RESUMO
Length-weight relationships (LWR) and relative condition factor were described for species of deep-water crustaceans caught with bottom trawls in a depth range between 150 and 535 m during August and December of 2009, and March and May of 2010 in the Colombian Caribbean Sea. A linear regression was performed using the logarithmically transformed data to calculate the a and b coefficients of the LWR for 22 crustacean species corresponding to 13 families and 19 genera and three types of crustaceans (shrimp, crab, lobster). Six crustacean species showed a maximum total length greater than that reported in SeaLifeBase: Garymunida longipes (77.00 mm), Eunephrops bairdii (220.00 mm), Metanephrops binghami (197.46 mm), Penaeopsis serrata (149.00 mm), Polycheles typhlops (196.27 mm) and Pleoticus robustus (240.00 mm). A total of 11 species (50.0%) exhibited isometric growth, five species (22.7%) negative allometric and six species (27.3%) positive allometric. This study shows the first estimates of LWR for 12 species of deep-water crustaceans in the Colombian Caribbean Sea. We demonstrate for the first time that the growth parameters (intercept and slope) of the LWR varying significantly as a function of the body shape of crabs, lobsters and shrimps in deep-water crustaceans.
Assuntos
Braquiúros , Decápodes , Animais , Região do Caribe , Colômbia , Nephropidae , Somatotipos , ÁguaRESUMO
INTRODUCTION: The lobster cockroach Nauphoeta cinerea (N. cinerea) is indicated as a promising non-mammalian model, because it presents behavioral and biochemical alterations also observed in conventional models. In this research, we identified and characterized the distribution of protozoa that inhabit the digestive system (DS) of N. cinerea cockroaches. METHODS: The adult specimens of N. cinerea used in this study (n = 32) were obtained at the Federal University of Santa Maria, dissected and had their visceral contents observed in bright-field microscopy without staining and after application of lugol, Ziehl-Neelsen staining, EA36 trichrome and simulated dark-field microscopy with application of nankin ink. The presence of protozoa in different portions of the DS was semi-quantified by a system of crosses (+). RESULTS: The main taxa observed were: amoebas (Archaemebae:Entamoebida), gregarins (Apicomplexa:Eugregarinide), coccidia (Apicomplexa:Eucoccidiorida), kinetoplastids (Kinetoplastea:Kinetoplastida) and oxymonads (Preaxostyla:Oxymonadida). The highest prevalence of amoebas and gregarines was observed in the medial portion of the DS, while for the other groups, this was seen in the final portion, and in the case of coccidia, such prevalence was specially evidenced by the alcohol-acid coloration. In the present work, the great biological diversity that exists in the microbiota of the digestive system of Nauphoeta cinerea was demonstrated, being possible to find several pathogenic species for humans such as Entamoeba histolytica/dispar/moshkovskii, Cryptosporidium sp. and Cyclospora cayetanensis. There is still a lot to know about the interactions between endocommensal protozoa and their respective invertebrate hosts, so the best way to clarify such relationships is through molecular and genetic test.
