RESUMO
The spindle checkpoint complex is a key surveillance mechanism in cell division that prevents premature separation of sister chromatids. Mad2 is an integral component of this spindle checkpoint complex that recognizes cognate substrates such as Mad1 and Cdc20 in its closed (C-Mad2) conformation by fastening a "seatbelt" around short peptide regions that bind to the substrate recognition site. Mad2 is also a metamorphic protein that adopts not only the fold found in C-Mad2, but also a structurally distinct open conformation (O-Mad2) which is incapable of binding substrates. Here, we show using chemical exchange saturation transfer (CEST) and relaxation dispersion (CPMG) NMR experiments that Mad2 transiently populates three other higher free energy states with millisecond lifetimes, two in equilibrium with C-Mad2 (E1 and E2) and one with O-Mad2 (E3). E1 is a mimic of substrate-bound C-Mad2 in which the N-terminus of one C-Mad2 molecule inserts into the seatbelt region of a second molecule of C-Mad2, providing a potential pathway for autoinhibition of C-Mad2. E2 is the "unbuckled" conformation of C-Mad2 that facilitates the triage of molecules along competing fold-switching and substrate binding pathways. The E3 conformation that coexists with O-Mad2 shows fluctuations at a hydrophobic lock that is required for stabilizing the O-Mad2 fold and we hypothesize that E3 represents an early intermediate on-pathway towards conversion to C-Mad2. Collectively, the NMR data highlight the rugged free energy landscape of Mad2 with multiple low-lying intermediates that interlink substrate-binding and fold-switching, and also emphasize the role of molecular dynamics in its function.
RESUMO
Macrobrachium nipponense is an important commercial freshwater species in China. However, the ability of alkali tolerance of M. nipponense is insufficient to culture in the major saline-alkali water source in China. Thus, it is urgently needed to perform the genetic improvement of alkali tolerance in this species. In the present study, we aimed to analyse the effects of alkali treatment on gills in this species after 96 h alkalinity exposure under the alkali concentrations of 0 mmol/L, 4 mmol/L, 8 mmol/L, and 12 mmol/L through performing the histological observations, measurement of antioxidant enzymes, metabolic profiling analysis, and transcriptome profiling analysis. The results of the present study revealed that alkali treatment stimulated the contents of malondialdehyde, glutathione, glutathione peroxidase in gills, indicating these antioxidant enzymes plays essential roles in the protection of body from the damage, caused by the alkali treatment. In addition, high concentration of alkali treatment (> 8 mmol/L) resulted in the damage of gill membrane and haemolymph vessel, affecting the normal respiratory function of gill. Metabolic profiling analysis revealed that Metabolic pathways, Biosynthesis of secondary metabolites, Biosynthesis of plant secondary metabolites, Microbial metabolism in diverse environments, Biosynthesis of amino acids were identified as the main enriched metabolic pathways of differentially expressed metabolites, which are consistent with the previous publications, treated by the various environmental factors. Transcriptome profiling analyses revealed that the alkali concentration of 12 mmol/L has more regulatory effects on the changes of gene expression than the other alkali concentrations. KEGG analysis revealed that Phagosome, Lysosome, Glycolysis/Gluconeogenesis, Purine Metabolism, Amino sugar and nucleotide sugar metabolism, and Endocytosis were identified as the main enriched metabolic pathways in the present study, predicting these metabolic pathways may be involved in the adaption of alkali treatment in M. nipponense. Phagosome, Lysosome, Purine Metabolism, and Endocytosis are immune-related metabolic pathways, while Glycolysis/Gluconeogenesis, and Amino sugar and nucleotide sugar metabolism are energy metabolism-related metabolic pathways. Quantitative PCR analyses of differentially expressed genes (DEGs) verified the accuracy of the RNA-Seq. Alkali treatment significantly stimulated the expressions of DEGs from the metabolic pathways of Phagosome and Lysosome, suggesting Phagosome and Lysosome play essential roles in the regulation of alkali tolerance in this species, as well as the genes from these metabolic pathways. The present study identified the effects of alkali treatment on gills, providing valuable evidences for the genetic improvement of alkali tolerance in M. nipponense.
Assuntos
Álcalis , Brânquias , Palaemonidae , Animais , Brânquias/metabolismo , Brânquias/efeitos dos fármacos , Palaemonidae/genética , Palaemonidae/efeitos dos fármacos , Palaemonidae/metabolismo , Perfilação da Expressão Gênica , Transcriptoma/efeitos dos fármacos , Redes e Vias Metabólicas/efeitos dos fármacosRESUMO
Solution NMR spectroscopy is a particularly powerful technique for characterizing the functional dynamics of biomolecules, which is typically achieved through the quantitative characterization of chemical exchange processes via the measurement of spin relaxation rates. In addition to the conventional nuclei such as 15N and 13C, which are abundant in biomolecules, fluorine-19 (19F) has recently garnered attention and is being widely used as a site-specific spin probe. While 19F offers the advantages of high sensitivity and low background, it can be susceptible to artifacts in quantitative relaxation analyses due to a multitude of dipolar and scalar coupling interactions with nearby 1H spins. In this study, we focused on the ribose 2'-19F spin probe in nucleic acids and investigated the effects of 1H-19F spin interactions on the quantitative characterization of slow exchange processes on the millisecond time scale. We demonstrated that the 1H-19F dipolar coupling can significantly affect the interpretation of 19F chemical exchange saturation transfer (CEST) experiments when 1H decoupling is applied, while the 1H-19F interactions have a lesser impact on Carr-Purcell-Meiboom-Gill relaxation dispersion applications. We also proposed a modified CEST scheme to alleviate these artifacts along with experimental verifications on self-complementary RNA systems. The theoretical framework presented in this study can be widely applied to various 19F spin systems where 1H-19F interactions are operative, further expanding the utility of 19F relaxation-based NMR experiments.
