distribution was found for those antibiotics and anthelmintics with detection frequency higher than 50%, especially for the antibiotics, indicating regional differences in drug application. Based on the current study, exposure to antibiotics and anthelmintics through whole cow milk consumption are lower than the acceptable daily intake values suggested by the China Institute of Veterinary Drug Control. However, long-term exposure to low doses of antibiotics and anthelmintics still needs attention and merits further study.

Size matters either way: Differently-sized microplastics affect amphibian host and symbiotic microbiota discriminately.

Concerns about the implications of microplastics (MPs) on aqueous animals have gained widespread attention. It has been postulated that the magnitude of MPs can influence its toxicity. However, little is known about how MPs toxicity changes with particle size. Amphibians are reliable bioindicators of ecosystem health due to their complex life cycles. In this study, we compared the influences of two sizes nonfunctionalized polystyrene microspheres (1 and 10 µm) on the metamorphosis of Asiatic toad (Bufo gargarizans). Acute exposure to MPs at high concentrations led to bioaccumulation in the digestive track and internal organs (i.e., liver and heart) of tadpoles. Long-term exposure to either size, at environmentally-related concentrations (1 and 4550 p/mL), led to growth and development delay in pro-metamorphic tadpoles. Remarkably, developmental plasticity mitigated these deleterious effects prior to the onset of metamorphic climax without compromising survival rate in later stages. MPs with a diameter of 10 µm dramatically altered the gut microbiota (e.g., abundance of Catabacter and Desulfovibrio) of pro-metamorphic tadpoles, whereas MPs with a diameter of 1 µm induced much more intensive transcriptional responses in the host tissues (e.g., upregulation of protein synthesis and mitochondrial energy metabolism, and downregulation of neural functions and cellular responses). Given that the two MPs sizes induced similar toxic outcomes, this suggests that their principal toxicity mechanisms are distinct. Small-sized MPs can travel easily across the intestinal mucosa and cause direct toxicity, while large-sized MPs accumulate in gut and affect the host by changing the homeostasis of digestive track. In conclusion, our findings indicate that MPs can affect the growth and development of amphibian larvae, but their developmental plasticity determines the ultimate detrimental effects. Multiple pathways of toxicity may contribute to the size-dependent toxicity of MPs. We anticipate that these findings will increase our understanding of the ecological effects of MPs.

Cascading effects of Pb on the environmental and symbiotic microbiota and tadpoles' physiology based on field data and laboratory validation.

Heavy metal pollution poses a serious threat to ecosystems. Currently, there is a lack of field data that would enable us to gain a systematic understanding of the influences of heavy metals on aquatic ecosystems, especially the interactions between environments and animals. We studied the relationships between the variations in heavy metal concentrations (10 species including Pb in sediments and surface water), the community structure of environmental and symbiotic microbiota, and the gut traits of Bufo gargarizans tadpoles across 16 sampling sites on the Chengdu Plain through rigorous statistical analysis and laboratory validation. The results show that heavy metal concentrations, especially the Pb concentration of the sediment, are linked to the variations in sediment and tadpoles' gut microbiomes but not to water microbiota. For the sediment microbiota, Pb causes a trade-off between the proportions of Burkholderiales and Verrucomicrobiae and affects the methane, sulfide, and nitrate metabolisms. For tadpoles, a high sediment Pb content leads to a low abundance of gut aerobic bacteria and a large relative gut weight under both field and laboratory conditions. In addition, Pb promotes the growth of B. gargarizans tadpoles under laboratory conditions. These effects seem to be beneficial to tadpoles. However, a high Pb content leads to a low abundance of probiotic bacteria (e.g., Verrucomicrobiae, Eubacteriaceae, and Cetobacterium) and a high abundance of pathogenic bacteria in the gut and environment, suggesting potential health risks posed by Pb. Interestingly, there is a causal relationship between Pb-induced variations in sediment and symbiotic microbiotas, and the latter is further linked to the variation in relative gut weight of tadpoles. This suggests a cascading effect of Pb on the ecosystem. In conclusion, our results indicate that among the heavy metals, the Pb in sediment is a critical factor affecting the aquatic ecosystem through an environment-gut-physiology pathway mediated by microbiota.

Albinism in the largest extant amphibian: A metabolic, endocrine, or immune problem?

