1H NMR spectroscopic survey of plasma and erythrocytes from selected marsupials and domestic animals of Australia.

1. 1H NMR spectra were acquired from whole plasma, intact erythrocytes, and ultrafiltrates of erythrocytes from nine native and eight introduced (domestic) Australian animals; single-pulse, spin-echo and 2-dimensional spectra were obtained. The aim was to detect and at least semi-quantify metabolites in the samples and compare the profiles amongst the species. 2. The Australian natives that were studied were all marsupials: greater brown bandicoot; bettong; eastern grey kangaroo; red kangaroo; koala; possum; red necked pademelon; Tammar wallaby; and wombat. The introduced mammals that were studied were: cat; cattle; dog; goat; horse; pig; rabbit; and sheep. 3. Because of the range of habitats and diets amongst the animals, it was postulated that the concentrations of the common metabolites in the blood would show marked differences and that there would also be some metabolites that were peculiar to a given animal. There were several major differences in the spectra: in the spectra of plasma, the glycoprotein and lipoprotein resonances showed the largest inter-species variation, whereas the most dramatic finding from the spectra of erythrocytes was a very high concentration of lysine in the cells from the Tammar wallaby.     

Immunochemical characteristics of the ovine polymeric haptoglobin HpC

It was found that haptoglobins of camel, cattle, horse, pig, rat, guinea pig and man form upon immunoelectrophoresis precipitation arcs with antibodies against polymeric sheep haptoglobin C, corresponding to alpha 2-globulins. The immunocrossreactivity of haptoglobins of man and various animal species towards antibody to sheep haptoglobin (100, 88.0, 75.2, 72.1, 56.3, 51.0, 41.3 and 28.0% for haptoglobin of sheep, camel, pig, cattle, man, rat, guinea pig and horse, respectively) was determined. The intensity of crossreactions between sheep haptoglobin and the proteins under study towards antibody to haptoglobin C reflects the similarity of their primary structure and, consequently, the immune homology of their molecules. Using quantitative titration, the antigenic valency values for human (6), sheep (5), cattle (4) and horse (3) haptoglobins were determined.     

Plasma membrane nadh dehydrogenase and Ca2+-dependent potassium transport in erythrocytes of several animal species

Ca2+-dependent K+ transport and plasma membrane NADH dehydrogenase activities have been studied in several 'high-K+' (human, rabbit and guinea pig) and 'low-K+' (dog, cat and sheep) erythrocytes. All the species except sheep showed Ca2+-dependent K+ transport. NADH-ferricyanide reductase was detected in all the species and showed positive correlation with the flavin contents of the membranes. NADH-cytochrome c reductase was very low or absent in dog, sheep and guinea pig membranes. No correlation was found between NADH dehydrogenase and Ca2+-dependent K+ channel activities in the species studied. Nor were any of the above activities correlated with (Na+ + K+)-ATPase activity.     

Hemagglutinating activity inCorynebacterium pyogenes

Thirty-eight strains ofCorynebacterium pyogenes isolated from cases of heifer- and dry-cow mastitis and from other infections of sheep, cows, pigs, and man were screened for agglutination of sheep erythrocytes. Bacteria grown either in serum broth or on blood agar in the presence of CO₂ hemagglutinated. Performance of titrations at 4°C avoided the hemolytic effects ofC. pyogenes. Erythrocytes of cat, chicken, cow, dog, guinea pig, horse, man (Group A), pig, and rabbit were also agglutinated. Pretreatment of sheep erythrocytes with trypsin, pepsin, A1 proteinase or pronase had no effect on agglutinability. Pretreatment ofC. pyogenes with pronase, but not with trypsin, A1 proteinase, or pepsin, abolished hemagglutinating capacity. The hemagglutinin was inactivated by exposure to 60°C for 10 min. Agglutination of sheep erythrocytes was inhibited by five glycoproteins. None of 12 mono-, di-, or trisaccharides nor heparin, chondroitin sulfate, or dextrin inhibited hemagglutination. These data suggest that the receptor may possibly be an oligohexosyl group of a glycoconjugate of lipid nature. Although a few cells of three mastitic strains ofC. pyogenes possessed fimbriae-like surface structures, no correlation between fimbriation and hemagglutinating activity was apparent.     

