An investigation of gill and blood carbonic anhydrase characteristics in three basal actinopterygian species: alligator gar (Atractosteus spatula), white sturgeon (Acipenser transmontanus) and Senegal bichir (Polypterus senegalus).

Many teleosts possess a unique set of respiratory characteristics allowing enhanced oxygen unloading to the tissues during stress. This system comprises three major components: highly pH sensitive haemoglobins (large Bohr and Root effects), rapid red blood cell (RBC) intracellular pH (pHi) protection, and a heterogeneous distribution of membrane-bound plasma-accessible carbonic anhydrase (paCA; absence in the gills). The first two components have received considerable research effort; however, the evolutionary loss of branchial paCA has received little attention. In the current study, we investigated the availability of branchial membrane-bound CA, along with several other CA-related characteristics in species belonging to three basal actinopterygian groups: the Lepisosteiformes, Acipenseriformes and Polypteriformes to assess the earlier hypothesis that Root effect haemoglobins constrain branchial paCA availability. We present the first evidence suggesting branchial membrane-bound CA presence in a basal actinopterygian species: the Senegal bichir (Polypterus senegalus) and show that like the teleosts, white sturgeon (Acipenser transmontanus) and alligator gar (Atractosteus spatula) do not possess branchial membrane-bound CA. We discuss the varying respiratory strategies for these species and propose that branchial paCA may have been lost much earlier than previously thought, likely in relation to the changes in haemoglobin buffer capacity associated with the increasing magnitude of the Bohr effect. The findings described here represent an important advancement in our understanding of the evolution of the unique system of enhanced oxygen unloading thought to be present in most teleosts, a group that encompasses half of all vertebrates.

Myxobolus spp. (Cnidaria: Myxobolidae) in the circulating blood of fishes from Goiás and Mato Grosso States, Brazil: case report / Myxobolus spp. (Cnidaria: Myxobolidae) no sangue circulante de peixes dos estados de Goiás e Mato Grosso, Brasil: relato de caso

Myxosporidiosis is an infectious disease caused by myxozoans of the Phylum Cnidaria, Class Myxosporea, and Order Bivalvulida, considered a common parasite in fresh and saltwater fishes that parasitize many organs, especially gills. In the present study, 49 specimens of fishes belonging to eight genera: Tetragonopterus, Leporinus, Myleus, Pirinampus, Rhapiodon, Pygocentrus, Ageneiosus, and Serrasalmus were collected and blood smears were made, fixed with absolute methanol, and stained with Giemsa 10% to survey hemoparasites. However, myxospores were found in the circulating blood of five (10.20%) fishes belonging to genus Tetragonopterus, Myleus, and Pygocentrus. Two morphological types of Myxobolus spp. were identified in all the five fish specimens analyzed. Usually, investigations on myxozoans in fish are carried out with the search for plasmodia or cysts in the fish organs and observation of the cavity of organs. Nevertheless, this study highlights the importance of also examining the blood of these animals, since these parasites can cause severe pathogenic diseases in fish. Thus, the blood analyses can proportionate preventive sanitary control for commercial fish avoiding economic loss.

A mixosporidiose é considerada uma doença infecciosa causada por mixozoários pertentences ao Filo Cnidaria, Classe Myxosporea e Ordem Bivalvulida, considerados parasitos comuns de peixes de água doce e salgada, parasitando vários órgãos, principalmente as brânquias. No presente estudo, 49 espécimes de peixes pertencentes a oito gêneros: Tetragonopterus, Leporinus, Myleus, Pirinampus, Rhapiodon, Pygocentrus, Ageneiosus and Serrasalmus foram coletados e extensões sanguíneas foram feitas com a finalidade de encontrar hemoparasitos. Entretanto, mixoesporos foram observados em cinco (10.20%) espécimes de peixes, pertencentes aos gêneros Tetragonopterus, Myleus e Pygocentrus. Dois morfotipos de Myxobolus spp. foram identificados parasitando esses peixes. Normalmente, as investigações sobre mixozoários em peixes é realizada com a procura de plasmódios ou cistos nos órgãos dos peixes e observação da cavidade de órgãos. Porém, esse trabalho ressalta a importância de se examinar também o sangue desses animais, já que esses parasitos podem ocasionar patogenias severas em peixes. Sendo assim, a análise sanguínea pode proporcionar controle sanatório preventivo para peixes comerciais e evitar perdas econômicas.

A new experimental model for the investigation of sequential hermaphroditism.