Assuntos
Baratas , Criptosporidiose , Cryptosporidium , Entamoeba , Microbiota , Adulto , Animais , Sistema Digestório , Humanos , Insetos , NephropidaeRESUMO
Resumen Introducción: En el hábitat natural, el estado nutricional insatisfactorio de un número elevado de ejemplares es expresión de la influencia de factores desfavorables. Objetivo: Determinar variaciones espacio-temporales en el estado nutricional de langostas Panulirus argus, relacionarlas con variaciones reportadas en la comunidad bentónica y demostrar, mediante la aplicación de indicadores analíticos y morfométricos, que la variación de factores ambientales afecta a ambos grupos de índices; y que no todos los índices de condición nutricional evidencian por igual el impacto generalizado del proceso de deterioro ambiental en este golfo. Métodos: Las langostas se capturaron en seis áreas de pesca. La condición nutricional se estimó mediante tres índices no destructivos, poco costosos y de fácil y rápida aplicación (extensivos): índice de refracción de la hemolinfa (IRH), relación peso total / largo total = Klt, y factor de condición (FCA). Las variaciones a largo plazo (60 años) se determinaron mediante índices morfométricos (Klt y FCA). Se analizaron datos de los períodos 1963-1964 (N = 29 001), 1983-1993 (N = 3 123) y 2011-2017 (N = 3 600), separándolos en épocas de Lluvia y Seca. Resultados: En todos los períodos la condición nutricional varió significativamente entre áreas, pero sin similitudes entre períodos. Esto indica que los factores que impactan en el estado nutricional tienen una influencia estocástica más característica de factores ambientales. Aunque los tres índices fueron menores en 2017, sólo IRH disminuyó gradualmente entre 2011 y 2017, lo que sugiere que este índice, y los morfométricos, expresan diferente información. Todas las correlaciones fueron estadísticamente significativas, los coeficientes más altos se establecieron entre los índices morfométricos. El peso total y el Klt no mostraron diferencias entre Seca y Lluvia. Sin embargo, IRH y FCA resultaron mayores en Seca, hecho que se constató para cada sexo. FCA y Klt no presentaron diferencias entre el período 1963-1964 y el período 1983-1993, pero aumentaron (P < 0.05) en el período 2011-2017, lo que, dado el carácter morfométrico de estos índices, se atribuye a la presencia de langostas de mayor peso en las áreas de pesca. Conclusiones: Los menores valores en 2017 y la tendencia gradual al decrecimiento de IRH, corroboran el carácter generalizado del deterioro ambiental en el golfo de Batabanó, lo cual no fue igualmente expresado por los tres índices.
Abstract Introduction: Nutritional condition indices are morphophysiological estimators that quantify the nutritional status of organisms. In the natural habitat, unsatisfactory nutritional stage of numerous individuals is an expression of unfavorable factors. Objectives: Determine spatio-temporal variations in the nutritional condition of lobsters Panulirus argus, relate them to variations reported in the benthic community, and to demonstrate, through the application of analytical and morphometric indices, that variations of environmental factors affects both groups of indices due to its potential magnitude and its generalized nature. Methods: Lobsters were captured at six sites. Nutritional condition was estimated by three non-destructive, inexpensive and quick indices also easy to apply, which it means extensive indices: blood refractive index (BRI), relationship between total weight and total length (K), and condition factor (CFA). Because there are no historical records of BRI, long-term variations (60 years) were determined using morphometric indices (Klt and CFA). Data from the periods 1963-1964 (N = 29 001), 1983-1993 (N = 3 123) and 2011-2017 (N = 3 600) were analyzed, separating them in the rainy and dry seasons. Results: In all the periods the nutritional condition varied significantly between sites, but without similarities between periods. Despite this, it increased progressively in three sites. This indicates that factors impacting nutritional condition have a stochastic influence that is constantly changing, which is more characteristic of environmental factors than anthropogenic factors. Although the three indices showed a lower nutritional condition in 2017, only BRI gradually decreased between 2011 (16.6) and 2017 (14.2), which suggests that this index, and the morphometrics (Klt and CFA), express different information. All correlations were statistically significant but greatest coefficients were established between morphometric indices. The total weight and the Klt index did not show differences between rainy and dry seasons. However, BRI and CFA were statistically greater in dry season, a fact that was found for each sex. Both CFA and Klt decreased slightly (P > 0.05) between the first (1963-1964) and the second (1983-1993) period and then increased (P < 0.05) in the current period (2011-2017). Because they both are morphometric indices, this was attributed to the presence of greater weight lobsters. Conclusions: The lowest values in 2017 and gradually decreasing trend in BRI, corroborate the generalized nature of the environmental deterioration in the Gulf of Batabanó, but this was not equally expressed by the three indices.
Assuntos
Animais , Alimentos, Dieta e Nutrição , Nephropidae/crescimento & desenvolvimento , CubaRESUMO
A new species of Pseudione Kossmann, 1881 is described from two parasitized specimens of the clawed lobster Nephropsis aculeata Smith, 1881, collected from Brazilian waters in the southwestern Atlantic. It is the second species of the genus from Brazil and is the fourth species parasitizing nephropid lobsters in the world.
