Hidden in the sand: Phylogenomics unravel an unexpected evolutionary history for the desert-adapted vipers of the genus Cerastes.

The desert vipers of the genus Cerastes are a small clade of medically important venomous snakes within the family Viperidae. According to published morphological and molecular studies, the group is comprised by four species: two morphologically similar and phylogenetically sister taxa, the African horned viper (Cerastes cerastes) and the Arabian horned viper (Cerastes gasperettii); a more distantly related species, the Saharan sand viper (Cerastes vipera), and the enigmatic Böhme's sand viper (Cerastes boehmei), only known from a single specimen in captivity allegedly captured in Central Tunisia. In this study, we sequenced one mitochondrial marker (COI) as well as genome-wide data (ddRAD sequencing) from 28 and 41 samples, respectively, covering the entire distribution range of the genus to explore the population genomics, phylogenomic relationships and introgression patterns within the genus Cerastes. Additionally, and to provide insights into the mode of diversification of the group, we carried out niche overlap analyses considering climatic and habitat variables. Both nuclear phylogenomic reconstructions and population structure analyses have unveiled an unexpected evolutionary history for the genus Cerastes, which sharply contradicts the morphological similarities and previously published mitochondrial approaches. Cerastes cerastes and C. vipera are recovered as sister taxa whilst C. gasperettii is a sister taxon to the clade formed by these two species. We found a relatively high niche overlap (OI > 0.7) in both climatic and habitat variables between C. cerastes and C. vipera, contradicting a potential scenario of sympatric speciation. These results are in line with the introgression found between the northwestern African populations of C. cerastes and C. vipera. Finally, our genomic data confirms the existence of a lineage of C. cerastes in Arabia. All these results highlight the importance of genome-wide data over few genetic markers to study the evolutionary history of species.

Genome Characterization and Spaciotemporal Dispersal Analysis of Bagaza Virus Detected in Portugal, 2021.

In September 2021, Bagaza virus (BAGV), a member of the Ntaya group from the Flavivirus genus, was detected for the first time in Portugal, in the heart and the brain of a red-legged partridge found dead in a hunting ground in Serpa (Alentejo region; southern Portugal). Here we report the genomic characterization of the full-length sequence of the BAGV detected (BAGV/PT/2021), including phylogenetic reconstructions and spaciotemporal analyses. Phylogenies inferred from nucleotide sequence alignments, complemented with the analysis of amino acid alignments, indicated that the BAGV strain from Portugal is closely related to BAGV strains previously detected in Spain, suggesting a common ancestor that seems to have arrived in the Iberia Peninsula in the late 1990s to early 2000s. In addition, our findings support previous observations that BAGV and Israel turkey meningoencephalitis virus (ITV) belong to the same viral species.

Phytase Supplementation under Commercially Intensive Rearing Conditions: Impacts on Nile Tilapia Growth Performance and Nutrient Digestibility.

This study evaluated the effects of phytase supplementation on growth performance and apparent digestibility of Nile tilapia (Oreochromis niloticus) in a commercial fish farm setting. Nile tilapia (6300 male, 57.48 ± 1.04 g) were randomly stocked into 42 floating cages. The experimental design was completely randomized, comprising six treatments and seven replications. Fish were fed five phosphorus deficient plant-based diets with graded levels of phytase supplementation (0, 500, 1000, 1500, 2000 UF kg-1) and an additional diet containing phosphorus supplementation to meet the requirement of this fish species (positive control). After 97 days of feeding, growth performance data were collected and 900 fish (500 ± 10 g) were relocated to 6 floating cages for the digestibility assessment. Quadratic polynomial regression analysis indicated 1537.5 and 1593.2 UF kg-1 as the optimum dietary levels for daily weight gain and feed conversion rate, respectively. Including 2000 UF kg-1 resulted in the higher dry matter, crude protein, energy, and ash apparent digestibility coefficient values. Therefore, phytase supplementation from 1500 to 2000 UF kg-1 is recommended to enhance growth performance and nutrient bioavailability of Nile tilapia reared according to industry practices.

Ascorbic Acid Supplementation Improves Skeletal Muscle Growth in Pacu (Piaractus mesopotamicus) Juveniles: In Vivo and In Vitro Studies.