The stunning sexual transformation commonly triggered by age, size or social context in some fishes is one of the best examples of phenotypic plasticity thus far described. To date our understanding of this process is dominated by studies on a handful of subtropical and tropical teleosts, often in wild settings. Here we have established the protogynous New Zealand spotty wrasse, Notolabrus celidotus, as a temperate model for the experimental investigation of sex change. Captive fish were induced to change sex using aromatase inhibition or manipulation of social groups. Complete female-to-male transition occurred over 60 days in both cases and time-series sampling was used to quantify changes in hormone production, gene expression and gonadal cellular anatomy. Early-stage decreases in plasma 17ß-estradiol (E2) concentrations or gonadal aromatase (cyp19a1a) expression were not detected in spotty wrasse, despite these being commonly associated with the onset of sex change in subtropical and tropical protogynous (female-to-male) hermaphrodites. In contrast, expression of the masculinising factor amh (anti-Müllerian hormone) increased during early sex change, implying a potential role as a proximate trigger for masculinisation. Collectively, these data provide a foundation for the spotty wrasse as a temperate teleost model to study sex change and cell fate in vertebrates.

Evidence for a Novel Antiviral Mechanism of Teleost Fish: Serum-Derived Exosomes Inhibit Virus Replication through Incorporating Mx1 Protein.

Exosomes are associated with cancer progression, pregnancy, cardiovascular diseases, central nervous system-related diseases, immune responses and viral pathogenicity. However, study on the role of exosomes in the immune response of teleost fish, especially antiviral immunity, is limited. Herein, serum-derived exosomes from mandarin fish were used to investigate the antiviral effect on the exosomes of teleost fish. Exosomes isolated from mandarin fish serum by ultra-centrifugation were internalized by mandarin fish fry cells and were able to inhibit Infectious spleen and kidney necrosis virus (ISKNV) infection. To further investigate the underlying mechanisms of exosomes in inhibiting ISKNV infection, the protein composition of serum-derived exosomes was analyzed by mass spectrometry. It was found that myxovirus resistance 1 (Mx1) was incorporated by exosomes. Furthermore, the mandarin fish Mx1 protein was proven to be transferred into the recipient cells though exosomes. Our results showed that the serum-derived exosomes from mandarin fish could inhibit ISKNV replication, which suggested an underlying mechanism of the exosome antivirus in that it incorporates Mx1 protein and delivery into recipient cells. This study provided evidence for the important antiviral role of exosomes in the immune system of teleost fish.

Gonadal steroid fluctuations and reproduction results of European grayling (Thymallus thymallus L.) from broodstock farmed in a recirculation aquaculture system.

The initial experiment of this study was conducted to determine whether sex steroid concentrations varied before, during and after the reproductive season of European grayling (Thymallus thymallus L.) farmed broodstock confined in a recirculation aquaculture system (RAS). The results indicated that the plasma sex steroid (testosterone - T, progesterone - P4 and estradiol - E2) concentrations varied (P < 0.05) during these reproductive periods. There were greater concentrations of T, P4 and E2 before and during the grayling reproductive season indicating there are important functions of these steroid hormones associated with gonadal maturation, gamete production and reproductive behavior during the spawning period. In the second experiment of this study, the effectiveness of European grayling controlled reproduction treatment regimens was analyzed and there were 34.69 % and 68.18 % having ovulations in the 2 and 3-year-old broodfish, respectively. Of the embryos developing to the eyed-egg stage, there were 3.70 % and 6.87 % derived from eggs of 2- and 3-year-old grayling females, respectively. Proportions of embryos developing from the eyed-egg stage to hatching were 38.1 % and 52.1 % from eggs of 2- and 3-year-old grayling females, respectively, when there was culturing in a RAS. The results indicate that with grayling broodfish there is greater efficacy in induction of reproduction when there is imposing of the controlled reproduction treatment regimen on 3- rather than 2-year-old broodfish.