RESUMO
Hagfishes are characterized by omo- and iono-conforming nature similar to marine invertebrates. Conventionally, hagfishes had been recognized as the most primitive living vertebrate that retains plesiomorphic features. However, some of the "ancestral" features of hagfishes, such as rudimentary eyes and the lack of vertebrae, have been proven to be deceptive. Similarly, by the principle of maximum parsimony, the unique body fluid regulatory strategy of hagfishes seems to be apomorphic, since the lamprey, another cyclostome, adopts osmo- and iono-regulatory mechanisms as in jawed vertebrates. Although hagfishes are unequivocally important in discussing the origin and evolution of the vertebrate osmoregulatory system, the molecular basis for the body fluid homeostasis in hagfishes has been poorly understood. In the present study, we explored this matter in the inshore hagfish, Eptatretus burgeri, by analyzing the transcriptomes obtained from the gill, kidney, and muscle of the animals acclimated to distinct environmental salinities. Together with the measurement of parameters in the muscular fluid compartment, our data indicate that the hagfish possesses an ability to conduct free amino acid (FAA)-based osmoregulation at a cellular level, which is in coordination with the renal and branchial FAA absorption. We also revealed that the hagfish does possess the orthologs of the known osmoregulatory genes and that the transepithelial movement of inorganic ions in the hagfish gill and kidney is more complex than previously thought. These observations pose a challenge to the conventional view that the physiological features of hagfishes have been inherited from the last common ancestor of the extant vertebrates.
Assuntos
Brânquias , Feiticeiras (Peixe) , Osmorregulação , Animais , Feiticeiras (Peixe)/genética , Feiticeiras (Peixe)/fisiologia , Osmorregulação/genética , Brânquias/metabolismo , Rim/metabolismo , Salinidade , Transcriptoma , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Equilíbrio Hidroeletrolítico , Aminoácidos/metabolismo , Aclimatação/genéticaRESUMO
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.
RESUMO
The occurrence of regeneration of the organs involved in respiratory gas exchange amongst vertebrates is heterogeneous. In some species of amphibians and fishes, the gills regenerate completely following resection or amputation, whereas in mammals, only partial, facultative regeneration of lung tissue occurs following injury. Given the homology between gills and lungs, the capacity of gill regeneration in aquatic species is of major interest in determining the underlying molecular or signalling pathways involved in respiratory organ regeneration. In the present study, we used adult zebrafish (Danio rerio) to characterize signalling pathways involved in the early stages of gill regeneration. Regeneration of the gills was induced by resection of gill filaments and observed over a period of up to 10â days. We screened for the effects on regeneration of the drugs SU5402, dorsomorphin and LY411575, which inhibit FGF, BMP or Notch signalling pathways, respectively. Exposure to each drug for 5â days significantly reduced regrowth of filament tips in regenerating tissue, compared with unresected controls. In separate experiments under normal conditions of regeneration, we used reverse transcription quantitative PCR and observed an increased expression of genes encoding for the bone morphogenetic factor, Bmp2b, fibroblast growth factor, Fgf8a, a transcriptional regulator (Her6) involved in Notch signalling, and Sonic Hedgehog (Shha), in regenerating gills at 10â day post-resection, compared with unresected controls. In situ hybridization confirmed that all four genes were expressed in regenerating gill tissue. This study implicates BMP, FGF, Notch and Shh signalling in gill regeneration in zebrafish.
Assuntos
Brânquias , Peixe-Zebra , Animais , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Brânquias/metabolismo , Proteínas Hedgehog , Transdução de Sinais/genética , Fatores de Crescimento de Fibroblastos/genética , Fatores de Crescimento de Fibroblastos/metabolismo , Proteínas de Peixe-Zebra/genética , Mamíferos/metabolismoRESUMO
Natural temperature variation in many marine ecosystems is stochastic and unpredictable, and climate change models indicate that this thermal irregularity is likely to increase. Temperature acclimation may be more challenging when conditions are highly variable and stochastic, and there is a need for empirical physiological data in these thermal environments. Using the hermaphroditic, amphibious mangrove rivulus (Kryptolebias marmoratus), we hypothesized that compared with regular, warming diel thermal fluctuations, stochastic warm fluctuations would negatively affect physiological performance. To test this, we acclimated fish to: (1) non-stochastic and (2) stochastic thermal fluctuations with a similar thermal load (27-35°C), and (3) a stable/consistent control temperature at the low end of the cycle (27°C). We determined that fecundity was reduced in both cycles, with reproduction ceasing in stochastic thermal environments. Fish acclimated to non-stochastic thermal cycles had growth rates lower than those of control fish. Exposure to warm, fluctuating cycles did not affect emersion temperature, and only regular diel cycles modestly increased critical thermal tolerance. We predicted that warm diel cycling temperatures would increase gill surface area. Notably, fish acclimated to either thermal cycle had a reduced gill surface area and increased intralamellar cell mass when compared with control fish. This decreased gill surface area with warming contrasts with what is observed for exclusively aquatic fish and suggests a preparatory gill response for emersion in these amphibious fish. Collectively, our data reveal the importance of considering stochastic thermal variability when studying the effects of temperature on fishes.