Microquantitative determination of lactate dehydrogenase isoenzymes in the myocardium and the conducting system

Summary A newly developed technique was used for the electrophoretic separation of lactate dehydrogenase (LDH) isoenzymes from lyophilized tissue samples in the nanogram range. In this study portions of 10–200 ng from the myocardium and the conducting system of cattle, sheep, pig and man were microdissected and analysed.In the heart tissues of cattle, sheep and pig, the isoforms LDH1, LDH2 and LDH3 were detected in species-specific varying amounts. In all these animals, the conducting system is marked by high LDH1 activity, which is present at a ratio of about 2:1 compared with the myocardium. The values in man, however, differ from these values, but this might be due to post-mortem changes. The findings are discussed with respect to possible aerobic-anaerobic functions.     

Haemolysis of various mammalian erythrocytes in sodium chloride, glucose and phosphate-buffer solutions.

Mouse, rat, rabbit, hamster, cow, pig, sheep, guinea-pig, dog and human erythrocytes were studied. A 0.9% or stronger solution of sodium chloride completely prevented haemolysis; sheep and pig erythrocytes appeared the more fragile, while human and dog erythrocytes were not haemolized in concentrations of 0.4% or more. Haemolysis of human, rabbit, cow, hamster, guineapig, pig and sheep erythrocytes was not observed in solutions of 0.4% or more of glucose. Except for sheep, human and dog erythrocytes, haemolysis was depressed in rate but not completely prevented by phosphate-buffer solution of pH 7.0.     

Effects of heterologous sera on fertilized rabbit ova

Undiluted blood serum of various species was used to culture two-celled rabbit ova for 24 hours. It was found that there is an ovocidal factor present in the serum of man, sheep, cattle, goat, and fowl. The factor is lethal rather than inhibitory; exposure to it for 10 minutes will cause the death of the ova. This factor is unstable, thermolabile (destroyed at 55 degrees C. in 30 minutes), and of large molecular size. The strength or concentration of this factor varies according to the origin of the serum, increasing in the order man, sheep, cattle, goat, fowl. The blood serum of rabbit, horse, dog, guinea pig, rat, and pig contains no ovocidal factor against rabbit ova. The ovocidal factor differs from the spermicidal factor, which is present in all the sera of the different species studied with rabbit spermatozoa. Immunization of the guinea pig against rabbit ova is possible. Normal development of young rabbits was obtained by transplantation of ova cultured in the heated or normal serum of other species after 24 hours.     

Differential affinity of natural haemagglutinin of Macrobrachium rosenbergii towards vertebrate erythrocytes: effect of sex, size and moult stage on haemagglutination titre

Lectins play important role in innate immunity of animals. The affinity of the natural haemagglutinin of the giant freshwater prawn Macrobrachium rosenbergii towards vertebrate erythrocytes and its level with relation to sex, size and moult stages were studied. The strongest agglutinating titres in haemolymph of prawns were marked against guinea pig, chicken, Clarias batrachus, and rabbit erythrocytes, and the weakest towards cattle, dog, horse and goat erythrocytes. A moderately agglutinating titre was evident in duck and human erythrocytes. The haemolymph of adult, male or intermoult stage prawns weighing more than 100 g had the highest haemagglutinating activity as compared to their respective counterparts with varied responses observed towards various erythrocytes.     