Haematology of Mugil curema from the north coast estuary of Santa Catarina state, Brazil / Hematologia de Mugil curema em estuário no litoral norte do estado de Santa Catarina, Brasil

Mugil curema is a teleost fish often found in coastal, estuarine and lagoon regions throughout the Brazilian coast. Blood parameters may be used as health and welfare indicators of the fish, as well as the environment they live in. The aim of this study was to verify the hematologic profile of the M.curema along 18 months in estuary waters from the north coast of the state of Santa Catarina in the south of Brazil. A total of 133 M. curema fish were captured from March 2016 to August 2017. Erythrogram, leukogram, thrombocytogram, hematocrit percentage, hemoglobin dosage, total plasma protein dosage, mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH) and medium corpuscular hemoglobin concentration (MCHC) were analyzed. There was no significant difference (P>0.05) on the total number of erythrocytes between the fish gender. However, there was difference on the hematocrit between males and indefinite gender fish. The influences of temporal and geographical space, among the values of hematological parameters, in the different studies of the Brazilian coast are discussed.. The present research is an important contribution to establish a hematological pattern for M. curema. The local population of the studied fish has a proper behavior in hematological descriptors, which is different from other Brazilian coast studies with the same fish species.

Mugil curema é um peixe teleósteo, abundante em regiões costeiras, estuarinas e lagunares, em todo o litoral brasileiro. Os parâmetros sanguíneos podem ser usados como indicadores da saúde e do bem-estar dos peixes, bem como do ambiente onde estes vivem. O objetivo deste estudo foi verificar o perfil hematológico de M. curema ao longo de 18 meses, em águas estuarinas, no Litoral norte do estado de Santa Catarina, sul do Brasil. Um total de 133 peixes foram capturados, entre março de 2016 e agosto de 2017, para realização de eritrograma, leucograma, trombocitograma, porcentagem de hematócrito, dosagem de hemoglobina, dosagem de proteína plasmática total, volume corpuscular médio (VCM), hemoglobina corpuscular média (HCM) e concentração de hemoglobina corpuscular média (CHCM). Não houve diferença significativa (P>0,05) entre o número total de eritrócitos entre machos e fêmeas. No entanto, houve diferença no hematócrito, entre machos e peixes de sexo indeterminado. As influências do espaço temporal e geográfico, entre os valores dos parâmetros hematológicos, nos diferentes estudos do litoral brasileiro são discutidas. A presente investigação é importante contribuição para o estabelecimento de um padrão hematológico para M. curema. A população local de peixes estudada tem comportamento próprio em descritores hematológicos, diferenciado de outros estudos no litoral brasileiro para a mesma espécie.

Erythrocyte profile of circulating blood of Neogobius melanostomus (Pallas, 1814) under conditions of experimental hypothermia.

The influence of hypothermia on erythrocyte profile of thermophile teleost species round goby, Neogobius melanostomus (Pallas, 1814), has been studied. Fish were acclimated to temperature 1-2оС, 15-16оС and 19-20оС (control group) and held at given conditions for 5 days. The number of red blood cell precursors (pronormoblasts, basophilic and polychromatophilic normoblasts) in circulating blood has been estimated. Also, the number of abnormal erythrocytes, i.e. cells with micronuclei, nuclei invaginations, red blood cell shades, dacryocytes and cells undergoing amitosis has been determined on smears. The number of immature erythrocytes increased more than two times (p < 0,001) at 1-2оС. The number of low-differentiated precursors, pronormoblasts and early basophilic normoblasts, increased for the most part. The number of abnormal erythrocytes did not change substantially, The changes in cellular blood composition were accompanied with the increase of plasma lactate concentration, indicating hypoxic state of fish. The results of the present work indicate that hematopoietic tissue remains sensitive to controlling factors at hypothermia, such as hypoxia, and may enhance proliferation and differentiation of erythroid cells.

Development of specific enzyme-linked immunosorbent assays for multiple vitellogenins in marbled sole, Pleuronectes yokohamae.

Non-competitive, enzyme-linked immunosorbent assays (ELISAs) for three distinct sole vitellogenins (VtgAa, VtgAb and VtgC) were designed using their purified lipovitellin (Lv) products and corresponding digoxigenin-labeled, anti-Lv polyclonal antibodies, primarily for employment in monitoring estrogenic pollution of the environment. The working range of the ELISAs was from 0.97 to 1,000 ng/mL for all Vtg subtypes. Each ELISA appeared to be specific to the targeted Vtg subtype. Intra- and inter-assay coefficients of variation in the developed ELISAs were lower than 10%. Three Vtg subtypes were induced in serum of immature fish by estradiol-17ß (E2) injection (0.5 mg/kg body weight). All Vtg subtypes were induced one day after the injection, reaching peak levels (Lv equivalents) within three days, as follows: 39.1 ±â€¯28.9 µg/mL (VtgAa), 57.9 ±â€¯30.7 µg/mL (VtgAb) and 12.6 ±â€¯4.8 µg/mL (VtgC). In wild-caught males, VtgAa, VtgAb and VtgC were detected in ranges from 0.26 to 1.21, 0.19 to 8.69, and 0.17 to 53.50 µg/mL, respectively, over various sampling periods. In vitellogenic females sampled in January, the average level of VtgAb (8,744.43 ±â€¯733.93 µg/mL) was significantly higher than for VtgAa (150.33 ±â€¯22.35 µg/mL) or VtgC (57.08 ±â€¯6.00 µg/mL); thus VtgAb appeared to be the most dominant Vtg subtype. The present study entails the first report on development of subtype-specific Vtg ELISAs in marbled sole, which empowers us to detect and monitor estrogenic contamination in aquatic environments inhabited by this species.

