Hematology and blood biochemistry profile of the freshwater stingray Potamotrygon magdalenae as a tool for population assessment in artificial environments / Perfil hematológico e bioquímico do sangue da arraia de água doce Potamotrygon magdalenae como ferramenta para avaliação da população em ambientes artificiais

Hematological and blood biochemical reference information is important to establish physiological status of freshwater stingray populations and improve care and management protocols in artificial environments. Here, we used a commercial freshwater stingray with high mortality rates in the market (Potamotrygon magdalenae), as an example to understand how artificial environments and handling protocols influence physiological status of captive freshwater stingrays. To this purpose, blood from five adult males and six adult females was collected to perform complete blood counts and blood chemistry analyses. All sampled animals showed good body condition with no differences between sexes. Differences between sexes were only found for the differential count of lymphocytes. Red blood results were consistent with previously studied potamotrygonids while white blood results showed higher values of leukocytes, thrombocytes, heterophils and lymphocytes in P. magdalenae compared to other Potamotrygonids. All types of leukocytes described for elasmobranchs were found except neutrophils and basophils. Blood metabolites showed an influence of ex situ diet in total protein, triglycerides and cholesterol. Glucose results were consistent while urea showed lower levels than those recorded for other freshwater stingrays. These results highlight the importance of physical, physiological and health analysis in freshwater stingrays as a part of welfare assessment to improve monitoring protocols and survival rates in public or private aquaria.

A informação de referência hematológica e bioquímica do sangue é importante para estabelecer o estado fisiológico das populações de arraias de água doce e melhorar os protocolos de cuidado e manejo em ambientes artificiais. Aqui, usamos uma espécie comercial de arraia de água doce com elevadas taxas de mortalidade no mercado(Potamotrygon magdalenae) como espécie exemplo para compreender de que modo os ambientes artificiais e os protocolos de manipulação influenciam o estado fisiológico das arraias de água doce em cativeiro. Para este fim, foi coletado sangue de cinco machos adultos e seis fêmeas adultas para realizar contagens completas de células sanguíneas e análises bioquímicas de sangue. Todos os animais amostrados mostraram boa condição corpórea, sem diferenças entre os sexos. Diferenças entre os sexos foram encontradas só na contagem diferencial de linfócitos. Os resultados de células sanguíneas vermelhas foram consistentes com potamotrigonídeos previamente estudados, enquanto o leucograma revelou valores mais elevados de leucócitos, trombócitos, heterófilos e linfócitos em P. magdalenae, em comparação com outros potamotrigonídeos. Todos os tipos de leucócitos descritos para elasmobrânquios foram encontrados, exceto para neutrófilos e basófilos. Todos os tipos de leucócitos descritos para elasmobrânquios foram encontrados, exceto para neutrófilos e basófilos. Os metabólitos do sangue mostraram influência da dieta ex situ nas proteínas totais, triglicerídeos e colesterol. Os resultados da glicose foram consistentes, enquanto a ureia mostrou níveis mais baixos do que os registrados para outras espécies de arraias de água doce. Os resultados da glicose foram consistentes, enquanto a ureia mostrou níveis mais baixos em P. magdalenae. Estes resultados enfatizam a importância da análise física, fisiológica e de saúde em arraias de água doce como parte da avaliação do bem-estar para melhorar os protocolos [...].

Environmentally-induced osmoregulation in Neotropical freshwater stingrays (Myliobatiformes: Potamotrygoninae) after controlling for phylogeny.

The osmotic physiology of freshwater stingrays was investigated in fifteen species from white (WW), black (BW), and clearwater (CW) rivers of Brazilian hydrographic basins. Regardless of phylogeny, potamotrygonids collected in the BW (Negro, Jutai, Nhamunda, and Manacapuru rivers), and CW (Tapajos, Parana, Mutum, Demeni, and Branco rivers) exhibited lower levels of osmolytes and plasma osmolality than those from WW (Amazon estuary, Solimoes, and Tarauaca rivers). However, the gill and kidney Na+/K+-ATPase activities were higher in the potamotrygonid species from BW and CW than those from WW. These results may be related to the ability of the potamotrygonids to achieve high ion uptake from ion-poor waters, such as those of BW and/or CW. Additionally, the high kidney Na+/K+-ATPase activity may help to minimize ion loss and generate diluted urine. Thus, diffusional losses of salts are balanced by uptake of ions in the gill, and reabsorption by the kidney. The physiological traits showed a weak phylogenetic signal, which indicates a strong evolutionary convergence. Multivariate analyses revealed that variations in physiological traits has a significant association with the type of water, as well as its physical and chemical characteristics such as electric conductivity and pH. Therefore, the South American Neotropical freshwater stingrays adjust their osmoregulatory mechanisms according to the environment in which they live.

