Changes in serum immunoglobulin M (IgM) concentrations during early development of chum salmon (Oncorhynchus keta) as determined by sensitive elisa technique

• 1.1. A specific and sensitive sandwich enzyme-linked immunosorbent assay (ELISA) was developed for the measurement of low levels of serum immunoglobulin M (IgM) of chum salmon Oncorhynchus keta.
• 2.2. In this assay, 5 μl serum was enough to measure the concentration of IgM and the minimum detectable concentration of serum IgM was about 5 ng/ml.
• 3.3. Coefficients of variation within and between assays ranged from 2.90 to 9.61%.
• 4.4. IgM concentrations remained at low level (< 300 ng/ml) until 40 days after hatching and then increased rapidly at the period of emergence (48 days after hatching).

     

Study of the heme catabolism of fish

• 1.1. The composition of bile pigments in the blood and bile of 39 species were studied.
• 2.2. Conjugated bilirubin (trace to 4.62 mg/100 ml) was detected in the serum of most fish, while biliverdin (trace to 2.0 mg/100ml) was detected only in Anguilla Japonica, Thalassoma lunare and Clinocottus analis.
• 3.3. Analysis showed tht there are two types of bile pigments excretion pattern in these fishes. The first pattern excretes bilirubin (most conjugate) predominantly, the other excretes mostly biliverdin with some bilirubin. However, during starvation, the excretion of conjugate bilirubin gradually shifted to unconjugated biliverdin. The rate of shifting varies with species.
• 4.4. Introduction of bilirubin into Anguilla japonica produced an initial excretion of mono-conjugates, followed by di-conjugates. Introduction of biliverdin caused an increased in the excretion of unconjugated biliverdin, but no significant increase of bilirubin in the bile was detected.
• 5.5. A binary excretion pathway of bile pigments in fish is proposed. The evolutionary characteristics of heme catabolism in terrestrial animals with respect to this pathway is discussed.

     

Miniature gel filtration: a simple and inexpensive method for the measurement of protein- and non-protein-bound cortisol: application to guinea pig plasma

• 1.1. A method is described for the accurate and rapid measurement of protein- and non-protein-bound cortisol by miniature gel filtration in small volumes of plasma, e.g. of rodents.
• 2.2. Binding of cortisol by guinea pig plasma proteins is strongly reduced at elevated temperature (4°C: 102 ± 12ng/ml; 40°C: 5 ± 2 ng/ml).
• 3.3. Incubation of guinea pig plasma with 1–5000 ng cortisol resulted in a dose-dependent increase in cortisol bound to proteins (specific binding by corticosteroid binding globulin: 230 ± 12 ng/ml).
• 4.4. Administration of 20 IU (1–24)ACTH induced a significant increase of total protein-bound and non-protein-bound cortisol.
• 5.5. Values reported in this study agree well with those of previous investigations, in which bound and non-bound glucocorticosteroids were separated by gel filtration on large Sephadex® columns.

     

Atrial natriuretic peptides in the heart and hemolymph of the oyster,Crassostrea virginica: A comparison with vertebrates

• 1.1. The content of atrial natriuretic peptides (ANPs) in the auricles of oysters, Crassostrea virginica, was significantly (P < 0.01) greater than in their ventricles.
• 2.2. High-performance gel permeation chromatography (HP-GPC) followed by ANF radioimmunoassay revealed two peaks in both oyster and vertebrate (rat) hearts—a major peak where the 12.6–14 kDa ANF prohormone elutes and a smaller peak where the pure human form of ANF elutes.
• 3.3. HP-GPC evaluation followed by proANF 31–67 radioimmunoassay revealed only an ANF-like prohormone while HP-GPC followed by proANF 1–30 radioimmunoassay revealed the ANF prohormone and a proANF 1–30-like peptide in oyster and rat hearts.
• 4.4. ANPs concentrations in hemolymph were 940 ± 129, 225 ± 25, and 100 ± 10 pg/ml by the proANF 1–30, proANF 31–67, and ANF radioimmunoassays, respectively.
• 5.5. Atrial natriuretic-like peptides are present in the oyster heart in molecular species similar to vertebrate species and these peptides are also present in hemolymph.

     

Changes in free amino acid composition in haemolymph of larvae of the wax moth,Galleria mellon ella L., during cold acclimation

• 1.1. Free amino acids were analysed in the haemolymph of Galleria mellonella larvae by HPLC chromatography with o-phthaldialdehyde (OPA)-l-thio-β-d-glucose as derivatization agent.
• 2.2. Fourteen primary amino acids were detected among which glutamine, alanine, γ-aminobutyric acid (GABA) and glycine predominated and constituted 67.7% of the amino acids found.
• 3.3. The concentration of GABA increased significantly with the age of larvae entering the wandering phase and reached a maximum during metamorphosis.
• 4.4. Analysis of cold-acclimated larvae revealed a net increase of free primary amino acids from 96 to 151.8 μmol/ml during consecutive acclimation to 0°C within 20 days and to 205.4μmol/ml during cold shock injury at 0°C (3 hr).
• 5.5. The bulk of this increase was accounted for by alanine, glycine, phenylalanine and lysine.