Assuntos
Aclimatação , Brânquias , Processos Estocásticos , Animais , Brânquias/fisiologia , Aclimatação/fisiologia , Ciprinodontiformes/fisiologia , Temperatura , Mudança Climática , Organismos Hermafroditas/fisiologia , Temperatura AltaRESUMO
In studies of infectious disease prevention, the level of protective efficacy of medicinal products such as vaccines and prophylactic drugs tends to vary over time. Many products require administration of multiple doses at scheduled times, as opposed to one-off or continual intervention. Accurate information on the trajectory of the level of protective efficacy over time facilitates informed clinical recommendations and implementation strategies, for example, with respect to the timing of administration of the doses. Based on concepts from pharmacokinetic and pharmacodynamic modeling, we propose a non-linear function for modeling the trajectory after each dose. The cumulative effect of multiple doses of the products is captured by an additive series of the function. The model has the advantages of parsimony and interpretability, while remaining flexible in capturing features of the trajectories. We incorporate this series into the Andersen-Gill model for analysis of recurrent event time data and compare it with alternative parametric and non-parametric functions. We use data on clinical malaria disease episodes from a trial of four doses of an anti-malarial drug combination for chemoprevention to illustrate, and evaluate the performance of the methods using simulation. The proposed method out-performed the alternatives in the analysis of real data in terms of Akaike and Bayesian Information Criterion. It also accurately captured the features of the protective efficacy trajectory such as the area under curve in simulations. The proposed method has strong potential to enhance the evaluation of disease prevention measures and improve their implementation strategies.
Assuntos
Antimaláricos , Doenças Transmissíveis , Malária , Humanos , Teorema de Bayes , Malária/tratamento farmacológico , Simulação por ComputadorRESUMO
Mytilus unguiculatus is an important economic bivalve species with wide consumption and aquaculture value. Disease is one of the primary limiting factors in mussel aquaculture, thus understanding the response of different tissues of M. unguiculatus to pathogens is crucial for disease prevention and control. In this study, we investigated the physiological and transcriptomic responses of the gills, adductor muscle, and mantle of M. unguiculatus infected with Vibrio alginolyticus. The results showed that V. alginolyticus infection caused inflammation and tissue structure changes in the gill, adductor muscle and mantle of M. unguiculatus. Meanwhile, the activities of superoxide dismutase and catalase in the three tissues increased, while the total antioxidant capacity decreased, suggesting that M. unguiculatus have an activated defense mechanism against infection-induced oxidative stress, despite a compromised total antioxidant capacity. Transcriptomic studies reveal that infected M. unguiculatus exhibits upregulation of endocytosis, lysosome activity, cellular apoptosis, and immune-related signaling pathways, indicating that M. unguiculatus responds to pathogen invasion by upregulating efferocytosis. Compared with the gill and adductor muscle, the mantle had a higher level of mytimycin, mytilin and myticin, and the three tissues also increased the expression of mytimycin to cope with the invasion of pathogens. In addition, the analysis of genes related to taste transduction pathways and muscle contraction and relaxation found that after infection with V. alginolyticus, M. unguiculatus may reduce appetite by inhibiting taste transduction in the gill, while improving muscle contraction of the adductor muscle and keeping the shell closed, to resist further invasion of pathogens and reduce the risk of pathogen transmission in the population.
Assuntos
Mytilus , Vibrioses , Vibrio , Animais , Mytilus/genética , Vibrio alginolyticus/fisiologia , Antioxidantes , Vibrioses/veterinária , Perfilação da Expressão Gênica/veterinária , Vibrio/fisiologiaRESUMO
The immune system of bony fish closely resembles that of mammals, comprising both specific (adaptive) and non-specific (innate) components. Notably, the mucosa-associated lymphoid tissue (MALT) serves as the first line of defense within the non-specific immune system, playing a critical role in protecting these aquatic organisms against invading pathogens. MALT encompasses a network of immune cells strategically distributed throughout the gills and intestines, forming an integral part of the mucosal barrier that interfaces directly with the surrounding aquatic environment. Spring Viremia of Carp Virusï¼SVCVï¼, a highly pathogenic agent causing substantial harm to common carp populations, has been designated as a Class 2 animal disease by the Ministry of Agriculture and Rural Affairs of China. Utilizing a comprehensive array of research techniques, including Hematoxylin and Eosin (HE)ãAlcian Blue Periodic Acid-Schiff (AB-PAS)ãtranscriptome analysis for global gene expression profiling and Reverse Transcription-Polymerase Chain Reaction (RT-qPCR), this study uncovered several key findings: SVCV is capable of compromising the mucosal architecture in the gill and intestinal tissues of carp, and stimulate the proliferation of mucous cells both in gill and intestinal tissues. Critically, the study revealed that SVCV's invasion elicits a robust response from the carp's mucosal immune system, demonstrating the organism's capacity to resist SVCV invasion despite the challenges posed by the pathogen.