Assuntos
Isópodes , Nephropidae , Animais , BrasilRESUMO
Suitable peptidases for biotechnological applications are those active at low temperature, in organic solvents, detergents or proteolytic additives. American lobster cathepsin D1 (CD1) is an enzyme highly efficient at 5-50°C and at pH 2.5-5.5. We assessed the effect of common industrial additives on CD1 activity. CD1 was isolated from lobster gastric fluid by chromatography. The proteolytic activity was measured using a fluorogenic specific substrate and the conformation by intrinsic fluorescence. Non-ionic detergents Tween-20 and Triton X-100 stabilize the peptidase activity. Ethanol, methanol and isopropanol [5-15% (v/v)] increased the enzyme activity up to 80%. The enzyme is active until 2.5M urea and is resistant to proteolysis by papain and renin. In this work, a crustacean peptidase that remains active when exposed to different chemical and proteolytic additives is reported, evincing that crustaceans are a good model for discovery of novel stable peptidases for future pharmaceutical, cosmetic and alimentary applications.
Assuntos
Catepsina D/metabolismo , Detergentes/farmacologia , Nephropidae/enzimologia , Proteólise/efeitos dos fármacos , Sais/farmacologia , Solventes/química , Animais , Catepsina D/química , Catepsina D/isolamento & purificação , Estabilidade Enzimática , Fluorescência , Glicerol/farmacologia , Papaína/farmacologia , Conformação Proteica , Renina/farmacologia , Cloreto de Sódio/farmacologia , Tensoativos/farmacologia , Ureia/farmacologiaRESUMO
Gastroliths are highly calcified structures formed in the cardiac stomach wall of crustaceans for the temporary storage of amorphous CaCO3 (ACC). The gastrolithic ACC is stabilized by the presence of biomolecules, and represents a novel model for research into biomineralization. For the first time, an in vitro biomimetic retrosynthesis of scaffolds of gastrolithic matrices with CaCO3 is presented. With the help of synthetic polyacrylic (PAA) and phytic (PA) acids, amorphous precursor particles were stabilized in double (DD) and gas (GD) diffusion crystallization assays. The presence of these synthetic molecules as efficient inhibitors of nucleation and growth of CaCO3, and the use of biological gastrolith scaffolds as confined reaction environments determined the kinetics of crystallization, and controlled the morphogenesis of CaCO3. The formation of ACC particles was demonstrated and their crystallization was followed by light microscopy, scanning and transmission electron microscopy, and electron diffraction.
Assuntos
Biomimética/métodos , Calcificação Fisiológica , Carbonato de Cálcio/química , Nephropidae/anatomia & histologia , Animais , Cristalização , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de TransmissãoRESUMO
The Galapagos slipper lobster Scyllarides astori was found for the first time off western Mexico mainland. A male and two female specimens were collected in south Jalisco and north Colima. This represents a new record from central Pacific coast of Mexico and a range extension of 700 km southwards, as the species was previously reported from the Gulf of California, 780 km eastwards from Revillagigedo Archipelago and 2400 km northwestwards from Isla del Coco and Galapagos Archipelago.
Assuntos
Nephropidae , Animais , California , Feminino , Masculino , Oceano PacíficoRESUMO
The genus Nephropsis Wood-Mason, 1872 has been reported from Brazil by Tavares (1998), Tavares & Young (2002), Silva et al. (2003), Dall´Occo et al. (2007) and Serejo et al. (2007), recording Nephropsis aculeata Smith, 1881, N. rosea Bate, 1888 and N. agassizii A. Milne-Edwards, 1880, the last of which occurs in both northeastern and southeastern of Brazil.