     

Acute stress suppresses plasma estradiol levels in female alligators (Alligator mississippiensis)

• 1.1. Five adult, female alligators (Alligator mississippiensis) were captured at night during the breeding season, and a blood sample taken within 5 min of capture.
• 2.2. The alligators were physically restrained (tied to boards) and additional blood samples taken at 4, 8, 12, 16, 22, 28, 38, and 48 hr after capture. After the last blood sample was collected the animals were released.
• 3.3. Plasma estradiol-17β and corticosterone were measured by radioimmunoassay. Estradiol declined significantly from initial values by 22 hr post capture, but remained unchanged for 48 hr.
• 4.4. Plasma corticosterone rose from a mean of 0.8 ng/ml at capture to 12.6 ng/ml after 4 hr. Corticosterone continued to rise up to 16 hr then declined after 22 hr. From 28 until 48 hr corticosterone again increased significantly.
• 5.5. These results demonstrate that acute stress in female alligators causes significant suppression of plasma estradiol and a biphasic pattern of corticosterone secretion.

     

Hemolymph ferritin and radula structure in the limpets Patelloida alticostata and Patella peronii (Mollusca: Gastropoda)

• 1.1. The mean concentration of total hemolymph iron was 3060μg/100 ml in Patella peronii and 2950μg/100 ml in Patelloida alticostata.
• 2.2. Ferritin was found to act as a major iron-binding protein in the hemolymph of both P. peronii and P. alticostata.
• 3.3. P. alticostata ferritin has a molecular weight of approximately 505,000, while that of P. peronii has a mol wt. of approximately 520,000.
• 4.4. The lateral radula teeth of both species are mineralized by deposits of silica (SiO₂) and iron in the form of goethite (α-FeOOH).
• 5.5. Hemolymph ferritin is suggested to act as a high capacity transport system to supply iron to the mineralizing front of the radula.

     

Biochemical aspects of Balanus hameri haemolymph,together with some comparative haemolymph data for Balanus balanus and Lepas anatifera

• 1.1. The concentration of protein in the haemolymph of Balanus hameri ranged from 2.0 to 17.3 mg/ml, and the lipid from 1.4 to 7.7 mg/ml; the haemolymph protein and lipid levels increased significantly prior to cross-fertilization.
• 2.2. The protein and lipid concentrations in Balanus balanus haemolymph were 8.1 and 1.7 mg/ml respectively.
• 3.3. The lipid concentration of Lepas anatifera haemolymph was 1.2 mg/ml.
• 4.4. The neutral lipid and phospholipid components of B. hameri and L. anatifera haemolymph were the same, with the major components of the phospholipid fraction being phosphatidyl ethanolamine and phosphatidyl choline.
• 5.5. The osmolarity (970.4 mOsm), chloride ion concentration (501.3 m-eq/l) and pH (7.29) of B. hameri haemolymph were also determined.

     

A novel inhibitor of glycogen phosphorylase from crayfish hepatopancreas

• 1.1. A novel glycogen phosphorylase inhibitor was partially purified from crayfish hepatopancreas.
• 2.2. The inhibitor was found only in two species of crayfish examined, and not in lobster, fresh and salt water clams, mussels or cockroaches.
• 3.3. The inhibitor is a small protein (M_r = 23,000) which did not show proteolytic activity.
• 4.4. Preliminary kinetic analysis of the inhibitory mechanism indicated that it bound to both glycogen and the glycogen phosphorylase protein.
• 5.5. Inhibitor binding to glycogen resulted in a competitive inhibition pattern with respect to glycogen phosphorylase (inhibition constant of ca 10 μg/ml).
• 6.6. The inhibitor also bound glycogen phosphorylase directly with a binding coefficient of 100 μg/ml resulting in a partially non-competitive inhibition pattern with respect to phosphate.

     

Determination of β-phenylethylamine in catfish (Parasilurus asotus) tissues by gas chromatography/mass spectrometry

• 1.1. β-Phenylethylamine (PEA) was detected and quantitated in tissues of the catfish, Parasilurus asotus, by very specific and sensitive gas chromatography/mass spectrometry.
• 2.2. The selected ion monitoring was made with a strong quasi-molecular ion of the pentafluoropropionic derivative of PEA in the positive chemical ionization mode.
• 3.3. PEA was found in all tissues tested ranging from 2.8 to 38.2 ng/g wet wt tissue. It was highest in the spinal cord, followed by the skin, brain and intestine.