Immuno-physiological and antioxidant responses of Siberian sturgeon (Acipenser baerii) fed with different levels of olive pomace.

Nowadays, use of the natural plant derivatives as an immunostimulant in aquaculture is becoming more useful than antibiotics which are destructive to target animals, consumers, and the environment. The present study aimed to evaluate the efficacy of dietary substitution of wheat flour at 2, 5, 7.5, and 10% of olive pomace (OP2, OP5, OP7.5, and OP10) in some immune-physiological and antioxidant variables of yearling Siberian sturgeon (Acipenser baerii) weighing 165 ± 6.8 g at 19.1 ± 1.52 °C for 56 days. Fifteen fish per tank were randomly accommodated in triplicates per treatment and the control group was also included. No significant difference was seen in hematological indices including red blood cells (RBS) and immunocompetent cell population sizes among the treated groups (P > 0.05). Also, malondialdehyde (MDA) activity and superoxide dismutase (SOD) in serum remained unaffected in different treatments (P > 0.05). Total protein (TP), high-density lipoprotein (HDL), catalase (CT), and lysozyme were affected by dietary OP levels (P < 0.05), and the highest value was observed at the OP5 diet, but no influence was seen in serum IgM by OP. These data show that application of olive pomace as a partial substitute of wheat flour in the diet of Siberian sturgeon is feasible with stimulatory and antioxidant functions.

Long-term exposure to waterborne nonylphenol alters reproductive physiological parameters in economically important marine fish.

Low concentrations of nonylphenol (NP) in aquatic environment can induce drastic effects on the endocrine system in animals. In this study, we examined the modulatory effects of NP on reproductive and physiological parameters in juveniles of the red seabream and black rockfish following waterborne NP exposure (0, 1, 10, and 50 µg L-1) for 60 days. In red seabream exposed to 50 µg L-1 NP, plasma levels of 17ß-estradiol (E2) and 11-ketotestosterone (11-KT) were significantly lower at 30 and 60 days, while E2 levels were slightly higher in 10 µg L-1-exposed individuals at day 30. Similarly, significantly lower levels of E2 and 11-KT were observed in 10 and 50 µg L-1-exposed black rockfish at 60 days, whereas the E2 level was higher in 1 µg L-1-exposed individuals at day 30. After exposure to NP, plasma and mRNA levels of vitellogenin (VTG) were significantly higher in both species at 30 and 60 days, similar to the inducible effects from synthetic estrogen. Plasma cortisol levels were significantly elevated by relatively higher concentrations of NP (10 and 50 µg L-1) at 30 and 60 days. Finally, 60 days of exposure of 50 µg L-1 NP significantly decreased the gonadosomatic index (GSI) and increased the hepatosomatic index (HSI) in both species. The results obtained from this study provide an evidence of the endocrine disrupting potential of waterborne NP on early stages of economically important marine fish. The NP-triggered endocrine modulation can induce effects on the development of reproductive and metabolic organs in fish species.

Bioavailability and palatability of praziquantel incorporated into solid-lipid nanoparticles fed to yellowtail kingfish Seriola lalandi.