TEMPORAL STABILITY OF IN VITRO VENOUS BLOOD GAS, pH, AND LACTATE VALUES OF COWNOSE RAYS (RHINOPTERA BONASUS) AND RED-EARED SLIDER TURTLES (PSEUDEMYS SCRIPTA ELEGANS).

This study assessed the in vitro temporal changes that occur in blood pH and lactate concentrations for an elasmobranch species and a chelonian species, as well as blood gases (partial pressures of carbon dioxide [pCO2] and oxygen [pO2]) for a chelonian species, with a portable clinical point-of-care analyzer. Blood samples were collected from 10 cownose rays (Rhinoptera bonasus) and 10 red-eared sliders (Pseudemys scripta elegans), stored on ice, and serially analyzed at six time points up to 90 min postcollection. Results indicate that analysis should be conducted as soon as possible after blood collection for these species, with immediate analysis being preferred. However, if analysis must be delayed, syringes may be capped, placed on ice, and analyzed at a later time. Analysis within 90 min provided clinically acceptable results for pH and lactate in both species and for pCO2 in red-eared sliders, whereas substantial artifactual increases of pO2 were seen in red-eared sliders.

Ultrasonographic and hormonal characterization of reproductive health and disease in wild, semiwild, and aquarium-housed southern stingrays (Hypanus americanus).

OBJECTIVE: To characterize physical examination, plasma biochemical, and ultrasonographic findings in aquarium-housed, managed semiwild, and wild southern stingrays (Hypanus americanus) with and without reproductive disease. ANIMALS: Southern stingrays from aquarium (n = 48), lagoon (managed semiwild; 34), and wild (12) habitats. PROCEDURES: Limited, opportunistic prosections were performed of presumed anatomically normal wild southern stingrays and compared with findings for aquarium-housed stingrays with reproductive disease. Ultrasonographic video data from both groups were used to assign a score (1 to 5) indicating increasing severity of ovarian and uterine reproductive disease. Plasma total 17ß-estradiol, estrone, progesterone, and testosterone concentrations were measured with enzyme immunoassays validated for use in southern stingrays. RESULTS: Ultrasonographic ovarian scores were significantly correlated with uterine scores. No reproductive disease was detected in semiwild or wild stingrays, but 65% (31/48) of aquarium-housed stingrays had developing or advanced reproductive disease (ie, ultrasonographic ovarian or uterine score of 4 or 5). Significant correlations were identified between ovarian and uterine disease status and plasma concentrations of all steroid hormones except testosterone. CONCLUSIONS AND CLINICAL RELEVANCE: Findings suggested that ultrasonography and plasma hormone concentrations may be useful in the identification of reproductive disease and determination of disease severity in southern stingrays.

Steroid concentrations in maternal serum and uterine histotroph in round stingrays (Urobatis halleri).

Despite a wide range of elasmobranch (sharks, skates and rays) matrotrophic strategies, and thus potentially diverse pathways for maternal-fetal hormone exchange, little attention has been given to uterine steroids during development. Round Stingrays (Urobatis halleri) with matrotrophic histotrophy were captured during every month of their annual reproductive season from post-ovulation to near parturition, and paired samples of plasma and histotroph were analyzed for a suite of steroid hormones using LC-ESI/MRM. Hormone concentrations within and between maternal and uterine compartments were compared using two markers of embryo development. Histotroph had consistently higher detection rates and concentrations of hormones than maternal plasma, especially during early pregnancy when embryos are yolk sac-dependent for nutrition. Peaks in histotroph testosterone concentrations preceded maternal plasma, suggesting that hormones were locally produced within the uterine compartment. Embryonic sexual differentiation based on the presence of visible claspers (male copulatory organs) coincided with peaks in histotroph progesterone, testosterone, 17-hydroxyprogesterone and estradiol, suggesting that, like mammalian pregnancy, elasmobranch embryonic steroids also contribute to their own developmental environment.

HEMATOLOGIC AND BIOCHEMICAL SUMMARY STATISTICS IN AQUARIUM-HOUSED SPOTTED EAGLE RAYS ( AETOBATUS NARINARI).