     

Characteristics of Crassostrea virginica crystalline style chitin digestion

• 1.1. Fundamental chitin digestion characteristics of Crassostrea virginica crystalline style were investigated.
• 2.2. Optimum temperature and pH were 34°C and 4.8. respectively.
• 3.3. The colloidal regenerated chitin (0.56mol/0.5 ml: GlcNAc equivalents) was saturating under all enzyme levels encountered.
• 4.4. There was no evidence of end product inhibition, even after 100 hr incubation.
• 5.5. Calculated K_m for the chitinase complex was 1.19mM when determined using a 30 min assay, but was only 0.70 mM when determined using a 4.6 hr assay.
• 6.6. Both K_m values are lower than reported for similar assays in other molluscs and for most bacteria.
• 7.7. Effect of substrate preparation on the kinetics are discussed.
• 8.8. Eight peaks of chitinase activity were resolved by DEAE-Fractogel ion exchange chromatography.

     

Bovine serum sialic acid: Age-related changes in type and content

• 1.1. The sialic acid content of newborn calf serum (4.8 μmol/ml) is approx. 3-fold higher than that of mature animals (1.4 μmol/ml) and decreases to 2.4 μmol/ml at 20 days of age. Colostrum-fed and colostrum-deprived calves have similar levels of sialic acid from birth to 14 days of age.
• 2.2. The high level of sialic acid in newborn calf serum is due predominantly to N-acetylneuraminic acid, since this sialic acid accounts for 93% of the total and since <5% of the sialic acid is O-acetylated.
• 3.3. Comparison of day 0 and day 20 serum by gel filtration and by SDS polyacrylamide gel electrophoresis demonstrates that the increase in sialic acid is associated with increased production and/or sialylation of components with MW of 45–60 kDa.
• 4.4. A high percentage (64%) of the sialic acid in newborn calf serum is detected with the lipid-linked sialic acid assay, relative to 20 day old (25%) or mature (18%) animals.
• 5.5. This indicates that the glycoproteins of newborn calf serum are more efficiently extracted under the conditions of this assay than glycoproteins of mature serum.

     

Comparative study of proteolytic enzymes in the digestive tracts of the european sea bass and hybrid striped bass reared in freshwater

• 1.1. A comparative study of the proteolytic activity in four different sections of the digestive tracts of the European sea bass (Dicentrarchus labrax) and hybrid striped bass (Morone chrysops × M. saxatilis) reared in freshwater revealed minor differences between these fish.
• 2.2. Tryptic activity plays a major role in the proteolytic process in both fish.
• 3.3. The activity of seven intestinal proteolytic enzymes was detected utilizing a combination of specific substrates and inhibitors.
• 4.4. High levels of proteolytic activity were detected in both the proximal and distal sections of the fish intestine at a high pH range (9–10).
• 5.5. In situ monitoring of pH levels revealed a lower pH level in the intestinal proximal section of hybrid striped bass compared with the distal section.
• 6.6. In contrast, higher pH levels were detected at the proximal compared with the distal sections of D. labrax intestine.

     

Immunochemical and immunohistochemical determination of ubiquitin in molluscan (Mytilus edulis) smooth muscle

• 1.1. Immunochemical and immunohistochemical distribution of ubiquitin in the anterior byssus retractor muscle (ABRM) of Mytilus edulis was investigated.
• 2.2. In immunostaining, specific ubiquitin immunoreactivity was observed in the cross-sectioned ABRM, and was uniformly localized in this section.
• 3.3. The amount of free ubiquitin in the ABRM homogenate was 130 ± 4.6 ng/mg protein by western blot analysis, and ubiquitin conjugates were found at about 25, 29 and 200–230 kDa.
• 4.4. These findings were similar to those obtained in the skeletal muscle of rat.

     

Alteration of the protein composition of bothrops jararaca venom and venom gland by isoproterenol treatment

• 1.1. The protein composition of Bothrops jararaca venom and venom gland was analyzed through SDS-PAGE, after isoproterenol (IPR) treatment.
• 2.2. Some proteins (47, 48, 57 and 72 kDa) were detected in the gland homogenate from the control but not from the IPR-treated samples.
• 3.3. Three proteins (26.5, 44.5 and 53 kDa) were detected in the venom gland from IPR-treated snakes but not from the venom gland from the control.
• 4.4. In the venom samples proteins of 41 and 74 kDa were detected only in the IPR treated samples, while proteins of 17 and 28 kDa were detected only in the control.
• 5.5. The biological activity of the venom did not change with IPR treatment.