In an effort to overcome the palatability issues currently constraining the effective delivery of praziquantel (PZQ) via feed to treat monogenean parasites in yellowtail kingfish, this study compared the bioavailability and palatability of PZQ in hydrogenated castor oil (HCO) solid lipid nanoparticles (SLN) against pure PZQ in this species. Improving bioavailability would facilitate lower dietary inclusion levels to achieve the same therapeutic dose and therefore reduce the bitterness of feeds containing PZQ. Bioavailability was determined by co-administering feed with either pure PZQ, HCO-SLN or HCO-SLN coated with chitosan via intubation and quantifying the pharmacokinetics response. In contrast to studies with mammals, the results demonstrated that PZQ in HCO-SLN had equal bioavailability to pure PZQ in yellowtail kingfish, including when HCO-SLN were coated with chitosan. We hypothesise that the lack of improvement in bioavailability may be due to the lack of M cells and Peyer's patches in fish and the subsequent inability of fish to take nanoparticles directly into the lymphatic system. Furthermore, palatability of the feeds medicated with PZQ was not improved when the PZQ was incorporated into HCO-SLN, possibly due to the low loading rate of PZQ within the HCO-SLN and the subsequent thick coating of nanoparticles that was required on the surface of the feed pellets. Combined, these data demonstrate that the SLN used in the current study are not capable of delivering the benefits required to enable effective in-feed treatment of PZQ against monogenean parasites in yellowtail kingfish.

Effects of dietary soybean lecithin on growth performance, blood chemistry and immunity in juvenile stellate sturgeon (Acipenser stellatus).

An eleven weeks feeding trial was conducted to determine the effects of different levels of dietary soybean lecithin (SBL) on growth performance, blood chemistry and immunity in juvenile stellate sturgeon (Acipenser stellatus). Fish were fed seven isoproteic (44% crude protein) and isolipidic (17% crude fat) diets containing graded levels of SBL: 0 (control), 1, 2, 4, 6, 8 and 10%. Results showed that dietary SBL supplementation significantly improved the final body weight (BW) and weight gain (WG). Fish fed 6% SBL showed the highest BW and WG values in comparison to fish fed the control diet (P < 0.05), whereas increasing SBL levels above 6% had little practical benefit in terms of somatic growth performance. The inclusion of SBL in diets significantly improved the immune response as data from lysozyme, total Ig levels, alternative complement, phagocytic and bactericidal activities indicated (P < 0.05). The broken-line regression analysis of immunological variable revealed that depending on the parameter considered, the optimal SBL levels in diets for stellate sturgeon juveniles varied. In particular, dietary SBL levels requirements in stellate sturgeon when considering the phagocytic activity rate were determined at 3.3%, whereas 4.1-4.2% were recommended when considering data from lysozyme, alternative complement and bactericidal activities. In contrast, the highest minimum dietary SBL content was estimated at 6.9% when data from total Ig levels were considered. These results indicated that dietary PLs are required for boosting innate immunity in stellate sturgeon, although their minimal level changed depending on the immunological parameter considered. Therefore, we assume that SBL levels comprised between 3.3 and 6.9% may be used as a prophylactic measure to improve the health status in stellate sturgeon. Red blood cell count, hemoglobin and hematocrit levels increased with increasing dietary SBL levels, especially in those sturgeons fed the diet with 6% SBL (P < 0.05). In addition, white blood cell counts significantly increased as dietary SBL levels increased from 4 to 8% in comparison to the control group. Blood biochemistry was also affected by different dietary SBL levels. In particular, significantly higher levels of glucose, cholesterol, HDL and triglycerides were detected in fish fed >6%, >4%, >2% and 2% SBL, respectively (P < 0.05). Based on somatic growth parameters, blood chemistry and systemic immunity parameters, diets containing ca. 6% SBL are recommended for juvenile stellate sturgeon.

Comparison of proteomic profiles in the ovary of Sterlet sturgeon (Acipenser ruthenus) during vitellogenic stages.

One of the challenges of sturgeon aquaculture is that sturgeon takes an extended amount of time to reach sexual maturity. The pattern of the protein expression in relation to the late maturity of sturgeon can help to better understand changes in sexual maturity. 17ß-estradiol (E2), testosterone (T) and vitellogenin (Vtg) levels were examined at all stages of sexual maturation in Sterlet sturgeon (Acipenser ruthenus). Two-dimensional gel electrophoresis and mass spectrometry analysis were used to show the pattern of the ovarian proteins. The T levels increased from the previtellogenic to the postvitellogenic stages (P < 0.05) and Vtg showed a decremental pattern in pre- and postvitellogenic, and atresia (not significantly). The analysis showed 900 protein spots, 19 of which were successfully identified and had significant differences between the previtellogenic and the vitellogenic groups (P < 0.05). Among the identified proteins, 40% involved in cell defense (heat shock protein, Glutathione peroxidase, natural killer enhancing factor, peroxiredoxin-2), 30% in transcription and translation (constitutive photomorphogenesis 9 and Ybx2), 20% in metabolism and energy production (triose-phosphate isomerase (TPI)) and 10% in transport (glycolipid transfer protein). In the vitellogenic stage, the proteins were related to metabolism and energy production (TPI, ES1, creatin kinase, enolase, nucleoside diphosphate kinase, 50%), cell defense (thioredoxin and dislophid isomerase, 20%) and transport (fatty acid binding protein, 10%). Our findings show changes in protein expression pattern from cell defense to metabolism during egg development.

