bluuebunny/crossref_metadata_2025 huggingface dataset:
Distribution and density ofα– andβ–adrenergic receptor binding sites in the bovine mammary gland
Authors: Hammon Harald M. , Bruckmaier Rupert M. , Honegger Ulrich E. , Blum Jürg W.
Published: June 2009

Radioreceptor binding studies were designed to localize and determine the number of α and β-adrenergic receptors in the mammary gland of lactating cows.3H-prazosin,3H-rauwolscine and3H-dihydroalprenolol were used for the regional characterization of α1, α2- and β-adrenergic receptors by competitive inhibition of binding of3H-ligands with unlabelled adrenergic agonists and antagonists. The α1-, α2- and β2-adrenergic receptor subtypes could thus be demonstrated in the regions of the teats, large mammary ducts and parenchyma. Tissues of the teat wall, of the large mammary ducts above the gland cistern and of the mammary parenchyma were prepared to determine the density of α1, α2- and β-receptors by saturation binding assays using3H-prazosin,3H-rauwolscine and3H-dihydroalprenolol respectively. Binding to high affinity sites was reversible within minutes and saturable. Equilibrium was reached within minutes. The number of α1-and α2-adrenergic receptors decreased from the teat to the mammary ducts to the parenchyma. Most of the α1- and α2-adrenergic receptors were found in the teat wall, whereas in the parenchyma α-adrenergic receptors were absent or barely detectable. The density of β-adrenergic receptors was similar in the teat wall and the large mammary ducts, but much lower in the parenchyma. Thus, α1, α2- and β-adrenergic receptors were found mainly in the milk purging system and hardly at all in mammary parenchyma. Inhibition of milk removal by α-adrenergic stimulation is possibly due to constriction of teat wall and to constriction of the mammary ducts, whereas enhanced milk flow after β-adrenergic stimulation is possibly due to relaxation not only of the teat sphincter and teat wall, but probably also of the large mammary ducts.
Rapid, sensitive two-site ELISA for detection of cows' milk in goats' or ewes' milk using monoclonal antibodies
Authors: Levieux Didier , Venien Annie
Published: June 2009

A sandwich ELISA (enzyme-linked immunosorbent assay) of the two-site type has been successfully developed for the detection of cows' milk in goats' or ewes' milk. The assay uses two monoclonal antibodies (MAb) raised in mice against cows' β-lactoglobulin (β-lg). These MAb recognize different epitopes of the β-lg, which are sufficiently distinct to allow simultaneous binding of the corresponding antibodies. One of the MAb recognizes a species-specific epitope of the bovine β-lg and was adsorbed to a plastic microtitration plate (capture antibody). The second MAb was labelled with peroxidase and used to detect the captured cows' β-lg. Factors affecting assay performance were investigated. The optimized assay is highly specific, reproducible (intra- and inter-assay CV were 8 and 13% respectively) and sensitive: as little as 5 ng β-lg/ml or 1 part cows' milk per 100000 parts goats' or ewes' milk can be detected. The technique is robust, cheap, rapid, reliable and suitable for high sample throughput, semi-automation and screening surveys. The MAb used guarantee the high specificity of the assay and indefinite reagent supply of constant quality once approved by collaborative national or international trials.
Some aspects of measuring the size and rate of dispersion of milk powder agglomerates using the Malvern Particle Sizer 2600c
Authors: Chen X. Dong , Lloyd Richard J.
Published: June 2009

The Malvern Particle Sizer, which applies Fraunhofer light diffraction theory, has recently become popular for measuring particle size distributions in dried milk and other powders. The size distribution is obtained by dispersing the powder in air or a suitable liquid in the laser light beam of the instrument. The suitability of the dry powder dispersion system (DPD system) of the Malvern Particle Sizer 2600c for characterizing agglomerated milk powders was examined. It was found that, although the instrument gave reproducible and reliable results for non-agglomerated powders (standard milk powders), milk powder agglomerates could be broken in the standard DPD system and erroneous size distributions might be obtained. However, the DPD system had potential for providing an objective test of the mechanical stability of milk powder agglomerates. In addition, the transient size distribution during dispersion of an instantized agglomerated milk powder in water was measured at intervals of about 20 s.
Thermostability of yeast lactase (Kluyveromyces marxianus) in milk
Authors: Mahoney Raymond R. , Wilder Teresa
Published: June 2009