During this retrospective study, 18 plasma blood chemistry and 17 complete blood count (CBC) samples were analyzed from clinically healthy spotted eagle rays ( Aetobatus narinari) at Georgia Aquarium in order to generate hematological ranges for complete blood count (CBC) and biochemical profiles. Summary statistics were generated according to the American Society for Veterinary Clinical Pathology guidelines for the determination of reference intervals in veterinary species. 4 The mean packed cell volume (PCV) was 28.09% with a range of 23-35%. Mean total solids were 5.72 g/dl with a range of 5-7.0 g/dl. Lymphocytes were the dominant leukocyte observed on differential (67.35%), followed by fine eosinophilic granulocytes (FEGs) (15.41%), coarse eosinophilic granulocytes (CEGs) (10.24%), monocytes (1.88%), and basophils (1.24%). Chemistry samples were analyzed at two diagnostic laboratories, Michigan State University (MSU) and University of Miami (UMiami), and the results were compared. Both labs have the capacity to run blood chemistries on zoo and aquatic species, but utilize different methods to obtain chemistry analyte values. UMiami uses a thin-film dry-slide technology, whereas MSU uses an ion-selective electrode (ISE) and Beckman Coulter AU 640 analyzer. There is poor agreement between the analyzers used by the two laboratories for both alkaline phosphatase and BUN, because of proportional error. Establishing hematological ranges in spotted eagle rays and in elasmobranchs in general may enhance the understanding of the species and their health. This information may aid clinicians in deciding when and how to treat elasmobranchs.

REPRODUCTIVE HORMONE PATTERNS IN MALE AND FEMALE COWNOSE RAYS ( RHINOPTERA BONASUS) IN AN AQUARIUM SETTING AND CORRELATION TO ULTRASONOGRAPHIC STAGING.

Reproductive management of cownose rays ( Rhinoptera bonasus) under professional care plays an important role in conservation of the species, but hormone and ultrasonographic analyses of their 12-mo reproductive cycle have not been documented previously. Plasma reproductive hormone concentrations (17B-estradiol, progesterone, testosterone, and androstenedione) were measured monthly via radioimmunoassay for 1 yr in an aquarium-managed population of adult females ( n = 15) and males ( n = 5). Ultrasounds of the uterus were performed each month at the time of sample collection to identify gestation stage (0-5) based on a previously developed in-house staging system. Stages were correlated to hormone concentrations to track progression through pregnancy. Thirteen females were reproductively active, and each produced one pup in March-June, similar to timing for free-ranging populations. Female estradiol increased steadily throughout gestation from stages 0 to 5, while progesterone, testosterone, and androstenedione were increased only in early gestation (stages 1 and 2). Unlike month of year, gestation stage strongly predicted hormone concentration, but specific values to predict parturition date were not identified. Male testosterone and progesterone were higher in March-June (mating season) than July-January, while estradiol and androstenedione did not exhibit a seasonal pattern. Aquarium-managed cownose rays have similar reproductive patterns to what is reported in wild populations. Ultrasonographic monitoring with serial hormone analysis and accurate mating records will provide the most useful information for managing a reproductive population of cownose rays in an aquarium setting.

Ecophysiological interactions and water-related physicochemical parameters among freshwater stingrays / Interações ecofisiológicas e dos parâmetros físico-químico da água em arraias de água doce

Abstract The objective of this study was to compare and correlate the ecology of neonates and young individuals of Potamotrygon wallacei, Potamotrygon motoro and Paratrygon aiereba with regard to their hematological profile and the physicochemical parameters of the water that they inhabit. Principal component analysis (PCA) on the complete blood count revealed total variation of 72.92%, thus demonstrating a differentiation system for oxygen demand. On the other hand, P. motoro was considered to be an intermediate species, given that its complete blood count characteristics interacted with both P. wallacei and with P. aiereba. The interaction among the biochemical variables was shown to total 64.67% of the factors. This allowed differentiation of P. wallacei from P. aiereba, while P. motoro maintained an intermediate position. These characteristics of differentiation within the preferred environment corroborate the PCA of the present study and confirm that these species can be differentiated through considering the complete blood count and biochemical parameters. The PCA on water properties showed 68.57% differentiation, mainly comprising the x axis (49.44%). It can be affirmed that P. motoro has the capacity to inhabit the preferential areas of P. wallacei and P. aiereba, as well as occupying localities in which other stingrays are not found. In conclusion, P. wallacei presents patterns differentiating it from P. aiereba, while P. motoro is a species that presents intermediate characteristics. The latter can be considered to be a more broadly distributed species regarding its ecophysiological characteristics.