Standardization of plasma vitellogenin level for surveying environmental estrogen pollution using the Japanese common goby Acanthogobius flavimanus.

Field surveys of the impact of environmental estrogen (EE) pollution in aquatic wildlife have been conducted using vitellogenin (VTG) as a biomarker to evaluate the influence of EE. However, a standard baseline of VTG level that can be used to evaluate EE pollution has not been fully determined. To the best of our knowledge, the present study is the first to determine the standard baseline VTG level for evaluating the biological effects of EE pollution using the Japanese common goby (Acanthogobius flavimanus) as the target model fish. Plasma VTG and estradiol-17ß (E2 ) levels associated with the reproductive cycle of wild goby inhabiting an unpolluted environment were measured. Mean plasma VTG and E2 levels exhibited similar changes, increasing in the yolk vesicle stage and peaking in the tertiary yolk stage in females. However, plasma VTG and E2 levels showed no significant changes in males, remaining at low levels throughout the reproductive cycle. The highest VTG levels in females and males were 1.6 mg ml-1 and 124.87 ng ml-1 , respectively. These results indicate that the baseline level (normal level) in males was approximately 130 ng ml-1 at most. We concluded that the threshold between normal and abnormal levels with a 10% risk rate was 150 ng ml-1 in the wild male goby. Plasma VTG levels in males captured from Nagasaki Harbor were higher than the threshold in each reproductive developmental stage, indicating the possibility of EE pollution at this site. The biological standard baseline for VTG established in this study is useful for assessing EE pollution in natural waters.

Haematological assessment of four amazonian ornamental armoured catfish (Teleostei, Loricariidae) / Avaliação hematológica em quatro acaris ornamentais amazônicos (Teleostei, Loricariidae)

The aim of this study was to characterize the haematological profile of four Amazonian ornamentalfreshwater armoured catfish: acari-bola (Peckoltia oligospila - L06), acari-pleco (Cochliodon sp. - L145), acari-canoa(Lasiancistrus saetiger - L323) and acari-assacu (Pseudocanthicus spinosus - L160). The highest blood glucose levels(72.47 ± 28.7 mg dL-1) and erythrocyte counts (0.51 ± 0.2 x 106 cel. μL-1) were recorded for acari-canoa. Theacari-bola and acari-pleco presented similar concentrations of total plasma protein (TPP) (7.96 ± 1.8 and 7.93 ±1.8 g dL-1, respectively) against lower TPP concentrations observed in acari-canoa (4.87 ± 1.5 g dL-1) and acariassacu(6.55 ± 1.5 g dL-1). The acari-assacu had lower total haemoglobin concentration (5.88 ± 1.7 g dL-1) andhaematocrit (12.66 ± 4.6%). No interspecific differences were observed in mean corpuscular volume (MCV) andmean corpuscular haemoglobin concentration (MCHC). The acari-pleco and acari-assacu presented the highestcounts for neutrophils (4142.42 ± 3280.1 cel. μL-1) and thrombocytes (4778.33 ± 1224.8 cel. μL-1), respectively.The haematological profiles were similar to those reported in the literature for freshwater fish and the interspecificdifferences observed were discussed.