Background: Pigment regression is an intriguing phenomenon that can be caused by disorders in melanin metabolism or endocrine regulation, or by autoimmune disorders. Albino animals serve as excellent models for the study of the genetic determination of morphology, particularly the evolution of and molecular mechanisms underlying chromatophore-related diseases in animals and humans. Material and Methods: The artificial culture of Andrias davidianus, the largest extant amphibian, is flourishing in China due to the great ecological and economic value of this animal. Approximately 0.1% of individuals express an albino phenotype accompanied by delayed somatic growth and mortality at early developmental stages. In this study, brain and skin transcriptomics were conducted to study the underlying molecular basis of the phenotype. Results: The results indicated decreased transcription of genes of melanin synthesis. Interestingly, MHC I isotypes and immune-related pathways accounted for the primary transcriptional differences between groups, suggesting that the albino phenotype represents a systematic immune problem to a far greater extent than a pigmentation defect. Albino individuals exhibited shifted transcription of MHC I isotypes, and the albino-specific isotype was characterized by increased charges and decreased space in the antigen- binding pocket, implying a drastic change in antigen specificity and a potential risk of autoimmune disorders. Conclusion: These results suggest an association between the albino phenotype and MHC I variants in A. davidianus, which could serve as a convenient model for vitiligo or other autoimmune diseases.

Multi-omics provide mechanistic insight into the Pb-induced changes in tadpole fitness-related traits and environmental water quality.

Water pollution from lead/Pb2+ poses a significant threat to aquatic ecosystems, and its repercussions on aquatic animals have received considerable attention. Although Pb2+ has been found to affect numerous aspects of animals, including individual fitness, metabolic status, and symbiotic microbiota, few studies have focused on the associations between Pb2+-induced variations in fitness, metabolome, symbiotic microbiome, and environmental parameters in the same system, limiting a comprehensive understanding of ecotoxicological mechanisms from a holistic perspective. Moreover, most ecotoxicological studies neglected the potential contributions of anions to the consequences generated by inorganic lead compounds. We investigated the effects of Pb(NO3)2 at environmentally relevant concentrations on the Rana omeimontis tadpoles and the water quality around them, using blank and NaNO3-treated groups as control. Results showed that Pb(NO3)2 not only induced a rise in water nitrite level, but exposure to this chemical also impaired tadpole fitness-related traits (e.g., growth and development). The impacts on tadpoles were most likely a combination of Pb2+ and NO3-. Tissue metabolomics revealed that Pb(NO3)2 exposure influenced animal substrate (i.e., carbohydrate, lipid, and amino acid) and prostaglandin metabolism. Pb(NO3)2 produced profound shifts in gut microbiota, with increased Proteobacteria impairing Firmicutes, resulting in higher aerobic and possibly pathogenic bacteria. NaNO3 also influenced tadpole metabolome and gut microbiome, in a manner different to that of Pb(NO3)2. The presence of NO3- seemed to counteract some changes caused by Pb2+, particularly on the microbiota. Piecewise structural equation model and correlation analyses demonstrated connections between tissue metabolome and gut microbiome, and the variations in tadpole phenotypic traits and water quality were linked to changes in tissue metabolome and gut microbiome. These findings emphasized the important roles of gut microbiome in mediating the effects of toxin on aquatic ecosystem. Moreover, it is suggested to consider the influences of anions in the risk assessment of heavy metal pollutions.

Rising floor and dropping ceiling: organ heterogeneity in response to cold acclimation of the largest extant amphibian.

Low temperature imposes strong selective pressure on ectotherms. To maximize their overall fitness under cold conditions, the ectotherms may either try to maintain their physiological activities through metabolic compensation or enter into metabolic depression; however, some species adopt both strategies to cope with different degrees of cold. Nevertheless, how these two seemingly opposite strategies are coordinated has rarely been elucidated. Here, we investigated the molecular strategy underlying the cold acclimation of Andrias davidianus, the largest extant amphibian, using multi-organ metabolomics and transcriptomics. The results showed remarkable organ heterogeneity in response to cold. While most organs showed transcriptional upregulation of metabolic processes, the heart exhibited downregulation. This heterogeneity explained the adaptive reorganization in resource allocation, which compensates for metabolic maintenance by compromising growth. Importantly, the cardiac function might constitute a 'ceiling' to constrain the space for compensation, especially under colder conditions. Additionally, the opposite transcriptional regulation of oxidative phosphorylation and other pathways might also shape the overall metabolic capacity under cold conditions. The heterogeneity in cold responses may have directed a shift in cold adaptive strategy from compensation to depression with a drop in temperature. These results provide a novel insight into the regulatory mechanisms underlying cold survival strategies of ectotherms.

Multi-Omics Approaches Revealed the Associations of Host Metabolism and Gut Microbiome With Phylogeny and Environmental Adaptation in Mountain Dragons.