In fish, fasting leads to loss of muscle mass. This condition triggers oxidative stress, and therefore, antioxidants can be an alternative to muscle recovery. We investigated the effects of antioxidant ascorbic acid (AA) on the morphology, antioxidant enzyme activity, and gene expression in the skeletal muscle of pacu (Piaractus mesopotamicus) following fasting, using in vitro and in vivo strategies. Isolated muscle cells of the pacu were subjected to 72 h of nutrient restriction, followed by 24 h of incubation with nutrients or nutrients and AA (200 µM). Fish were fasted for 15 days, followed by 6 h and 15 and 30 days of refeeding with 100, 200, and 400 mg/kg of AA supplementation. AA addition increased cell diameter and the expression of anabolic and cell proliferation genes in vitro. In vivo, 400 mg/kg of AA increased anabolic and proliferative genes expression at 6 h of refeeding, the fiber diameter and the expression of genes related to cell proliferation at 15 days, and the expression of catabolic and oxidative metabolism genes at 30 days. Catalase activity remained low in the higher supplementation group. In conclusion, AA directly affected the isolated muscle cells, and the higher AA supplementation positively influenced muscle growth after fasting.

The combination of resveratrol and exercise enhances muscle growth characteristics in pacu (Piaractus mesopotamicus).

Pacu is a tropical fish with important value to aquaculture. During cellular metabolism, reactive oxygen species (ROS) are produced, which can influence muscle growth. Resveratrol is an effective antioxidant that scavenges ROS and can modulate physical performance preventing oxidative stress. We investigated the effects of resveratrol and exercise on pacu muscle growth characteristics. Four groups were used: fish fed with control diet /without exercise (C); fish fed with control diet/subjected to exercise (CE); fish fed resveratrol-supplemented diet/without exercise (R); and fish fed resveratrol-supplemented diet/subjected to exercise (RE). At 30 days, the RE group presented a significant increase in body weight, fewer muscle fibers in the 20-40 µm and more fibers in the >60 µm diameter class compared to the C group. At day 7, catalase activity decreased in CE and RE groups. Superoxide dismutase activity decreased only in the CE group. Myod and mtor gene expression was higher in R and RE and igf-1 was up-regulated in the RE group. Murf1a level decreased in CE, R, and RE, while sdha expression was higher in the RE group. We suggest that resveratrol in combination with exercise was beneficial for muscle growth and metabolism, increasing the expression levels of genes related to muscle anabolism and oxidative metabolism, besides the decrease of catabolic gene expression. Notably, all of these changes occurred together with muscle hypertrophy and increased body weight. Our results show a positive application for resveratrol in association with exercise as a strategy to improve the growth performance of juvenile pacus.

l-glutamine in vitro supplementation enhances Nile tilapia Oreochromis niloticus (Linnaeus, 1758) leukocyte function.

Under appropriate conditions, glutamine (Gln) is an essential nutrient for immunological responses, acting as a metabolic substrate for proliferation of enterocytes and lymphocytes, and having positive effects on the function of stimulated immune cells. Thus, specific components of both innate and adaptive immune systems of Nile tilapia were evaluated after supplementing Gln to cell culture media. Primary cell cultures of kidney leukocytes were used for respiratory burst and phagocytic activity assessment. The ability of macrophages to kill Streptococcus iniae also was evaluated. Additionally, a proliferation assay was conducted with peripheral blood lymphocytes (PBL) exposed to non-specific mitogens. Results showed that macrophage phagocytosis, anion superoxide production, and bactericidal capacity were significantly (P < 0.05) enhanced by Gln supplementation to the culture media. The proliferation of lymphocytes upon mitogenic exposure also was significantly (P < 0.05) enhanced by Gln supplementation to the media. Our results suggest that in vitro, different levels of Gln were necessary for optimal immunological responses of leukocytes and lymphocytes. As such, Gln supplementation was able to enhance and modulate both innate and adaptive responses of Nile tilapia leukocytes, highlighting its potential application as an immunonutrient.

Highly confined stacks of graphene oxide sheets in water.

Since the discovery of graphene oxide (GO), the most accessible of the precursors of graphene, this material has been widely studied for applications in science and technology. In this work, we describe a procedure to obtain GO dispersions in water at high concentrations, these highly dehydrated dispersions being in addition fully redispersible by dilution. With the availability of such concentrated samples, it was possible to investigate the structure of hydrated GO sheets in a previously unexplored range of concentrations, and to evidence a structural phase transition. Tentatively applying models designed for describing the small-angle scattering curve in the Smectic A (or [Formula: see text]) phase of lyotropic systems, it was possible to extract elastic parameters characterising the system on the dilute side of the transition, thereby evidencing the relevance of both electrostatic and steric (Helfrich) interactions in stabilising aqueous lamellar stacks of GO sheets.