Resumo Este trabalho tem por objetivo investigar o perfil hematológico e os parâmetros físico-químicos da água, comparando e correlacionando ecologicamente entre neonatos e jovens de Potamotrygon wallacei (arraia cururu), Potamotrygon motoro e Paratrygon aiereba. A análise de componentes principais (PCA) do hemograma revelou um total 72,92% de variação, constituindo-se em um sistema de diferenciação na demanda por oxigênio. P. wallacei apresenta diferenciação no eixo X quando comparada a P. aiereba, por outro lado P. motoro constitui-se como uma espécie intermediária que apresenta as características do hemograma interagindo tanto com P. wallacei quanto com P. aiereba. A interação entre as variáveis bioquímica demonstram um total de 64,67% dos fatores, no qual foi possível diferenciar, a arraia P. wallacei de P. aiereba, tendo P. motoro uma aspecto de espécie intermediária entre as demais. Esses aspectos de diferenciação de ambiente de preferência corroboraram a PCA obtida no presente estudo e confirmam que essas espécies podem ser diferenciadas quando se considerar as variáveis referentes ao hemograma e a bioquímica. Nos íons, no trombograma e no leucograma, não foi possível diferenciar as espécies. O PCA das propriedades da água foi constituído por 68,57% de diferenciação que se constituiu principalmente no eixo x (49,44%). É possível confirmar que P. motoro tem a capacidade de habitar as áreas preferências de P. wallacei e P. aiereba, além do mais esta possui uma localidade que as demais arraias não são encontradas. Conclui-se que P. wallacei, apresenta padrões diferenciados de P. aiereba, além do mais P. motoro é uma espécie que apresenta características intermediárias entre as descritas, o qual pode ser considerado uma espécie com distribuição mais ampla em seus aspectos ecofisiológicos.

Molecular characterization of two Rhesus glycoproteins from the euryhaline freshwater white-rimmed stingray, Himantura signifer, and changes in their transcript levels and protein abundance in the gills, kidney, and liver during brackish water acclimation.

Himantura signifer is a freshwater stingray which inhabits rivers in Southeast Asia. It is ammonotelic in fresh water, but retains the capacities of urea synthesis and ureosmotic osmoregulation to survive in brackish water. This study aimed to elucidate the roles of Rhesus glycoproteins (Rhgp), which are known to transport ammonia, in conserving nitrogen (N) in H. signifer during brackish water acclimation when N became limited resulting from increased hepatic urea synthesis. The complete coding sequence of rhbg from H. signifer consisted of 1383 bp, encoding 460 amino acids with an estimated molecular mass of 50.5 kDa, while that of rhcg comprised 1395 bp, encoding for 464 amino acids with an estimated molecular mass of 50.8 kDa. The deduced amino sequences of Rhbg and Rhcg contained ammonia binding sites, which could recruit NH4+ to be deprotonated, and a hydrophobic pore with two histidine residues, which could mediate the transport of NH3. Our results indicated for the first time that brackish water acclimation resulted in significant decreases in the expression levels of rhbg/Rhbg and rhcg/Rhcg in the gills of H. signifer, which offered a mechanistic explanation of brackish water-related decreased ammonia excretion reported elsewhere. Furthermore, rhbg/Rhbg expression levels increased significantly in the liver of H. signifer during brackish water acclimation, indicating that the ammonia produced by extra-hepatic tissues and released into the blood could be channeled into the liver for increased urea synthesis. Overall, these results lend support to the proposition that H. signifer becomes N-limited upon utilizing urea as an osmolyte in brackish water.

Ecophysiological interactions and water-related physicochemical parameters among freshwater stingrays.

The objective of this study was to compare and correlate the ecology of neonates and young individuals of Potamotrygon wallacei, Potamotrygon motoro and Paratrygon aiereba with regard to their hematological profile and the physicochemical parameters of the water that they inhabit. Principal component analysis (PCA) on the complete blood count revealed total variation of 72.92%, thus demonstrating a differentiation system for oxygen demand. On the other hand, P. motoro was considered to be an intermediate species, given that its complete blood count characteristics interacted with both P. wallacei and with P. aiereba. The interaction among the biochemical variables was shown to total 64.67% of the factors. This allowed differentiation of P. wallacei from P. aiereba, while P. motoro maintained an intermediate position. These characteristics of differentiation within the preferred environment corroborate the PCA of the present study and confirm that these species can be differentiated through considering the complete blood count and biochemical parameters. The PCA on water properties showed 68.57% differentiation, mainly comprising the x axis (49.44%). It can be affirmed that P. motoro has the capacity to inhabit the preferential areas of P. wallacei and P. aiereba, as well as occupying localities in which other stingrays are not found. In conclusion, P. wallacei presents patterns differentiating it from P. aiereba, while P. motoro is a species that presents intermediate characteristics. The latter can be considered to be a more broadly distributed species regarding its ecophysiological characteristics.