The molecular basis enabling the adaptation of animals to spatially heterogeneous environments is a critical clue for understanding the variation, formation, and maintenance of biodiversity in the context of global climate change. Mountain dragons (Agamidae: Diploderma) thrive in the Hengduan Mountain Region, a biodiversity hotspot and a typical spatially heterogeneous environment. Here, we compare the liver and muscle metabolome and gut microbiome of 11 geographical populations from three Diploderma species (D. iadinum, D. yulongsense, and D. vela) after 7 days acclimation in the same laboratory conditions. Amino acid metabolism, particularly the products of the glutathione cycle, accounted for major interspecies variations, implying its significance in genetic differentiation among mountain dragons. Notably, the cold-dwelling D. vela and D. yulongense populations tended to have higher glycerophosphate, glycerol-3-phosphocholine, and kinetin levels in their liver, higher carnosine levels in their muscle, and higher Lachnospiraceae levels in their gut. Phylogeny, net primary productivity (NPP), and the temperature had the highest explanation rate to the variations in muscle metabolome, liver metabolome, and gut microbiome, respectively, suggesting heterogeneity of biological systems in response to climatic variations. Therefore, we suggested that the organ heterogeneity in environmental responsiveness might be substantial for mountain dragons to thrive in complicated environments.

Mesenchymal Stem Cell-derived Extracellular Vesicles Prevent Experimental Bronchopulmonary Dysplasia Complicated By Pulmonary Hypertension.

Mesenchymal stem cell (MSC) extracellular vesicles (EVs) have beneficial effects in preclinical bronchopulmonary dysplasia and pulmonary hypertension (BPD-PH) models. The optimal source, dosing, route, and duration of effects are however unknown. The objectives of this study were to (a) compare the efficacy of GMP-grade EVs obtained from Wharton's Jelly MSCs (WJ-MSCs) and bone marrow (BM-MSCs), (b) determine the optimal dosing and route of administration, (c) evaluate its long-term effects, and (d) determine how MSC EVs alter the lung transcriptome. Newborn rats exposed to normoxia or hyperoxia (85% O2) from postnatal day (P)1-P14 were given (a) intra-tracheal (IT) BM or WJ-MSC EVs or placebo, (b) varying doses of IT WJ-MSC EVs, or (c) IT or intravenous (IV) WJ-MSC EVs on P3. Rats were evaluated at P14 or 3 months. Early administration of IT BM-MSC or WJ-MSC EVs had similar beneficial effects on lung structure and PH in hyperoxia-exposed rats. WJ-MSC EVs however had superior effects on cardiac remodeling. Low, medium, and high dose WJ-MSC EVs had similar cardiopulmonary regenerative effects. IT and IV WJ-MSC EVs similarly improved vascular density and reduced PH in hyperoxic rats. Gene-set enrichment analysis of transcripts differentially expressed in WJ-MSC EV-treated rats showed that induced transcripts were associated with angiogenesis. Long-term studies demonstrated that a single early MSC EV dose has pulmonary vascular protective effects 3 months after administration. Together, our findings have significant translational implications as it provides critical insight into the optimal source, dosing, route, mechanisms of action, and duration of effects of MSC-EVs for BPD-PH.

Effects of Habitat River Microbiome on the Symbiotic Microbiota and Multi-Organ Gene Expression of Captive-Bred Chinese Giant Salamander.

The reintroduction of captive-bred individuals is a primary approach to rebuild the wild populations of the Chinese giant salamander (Andrias davidianus), the largest extant amphibian species. However, the complexity of the wild habitat (e.g., diverse microorganisms and potential pathogens) potentially threatens the survival of reintroduced individuals. In this study, fresh (i.e., containing environmental microbiota) or sterilized river sediments (120°C sterilized treatment) were added to the artificial habitats to treat the larvae of the Chinese giant salamander (control group-Cnt: 20 individuals, treatment group 1 with fresh river sediments-T1: 20 individuals, and treatment group 2 with sterilized river sediments-T2: 20 individuals). The main objective of this study was to test whether this procedure could provoke their wild adaptability from the perspective of commensal microbiotas (skin, oral cavity, stomach, and gut) and larvae transcriptomes (skin, spleen, liver, and brain). Our results indicated that the presence of habitat sediments (whether fresh or sterilized) reshaped the oral bacterial community composition. Specifically, Firmicutes decreased dramatically from ~70% to ~20-25% (mainly contributed by Lactobacillaceae), while Proteobacteria increased from ~6% to ~31-36% (mainly contributed by Gammaproteobacteria). Consequently, the proportion of antifungal operational taxonomic units (OTUs) increased, and the function of oral microbiota likely shifted from growth-promoting to pathogen defense. Interestingly, the skin microbiota, rather than the colonization of habitat microbiota, was the major source of the pre-treated oral microbiota. From the host perspective, the transcriptomes of all four organs were changed for treated individuals. Specifically, the proteolysis and apoptosis in the skin were promoted, and the transcription of immune genes was activated in the skin, spleen, and liver. Importantly, more robust immune activation was detected in individuals treated with sterilized sediments. These results suggested that the pathogen defense of captive-bred individuals was improved after being treated, which may benefit their survival in the wild. Taken together, our results suggested that the pre-exposure of captive-bred Chinese giant salamander individuals to habitat sediments could be considered and added into the reintroduction processes to help them better adapt to wild conditions.