Activity of Brazilian propolis against Aeromonas hydrophila and its effect on Nile tilapia growth, hematological and non-specific immune response under bacterial infection

ABSTRACT The effect of the ethanolic extract of propolis (EEP) on Aeromonas hydrophila was analyzed by determination of minimum inhibitory concentration (MIC). Then, the effects of crude propolis powder (CPP) on growth, hemato-immune parameters of the Nile tilapia, as well as its effects on resistance to A. hydrophila challenge were investigated. The CPP (0.5, 1.0, 1.5, 2.0, 2.5 and 3.0%) was added to the diet of 280 Nile tilapia (50.0 ± 5.7 g fish-1). Hemato-immune parameters were analyzed before and after the bacterial challenge. Red blood cell, hematocrit, hemoglobin, mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC), and hydrogen peroxide (H2O2) and nitric oxide (NO) were evaluated. The MIC of the EEP was 13% (v/v) with a bactericidal effect after 24 hours. Growth performance was significantly lower for those fish fed diets containing 2.5 and 3% of CPP compared to the control diet. Differences in CPP levels affected fish hemoglobin, neutrophils number and NO following the bacterial challenge. For others parameters no significant differences were observed. Our results show that although propolis has bactericidal properties in vitro, the addition of crude propolis powder to Nile tilapia extruded diets does not necessarily lead to an improvement of fish health.

Activity of Brazilian propolis against Aeromonas hydrophila and its effect on Nile tilapia growth, hematological and non-specific immune response under bacterial infection.

The effect of the ethanolic extract of propolis (EEP) on Aeromonas hydrophila was analyzed by determination of minimum inhibitory concentration (MIC). Then, the effects of crude propolis powder (CPP) on growth, hemato-immune parameters of the Nile tilapia, as well as its effects on resistance to A. hydrophila challenge were investigated. The CPP (0.5, 1.0, 1.5, 2.0, 2.5 and 3.0%) was added to the diet of 280 Nile tilapia (50.0 ± 5.7 g fish-1). Hemato-immune parameters were analyzed before and after the bacterial challenge. Red blood cell, hematocrit, hemoglobin, mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC), and hydrogen peroxide (H2O2) and nitric oxide (NO) were evaluated. The MIC of the EEP was 13% (v/v) with a bactericidal effect after 24 hours. Growth performance was significantly lower for those fish fed diets containing 2.5 and 3% of CPP compared to the control diet. Differences in CPP levels affected fish hemoglobin, neutrophils number and NO following the bacterial challenge. For others parameters no significant differences were observed. Our results show that although propolis has bactericidal properties in vitro, the addition of crude propolis powder to Nile tilapia extruded diets does not necessarily lead to an improvement of fish health.

Novel inexpensive fungi proteases: Production by solid state fermentation and characterization.

A comparative study was carried out for proteases production using agroindustrial residues as substrate for solid state fermentation (SSF) of several fungal strains. High protease production was observed for most of the microorganisms studied, as well as very different biochemical characteristics, including activities at specific temperatures and a wide range of pH values. The enzymes produced were very different regarding optimum pH and they showed stability at 50 °C. Aspergillus oryzae showed stability at all pH values studied. Penicillium roquefortii and Aspergillus flavipes presented optimum activity at temperatures of 50 °C and 90 °C, respectively. Lyophilized protease from A. oryzae reached 1251.60 U/g and yield of 155010.66 U/kg of substrate. Therefore, the substrate as well as the microorganism strain can modify the biochemical character of the enzyme produced. The high protease activity and stability established plus the low cost of substrates, make these fungal proteases potential alternatives for the biotechnological industry.

Non-specific immune parameters and physiological response of Nile tilapia fed ß-glucan and vitamin C for different periods and submitted to stress and bacterial challenge.

This study attempts to describe the effects of different administration periods of dietary ß-glucan and Vit C on the non-specific immune response, physiological parameters and disease resistance of Nile tilapia against Aeromonas hydrophila infection. Therefore, a feeding trial (288 fish) was conducted to determine the best administration period (7, 15, 30 and 45 days) for a Nile tilapia diet supplemented with 0.1% ß-glucan and 600 mg Vit C/kg diet. After the administration period, three different groups of 96 fish were exposed to one of the following three stresses: cold-induced stress, transport-induced stress, and A. hydrophila challenge. Hematological, biochemical and immunological responses were analyzed before and/or after stress. Cold-induced stress increased cortisol levels and reduced the leukocyte count in fish fed the test diet for seven days compared with the other periods. After transport-induced stress, fish fed the test diet for seven days required more hours to return to the baseline levels of cortisol and neutrophils. Moreover, independently of the administration period, fish needed 24 h for leukocyte and glucose levels to return to the initial values. The lowest survival after bacterial infection was observed in fish test diet for seven days. Based on fish hematological and biochemical responses, diet supplemented with 0.1% of ß-glucan and 600 mg of Vit C/kg fed for at least 15 days is recommended for Nile tilapia especially when fish are likely to encounter transport-induced stress, and this stress was more severe than cold-induced stress or bacterial challenge.