Assuntos
Nephropidae/classificação , Distribuição Animal , Estruturas Animais/anatomia & histologia , Estruturas Animais/crescimento & desenvolvimento , Animais , Tamanho Corporal , Brasil , Ecossistema , Feminino , Masculino , Nephropidae/anatomia & histologia , Nephropidae/crescimento & desenvolvimento , Tamanho do ÓrgãoRESUMO
The amount of Uruguayan lobster Metanephrops rubellus landed in the state of São Paulo has shown considerable annual variation, at first due to ups and downs in the fishing sector interest. However, biological factors may also be influencing, since little is known about the biology of M. rubellus, and most of the available literature on Nephropidae refer to the Norwegian lobster (Nephrops norvegicus), which is morphologically and behaviorally similar. The morphometric relations found characterize the relative growth of M. rubellus as positive allometric, except for Ct x Cc x Pt and Pa, which showed isometric growth. The length at first maturity (L50) and the length frequencies distribution indicate that fisheries are focusing preferably on the portion of population able to reproduce, saving the immature ones. Due to this and to the biological and behavioral similarity between N. norvegicus and M. rubellus, fishing closure or minimum size of capture are not recommended as fishery management tools. Instead, additional research is proposed in order to define the distribution area and density of population, to establish fishing exclusion areas, which function as reservoirs for replenishment of stocks. It is also essential monitoring the population dynamics parameters, in order to control fishing effort.(AU)
A estatística pesqueira paulista mostra considerável oscilação anual na quantidade de lagostim desembarcado, a princípio devido a altos e baixos no interesse do setor pesqueiro. No entanto, fatores biológicos também podem estar influenciando, já que pouco se conhece da biologia de M. rubellus, pois as principais referências disponíveis sobre nefropídeos referem-se ao lagostim norueguês (Nephrops norvegicus), espécie europeia similar morfológica e comportamentalmente. As relações morfométricas encontradas caracterizam o crescimento relativo de M. rubellus como alométrico positivo, exceto para Ct x Cc e Pt x Pa, que mostraram isometria. O comprimento de início da fase de maturidade (L50) e a distribuição de comprimentos dos exemplares capturados indicam que a pesca incidiu preferencialmente sobre indivíduos aptos à reprodução. Devido a isto e à similitude biológica e comportamental entre N. norvegicus e M. rubellus não são indicados o defeso ou o tamanho mínimo de captura como instrumentos de gestão da pescaria, propondo-se a realização de pesquisas para definir a área de distribuição e densidade da população, visando estabelecer áreas de exclusão de pesca que funcionem como reservatórios para a reposição dos estoques. É essencial ainda monitorar o comportamento dos parâmetros de dinâmica populacional, visando ao controle do esforço de pesca.(AU)
Assuntos
Animais , Nephropidae/anatomia & histologia , Nephropidae/crescimento & desenvolvimento , Maturidade Sexual , Decápodes/anatomia & histologia , Decápodes/crescimento & desenvolvimentoRESUMO
The amount of Uruguayan lobster Metanephrops rubellus landed in the state of São Paulo has shown considerable annual variation, at first due to ups and downs in the fishing sector interest. However, biological factors may also be influencing, since little is known about the biology of M. rubellus, and most of the available literature on Nephropidae refer to the Norwegian lobster (Nephrops norvegicus), which is morphologically and behaviorally similar. The morphometric relations found characterize the relative growth of M. rubellus as positive allometric, except for Ct x Cc x Pt and Pa, which showed isometric growth. The length at first maturity (L50) and the length frequencies distribution indicate that fisheries are focusing preferably on the portion of population able to reproduce, saving the immature ones. Due to this and to the biological and behavioral similarity between N. norvegicus and M. rubellus, fishing closure or minimum size of capture are not recommended as fishery management tools. Instead, additional research is proposed in order to define the distribution area and density of population, to establish fishing exclusion areas, which function as reservoirs for replenishment of stocks. It is also essential monitoring the population dynamics parameters, in order to control fishing effort.
A estatística pesqueira paulista mostra considerável oscilação anual na quantidade de lagostim desembarcado, a princípio devido a altos e baixos no interesse do setor pesqueiro. No entanto, fatores biológicos também podem estar influenciando, já que pouco se conhece da biologia de M. rubellus, pois as principais referências disponíveis sobre nefropídeos referem-se ao lagostim norueguês (Nephrops norvegicus), espécie europeia similar morfológica e comportamentalmente. As relações morfométricas encontradas caracterizam o crescimento relativo de M. rubellus como alométrico positivo, exceto para Ct x Cc e Pt x Pa, que mostraram isometria. O comprimento de início da fase de maturidade (L50) e a distribuição de comprimentos dos exemplares capturados indicam que a pesca incidiu preferencialmente sobre indivíduos aptos à reprodução. Devido a isto e à similitude biológica e comportamental entre N. norvegicus e M. rubellus não são indicados o defeso ou o tamanho mínimo de captura como instrumentos de gestão da pescaria, propondo-se a realização de pesquisas para definir a área de distribuição e densidade da população, visando estabelecer áreas de exclusão de pesca que funcionem como reservatórios para a reposição dos estoques. É essencial ainda monitorar o comportamento dos parâmetros de dinâmica populacional, visando ao controle do esforço de pesca.