Assuntos
Carpas , Doenças dos Peixes , Perfilação da Expressão Gênica , Brânquias , Intestinos , Infecções por Rhabdoviridae , Rhabdoviridae , Animais , Brânquias/imunologia , Brânquias/virologia , Rhabdoviridae/fisiologia , Doenças dos Peixes/imunologia , Doenças dos Peixes/virologia , Carpas/imunologia , Carpas/genética , Perfilação da Expressão Gênica/veterinária , Infecções por Rhabdoviridae/imunologia , Infecções por Rhabdoviridae/veterinária , Infecções por Rhabdoviridae/virologia , Intestinos/imunologia , Intestinos/virologia , Imunidade Inata/genética , Transcriptoma/imunologia , Imunidade nas MucosasRESUMO
Cobia (Rachycentron canadum), a commercially important marine fish, has been used to develop a novel gill cell line, designated CG, for the first time. The CG cell line was cultured in Leibovitz's-15 medium with 5% fetal bovine serum (FBS) and successfully sub-cultured more than 110 passages. It underwent verification through sequencing of the mitochondrial cytochrome C oxidase subunit I (COI) gene. Optimal growth rate was achieved when the CG cell line was cultured in a medium supplemented with 5% FBS, 1% Penicillin-Streptomycin (P/S), and 5 parts per thousand (ppt) of coral sea salt water, maintained at a temperature of 27 °C. The addition of 5 ppt of salt in the growth medium suggests that this cell line could be a viable in vitro tool for marine ecosystem toxicological studies or for culturing marine parasitic microorganisms. The CG cell line was also successfully transfected using the pTurbo-GFP plasmids, showing an 18% efficiency, with observable GFP expression. Furthermore, the cell line has been effectively cryopreserved. Gene expression analysis indicated that the CG cell line exhibits responsive regulation of immune gene expression when exposured to various stimulants, highlighting its potential as an in vitro platform for immune response studies. This makes it suitable for exploring dynamic immune signaling pathways and host-pathogen interactions, thereby offering valuable insights for therapeutic development.
Assuntos
Brânquias , Perciformes , Animais , Ecossistema , Perciformes/metabolismo , Linhagem Celular , ImunidadeRESUMO
Mobulid species are endangered globally, and the market trade for gill plates is believed to be a major threat. Successful conservation and the sustainable use of mobulids therefore require an objective understanding of consumer characteristics and preferences for gill plates. Previous studies focused on qualitative descriptions, and reliable quantitative analyses are currently lacking. We used a latent class choice experiment method and a semistructured questionnaire to provide important new quantitative information about gill plate consumer characteristics and the heterogeneous nature of demand for gill plates. From May to July 2019, we conducted a field study in Guangzhou, the primary consumption hub for mobulid gill plates in mainland China. Utilizing a simple random sampling method, we engaged in face-to-face interviews with 428 consumers of gill plates in the major trading markets in Guangzhou. Our results showed that 59.8% of consumers of gill plates were over 40 years old, 62.6% were female, 80.7% had annual household incomes of <200,000 yuan, and 84.5% recognized the medical and health value of gill plates and purchased them. About seventy-two percent of consumers preferred to purchase imported and less expensive gill plates from unprotected species, but they had a strong preference for large gill plates from protected species, such as Mobula birostris. This contradiction arose from consumers' lack of knowledge of mobulids and their conservation status. We found, for example, female consumers over 40 years old had the least understanding of conservation status of mobulid species and the link between size of gill plates and rarity of mobulids. This suggests there may be opportunities to promote mobulid conservation through education and marketing targeted at this demographic. Consumers who had a positive preference for gill plates from protected species (regardless of price) (10%) may be harder to influence. Overall, we believe education alone is not enough and that the conservation of mobulids would benefit from an integrated approach that involves conservation education and strengthened trade regulations, such as the introduction of traceability systems and a stiffer legal framework for consumption of protected species.