Levels of copper in Nile tilapia from Brazil.

The purpose of this work was to determine the concentration of copper in samples of farmed Nile tilapia (Oreochromis niloticus L.) fillets purchased in the city of Botucatu (São Paulo, Brazil) and in fillet and liver samples of Tilapia fed diets supplemented with different concentrations of Cu from the Laboratory of Aquatic Organism Nutrition/FMVZ-UNESP (Botucatu, Brazil). The fillet samples were prepared by lyophilisation and cryogenic grinding into particles smaller than 60 µm, and copper was extracted ultrasonically using 0.10 mol l(-1) HCl as extraction solution. Copper determination was performed by graphite furnace atomic absorption spectrometry (GFAAS) with optimised temperatures of drying, pyrolysis, atomisation and cleaning. Palladium nitrate was injected into the samples as a chemical modifier and tungsten as a permanent modifier. Copper concentrations of 0.70-1.60 mg kg(-1) were found, which are in line with Brazilian regulations. The accuracy and precision of the copper concentrations determined in this study were evaluated using certified standard Lake Michigan fish tissue (NIST SRM 1947).

Determination of selenium by GFAAS in slurries of fish feces to estimate the bioavailability of this micronutrient in feed used in pisciculture.

This paper presents a simple, fast and sensitive method to determine selenium in samples of feces and of fish feed by graphite furnace atomic absorption spectrometry (GFAAS) through the direct introduction of slurries of the samples into the spectrometer's graphite tube. The limits of detection (LOD) and quantification (LOQ) calculated for 20 readings of the blank of the standard slurries (0.50% m/v of feces or feed devoid of selenium) were 0.31 microg l(-1) and 1.03 microg l(-1), respectively, for the standard feces slurries and 0.35 microg l(-1) and 1.16 microg l(-1), respectively, for the standard feed slurries. The proposed method was applied in studies of bioavailability of selenium in different fish feeds and the results proved consistent with that obtained from samples mineralized by acid digestion using the microwave oven.

Exigência em fósforo disponível para alevinos de tilápia do Nilo / Dietary phosphorus requirement for fingerlings Nile Tilápia

O fósforo se destaca como um dos minerais mais importantes na nutrição animal. Apresenta-se pouco disponível em alimentos de origem vegetal por estar quimicamente ligado a outros minerais e nutrientes, em que se denomina fósforo fítico ou fitato. Desta forma, objetivou-se avaliar diferentes níveis de fósforo disponível em rações para alevinos de tilápia do Nilo (Oreochromis niloticus), por meio do ganho em peso, do consumo de ração aparente, da conversão alimentar aparente, da eficiência alimentar, da sobrevivência e da composição química da carcaça. O delineamento experimental foi inteiramente casualizado, com seis tratamentos (0; 0,25; 0,50; 0,75; 1,00 e 1,25 por cento de fósforo disponível) e cinco repetições. As rações, utilizadas para avaliar os diferentes níveis de fósforo disponível, foram formuladas exclusivamente com alimentos de origem vegetal, sendo isoprotéicas (30,0 por cento PD), isoenergéticas (3.300kcal de ED kg-1) e contendo a mesma relação cálcio:fósforo. Conclui-se que, para alevinos de tilápia do Nilo, o nível mínimo de 0,75 por cento de fósforo disponível nas rações proporciona melhor desempenho produtivo e composição química da carcaça.

Phosphorus is considered one of the most important mineral in animal nutrition. In vegetables feedstuffs this mineral can be less available because his chemistry connection with other minerals and nutrients, named the phosphorus phytic or phytate. Thus this research was aimed evaluating different levels of available phosphorus in diets for Nile tilapia (Oreochromis niloticus) fingerlings based on weight gain, feed consumption, feed conversion ratio, feed efficiency, survival and body composition. The experimental design was completely randomized with six treatment (0, 0.25, 0.50, 0.75, 1.00 e 1.25 percent of available phosphorus) and five replicates. The rations used to evaluate the different levels of available phosphorus were exclusively formulated, using vegetables feedstuffs, to contain 30.0 percent PD, 3300kcal of DE kg-1 and the same rate between calcium:phosphorus. Based on these results, 0.75 percent of available phosphorus in Nile tilapia fingerlings diets determines a better growth performance and body chemical composition.