Assuntos
Animais , Maturidade Sexual , Nephropidae/anatomia & histologia , Nephropidae/crescimento & desenvolvimento , Decápodes/anatomia & histologia , Decápodes/crescimento & desenvolvimentoRESUMO
Crustaceans are a diverse group, distributed in widely variable environmental conditions for which they show an equally extensive range of biochemical adaptations. Some digestive enzymes have been studied by purification/characterization approaches. However, global analysis is crucial to understand how digestive enzymes interplay. Here, we present the first proteomic analysis of the digestive fluid from a crustacean (Homarus americanus) and identify glycosidases and peptidases as the most abundant classes of hydrolytic enzymes. The digestion pathway of complex carbohydrates was predicted by comparing the lobster enzymes to similar enzymes from other crustaceans. A novel and unbiased substrate profiling approach was used to uncover the global proteolytic specificity of gastric juice and determine the contribution of cysteine and aspartic acid peptidases. These enzymes were separated by gel electrophoresis and their individual substrate specificities uncovered from the resulting gel bands. This new technique is called zymoMSP. Each cysteine peptidase cleaves a set of unique peptide bonds and the S2 pocket determines their substrate specificity. Finally, affinity chromatography was used to enrich for a digestive cathepsin D1 to compare its substrate specificity and cold-adapted enzymatic properties to mammalian enzymes. We conclude that the H. americanus digestive peptidases may have useful therapeutic applications, due to their cold-adaptation properties and ability to hydrolyze collagen.
Assuntos
Proteínas de Artrópodes/metabolismo , Suco Gástrico/química , Glicosídeo Hidrolases/metabolismo , Nephropidae/enzimologia , Peptídeo Hidrolases/metabolismo , Adaptação Fisiológica , Sequência de Aminoácidos , Animais , Proteínas de Artrópodes/genética , Temperatura Baixa , Digestão/fisiologia , Suco Gástrico/enzimologia , Expressão Gênica , Glicosídeo Hidrolases/genética , Anotação de Sequência Molecular , Nephropidae/genética , Peptídeo Hidrolases/genética , Proteólise , Proteômica , Especificidade por SubstratoRESUMO
Aspartic proteinases in the gastric fluid of clawed lobsters Homarus americanus and Homarus gammarus were isolated to homogeneity by single-step pepstatin-A affinity chromatography; such enzymes have been previously identified as cathepsin D-like enzymes based on their deduced amino acid sequence. Here, we describe their biochemical characteristics; the properties of the lobster enzymes were compared with those of its homolog, bovine cathepsin D, and found to be unique in a number of ways. The lobster enzymes demonstrated hydrolytic activity against synthetic and natural substrates at a wider range of pH; they were more temperature-sensitive, showed no changes in the K(M) value at 4°C, 10°C, and 25°C, and had 20-fold higher k(cat)/K(M) values than bovine enzyme. The bovine enzyme was temperature-dependent. We propose that both properties arose from an increase in molecular flexibility required to compensate for the reduction of reaction rates at low habitat temperatures. This is supported by the fast denaturation rates induced by temperature.