The half-life (∼ 2 h) of commercial yeast lactase (Kluyveromyces marxianus) in milk at 45 °C was 20 times greater than in milk salts and 50 times greater than in phosphate buffer. K was a more effective stabilizer than Na, both in milk salts and buffer. Stability was markedly reduced by the absence of divalent cations. Lactose and caseinate both stabilized the enzyme about 5-fold in milk salts, but together they stabilized it almost 50-fold. At equimolar concentrations, galactose was almost as effective as lactose as a stabilizer, but glucose was much less effective and sucrose had no effect. Stability in milk was independent of enzyme concentration, but varied with concentration of milk solids, reaching a maximum at about 25% solids. In milk, enzyme denaturation was more sensitive to changes in temperature in the range 42–53 °C than in the range 30–40 °C. At a concentration of 10O-nitrophenyl-β-d-galactopyranoside units/ml it took 19 min at 45 °C to achieve 80% hydrolysis of lactose in milk, compared to 87 min at 30 °C.
Influence of lactic cultures in denaturation of whey proteins during fermentation of milk
Authors: Vaitheeswaran Nataraja Iyer , Bhat Gajanan S.
Published: June 2009

Undenatured whey protein (UWP) content of skim milk acidified with lactic acid or cultured with lactic cultures was estimated by a dye-binding method. The UWP content decreased with increase in acidity and the denaturation was only partly reversible on neutralization to the original acidity. The decrease in UWP was higher in cultured milk than in the milk acidified to the same extent with lactic acid, indicating the effect of lactic cultures in denaturation of whey proteins during fermentation of milk. Among the lactic cultures the denaturation effect of Lactobacillus delbrueckii subsp. bulgaricus was highest, followed by Streptococcus salivarius subsp. thermophilus, Lactococcus lactis subsp. lactis and Lact. lactis biovar diacetylactis. Denaturation of whey proteins by lactic cultures was found to be partly irreversible.
Laboratory simulations of a commercial UHT treatment using an oil bath and a programmable, electronically temperature-controlled oven
Authors: O'Connor Robyn E. , O'Connor Colin
Published: June 2009

Heat stability of secreted bacterial proteinase is normally assessed using an oil bath test in which the sample is heated at a temperature and time representing the commercial UHT treatment of milk. The temperature/time (T/t) profiles of the oil bath and the commercial UHT process being represented are not taken into account. Two laboratory procedures were used to simulate a commercial UHT process in which milk was heated to 140°C and held for 3 s in a Spiroflo heat exchanger. They consisted of a conventional oil bath test and the use of a programmable, electronically temperature-controlled oven.T/tprofiles were established for each heat treatment. TheT/tprofile of the oil bath test was predicted accurately using the governing heat transfer equation.B*andC*values, which measure the severity of a heat process, were calculated from theT/tprofiles and used to compare the three different heat treatments.B*andC*values of 4·95 and 1·92 respectively were calculated for the Spiroflo heat exchanger. An oil bath test, in which the sample was immersed in a bath at 136°C for 73 s, gave approximately the sameB*andC*values as calculated for the Spiroflo heat exchanger.B*andC*values of 5·01 and 1·73 respectively were calculated for the oven procedure. The oven test gave the better laboratory simulation of the Spiroflo UHT treatment. Despite the slight difference in each of theB*andC*values between the oven and the Spiroflo, theT/tprofile of the oven test closely resembled that of the Spiroflo. TheT/tprofile of the oil bath was completely dissimilar. Although it was possible to use an oil bath test to replicate simultaneously both theB*andC*values of this particular UHT treatment, this would not be the case for other commercial processes.
Effect of plane of nutrition on insulin sensitivity during lactation in the ewe
Authors: Metcalf John A. , Weekes Timothy E. C.
Published: June 2009

The hyperinsulinaemic euglycaemic insulin clamp technique was used to compare insulin sensitivity in lactating ewes at two levels of feeding. Clamps were performed at two (restricted intake) or three (ad libitum intake) stages of lactation and also 30 d after drying off. Dose response curves for insulin were constructed using the glucose metabolic clearance rate (MCR) as the measure of glucose metabolism and these were statistically compared between stages of lactation within the feeding levels, and also between feeding regimes. Animals on a restricted feed intake showed a weight loss throughout lactation, coupled with a lower insulin sensitivity, as measured by the ED50 (concentration of insulin required to produce a half maximal increase in MCR), while those on ad libitum feeding showed a weight gain and a decreased insulin sensitivity in the dry period. Endogenous glucose production was less sensitive to inhibition by insulin in the animals subjected to restricted food intake, compared with those on the ad libitum regime, which suggested a major role for the plane of nutrition in adjusting the homeorhetic control of metabolism during lactation.
Bacteriological investigations of clinical mastitis in heifers in Sweden
Authors: Jonsson Per , Olsson Sven-Ove , Olofson Ann-Sophie , Fälth Christer , Holmberg Olof , Funke Hans
Published: June 2009