EVALUATION OF HEALTH PARAMETERS IN COWNOSE RAYS ( RHINOPTERA BONASUS) HOUSED IN A SEASONAL TOUCH POOL HABITAT COMPARED WITH AN OFF-EXHIBIT HABITAT.

Cownose rays ( Rhinoptera bonasus) are commonly displayed in zoo and aquarium touch pool exhibits; however, there is a gap in our understanding of how these practices might impact the health of these animals. The aim of this study was to evaluate and compare selected health parameters in cownose rays housed in a seasonal outdoor exhibit touch pool system with abundant public contact and an indoor off-exhibit holding system with minimal human contact. All animals underwent physical examination, ultrasound, cloacal wash and cytology, and blood collection for complete blood counts, point-of-care blood analysis, plasma protein electrophoresis, and plasma cholesterol electrophoresis in May and October 2014. Physical examination, ultrasound, and cloacal wash cytology findings were all unremarkable for both groups of animals. Significant differences in health parameters among animals by location and time point were few and included decreased heart rate ( F = 12.158, P = 0.001), increased lactate ( F = 6.838, P = 0.012), and increased low-density lipoproteins ( F = 19.961, P = 0.000) in touch pool animals over time. Based on these results, cownose rays inhabiting a touch pool exhibit and an off-exhibit system remained in comparable planes of health based on routine diagnostic modalities with few differences in measured health parameters.

WHOLE BLOOD TRANSFUSIONS TO TREAT SEVERE ANEMIA IN SEVEN COWNOSE RAYS ( RHINOPTERA BONASUS) AND ONE SHORT-TAIL STINGRAY ( DASYATIS BREVICAUDATA).

Blood transfusions can provide life-saving treatment to severely anemic animals. Due to limited availability and the difficulty of storing whole blood and blood products, such as fresh frozen plasma and packed red blood cells, exotic animals often receive fresh whole blood transfusions. Little is known about elasmobranch blood types and transfusions. Conspecific cross-matches within several different elasmobranch species were negative, indicating that in an emergency situation a single whole blood transfusion may be possible without causing a transfusion reaction. Experimental transfusions between healthy conspecific Atlantic rays ( Dasyatis sabina) showed no adverse reactions and autotransfusions in marbled electric rays ( Torpedo marmorata) were successful. There are no published reports of blood transfusions performed on clinically abnormal elasmobranchs. The following case series documents blood transfusions performed on seven cownose rays ( Rhinoptera bonasus) and one short-tail stingray ( Dasyatis brevicaudata). All rays were treated with the same protocol, which included pretreatment with steroids and antibiotics followed by an intravenous transfusion of freshly collected, heparinized, whole blood. Three animals survived and currently exhibit no abnormal clinical signs. Two animals died 55 days and 100 days post transfusion. Three animals died 2-22 days post transfusion. Although complications from blood transfusions could not be ruled out, all five animals that died had other health problems that likely contributed to their demise. All eight animals would almost certainly have died without a blood transfusion as they were severely anemic and moribund at the time of presentation. The methods described in this paper may be useful in the treatment of severely anemic elasmobranchs and this is the first report of blood transfusions in clinically abnormal elasmobranchs.

Comparison of hematology, plasma biochemistry, and blood gas variables between 2 venipuncture sites in Southern Stingrays (Dasyatis americana).