Comparison of the serum amylases of farm animals

1. Serum isoamylases with alpha-glucosidase activity from cattle, sheep, horses, goats, red deer, pigs and dogs were compared to one another. 2. The isoamylases from cattle and pigs were polymorphic. 3. In agarose gel electrophoresis the isoamylases behaved as alpha-1-globulins but in starch gel electrophoresis they were differentially retarded by affinity effects. 4. Molecular weights were estimated: cattle (417,000); sheep (402,000); horses (420,000); goat (399,000); red deer (405,000); pigs (375,000) and dogs (390,000). 5. Isoelectric points were estimated: cattle, sheep, goat and red deer (pH 3.5); pig (pH 3.6); dog (pH 3.7) and horse (pH 3.9).     

Haemolysins of Salmonella, their role in pathogenesis and subtyping of Salmonella serovars

Haemolysin patterns of 175 strains of different Salmonella enterica subspecies enterica serovars isolated from different animal sources and places were determined using 11 different blood agar media made with either non-washed horse/sheep erythrocytes or with washed erythrocytes of cattle, sheep, horse, goat, rabbit, guinea pig, and human A, O and B blood groups. Study on 47 strains belonging to 10 serovars of Salmonella from buffalo meat (buffen), 42 strains of 11 serovars from goat meat (chevon): 16 strains of Salmonella enterica serovar Paratyphi B and 25 of S. enterica serovar Paratyphi B var Java from fish, meat, meat products and clinical cases; 45 isolates of S. Abortusequi from aborted mares (18), fetal contents (21), aborted donkey mares (2) and 4 reference strains, revealed that all host restricted Salmonella namely, S. enterica serovar Gallinarum, S. enterica serovar Anatum, S. enterica serovar Abortusequi and S. enterica serovar Paratyphi B could be divided into different haemolysin types based on their inability to produce haemolysis on one or more types of blood agar, while strains of all zoonotic Salmonella serovars induced haemolysis on all the 9 types of blood agar made of washed erythrocytes. None of 175 Salmonella could produce hemolytic colonies on blood agar made of non-washed horse/ sheep erythrocytes. Haemolysin type I (lysing all types of washed erythrocytes) was the commonest one among all serovars except S. Abortusequi, none of which lysed horse erythrocytes. Salmonella enterica serovar Abortusequi having hemolytic activity against sheep erythrocytes were more invasive but had lesser ability to survive in sheep mononuclear cells than non-hemolytic strains. Multiplicity of haemolysins appeared significant epidemiological tool.     

Lipid peroxidation and haemoglobin degradation in red blood cells exposed to t-butyl hydroperoxide. Effects of the hexose monophosphate shunt as mediated by glutathione and ascorbate.

Lipid peroxidation and haemoglobin degradation were the two extremes of a spectrum of oxidative damage in red cells exposed to t-butyl hydroperoxide. The exact position in this spectrum depended on the availability of glucose and the ligand state of haemoglobin. In red cells containing oxy- or carbonmono-oxy-haemoglobin, hexose monophosphate-shunt activity was mainly responsible for metabolism of t-butyl hydroperoxide; haem groups were the main scavengers in red cells containing methaemoglobin. Glutathione, via glutathione peroxidase, accounted for nearly all of the hydroperoxide metabolizing activity of the hexose monophosphate shunt. Glucose protection against lipid peroxidation was almost entirely mediated by glutathione, whereas glucose protection of haemoglobin was only partly mediated by glutathione. Physiological concentrations of intracellular or extracellular ascorbate had no effect on consumption of t-butyl hydroperoxide or oxidation of haemoglobin. Ascorbate was mainly involved in scavenging chain-propagating species involved in lipid peroxidation. The protective effect of intracellular ascorbate against lipid peroxidation was about 100% glucose-dependent and about 50% glutathione-dependent. Extracellular ascorbate functioned largely without a requirement for glucose metabolism, although some synergistic effects between extracellular ascorbate and glutathione were observed. Lipid peroxidation was not dependent on the rate or completion of t-butyl hydroperoxide consumption but rather on the route of consumption. Lipid peroxidation appears to depend on the balance between the presence of initiators of lipid peroxidation (oxyhaemoglobin and low concentrations of methaemoglobin) and terminators of lipid peroxidation (glutathione, ascorbate, high concentrations of methaemoglobin).     