     

O2-binding by the blood of the landhopper,Arcitalitrus dorrieni (Hunt) (Crustacea: Amphipoda)

• 1.1. The O₂-binding characteristics of the blood of the euterrestrial amphipod (landhopper) Arcitalitrus dorrieni have been studied.
• 2.2. The blood exhibited a low O₂ affinity, with a p₅₀ (at pH = 7.8) of 21.4 torr (10°C). Affinity decreased with an increase in temperature at constant pH (ΔH = − 79.4kJ/mol) but the Bohr factor (ΔlogP₅₀/Δ pH = −0.67) was unaffected.
• 3.3. The O₂-carrying capacity of the blood was moderate (1.51 ml/100 ml)
• 4.4. The results support the hypothesis that the blood of terrestrial amphipods is characterized by having a low affinity pigment.

     

Radioimmunological quantification of C21, C19 and C18 steroids in the gonads of Locusta migratoria migratorioides R. and F.

• 1.1. Levels of progesterone, pregnenolone, testosterone, 5α-dihydrotestosterone, estrone and estradiol were measured by radioimmunoassay in purified gonadal extracts of larval and adult male and female Locusta migratoria.
• 2.2. The average steroid contents varied between less than 1 ng to more than 160 ng/g tissue.
• 3.3. Young adults were treated with precocene or ketoconazole in an attempt to influence the steroid contents in gonads.
• 4.4. Ketoconazole treatment had no effect on the steroid contents in gonads whereas precocene treatment resulted in higher contents of androgens.

     

Investigating Lactococcus lactis MG1363 Response to Phage p2 Infection at the Proteome Level

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Highlights

• •The proteomes of L. lactis MG1363 and phage p2 at different stages of infection were characterized.
• •16% (226/1412) of the bacterial proteins detected were unique to infected cultures.
• •A targeted approach using synthetic peptides improved the coverage of phage p2 proteome.
• •By means of proteogenomics, we uncovered a conserved phage protein coded by a previously unannotated gene.
• •Deletion of the bacterial gene llmg_0219 (unknown function) impedes phage p2 infection.

     

Lipid components and utilization in consumers of a seagrass community: An indication of carbon source

• 1.1. The lipid components of three animals, the rock crab Nectocarcinus integrifons, the rock flathead Platycephalus laevigatus and the southern garfish Hyporhamphus melanochir, feeding in the seagrass beds at Corner Inlet, Victoria, Australia have been examined in detail in order to provide further information on seagrass community structure.
• 2.2. Biological marker compounds detected within animal gut content material were used to recognize dietary sources and then utilized by community members.
• 3.3. Both H. melanochir and N. integrifons have been shown to ingest and to varying degrees incorporate seagrass lipid material, thus further confirming the importance of seagrass carbon in the Corner Inlet environment.
• 4.4. The southern sea garfish H. melanochir is observed to remove C₁₈ PUFAs (polyunsaturated fatty acids) from ingested seagrass material.
• 5.5. Seagrass sterols are altered during incorporation into the lipids of this fish.
• 6.6. Lipid-rich digestive juices play a role in the digestive processes of all three animals.
• 7.7. Components tentatively identified as (NMI) (non-methylene interrupted) fatty acids have been detected in the lipids of the garfish H. melanochir and the crab N. integrifons.
• 8.8. The fecal material of all three animals represent possible sources of these lipids (NMI acids) in Corner Inlet sediments.
• 9.9. Based on lipid compositional data, N. integrifons feeds on Posidonia australis detritus and associated epiphyte material.
• 10.10. The removal of both plant and epibiota cellular lipids along the digestive tract of the crab was observed, although structural components such as long chain mono- and α,ω-dicarboxylic acids, which have been previously recognized as seagrass marker lipids are not directly absorbed.

     

Effect of experimental ventilation of the skin on cutaneous oxygen uptake in the carp,Cyprinus carpio

• 1.1. Cutaneous O₂ uptake in the carp, Cyprinus carpio, was determined at various water flow rates across the skin (.V) ranging from 2.5 to 40 ml/min, using flow-through respirometers.
• 2.2. When thickness of water flow was 2mm, cutaneous O₂ uptake remained stable (about 3.8 nmol/cm²/min) at a .V of 20–40 ml/min and decreased with .V below 20 ml/min.
• 3.3. When thickness of water flow was 4 mm, cutaneous O₂ uptake decreased with .V below 40 ml/min.
• 4.4. Apparent water velocity (U') was calculated dividing .V by an area of a cross section of the water flow (0.5 and 1.0 cm² respectively). In both experiments, cutaneous O₂ uptake decreased with U' below 0.7 cm/sec.
• 5.5. This suggests that cutaneous O₂ uptake in the carp is limited at a low water velocity by a resistance of the hypoxic boundary layer.