Características y preferencias de los consumidores de placas branquiales de mobúlidos en China Resumen Las especies de mobúlidos están en peligro de extinción en todo el mundo y se considera al mercado de placas branquiales como una amenaza principal. Por lo tanto, la conservación exitosa y el uso sustentable de los mobúlidos requiere del entendimiento objetivo de las características y preferencias de los consumidores de estas placas branquiales. Los estudios previos se han enfocado en descripciones cualitativas, por lo que actualmente no hay suficientes análisis cuantitativos confiables. Usamos un método de experimento de elección de clase latente y un cuestionario semiestructurado para proporcionar información cuantitativa nueva e importante sobre las características de los consumidores de placas branquiales y la naturaleza heterogénea de la demanda de estas placas. Realizamos un estudio de campo entre mayo y julio de 2019 en Guangzhou, el principal centro de consumo de placas branquiales de mobúlidos en el interior de China. Utilizamos un método de muestreo aleatorio simple para entrevistar caraacara a 428 consumidores de placas branquiales en los principales mercados de Guangzhou. Nuestros resultados mostraron que el 59.8% de los consumidores son mayores a 40 años, 62.6% son mujeres, 80.7% tienen un ingreso doméstico anual mayor a 200,000 yuan y 84.5% reconocieron el valor médico y para la salud que tienen las placas branquiales, razones por las que las compran. El 72% de los consumidores prefirió comprar las placas importadas y menos caras de especies no protegidas, aunque tuvieron una mayor preferencia por las placas más grandes de las especies protegidas, como Mobula birostris. Esta contradicción se debe a la falta de conocimiento que tienen los consumidores sobre los mobúlidos y su estado de conservación. Descubrimos que, por ejemplo, las consumidoras de más de 40 años tienen el menor conocimiento del estado de conservación de los mobúlidos y la conexión entre el tamaño de las placas branquiales y la rareza de la especie. Lo anterior sugiere que podría haber oportunidad de promover la conservación de los mobúlidos por medio de la educación y la mercadotecnia enfocada en este grupo demográfico. Podría ser más difícil influir sobre el 10 % de los consumidores, el cual tiene una preferencia positiva por las placas branquiales de las especies protegidas (sin importar el precio). En general creemos que la educación por sí sola no es suficiente y que la conservación de los mobúlidos se beneficiaría de una estrategia integrada que involucre la educación para la conservación y regulaciones de mercado fortalecidas, como la introducción de los sistemas de trazabilidad y un marco legal más rígido para el consumo de las especies protegidas.
Assuntos
Conservação dos Recursos Naturais , Comportamento do Consumidor , China , Animais , Adulto , Feminino , Masculino , Conservação dos Recursos Naturais/métodos , Humanos , Pessoa de Meia-Idade , Espécies em Perigo de ExtinçãoRESUMO
Fish gills are highly sensitive organs for microplastic (MP) and nanoplastic (NP) invasions, but the cellular heterogeneity of fish gills to MPs and NPs remains largely unknown. We employed single-cell RNA sequencing to investigate the responses of individual cell populations in tilapia Oreochromis niloticus gills to MP and NP exposure at an environmentally relevant concentration. Based on the detected differentially expressed gene (DEG) numbers, the most affected immune cells by MP exposure were macrophages, while the stimulus of NPs primarily targeted T cells. In response to MPs and NPs, H+-ATPase-rich cells exhibited distinct changes as compared with Na+/K+-ATPase-rich cells and pavement cells. Fibroblasts were identified as a potential sensitive cell-type biomarker for MP interaction with O. niloticus gills, as evidenced by the largely reduced cell counts and the mostly detected DEGs among the 12 identified cell populations. The most MP-sensitive fibroblast subpopulation in O. niloticus gills was lipofibroblasts. Cell-cell communications between fibroblasts and H+-ATPase-rich cells, neurons, macrophages, neuroepithelial cells, and Na+/K+-ATPase-rich cells in O. niloticus gills were significantly inhibited by MP exposure. Collectively, our study demonstrated the cellular heterogeneity of O. niloticus gills to MPs and NPs and provided sensitive markers for their toxicological mechanisms at single-cell resolution.
Assuntos
Microplásticos , Plásticos , Animais , Microplásticos/toxicidade , Brânquias , ATPases Translocadoras de Prótons , Análise de Sequência de RNARESUMO
The compound Honokiol, derived from the bark of Magnolia officinalis, possesses the ability to induce apoptosis and inhibit cellular damage caused by reactive oxygen species. The objective of this study was to investigate the toxicological and histopathological effects of Honokiol on zebrafish (Danio rerio) through conducting a semistatic acute toxicity test involving immersion in an Honokiol-containing solution. The results showed that the toxic effects of Honokiol on zebrafish were primarily manifested in the liver and gills. When exposed to 0.6 mg/L of Honokiol, it could lead to liver hemorrhage as well as swelling and necrosis of gill tissues, and high concentrations of Honokiol could trigger inflammatory responses. Additionally, research found that Honokiol could induce apoptosis in liver and gill tissues through the P53 pathway and possessed the ability to enhance antioxidation. The present findings significantly contribute to a more profound understanding of the toxic impact of Honokiol and its underlying mechanism, thereby providing a valuable reference for the future safe utilization of Honokiol and related pharmaceutical advancements.