O objetivo deste estudo foi caracterizar os perfis hematológicos de quatro espécies de acarisornamentais da região amazônica: acari-bola (Peckoltia oligospila - L06), acari-pleco (Cochliodon sp. - L145), acaricanoa(Lasiancistrus saetiger - L323) e acari-assacu (Pseudocanthicus spinosus - L160). Os maiores níveis de glicose(72,47 ± 28,7 mg dL-1) e contagem de eritrócitos (0,51 ± 0,2 x 106 cel. μL-1) foram encontrados no acari-canoa. Oacari-bola e acari-pleco apresentaram concentrações semelhantes de proteína plasmática total (PPT) (7,96 ± 1,8 e7,93 ± 1,8 g dL-1, respectivamente) em oposição às menores concentrações de PPT no acari-canoa (4,87 ±1,5 g dL-1) e acari-assacu (6,55 ± 1,5 g dL-1). Essa última espécie apresentou menor concentração total dehemoglobina (5,88 ± 1,7 g dL-1) e hematócrito (12,66 ± 4,6%). Não foram observadas diferenças significativasentre as quatro espécies nos parâmetros volume corpuscular médio (VCM) e concentração de hemoglobinacorpuscular média (CHCM). O acari-pleco e o acari-assacu apresentaram os maiores valores de neutrófilos(4142,42 ± 3280,1 cel. μL-1) e de trombócitos (4778,33 ± 1224,8 cel. μL-1), respectivamente. Os perfishematológicos foram semelhantes aos relatados na literatura para peixes de água doce e as diferençasinterespecíficas observadas foram discutidas.

Dimorphic expression of sex-related genes in different gonadal development stages of sterlet, Acipenser ruthenus, a primitive fish species.

Molecular mechanism of sex determination and differentiation of sturgeon, a primitive fish species, is extraordinarily important due to the valuable caviar; however, it is still poorly known. The present work aimed to identify the major genes involved in regulating gonadal development of sterlet, a small species of sturgeon, from 13 candidate genes which have been shown to relate to gonadal differentiation and development in other teleost fish. The sex and gonadal development of sterlets were determined by histological observation and levels of sex steroids testosterone (T), 11-ketotestosterone (11-KT), and 17ß-estradiol (E2) in serum. Sexually dimorphic gene expressions were investigated. The results revealed that gonadal development were asynchronous in 2-year-old male and female sterlets with the testes in early or mid-spermatogenesis and the ovaries in chromatin nucleolus stage or perinucleolus stage, respectively. The levels of T and E2 were not significantly different between sexes or different gonadal development stages while 11-KT had the higher level in mid-spermatogenesis testis stage. In all the investigated gonadal development stages, gene dmrt1 and hsd11b2 were expressed higher in male whereas foxl2 and cyp19a1 were expressed higher in female. Thus, these genes provided the promising markers for sex identification of sterlet. It was unexpected that dkk1 and dax1 had significantly higher expression in ovarian perinucleolus stage than in ovarian chromatin nucleolus stage and in the testis, suggesting that these two genes had more correlation with ovarian development than with the testis, contrary to the previous reports in other vertebrates. Testicular development-related genes (gsdf and amh) and estrogen receptor genes (era and erb) differentially expressed at different testis or ovary development stages, but their expressions were not absolutely significantly different in male and female, depending on the gonadal development stage. Expression of androgen receptor gene ar or rspo, which was supposed to be related to ovarian development, presented no difference between gonadal development stages investigated in this study whenever in male or female.

Genotoxic and mutagenic assessment of iron oxide (maghemite-γ-Fe2O3) nanoparticle in the guppy Poecilia reticulata.

The environmental risk of nanomaterials (NMs) designed and used in nanoremediation process is of emerging concern, but their ecotoxic effects to aquatic organism remains unclear. In this study, the citrate-coated (maghemite) nanoparticles (IONPs) were synthesized and its genotoxic and mutagenic effects were investigated in the female guppy Poecilia reticulata. Fish were exposed to IONPs at environmentally relevant iron concentration (0.3 mg L-1) during 21 days and the animals were collected at the beginning of the experiment and after 3, 7, 14 and 21 days of exposure. The genotoxicity and mutagenicity were evaluated in terms of DNA damage (comet assay), micronucleus (MN) test and erythrocyte nuclear abnormalities (ENA) frequency. Results showed differential genotoxic and mutagenic effects of IONPs in the P. reticulata according to exposure time. The IONP induced DNA damage in P. reticulata after acute (3 and 7 days) and long-term exposure (14 and 21 days), while the mutagenic effects were observed only after long-term exposure. The DNA damage and the total ENA frequency increase linearly over the exposure time, indicating a higher induction rate of clastogenic and aneugenic effects in P. reticulata erythrocytes after long-term exposure to IONPs. Results indicated that the P. reticulata erythrocytes are target of ecotoxicity of IONPs.

Seasonal and ontogenetic changes modulate oxygen consumption and antioxidant defenses in the cutlassfish Trichiurus lepturus (Pisces, Trichiuridae).