Mechanisms of ascorbic acid recycling in human erythrocytes.

Vitamin C, or ascorbic acid, is efficiently recycled from its oxidized forms by human erythrocytes. In this work the dependence of this recycling on reduced glutathione (GSH) was evaluated with regard to activation of the pentose cycle and to changes in pyridine nucleotide concentrations. The two-electron-oxidized form of ascorbic acid, dehydroascorbic acid (DHA) was rapidly taken up by erythrocytes and reduced to ascorbate, which reached intracellular concentrations as high as 2 mM. In the absence of D-glucose, DHA caused dose-dependent decreases in erythrocyte GSH, NADPH, and NADH concentrations. In the presence of 5 mM D-glucose, GSH and NADH concentrations were maintained, but those of NADPH decreased. Reduction of extracellular ferricyanide by erythrocytes, which reflects intracellular ascorbate recycling, was also enhanced by D-glucose, and ferricyanide activated the pentose cycle. Diethylmaleate at concentrations up to 1 mM was found to specifically deplete erythrocyte GSH by 75-90% without causing oxidant stress in the cells. Such GSH-depleted erythrocytes showed parallel decreases in their ability to take up and reduce DHA to ascorbate, and to reduce extracellular ferricyanide. These results show that DHA reduction involves GSH-dependent activation of D-glucose metabolism in the pentose cycle, but that in the absence of D-glucose DHA reduction can also utilize NADH.     

Isoenzyme patterns of glutathione transferases from mammalian erythrocytes

The occurrence of glutathione transferase isoenzymes in mammalian erythrocytes was investigated. The enzymes present in the hemolysates of human, horse, beef, pig, and sheep erythrocytes were purified by a column of GSH-linked epoxy-activated Sepharose 6B and subjected to an isoelectric focusing run in the pH range 3.5-10. Human and horse preparations were resolved in a single peak of activity centered at pH 4.6 and 5.9, respectively. Two forms with a maximum of activity at pH 4.9 and 7.0 and four with a maximum at pH 5.9, 6.5, 7.1, and 8.1 were separated from bovine and porcine erythrocytes. At least six forms ranging from pH 4.3 to pH 7.1 were present in the ovine preparation, the neutral contributing more than 90% of total activity. The subunit composition of affinity-bound fractions was studied by sodium dodecyl sulfate-gel electrophoresis. The analysis revealed that erythrocyte glutathione transferases are composed of subunits of identical molecular weights. This result suggests that the polymorphism existing in beef, pig, and sheep may be due to charge isomers. The erythrocyte glutathione transferases did not express selenium-independent GSH peroxidase activity.     

Plasma membrane NADH dehydrogenase and Ca2+-dependent potassium transport in erythrocytes of several animal species

Ca²⁺-dependent K⁺ transport and plasma membrane NADH dehydrogenase activities have been studied in several ‘high-K⁺’ (human, rabbit and guinea pig) and ‘low-K⁺’ (dog, cat and sheep) erythrocytes. All the species except sheep showed Ca²⁺-dependent K⁺ transport. NADH-ferricyanide reductase was detected in all the species and showed positive correlation with the flavin contents of the membranes. NADH-cytochrome $\text{c}$ reductase was very low or absent in dog, sheep and guinea pig membranes. No correlation was found between NADH dehydrogenase and Ca²⁺-dependent K⁺ channel activities in the species studied. Nor were any of the above activities correlated with $\text{(}\text{Na}{}^{\mathrm{+}}\text{+}\text{K}{}^{\mathrm{+}}\text{)-}\text{ATPase}$ activity.     

Protein binding of cortisol by means of competitive adsorption: application to cortisol binding by serum of sixteen eutherian mammals.