Assuntos
Apoptose , Compostos de Bifenilo , Lignanas , Fígado , Peixe-Zebra , Lignanas/farmacologia , Lignanas/toxicidade , Animais , Compostos de Bifenilo/toxicidade , Fígado/efeitos dos fármacos , Fígado/patologia , Apoptose/efeitos dos fármacos , Brânquias/efeitos dos fármacos , Brânquias/patologia , Proteína Supressora de Tumor p53/metabolismo , Magnolia/química , Compostos Alílicos , FenóisRESUMO
BACKGROUND: Understanding the relationship between resident microbiota and disease in cultured fish represents an important and emerging area of study. Marine gill disorders in particular are considered an important challenge to Atlantic salmon (Salmo salar) aquaculture, however relatively little is known regarding the role resident gill microbiota might play in providing protection from or potentiating different gill diseases. Here, 16S rRNA sequencing was used to examine the gill microbiome alongside fish health screening in farmed Atlantic salmon. Results were used to explore the relationship between microbial communities and gill disease. RESULTS: Microbial community restructuring was observed throughout the sampling period and linked to varied drivers of change, including environmental conditions and severity of gill pathology. Taxa with significantly greater relative abundance on healthier gills included isolates within genus Shewanella, and taxa within family Procabacteriaceae. In contrast, altered abundance of Candidatus Branchiomonas and Rubritalea spp. were associated with damaged gills. Interestingly, more general changes in community richness and diversity were not associated with altered gill health, and thus not apparently deleterious to fish. Gross and histological gill scoring demonstrated seasonal shifts in gill pathology, with increased severity of gill damage in autumn. Specific infectious causes that contributed to observed pathology within the population included the gill disorder amoebic gill disease (AGD), however due to the uncontrolled nature of this study and likely mixed contribution of various causes of gill disease to observed pathology results do not strongly support an association between the microbial community and specific infectious or non-infectious drivers of gill pathology. CONCLUSIONS: Results suggest that the microbial community of farmed Atlantic salmon gills undergo continual restructuring in the marine environment, with mixed influences upon this change including environmental, host, and pathogenic factors. A significant association of specific taxa with different gill health states suggests these taxa might make meaningful indicators of gill health. Further research with more frequent sampling and deliberate manipulation of gills would provide important advancement of knowledge in this area. Overall, although much is still to be learnt regarding what constitutes a healthy or maladapted gill microbial community, the results of this study provide clear advancement of the field, providing new insight into the microbial community structure of gills during an annual production cycle of marine-stage farmed Atlantic salmon.
Assuntos
Aquicultura , Doenças dos Peixes , Brânquias , Microbiota , Salmo salar , Animais , Salmo salar/microbiologia , Brânquias/microbiologia , Brânquias/patologia , Doenças dos Peixes/microbiologia , Doenças dos Peixes/patologia , RNA Ribossômico 16S/genética , Estações do Ano , Bactérias/classificação , Bactérias/isolamento & purificação , Bactérias/genética , AmebíaseRESUMO
Free-living amoebae (FLA) are capable of inhabiting diverse reservoirs independently, without relying on a host organism, hence their designation as "free-living". The majority of amoebae that infect freshwater or marine fish are amphizoic, or free-living forms that may colonize fish under particular circumstances. Symphysodon aequifasciatus, commonly referred to as the discus, is widely recognized as a popular ornamental fish species. The primary objective of the present study was to determine the presence of pathogenic free-living amoebae (FLA) in samples of discus fish. Fish exhibiting clinical signs, sourced from various fish farms, were transferred to the ornamental fish clinic. The skin, gills, and intestinal mucosa of the fish were collected and subjected to culturing on plates containing a 1% non-nutrient agar medium. The detection of FLA was conducted through morphological, histopathological and molecular methods. The construction of the phylogenetic tree for Acanthamoeba genotypes was achieved using the maximum likelihood approach. The molecular sequence analysis revealed that all cultures that tested positive for FLA were T4 genotype of Acanthamoeba and Acanthamoeba sp. The examination of gill samples using histopathological methods demonstrated the presence of lamellar epithelial hyperplasia, significant fusion of secondary lamellae, and infiltration of inflammatory cells. A multitude of cysts, varying in shape from circular to elliptical, were observed within the gills. The occurrence of interlamellar vesicles and amoeboid organisms could be observed within the epithelial tissue of the gills. In the current study, presence of the Acanthamoeba T4 genotype on the skin and gills of discus fish exhibiting signs of illness in freshwater ornamental fish farms was identified. This observation suggests the potential of a transmission of amoebic infection from ornamental fish to humans, thereby highlighting the need for further investigation into this infection among ornamental fish maintained as pets, as well as individuals who interact with them and their environment.
Assuntos
Acanthamoeba , Amoeba , Ciclídeos , Humanos , Animais , Amoeba/genética , Filogenia , Irã (Geográfico)/epidemiologia , Funções Verossimilhança , Acanthamoeba/genéticaRESUMO
Proliferative gill disease (PGD), caused by the myxozoan Henneguya ictaluri, has been the most notorious parasitic gill disease in the US catfish aquaculture industry. In 2019, an unusual gill disease caused by massive burdens of another myxozoan, Henneguya exilis, was described in channel (Ictalurus punctatus) × blue (Ictalurus furcatus) hybrid catfish. Targeted metagenomic sequencing and in situ hybridization (ISH) were used to differentiate these conditions by comparing myxozoan communities involved in lesion development and disease pathogenesis between massive H. exilis infections and PGD cases. Thirty ethanol-fixed gill holobranchs from 7 cases of massive H. exilis infection in hybrid catfish were subjected to targeted amplicon sequencing of the 18S rRNA gene and compared to a targeted metagenomic data set previously generated from clinical PGD case submissions. Furthermore, serial sections of 14 formalin-fixed gill holobranchs (2 per case) were analyzed by RNAscope duplex chromogenic ISH assays targeting 8 different myxozoan species. Targeted metagenomic and ISH data were concordant, indicating myxozoan community compositions significantly differ between PGD and massive branchial henneguyosis. Although PGD cases often consist of mixed species infections, massive branchial henneguyosis consisted of nearly pure H. exilis infections. Still, H. ictaluri was identified by ISH in association with infrequent PGD lesions, suggesting coinfections occur, and some cases of massive branchial henneguyosis may contain concurrent PGD lesions contributing to morbidity. These findings establish a case definition for a putative emerging, myxozoan-induced gill disease of farm-raised catfish with a proposed condition name of massive branchial henneguyosis of catfish (MBHC).