Several oxidative stress markers and liver oxygen consumption were measured in different tissues of the marine fish Trichiurus lepturus in late summer and late winter, as well as in juveniles and adult females. Oxygen consumption in liver, superoxide dismutase (SOD) and catalase (CAT) activity in liver, red cells, lens and roe, vitamin E, ubiquinol10, ß-carotene in liver, red cells, and roe, as well as contents of reduced glutathione (GSH) and lipoperoxidation (TBARS) in red cells were evaluated. Regarding ontogeny, compared to adult fish, juveniles showed significant higher SOD activity in liver and lens, as well as higher liver contents of vitamin E. In contrast, adult females showed higher contents of vitamin E in roe, ubiquinol10 in liver and roe, and higher GSH levels in red cells, while the other markers remained unchanged. Regarding seasonal changes, no differences were detected in adult females for liver CAT and ubiquinol10, CAT in roe, vitamin E in roe and in red cells, liver and red cell ubiquinol10, and in GSH in red cells. However, and coinciding with the spawning period of late summer, liver oxygen consumption, SOD and CAT activity and ubiquinol10 contents in roe and SOD activity in red cells, and red cell TBARS contents were higher compared to late winter. These temporal antioxidant adjustments of Trichiurus lepturus seem to be parallel to the higher oxygen consumption typical of juvenile forms and also to the intense spawning and foraging activities of adult females in late summer.

Short-term effects of 17ß-estradiol on physiological responses in stellate sturgeon Acipenser stellatus.

There is no evidence of the effect of 17ß-estradiol (E2) administration on stress response and related metabolites after a short-period in chondrostean. In this study, we examined whether E2 is capable of inducing the hypothalamus-pituitary-interrenal axis activity. To accomplish this, sexually immature Acipenser stellatus were injected intraperitoneally with saline or E2 (5mg/kg body mass) and sampled 0, 1, 3, 6, 12, 24, 48, and 72h later. Plasma E2, cortisol, glucose, lactate, hematocrit (Hct), total protein (TP), cholesterol, triglyceride, alkaline phosphatase activity (ALP), calcium (Ca), and phosphorus concentrations were examined. Plasma levels of E2 significantly increased after 1h in E2-injected fish and remained high after 12h. E2 injection enhanced plasma TP and Ca concentrations, but had no effect on other parameters. Injection of either saline or E2 enhanced plasma glucose, lactate, TP, and Ca concentrations, but had no effect on Hct, cholesterol, triglyceride, and ALP. The results demonstrated that acutely elevated level of E2 did not lead to intense changes of stress metabolites and altered biochemical changes compared to the fish in control or saline treatment groups. Therefore, we found no exhibition of E2 on the rate of cortisol synthesis or stress indicators in this species, while E2 could modulate some other related metabolites on vitellogenesis pathway.

Gender and gonadal maturity stage identification of captive Chinese sturgeon, Acipenser sinensis, using ultrasound imagery and sex steroids.

Long lifespan and late maturation make it difficult to establish gamete maturity and breeding age of captive endangered Chinese sturgeon, Acipenser sinensis. This greatly handicaps timely breeding and future conservation stocking efforts. We used ultrasound imagery and sex steroids to determine the gender and gonadal maturity stage of captive Chinese sturgeon (age, 10-17years old). The echogenicity of the reproductive organs and the respective morphology of the gonads were described and two quantitative parameters po (proportion of the ovary to the entire reproductive organs) and d (thickness of the reproductive organs) were measured to characterize sex and maturity stage of Chinese sturgeon. Females were accordingly placed fish into several categories: FII (FII-, FII, FII+), FIII (FIII, FIII+) and FIV (FIV, FIV+) and FVI and males as MII, MIII, MIV, MV and MVI. The accuracy of gender and maturity stage determination provided by ultrasonographic methods was 72.7% for FII- ovary (n=11) and 76.2% for MII testis (n=42). Accuracy of sex and maturity determination using only serum sex steroid of testosterone (T) and estradiol-17ß (E2) was low (58-73%, depending on maturity stage). However, when the two methods were used together, accuracy increased sharply, especially for immature (II stage) females. In summary, of 151 Chinese sturgeon, whose sex and maturity stage were independently confirmed, 88.1% (n=133), 62.9% (n=95), and 96.7% (n=146) were successfully sexed and staged using ultrasound, sex steroids, or both methods, respectively. The results provide reliable non-invasive techniques for determining sex and gonadal maturation of captive Chinese sturgeon. These methods can track individual gonad characteristics over multi-year reproductive cycles, which will assist captive broodstock management, artificial reproduction, and future conservation stocking.