Determination of chromium by GFAAS in slurries of fish feces to estimate the apparent digestibility of nutrients in feed used in pisciculture.

This paper presents a simple, fast and sensitive method to determine chromic oxide (used as a biological marker of fish feed) in samples of fish feces by GFAAS through the direct introduction of slurries of the samples into the spectrometer's graphite tube. The standard samples of feces and of fish feed containing 0.10-1.00mgkg(-1) of Cr(2)O(3) were pre-frozen for 1min in liquid nitrogen and then ground a cryogenic mill for 2min, which reduced the samples' grain size to less than 60mum. The standard slurries were prepared by mixing 20mg of standard samples of fish feed or feces with 1mL of a solution containing 0.05% (v/v) of Triton X-100 and 0.50% (v/v) of suprapure HNO(3) directly in the spectrometer's automatic sampling glass. The final concentrations of Cr(2)O(3) present in the standard slurries were 2, 4, 8, 16 and 20mugL(-1). After sonicating the mixture for 20s, 10muL of standard slurries were injected into the graphite tube, whose internal wall was lined with a metallic palladium film that acted as a permanent chemical modifier. The limits of detection (LOD) and quantification (LOQ) calculated for 20 readings of the blank of the standard slurries (2%, m/v of feces or feed devoid of minerals) were 0.81 and 2.70mugL(-1) of Cr(2)O(3) for the standard feces slurries, 0.84 and 2.83mugL(-1) of Cr(2)O(3) for the standard feed slurries. The proposed method was applied in studies of nutrient digestibility of different fish feeds and its results proved compatible with the results obtained from samples pre-mineralized by acid digestion.

Whole yeast and yeast derivatives in Nile tilapia diets: hematological and histological aspects- DOI: 10.4025/actascibiolsci.161 / Levedura íntegra e derivados do seu processamento em rações para tilápia do Nilo: aspectos hematológicos e histológicos - DOI: 10.4025/actascibiolsci.v28i4.161

The effects of inclusion of whole yeast, autolyzed yeast and yeast cell wall on hematological parameters and gut villus perimeter were evaluated in juvenile Nile tilapia, after 80 experimental days. Isoproteic (32.0% DP) and isoenergetic (3200 kcal DE kg-1) practical diets were supplemented with three levels of whole yeast or autolyzed yeast (1.0, 2.0 and 3.0%) and three levels of yeast cell wall (0.1, 0.2 and 0.3%), plus a control diet (with no test microingredients). Red blood cell count, hemoglobin concentration, total plasmatic protein, hematocrit percentage, mean corpuscular volume, mean corpuscular hemoglobin concentration and gut villus perimeter were evaluated. Variations on hematological parameters in animals fed diets with whole yeast; autolyzed yeast and yeast cell wall were observed to be within normal ranges for this species. There was significant influence (p

Os efeitos da inclusão de levedura íntegra, levedura autolisada e parede celular para juvenis da tilápia do Nilo sobre parâmetros hematológicos e perímetro das vilosidades intestinais foram avaliados após 80 dias de experimento. Foram utilizadas rações práticas isoprotéicas (32,0% PD) e isoenergéticas (3.200 kcal ED kg-1 de ração) suplementadas com três níveis de levedura íntegra ou autolisada (1,0; 2,0 e 3,0%) e três níveis de parede celular (0,1; 0,2 e 0,3%), além de uma ração controle, isenta destes microingredientes. Foram avaliados a contagem de eritrócitos, taxa de hemoglobina, proteína plasmática total, porcentagem de hematócrito, volume globular médio, concentração de hemoglobina globular média e o perímetro das vilosidades intestinais. Constatou-se que as variações nos parâmetros hematológicos dos animais alimentados com levedura íntegra, levedura autolisada e parede celular estão dentro da faixa de normalidade para a espécie. Houve influência significativa (p 0,05) dos diferentes níveis de levedura e derivados sobre o perímetro das vilosidades intestinais. Pode-se concluir que o período experimental e os níveis adotados neste estudo para levedura íntegra, levedura autolisada e parede celular não provocam alterações prejudiciais nos padrões hematológicos de tilápia do Nilo e podem ser utilizados com segurança para comporem rações para esta espécie e que a suplementa