From Water to Land: The Structural Construction and Molecular Switches in Lungs during Metamorphosis of Microhyla fissipes.

Most anurans must undergo metamorphosis to adapt to terrestrial life. This process enhances the air-breathing ability of the lungs to cope with the change in oxygen medium from water to air. Revealing the structural construction and molecular switches of lung organogenesis is essential to understanding the realization of the air-breathing function. In this study, histology and transcriptomics were conducted in combination to explore these issues in Microhyla fissipes' lungs during metamorphosis. During the pro-metamorphic phase, histological structural improvement of the alveolar wall is accompanied by robust substrate metabolism and protein turnover. The lungs, at the metamorphic climax phase, are characterized by an increased number of cilia in the alveolar epithelial cells and collagenous fibers in the connective tissues, corresponding to the transcriptional upregulation of cilia and extracellular matrix-related genes. Post-metamorphic lungs strengthen their contracting function, as suggested by the thickened muscle layer and the upregulated expression of genes involved in muscle contraction. The blood-gas barrier is fully developed in adult lungs, the transcriptional features of which are tissue growth and regulation of differentiation and immunity. Importantly, significant transcriptional switches of pulmonary surfactant protein and hemoglobin facilitate air breathing. Our results illuminated four key steps of lung development for amphibians to transition from water to land.

Temperature and Diet Acclimation Modify the Acute Thermal Performance of the Largest Extant Amphibian.

The Chinese giant salamander (Andrias davidianus), one of the largest extant amphibian species, has dramatically declined in the wild. As an ectotherm, it may be further threatened by climate change. Therefore, understanding the thermal physiology of this species should be the priority to formulate related conservation strategies. In this study, the plasticity in metabolic rate and thermal tolerance limits of A. davidianus larvae were studied. Specifically, the larvae were acclimated to three temperature levels (7 °C, cold stress; 15 °C, optimum; and 25 °C, heat stress) and two diet items (red worm or fish fray) for 20 days. Our results indicated that cold-acclimated larvae showed increased metabolic capacity, while warm-acclimated larvae showed a decrease in metabolic capacity. These results suggested the existence of thermal compensation. Moreover, the thermal tolerance windows of cold-acclimated and warm-acclimated larvae shifted to cooler and hotter ranges, respectively. Metabolic capacity is not affected by diet but fish-fed larvae showed superiority in both cold and heat tolerance, potentially due to the input of greater nutrient loads. Overall, our results suggested a plastic thermal tolerance of A. davidianus in response to temperature and diet variations. These results are meaningful in guiding the conservation of this species.

Commensal microbiota and host metabolic divergence are associated with the adaptation of Diploderma vela to spatially heterogeneous environments.

Heterogeneous environment adaptation is critical to understand the species evolution and response to climate change. However, how narrow-range species adapt to micro-geographic heterogeneity has been overlooked, and there is a lack of insights from metabolism and commensal microbiota. Here, we studied the environmental adaptation for 3 geographic populations (>40 km apart) of Diploderma vela, a lizard endemic to dry-hot valleys of the Hengduan Mountain Region. The climatic boundary caused a cooler, droughtier, and barren environment for northernmost population (RM) than the middle (QZK) and southernmost populations (FS). Correspondingly, significant divergences in liver and muscle metabolism and commensal microbiota were detected between RM and QZK or FS individuals, but not between QZK and FS individuals. Phospholipid composition, coenzyme level (i.e. pyridoxal and NAD+ ), and cholesterol metabolism (e.g. androgen and estriol synthesis) constituted the major metabolic difference between RM and QZK/FS groups. FS and QZK individuals kept abundant Proteobacteria and antifungal strains, while RM individuals maintained more Firmicutes and Bacteroidota. Strong associations existed between varied host metabolite and gut microbes. How were these interpopulation variations associated to the environment adaptation were discussed. These results provided some novel insights into the environmental adaptation and implicated the consequence of climate change on narrow-range species.