RESUMO
Dr. Harold R. Griffith and Richard C. Gill figure prominently in curare's storied history. In 1938, Gill returned from an Amazon expedition with over 11 kg of curare. After scientists at E. R. Squibb & Sons identified a plant source (Chondrodendron tomentosum) and isolated a stable extract of uniform potency (marketed as Intocostrin), Griffith administered it in the operating room in 1942, showing its advantages and safety. In this article, we report correspondence between Griffith and Gill, heretofore not appreciated, after finding a letter from Gill to Griffith affixed to the inside back cover of a book contained in a private library.Following the serendipitous discovery of this previously unknown letter, we interrogated archived correspondence and material associated with Griffith and Gill in the Osler Library History of Medicine (McGill University, Montreal, QC, Canada), Arthur E. Guedel Memorial Anesthesia Center (University of California, San Francisco, CA, USA), the Wood Library Museum of Anesthesiology (Schaumburg, IL, USA), the Anaesthesia Heritage Centre (London, UK), and the Wellcome Collection (London, UK). Further, we searched for information on the historical background of curare via Google, Ovid MEDLINE, Adam Matthew Explorer, Project MUSE, and Latin American History databases.We found seven letters. The first is a letter to Gill dated 2 June 1943 (Wood Library) and an earlier draft dated 2 June 1943 (Osler Library). In this letter, Griffith praises Gill's success in procuring curare and informs him of its usefulness in anesthesia. The second letter is a letter from Gill to Griffith dated 10 July 1943 (found affixed to a book that was donated to the Osler Library). In this letter, Gill congratulates Griffith and claims he foresaw curare's use in the operating room and predicts its routine use to produce muscle relaxation during surgery. The third letter is a letter to Griffith dated 17 April 1945 (Osler Library). In this correspondence, Gill disputes Squibb's claim that curare derives solely from C. tomentosum and asks Griffith to retract published statements on this point. The fourth letter is a letter to Gill dated 25 April 1945 (Osler Library), in which Griffith declines to retract and emphasizes that Gill receive credit for making curare available to medicine. The fifth letter is a letter to Griffith dated 24 May 1945 (Osler Library), in which Gill accepts Griffith's retraction decision and indicates negotiations with another drug company. The sixth letter is a letter to Griffith dated 11 July 1945 (Osler Library), in which Gill requests anesthesia morbidity and mortality data and continues to remonstrate against Squibb's claim of curare's botanical source. The seventh and final letter is to Gill and dated 21 July 1945 (Osler Library). In this letter, Griffith indicates the lack of morbidity and mortality information, mentions a new Squibb curare product, and cites data suggesting curare may exert dose-dependent CNS effects.These seven letters between Dr. H. Griffith and R. Gill reveal a professional relationship heretofore not appreciated. We discuss and consider these letters in the context of curare's remarkable history.
RéSUMé: Le Dr Harold R. Griffith et Richard C. Gill occupent une place importante dans l'histoire du curare. En 1938, Gill revient d'une expédition en Amazonie avec plus de 11 kg de curare. Après l'identification, par les scientifiques de E. R. Squibb & Sons, d'une source végétale (Chondrodendron tomentosum) et l'isolement d'un extrait stable de puissance uniforme (commercialisé sous le nom d'Intocostrin), Griffith l'administre en salle d'opération en 1942, montrant ses avantages et son innocuité. Dans cet article, nous résumons la correspondance entre Griffith et Gill, jusque-là peu appréciée, après avoir trouvé une lettre de Gill à Griffith glissée à l'intérieur de la quatrième de couverture d'un livre provenant d'une bibliothèque privée.À la suite de la découverte fortuite de cette lettre jusque-là inconnue, nous avons consulté la correspondance et les documents archivés associés à Griffith et Gill à la bibliothèque Osler History of Medicine (Université McGill, Montréal, QC, Canada), au centre Arthur E. Guedel Memorial Anesthesia Center (Université de Californie, San Francisco, CA, États-Unis), au Wood Library Museum of Anesthesiology (Schaumburg, IL, États-Unis), au Anaesthesia Heritage Centre (Londres, Royaume-Uni) et à la Wellcome Collection (Londres, Royaume-Uni). De plus, nous avons recherché des informations sur le contexte historique du curare via Google, Ovid MEDLINE, Adam Matthew Explorer, Project MUSE et dans les bases de données d'histoire de l'Amérique latine.Nous avons trouvé sept lettres. La première est une lettre à Gill datée du 2 juin 1943 (bibliothèque Wood) et une ébauche antérieure datée du 2 juin 1943 (bibliothèque Osler). Dans cette lettre, Griffith fait l'éloge du succès de Gill dans l'obtention du curare et l'informe de son utilité en anesthésie. La deuxième lettre est une lettre de Gill à Griffith datée du 10 juillet 1943 (trouvée collée dans un livre donné à la bibliothèque Osler). Dans cette lettre, Gill félicite Griffith et affirme qu'il avait pressenti l'utilisation du curare en salle d'opération et prédit son utilisation de routine pour produire une relaxation musculaire pendant la chirurgie. La troisième lettre est une lettre