Assuntos
Aclimatação/fisiologia , Ácido Aspártico Proteases/metabolismo , Temperatura Baixa , Ácido Gástrico/enzimologia , Nephropidae/enzimologia , Animais , Ácido Aspártico Proteases/fisiologia , Catepsina D/metabolismo , Bovinos , Cromatografia de Afinidade , Concentração de Íons de Hidrogênio , Nephropidae/fisiologia , Pepstatinas , Especificidade da EspécieRESUMO
Naturally occurring antimicrobial peptides take place in the first line of host defense against pathogen as part of the humoral innate immune response. ß-defensins are among the most abundant antimicrobial peptides in mammals, and thought to be solely found in vertebrates until a recent report describing the cloning and sequencing of defensin like peptides in the spiny lobster Panulirus japonicus. In the current study, we cloned and sequenced two genes from the hemocytes of the spiny lobster Panulirus argus encoding for two isoforms of defensin-like peptides, thus confirming the presence of this protein in the Panulirus genus. The 44 amino acids mature peptides showed the conservation of cysteine pattern characterizing the ß-defensins, as well as known amino acids residues critical to exert their antimicrobial activity. They are also amphipathics, hydrophobics, and display an overall positive charge (+1) located at the C-terminus. The tertiary structure obtained by homology modeling indicated that likely conformations of lobster peptides are highly similar to ß-defensins from vertebrates. The phylogenetic study carried out by probabilistic methods confirmed the relation with ancestral ß-defensin from vertebrates. The finding of a putative defensin-like peptide in the expressed sequence tag (EST) of the lobster Homarus americanus with high homology with those of P. argus described in this study, would indicate the presence of this peptides in Palinuridae family. Taking into account all similarities between these peptides with ß-defensins from vertebrates, it is conceivable to further support the finding of a new family of ß-defensins in invertebrate.
Assuntos
Proteínas de Artrópodes/genética , Defensinas/genética , Palinuridae/genética , Sequência de Aminoácidos , Animais , Proteínas de Artrópodes/química , Proteínas de Artrópodes/metabolismo , Sequência de Bases , Clonagem Molecular , Simulação por Computador , Defensinas/química , Defensinas/metabolismo , Etiquetas de Sequências Expressas , Hemócitos/metabolismo , Dados de Sequência Molecular , Nephropidae/química , Nephropidae/genética , Palinuridae/metabolismo , Filogenia , Reação em Cadeia da Polimerase , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Estrutura Secundária de Proteína , Estrutura Terciária de Proteína , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , beta-Defensinas/química , beta-Defensinas/genética , beta-Defensinas/metabolismoRESUMO
Necrotizing hepatopancreatitis bacterium (NHPB) is an obligated intracellular bacteria causing severe hepatopancreatic damages and mass mortalities in penaeid shrimp. The worldwide distribution of penaeid shrimp as alien species threatens the life cycle of other crustacean species. The aim of the experiment was to evaluate the possibility of experimentally infecting the American lobster (Homarus americanus) with NHPB extracted from shrimp hepatopancreas. Homogenates from infected shrimp were fed by force to lobsters. Other group of lobsters was fed with homogenates of NHPB-free hepatopancreas. After the 15th day from initial inoculation, the presence of NHPB was detected by polymerase chain reaction in feces and hepatopancreas from lobsters inoculated with infected homogenates. Necrotized spots were observed in the surface of lobster hepatopancreas. In contrast, lobsters fed on NHPB-free homogenates resulted negative for NHPB. Evidence suggests the plasticity of NHPB which can infect crustacean from different species and inhabiting diverse latitudes. Considering the results, the American lobster could be a good candidate to maintain available NHPB in vivo.
Assuntos
Bactérias/metabolismo , Hepatopâncreas/microbiologia , Nephropidae/microbiologia , Ração Animal , Animais , Infecções Bacterianas/metabolismo , DNA/metabolismo , Ecossistema , Fezes , Genoma , Estágios do Ciclo de Vida , Necrose , Reação em Cadeia da Polimerase/métodos , TemperaturaRESUMO
Two forms of the squat lobster Pleuroncodes monodon can be found along the Pacific coast of South America: a smaller pelagic and a larger benthic form that live respectively in the northern and southern areas of the geographic distribution of the species. The morphological and life history differences between the pelagic and benthic forms could be explained either by genetic differentiation or phenotypic plasticity. In the latter case it would correspond to a heterochronic phenotypic plasticity that is fixed in different environments (phenotype fixation). The aim of this study was to evaluate whether the two forms are genetically differentiated or not; and thus to infer the underlying basis-heritable or plastic-of the existence of the two forms. Based on barcoding data of mitochondrial DNA (the COI gene), we show that haplotypes from individuals of the pelagic and benthic forms comprise a single genetic unit without genetic differentiation. Moreover, the data suggest that all studied individuals share a common demographic history of recent and sudden population expansion. These results strongly suggest that the differences between the two forms are due to phenotypic plasticity.