1. Binding of 3H-cortisol by serum proteins by means of competitive adsorption was relatively high by serum of the gerbil, human, rabbit, sheep, tree shrew, hamster, rhesus monkey and horse. 2. A somewhat lower binding was observed by serum proteins of the baboon, cattle, dog, rat and cat. 3. Serum taken from either the mouse, guinea pig or pig gave very flat binding curves, specific binding not exceeding 5% of added 3H-cortisol. 4. It is concluded that the measurement of protein-binding of 3H-cortisol by means of competitive adsorption is a reliable method for serum of most eutherian species but is unsuited if serum of the mouse, guinea pig or pig is used.     

Comparative studies on the gastric glycopeptide in eleven animal species.

1. Glycopeptides in the stomachs of eleven mammalian species, including human, rabbit, horse, cow, pig, goat, sheep, dog, cat, guinea pig and rat were assayed by determining the carbohydrate content of materials which remained after proteolysis. 2. The glycopeptide content was higher in the mucosa than in the muscular layer including serosa, especially in the porcine stomach and the fourth stomachs of the ruminants than in the stomachs of any other animals. 3. The glycopeptide, which was stained with both alcian blue and PAS, was absent or sparingly present in the mucosae of the human, rabbit, horse stomachs and in the mucosae of the first to third stomachs of the cow, goat and sheep, whereas in the mucosae of the pig, dog, cat, guinea pig and rat stomachs and in the mucosae of the fourth stomachs of the cow, goat and sheep, it was found in noticeable extents.     

Absence of endo-beta-N-acetylglucosaminidase activity in the kidneys of sheep, cattle and pig.

The kidneys of man, sheep, cattle and pig were all found to contain 1-aspartamido-beta-acetylglucosamine amidohydrolase activity. However, among these, only human kidney was found to contain endo-beta-N-acetylglucosaminidase activity. The absence of this enzyme in the kidneys of sheep and cattle explains why the oligosaccharides accumulated in, and excreted by, sheep and cattle afflicted with disorders of glycoprotein catabolism (i.e. alpha-mannosidosis and beta-mannosidosis) contain two N-acetylglucosamine residues at the reducing terminus instead of one, as is the case for human patients afflicted with similar disorders.     

Bovine Adenovirus

Nine strains of an adenovirus serotype were recovered in bovine testicle cell cultures from Japanese cattle suffering with an acute febrile illness accompanied by rhinorrhea and diarrhea. The isolated virus was shown to have the physicochemical properties of the adenovirus group such as the nucleic acid type, the size and ultrastructure of the virion, and the resistance to ether and chloroform. The isolated virus produced eosinophilic intranuclear inclusion bodies characteristic of adenoviruses and the group specific antigen of adenovirus in bovine testicle cell culture. According to the results of cross-neutralization tests the isolated virus represents a serological type distinct from bovine adenovirus types 1, 2 and 3 and from the Nagano virus. The isolated virus agglutinated erythrocytes of cattle, sheep, goat, guinea pig, rat, hamster and mouse, but not those of vervet monkey, horse, goose and chicken. HI test using cattle erythrocytes corroborated the results of serological typing by neutralization tests.     

Distribution of collagens type I,type III and type V in the pancreas of rat,dog, pig and man

The presence of collagens type I, type III and type V was determined immunohistochemically in pancreatic tissue of rat, pig, dog and man. The reaction to anti-collagen type I is weak (pig, dog) or moderate (rat, man) in the peri-insular region and in the lobar, lobular and acinar septa, whereas the reaction to anti-collagen type III is well developed. In rat and dog, the latter reaction deposit on the lobar and acinar septa is prominent. These elements only show a moderate reaction intensity in pig and man. The peri-insular region displays a weak (rat, dog, man) or very weak (pig) reaction against collagen type III. Anti-collagen type V reacts moderately (rat, dog, man) or weakly (pig) in the lobar and lobular septa. The acinar septa show a moderate (rat, dog, man) or very weak (pig) reaction. This information regarding the types and distribution of the collagenous compounds in pancreatic extracellular matrix could lead to differentiated enzymatic pancreas dissociation and, ultimately, increased islet yield and improved reproducibility of pancreatic islet isolation procedures for transplantation purposes.