Bacterial analyses were carried out of 2069 udder secretions, isolated from 1481 heifers with mastitis in eight veterinary districts in Sweden. Streptococci, e.g. Streptococcus dysgalactiae and Str. uberis, dominated the bacterial flora, being isolated from 34·4 and 19·5% respectively of heifers with clinical mastitis occurring from puberty up to 14 d post partum. Bacterial species generally regarded as important pathogens in the summer mastitis complex, e.g. Actinomyces pyogenes, Stuart–Schwan coccus and strictly anaerobic bacteria such as Peptostreptococcus indolicus, Fusobacterium necrophorum and Bacteroides melaninogenicus were isolated at low frequencies (13·2, 6·3, 9·4, 3·8 and 1·3% respectively). When the cases of mastitis studied were restricted to those appearing in heifers pre partum, May 15 to October 14 (summer mastitis), these bacterial species were isolated at higher percentages (27·1, 14·4, 21·4, 13·5 and 5·2% respectively). These figures were, nevertheless, still lower than those published in reports from other countries. Whether considered up to 14 d post partum or only pre partum, there were no significant differences in the frequencies of A. pyogenes isolated at different seasons. There were geographical differences in bacterial incidence, e.g. Staphylococcus aureus was isolated significantly more often in northern regions whereas Str. dysgalactiae was more common in the south. This and other Swedish investigations support the theory that A. pyogenes and strictly anaerobic bacteria are ‘secondary invaders’ that depend on Str. dysgalactiae to cause a primary infection. It is stressed that the udders of all heifers should be examined daily so that cases of mastitis can be treated immediately.
Influence of milk flow rate and streak canal length on new intramammary infection in dairy cows
Authors: Grindal Robert J. , Walton Andrew W. , Hillerton J. Eric
Published: June 2009

Eighteen cows with a wide range of quarter peak flow rates (0·35–2·22 kg/min) were inoculated with Streptococcus agalactiae and Str. dysgalactiae 4 mm into each streak canal every 3 d for 12 d. Thirty of the 72 quarters developed intramammary infection. Mean peak flow rate and length of streak canal of those quarters that became infected were 1·26 ± 0·08 kg/min (mean ± SEM) and 11·47 ± 0·41 mm respectively, compared with 1·01 ± 0·05 kg/min and 12·05 ± 0·35 mm for those that remained uninfected. Logistic regression analysis showed that the probability of infection increased significantly with the increase in peak flow rate (P = O·O1). The influence of streak canal length on new infection, after allowing for the effect of peak flow rate, was not significant at the 5% level (P = 0·07), suggesting that there may be an inverse relationship between teat duct length and infection, but that it is of secondary importance to peak flow rate. If increased mass of milk distends the teat duct by raising intramammary pressure, then quarter susceptibility to infection is further compromised. These results strongly suggest that the benefits of reduced infection from mastitis control, achieved despite dramatic increases in milk flow rate and milk yield, are significantly underestimated.
Effects of α- and β-adrenergic agonists on intramammary pressure and milk flow in dairy cows
Authors: Bruckmaier Rupert , Mayer Harald , Schams Dieter
Published: June 2009

α-Adrenergic mimetics inhibit milk removal whereas β-adrenergic mimetics have a favourable effect upon milk removal in dairy cows. This work aimed to relate these effects to the milk ejection reflex mechanism and/or milk flow from the cisternal cavities only. The α-adrenergic agonist phenylephrine and the β-adrenergic agonist isoproterenol were injected prior to and after milk ejection. Applied after milk ejection, the phenylephrine treatment reduced intramammary pressure (IMP) and milk yield markedly. Phenylephrine application before milk ejection prevented IMP from rising during teat stimulation or after oxytocin injection. Milking produced small amounts of milk at minimal milk flow rates. The very different responses to phenylephrine application prior to and after milk ejection indicate separate adrenergic inhibitory mechanisms for milk ejection and milk flow. The application of the β-adrenergic agonist isoproterenol did not alter the IMP. Nevertheless, this compound improved milk removal by increasing milk flow. The observation of bimodal milk flow at milkings without preceding teat stimulation indicates a delayed milk ejection. Isoproterenol injection without teat stimulation enhanced milk flow rates, but the bimodality remained unchanged. We conclude that isoproterenol enhances milk flow without affectiiig milk ejection.
Enzymic modification of αs1-casein with peptidylarginine deiminase: preparation of less acid-coagulable and less calcium-sensitive casein
Authors: Azuma Norihiro , Nara Kazutoshi , Kanno Choemon
Published: June 2009