BACKGROUND: The Southern Stingray (Dasyatis americana) is a batoid elasmobranch frequently exhibited in zoological institutions. Blood is commonly collected from the caudal hemal arch at the tail base in stingrays for the purpose of health assessment and clinical pathology tests. An alternative site that allows a dorsal or ventral approach without necessitating puncture of a cartilaginous structure has been identified between the cartilaginous pectoral fin rays (ceratotrichia). OBJECTIVES: The purpose of the study was to compare CBC, plasma biochemistry analytes, and blood gas variables between blood samples collected from the caudal and pectoral fin vasculature sites of the Southern Stingray. METHODS: Fifteen captive Southern Stingrays (10 females, 5 males) from 4 zoo and aquarium facilities were sampled. Lithium heparinized blood samples were collected from the caudal and pectoral venipuncture sites of each animal. Values from estimated total and differential leukocyte counts, plasma biochemistry analytes, and blood gas variables were compared. RESULTS: There were no statistically significant differences between venipuncture sites for the measured analytes except for CK activity, which was statistically significantly higher in the pectoral site samples. Levels of agreement between sites were good or moderate for 22 analytes and poor for ALT, AST, CK, pO2 , lactate, monocytes, and eosinophils. CONCLUSIONS: The good agreement between sampling sites for the majority of the measured analytes and the lack of differences that would alter clinical interpretation support the use of the pectoral site as an alternative to the traditional caudal fin venipuncture site in Southern Stingrays.

COMPARISON OF PRE- AND POSTQUARANTINE BLOOD CHEMISTRY AND HEMATOLOGY VALUES FROM WILD-CAUGHT COWNOSE RAYS (RHINOPTERA BONASUS).

Though one of the most widely kept elasmobranchs in human care, the cownose ray (CNR; Rhinoptera bonasus ), remains a species with minimal published information on hematologic reference intervals. As part of a larger study investigating the health and nutrition of the CNR, this study established a preliminary data set of plasma chemistry and hematology values specific to animals recently caught from the wild and compared this data set (intake sample) to values obtained following a period of quarantine (27-40 days) in an aquarium (exit sample). Blood samples were collected from 47 wild female (n = 46) and male (n = 1) CNR caught in pound nets off the coast of North Carolina and South Carolina. Differences between intake and exit values were analyzed. Due to the preponderance of female animals, data were not analyzed for sex differences. Plasma biochemical profiles were performed and analyzed. A select number of complete blood cell counts were performed (n = 24 from 12 animals). Statistically significant differences (P < 0.05) specific to time of sampling were determined for packed cell volume, total solids, blood urea nitrogen, sodium, chloride, potassium, phosphorus, cholesterol, glucose, and aspartate aminotransferase. Values reported are a significant expansion on the existing limited data for CNRs and will serve as a reference for health assessment of individuals both in the wild and in exhibit populations.

Na⁺/K⁺-ATPase α1 mRNA expression in the gill and rectal gland of the Atlantic stingray, Dasyatis sabina, following acclimation to increased salinity.

BACKGROUND: The salt-secreting rectal gland plays a major role in elasmobranch osmoregulation, facilitating ion balance in hyperosmotic environments in a manner analogous to the teleost gill. Several studies have examined the central role of the sodium pump Na(+)/K(+)-ATPase in osmoregulatory tissues of euryhaline elasmobranch species, including regulation of Na(+)/K(+)-ATPase activity and abundance in response to salinity acclimation. However, while the transcriptional regulation of Na(+)/K(+)-ATPase in the teleost gill has been well documented the potential for mRNA regulation to facilitate rectal gland plasticity during salinity acclimation in elasmobranchs has not been examined. Therefore, in this study we acclimated Atlantic stingrays, Dasyatis sabina (Lesueur) from 11 to 34 ppt salinity over 3 days, and examined changes in plasma components as well as gill and rectal gland Na(+)/K(+)-ATPase α1 (atp1a1) mRNA expression. RESULTS: Acclimation to increased salinity did not affect hematocrit but resulted in significant increases in plasma osmolality, chloride and urea. Rectal gland atp1a1 mRNA expression was higher in 34 ppt-acclimated D. sabina vs. CONTROLS: There was no significant change in gill atp1a1 mRNA expression, however mRNA expression of this gene in the gill and rectal gland were negatively correlated. CONCLUSIONS: This study demonstrates regulation of atp1a1 in the elasmobranch salt-secreting gland in response to salinity acclimation and a negative relationship between rectal gland and gill atp1a1 expression. These results support the hypothesis that the gill and rectal gland play opposing roles in ion balance with the gill potentially facilitating ion uptake in hypoosmotic environments. Future studies should further examine this possibility as well as potential differences in the regulation of Na(+)/K(+)-ATPase gene expression between euryhaline and stenohaline elasmobranch species.