à Griffith datée du 17 avril 1945 (bibliothèque Osler). Dans cette missive, Gill conteste l'affirmation de Squibb selon laquelle le curare est exclusivement dérivé de C. tomentosum et demande à Griffith de se rétracter sur les déclarations publiées à ce sujet. La quatrième lettre est une lettre à Gill datée du 25 avril 1945 (bibliothèque Osler), dans laquelle Griffith refuse de se rétracter et insiste sur le fait que Gill soit crédité d'avoir mis le curare à la disposition de la médecine. La cinquième missive est une lettre à Griffith datée du 24 mai 1945 (bibliothèque Osler), dans laquelle Gill accepte la décision de Griffith de se rétracter et indique des négociations avec une autre compagnie pharmaceutique. La sixième lettre est une lettre à Griffith datée du 11 juillet 1945 (bibliothèque Osler), dans laquelle Gill demande des données concernant la morbidité et la mortalité liées à l'anesthésie et continue de protester contre l'affirmation de Squibb sur la source botanique du curare. La septième et dernière lettre est adressée à Gill et datée du 21 juillet 1945 (bibliothèque Osler). Dans cette lettre, Griffith indique le manque d'informations sur la morbidité et la mortalité, mentionne un nouveau produit à base de curare de Squibb et cite des données suggérant que le curare pourrait exercer des effets dose-dépendants sur le SNC.Ces sept lettres entre le Dr H. Griffith et R. Gill révèlent une relation professionnelle jusque-là peu appréciée. Nous discutons et considérons ces lettres dans le contexte de l'histoire remarquable du curare.
RESUMO
Nodular gill disease (NGD) is an emerging condition associated with amoeba trophozoites in freshwater salmonid farms. However, unambiguous identification of the pathogens still must be achieved. This study aimed to identify the amoeba species involved in periodic NGD outbreaks in two rainbow trout (Oncorhynchus mykiss) farms in Northeastern Italy. During four episodes (February-April 2023), 88 fish were euthanized, and their gills were evaluated by macroscopic, microscopic and histopathological examination. The macroscopic and microscopic severity of the lesions and the degree of amoebae infestation were scored and statistically evaluated. One gill arch from each animal was put on non-nutrient agar (NNA) Petri dishes for amoeba isolation, cultivation and subsequent identification with SSU rDNA sequencing. Histopathology confirmed moderate to severe lesions consistent with NGD and mild to moderate amoeba infestation. The presence of amoebae was significantly correlated with lesion severity. Light microscopy of cultured amoebae strains and SSU rDNA analysis revealed the presence of a previously characterized amoeba Naegleria sp. strain GERK and several new strains: two strains from Hartmannelidae, three vannelid amoebae from the genus Ripella and cercozoan amoeba Rosculus. Despite the uncertainty in NGD etiopathogenesis and amoebae pathogenic role, identifying known and new amoebae leans towards a possible multi-aetiological origin.
Assuntos
Amebíase , Doenças dos Peixes , Brânquias , Oncorhynchus mykiss , Animais , Oncorhynchus mykiss/parasitologia , Doenças dos Peixes/parasitologia , Doenças dos Peixes/epidemiologia , Itália , Amebíase/veterinária , Amebíase/parasitologia , Brânquias/parasitologia , Brânquias/patologia , Amoeba/genética , Amoeba/isolamento & purificação , Amoeba/classificação , Aquicultura , Amebozoários/genética , Amebozoários/isolamento & purificação , Amebozoários/classificação , Amebozoários/fisiologia , FilogeniaRESUMO
The excretory mechanisms of stenohaline marine osmoconforming crabs are often compared to those of the more extensively characterized euryhaline osmoregulating crabs. These comparisons may have limitations, given that unlike euryhaline brachyurans the gills of stenohaline marine osmoconformers possess ion-leaky paracellular pathways and lack the capacity to undergo ultrastructural changes that can promote ion-transport processes in dilute media. Furthermore, the antennal glands of stenohaline marine osmoconformers are poorly characterized making it difficult to determine what role urinary processes play in excretion. In the presented study, ammonia excretory processes as well as related acid-base equivalent transport rates and mechanisms were investigated in the Dungeness crab, Metacarcinus magister - an economically valuable stenohaline marine osmoconforming crab. Isolated and perfused gills were found to predominantly eliminate ammonia through a microtubule network-dependent active NH4+ transport mechanism that is likely performed by cells lining the arterial pockets of the gill lamella where critical Na+/K+-ATPase detection was observed. The V-type H+-ATPase - a vital component to transbranchial ammonia excretion mechanisms of euryhaline crabs - was not found to contribute significantly to ammonia excretion; however, this may be due to the transporter's unexpected apical localization. Although unconnected to ammonia excretion rates, a membrane-bound isoform of carbonic anhydrase was localized to the apical and basolateral membranes of lamella suited for respiration. Urine was found to contain significantly less ammonia as well as carbonate species than the hemolymph, indicating that unlike those of some euryhaline crabs the antennal glands of the Dungeness crab reabsorb these molecules rather than eliminate them for excretion.