Assuntos
DNA Mitocondrial/genética , Evolução Molecular , Deriva Genética , Variação Genética , Genética Populacional , Nephropidae/genética , Animais , Fenótipo , FilogeniaRESUMO
An aspartic proteinase was isolated from American lobster gastric fluid. The purified cathepsin D runs as a single band on native-PAGE displaying proteolytic activity on a zymogram at pH 3.0, with an isoelectric point of 4.7. Appearance of the protein in SDS-PAGE, depended on the conditions of the gel electrophoresis. SDS treatment by itself was not able to fully unfold the protein. Thus, in SDS-PAGE the protein appeared to be heterogeneous. A few minute of boiling the sample in the presence of SDS was necessary to fully denature the protein that then run in the gel as a single band of ~50 kDa. The protein sequence of lobster cathepsin D1, as deduced from its mRNA sequence, lacks a 'polyproline loop' and ß-hairpin, which are characteristic of some of its structural homologues. A comparison of amino acid sequences of digestive and non-digestive cathepsin D-like enzymes from invertebrates showed that most cathepsin D enzymes involved in food digestion, lack the polyproline loop, whereas all non-digestive cathepsin Ds, including the American lobster cathepsin D2 paralog, contain the polyproline loop. We propose that the absence or presence of this loop may be characteristic of digestive and non-digestive aspartic proteinases, respectively.
Assuntos
Catepsina D/química , Modelos Moleculares , Nephropidae/enzimologia , Sequência de Aminoácidos , Animais , Catepsina D/classificação , Catepsina D/isolamento & purificação , Eletroforese em Gel de Poliacrilamida , Dados de Sequência Molecular , Filogenia , Alinhamento de Sequência , Estômago/enzimologiaRESUMO
The temporal distribution of behavioral programs throughout the 24-h day, known as temporal niche of a species, is determined by ecological factors that directly affect the adaptive value of the timing of specific behaviors. Temporal niche switching has been described in several species and is likely adaptive in habitats where the daily timing of those factors changes. Benthic species whose habitats span a wide range of water depths are exposed to considerable depth-dependent environmental changes. Temporally scheduled trawl surveys of the Norway lobster, Nephrops norvegicus, reveal that animals emerge from burrows at night on the shallow shelf (10-50 m deep), at crepuscular hours on the lower shelf (50-200 m), and at daytime on the slope (200-400 m). The mechanisms underlying nocturnality/diurnality switches are chiefly unknown, and Nephrops offers a unique model for their study. The depth-dependent decrease in luminance is a likely candidate determining the temporal distribution of behavior. The authors explored this possibility in the laboratory by exposing Nephrops to light:dark (LD) cycles of 470-nm monochromatic lighting that mimic conditions at the 100-m-deep shelf (10 lux) or the 300-m slope (0.1 lux). Two groups of animals were respectively exposed to each light intensity according to the following protocol: an initial 12:12 LD stage followed by constant darkness (DD), followed in turn by a second 12:12 LD stage. Activity at the burrow opening (door-keeping = DK), as well as full emergence (E), was continuously monitored. Under 10-lux LD cycles, most animals showed nocturnal DK activity-with some being crepuscular or diurnal-and all animals showed nocturnal E activity. In contrast, both behaviors were clearly diurnal in animals under 0.1-lux LD cycles. The phase of the nocturnal and diurnal DK rhythms detected respectively at 10 and 0.1 lux upon release into DD revealed that these rhythms are entrained circadian rhythms. The present data indicate that nocturnality/diurnality switches in Nephrops in its natural habitat, evidenced by captures at different depths, are likely determined by light intensity. This temporal niche switching involves different patterns of photic entrainment, leading to bona fide circadian diurnal or nocturnal phenotypes, as well as exogenous masking of behavioral outputs.