Hematology, biochemical profile and thyroid hormones of four species of freshwater stingrays of the genus Potamotrygon / Hematologia, perfil bioquímico e hormônios da tireoide em quatro espécies de arraias de água doce do gênero Potamotrygon

Potamotrygonidae is a family of freshwater stingray endemic in South America. We determined the hematological and serum biochemistry parameters of Potamotrygon motoro and Potamotrygon falkneri from Parana Basin and Potamotrygon orbignyi and Potamotrygon scobina from the Amazon Basin. Blood was collected from 55 specimens of Potamotrygon sp., and these parameters were evaluated: red blood cell count, hematocrit percent, hemoglobin concentration, leucocytes count, trombocytes count, total protein concentration, albumin, uric acid, urea, creatinine, triglycerides, cholesterol, high-density lipoprotein HDL, calcium, phosphorus, calcium/phosphorus relationship, creatine kinase CK, gamma-glutamyl transpeptidase GGT, Aspartate transaminase AST, alanine transaminase ALT, globulin, albumin/globulin relationship, alkaline phosphatase AP, sodium, chloride, potassium, triiodothyronine T3, and thyroxine T4 hormones. The hematological indices were similar for all species, except for levels of heterophils in P. orbignyi and P. scobina that were higher. Calcium, phosphorus, calcium/phosphorus relationship, sodium, potassium, chloride, AST, ALT, GGT, CK, albumin/globulin relationship, uric acid, creatinine, urea, triglycerides, HDL, T3, T4 levels had no significant difference between the species. These results suggest that there is low influence of habitat and feeding habits. AP, total protein, albumin, globulin, glucose, and total cholesterol had significant differences between the studied species. These results support the theory that stingrays migrated from the Atlantic Ocean, adapting to different conditions over time and placed themselves geographically distant from each other.

Potamotrygonidae e um grupo de raias endêmicas da América do Sul distribuídas nas principais bacias hidrográficas brasileiras, mas informações sobre suas variáveis hematológicas e bioquímicas são escassas. Este estudo teve como objetivo determinar estas variáveis em Potamotrygon motoro e P. falkneri na Bacia do Rio Paraná, Estado do Paraná e de P. orbignyi e P. scobina do Rio Piririm, na Bacia Amazônica, Estado do Amapá. Foi capturado um total de 53 espécimes de Potamotrygon spp. para colheita de sangue e avaliações do hemograma e variáveis bioquímicas séricas. Os valores do hemograma foram próximos para as quatro espécies e os resultados com diferenças significativas entre Potamotrygon falkneri, P. motoro, P. orbigni e P. scobina ocorreram quanto ao número de eritrócitos e heterófilos. Apesar da diversidade geográfica os resultados apresentaram pouca interferência dos diferentes habitats nas variáveis avaliadas. Os níveis de proteínas, globulinas, relação albumina:globulina, acido úrico, creatinina, ureia, HDL-C, relação cálcio:fósforo, sódio, cloreto, AST, ALT, creatinoquinase (CK), fosfatase alcalina, hormônios triiodotironina (T3) e tiroxina (T4) não diferiram entre as espécies de Potamotrygon. Porem, os níveis de albumina, glicose, colesterol total, triglicerídeos e gama glutamil transferase (GGT) apresentaram diferenças. Este fato sugere pouca interferência do habitat nas variáveis avaliadas apesar da diversidade geográfica de origem das raias de vida livre.

Diversity of bacteria cultured from the blood of lesser electric rays caught in the northern gulf of Mexico.

The prevalence and taxonomic diversity of bacteria cultured from the blood of apparently healthy Lesser Electric Rays Narcine bancroftii captured from open beach habitat in the north-central Gulf of Mexico are reported herein. The blood of 9 out of 10 Lesser Electric Rays was positive for bacteria, and bacterial isolates (n = 83) were identified by 16S rRNA gene sequencing. The majority of the isolates belonged to the phylum Proteobacteria (91.5%). Vibrio spp. comprised 53% of all isolates and were recovered from all Lesser Electric Rays with culture-positive blood. Among them, V. harveyi (n = 14) and V. campbellii (n = 11) were most common, followed by a group of unidentified Vibrio sp. (n = 10) related to V. nigripulchritudo. Isolates representing other species of Proteobacteria included Pseudoalteromonas (n = 13), Shewanella (n = 5), Amphritea (n = 3), Nautella (n = 3), and Arenibacter (n = 1). Higher bacterial diversity was observed in blood cultured on marine agar relative to blood agar, but gram-positive bacteria were isolated from the latter only. The 16S rRNA gene sequences of bacterial isolates were compared phylogenetically to those from related type strains. Most isolates were identified to the level of species, but some clustered independently from reference strains, likely representing new species of Vibrio, Amphritea, Shewanella, and Tenacibaculum. The present study is the first record of any bacterium from this ray species and reveals a taxonomically and phylogenetically diverse microbiota associated with its blood. Moreover, these data document that the presence of bacteria in elasmobranch blood is not coincident with clinical signs of disease, thereby rejecting the paradigm of septicemia indicating a disease condition in aquatic vertebrates.