Assuntos
Comportamento Animal/fisiologia , Atividade Motora , Nephropidae/fisiologia , Fotoperíodo , Animais , Ritmo Circadiano , MasculinoRESUMO
Acid digestive proteinases were studied in the gastric fluids of two species of clawed lobster (Homarus americanus and Homarus gammarus). An active protein was identified in both species as aspartic proteinase by specific inhibition with pepstatin A. It was confirmed as cathepsin D by mass mapping, N-terminal, and full-length cDNA sequencing. Both lobster species transcribed two cathepsin D mRNAs: cathepsin D1 and cathepsin D2. Cathepsin D1 mRNA was detected only in the midgut gland, suggesting its function as a digestive enzyme. Cathepsin D2 mRNA was found in the midgut gland, gonads, and muscle. The deduced amino acid sequence of cathepsin D1 and cathepsin D2 possesses two catalytic DTG active-site motifs, the hallmark of aspartic proteinases. The putatively active cathepsin D1 has a molecular mass of 36.4 kDa and a calculated pI of 4.14 and possesses three potential glycosylation sites. The sequences showed highest similarities with cathepsin D from insects but also with another crustacean cathepsin D. Cathepsin D1 transcripts were quantified during a starvation period using real-time qPCR. In H. americanus, 15 days of starvation did not cause significant changes, but subsequent feeding caused a 2.5-fold increase. In H. gammarus, starvation caused a 40% reduction in cathepsin D1 mRNA, and no effect was observed with subsequent feeding.
Assuntos
Catepsina D/metabolismo , Digestão , Sistema Digestório/enzimologia , Nephropidae/enzimologia , Sequência de Aminoácidos , Animais , Ácido Aspártico Proteases/genética , Ácido Aspártico Proteases/isolamento & purificação , Ácido Aspártico Proteases/metabolismo , Sequência de Bases , Catepsina D/genética , Catepsina D/isolamento & purificação , Dados de Sequência Molecular , Nephropidae/genética , RNA Mensageiro/metabolismo , Inanição/enzimologiaRESUMO
Gills are the first site of impact by metal ions in contaminated waters. Work on whole gill cells and metal uptake has not been reported before in crustaceans. In this study, gill filaments of the American lobster, Homarus americanus, were dissociated in physiological saline and separated into several cell types on a 30, 40, 50, and 80% sucrose gradient. Cells from each sucrose solution were separately resuspended in physiological saline and incubated in 65Zn2+ in order to assess the nature of metal uptake by each cell type. Characteristics of zinc accumulation by each kind of cell were investigated in the presence and absence of 10 mM calcium, variable NaCl concentrations and pH values, and 100 muM verapamil, nifedipine, and the calcium ionophore A23187. 65Zn2+ influxes were hyperbolic functions of zinc concentration (1-1,000 microM) and followed Michaelis-Menten kinetics. Calcium reduced both apparent zinc binding affinity (K (m)) and maximal transport velocity (J (max)) for 30% sucrose cells, but doubled the apparent maximal transport velocity for 80% sucrose cells. Results suggest that calcium, sodium, and protons enter gill epithelial cells by an endogenous broad-specificity cation channel and trans-stimulate metal uptake by a plasma membrane carrier system. Differences in zinc transport observed between gill epithelial cell types appear related to apparent affinity differences of the transporters in each kind of cell. Low affinity cells from 30% sucrose were inhibited by calcium, while high affinity cells from 80% sucrose were stimulated. 65Zn2+ transport was also studied by isolated, intact, gill filament tips. These intact gill fragments generally displayed the same transport properties as did cells from 80% sucrose and provided support for metal uptake processes being an apical phenomenon. A working model for zinc transport by lobster gill cells is presented.
Assuntos
Brânquias/metabolismo , Zinco/metabolismo , Animais , Transporte Biológico/efeitos dos fármacos , Calcimicina/farmacologia , Cloreto de Cálcio/farmacologia , Separação Celular , Centrifugação com Gradiente de Concentração , Células Epiteliais/metabolismo , Brânquias/citologia , Concentração de Íons de Hidrogênio , Cinética , Masculino , Nephropidae/fisiologia , Nifedipino/farmacologia , Verapamil/farmacologia , Radioisótopos de ZincoRESUMO
Deformation and compaction properties of native amino poly-saccharides chitin and chitosan were studied and compared with those obtained with established pharmaceutical direct compression excipients. An instrumented single-punch tablet machine was used for tablet compaction. The following compression parameters were evaluated: a ratio of crushing strength and compression pressure, plasticity and elasticity factor (PF and EF), tensile strength and R-value. Chitin and chitosan were found to have a marked tendency to plastic deformation, and both showed a good compression behaviour compared with the other direct compression excipients including microcrystalline cellulose. It is concluded that chitin and chitosan are potential co-excipients for direct compression applications.