Some euryhalinity may be more common than expected in marine elasmobranchs: the example of the South American skate Zapteryx brevirostris (Elasmobranchii, Rajiformes, Rhinobatidae).

Elasmobranchs are essentially marine, but ~15% of the species occur in brackish or freshwater. The Brazilian marine coastal skate Zapteryx brevirostris, non-reported in nearby estuaries, was submitted to 35, 25, 15, and 5 psu, for 6 or 12h (n=6). Plasma was assayed for osmolality, urea, and ions (Na(+), Cl(-), K(+), Mg(2+)). Muscle water content was determined, and the rectal gland, kidney and gills were removed for carbonic anhydrase (CA) and Na(+),K(+)-ATPase (NKA) activities. The skate survived to all treatments. Plasma osmolality and urea levels decreased respectively by 27% and 38% after 12h in 5 psu (with respect to levels when in seawater), but plasma Na(+), Cl(-), and Mg(2+) were well regulated. Plasma K(+) showed some conformation after 12h. Muscle hydration was maintained. Branchial CA and NKA did not respond to salinity. Rectal gland NKA decreased upon seawater dilution, while renal NKA increased. This skate was shown to be partially euryhaline. The analysis of plasma urea of elasmobranchs in brackish and freshwater versus salinity and time-allied to the widespread occurrence of some euryhalinity in the group-led us to revisit the hypothesis of a brackish water habitat for elasmobranch ancestors.

Differences in activity level between cownose rays (Rhinoptera bonasus) and Atlantic stingrays (Dasyatis sabina) are related to differences in heart mass, hemoglobin concentration, and gill surface area.

Aquatic animals are faced with the challenge of extracting oxygen from water, a medium that is metabolically expensive to ventilate and that contains just a fraction of the oxygen concentration relative to air, yet the physiologies of fishes have evolved to support a wide range of activity levels in nature. Oxygen delivery components, including gill surface area (oxygen uptake), blood chemistry (oxygen transport), and the heart (system pump), have been positively correlated to activity level in teleost fishes, yet relatively little is known about how these components are related to activity in elasmobranches. The current study addresses this question by examining heart mass, hemoglobin concentration, hematocrit level, and gill surface area in wild-caught representatives of the benthic Atlantic stingray (Dasyatis sabina) and active cownose ray (Rhinoptera bonasus). Allometric scaling exponents are similar for all four measures between the study species. Heart mass, gill surfaces areas, and hemoglobin concentrations were 2.1 times, approximately 7.1 times, and 2.0 times higher, respectively, in active cownose rays, when compared to benthic Atlantic stingrays, after correcting for differences in body mass. When considered in the context of functional plasticity within the oxygen delivery systems of benthic and active species, data from the current study indicate that higher activity levels in cownose rays are supported by modifications that, at least in part, are likely to enhance oxygen uptake.

Stress responses of the endemic freshwater cururu stingray (Potamotrygon cf. histrix) during transportation in the Amazon region of the Rio Negro.

Potamotrygon cf. histrix (cururu stingray) are endemic freshwater stingrays from the middle region of the Rio Negro in the Brazilian Amazon basin and are exported worldwide as ornamentals caught by artisanal fishermen. The transport process from capture to final destination is long and stressful. This study quantified stress related changes in corticosterone, blood and water samples (baseline, pre-transport, 3h, 12h and 24h) analyzed during a transport experiment which tested two water additives (tetracycline and the probiotic Efinol). There was a significant stepwise increase in corticosterone levels in stingrays over transport time in combination with osmoregulatory disturbances suggesting a stress related role of this corticosteroid. There were significant increases in water conductivity, Na(+) and K(+) losses and ammonia excretion. Blood parameters such as glucose, hematocrit, red blood count and urea did not change significantly during the experiment. Glucose levels did not increase significantly during transport and this may be due to the fact that other elasmobranchs have been shown to rely more on ketone bodies for energy rather than glucose and produce ammonia as their main nitrogenous waste. The mineralocorticoid action of this hormone has been shown in elasmobranchs and most likely plays a role in osmotic homeostasis. The use of probiotic and especially antibiotic should be avoided since no beneficial effects were observed.