Enzymic modification with peptidylarginine deiminase (EC 3.5.3.15) enabled five out of six arginyl residues in αs1-casein to be convcrted to citrullyl residues, only the N-terminal arginyl residue remaining unaffected. An increase in the net negative charge was confirmed by PAGE. The isoelectric point was decreased from 4·46 for the intact αsl-casein to 4·30 for the deiminated type, while simultaneously lowering the acid-precipitation starting point from pH 5·17 to pH 4·62. The deiminated αs1-casein self-associated less in the absence of Ca and was less Ca-sensitive than the native type, although its Ca-binding ability was slightly enhanced. In the presence of 25 mM-CaC12andK-casein, Ca-induced precipitation of αs1-casein did not occur, the solution of the mixture remaining transparent. Deimination of αs1-casein resulted in altering its characteristics, possibly by interfering with interactions through hydrophobicity and/or hydrogen bonding. The positive charge of the arginyl residues might play an important role in casein micelle formation.
Immunoadjuvant activity of oralLactobacillus casei: influence of dose on the secretory immune response and protective capacity in intestinal infections
Authors: Perdigón Gabriela , Alvarez Susana , de Ruiz Holgado Aída Pesce
Published: June 2009

Lactobacilli, often used as effectors of host functions, could play an important role in maintaining human health by controlling other intestinal microorganisms capable of producing harmful effects. Using an experimental model, we studied the effect of different oral doses ofLactobacillus caseion the secretory IgA response and the protective capacity of the microorganism in preventing intestinal infections. The optimization of the protective dose ofLb. caseiby previous feeding and the use of the lactobacillus as an immunological way to control enteric infections were investigated. We found that conventional mice were protected against infection withSalmonella typhimuriumandEscherichia coliby previous feeding for 2 consecutive days with a dailyLb. caseidose of 1·2 × 109cfu/mouse. Previous feeding for 7 d proved less effective, and feeding for 5 d afforded no protection at all. We were also able to demonstrate that the protective effect ofLb. caseiagainstSal. typhimuriumandEsch. coliwas connected mainly with the high level of IgA antipathogen antibodies present in intestinal secretions. β-Glucuronidase (EC 3.2.1.31) and β-galactosidase (EC 3.2.1.23) activities, measured both in the intestinal fluid and histological samples, showed a marked increase in intestinal inflammatory response on day 5 of feeding. These results show thatLb. caseiplays an important role in the prevention of enteric infections, a low dose being enough for protection against intestinal infections by increasing IgA secretion into the intestinal lumen, thus providing adequate defences for the mucosal surface. A previously administered dose of this magnitude could therefore be used as an oral adjuvant in preventing enteric infections.
Somatic cell counts in relation to infection status of the goat udder
Authors: Kalogridou-Vassiliadou Despina , Manolkidis Konstantinos , Tsigoida Afrodite
Published: June 2009

Bacteriological analyses, cell counts using the Fossomatic method and California Mastitis Test were performed on 1523 goat milk samples taken aseptically at monthly intervals throughout lactation from three goat herds. Of the goat udders, 81·4% were infected, minor pathogens being the most frequent isolates (65·7%). Differences in the level of infection by minor pathogens were found betwccn herds. Cell counts were influenced by stage of lactation and intramammary infection. Cell counts < 106cells/ml were found in 80% of milk samples infected by major pathogens and in 45% infected by minor pathogens. About 81% of udders infected with major pathogens gave California Mastitis Test scores of 2 and 3, compared with 20% for uninfected goats. A high proportion (65%) of udders infected with minor pathogens also produced scores of 2 and 3. A significant positive correlation was found between the California Mastitis Test and cell counts. The use of cell counts for the detection of abnormal goat milk is discussed.
Capture immunoassay for the diagnosis of bovine mastitis using a monoclonal antibody to polymorphonuclear granulocytes
Authors: O'Sullivan Catherine A. , Joyce Patrick J. , Sloan Teresa , Shattock Alan G.
Published: June 2009

A direct capture enzyme-linked immunosorbent assay (ELISA) was developed to measure elevated polymorphonuclear granulocyte (PMN) antigens using horseradish peroxidase (EC 1.11.1.7) conjugated rabbit polyclonal anti-PMN antisera and a monoclonal antibody specific for PMN cells. Optical densities obtained in the ELISA were used to predict the cell counts of milk samples. Predicted counts were not significantly different from actual somatic cell counts (SCC). In a total of 156 bovine milk samples the correlation coefficient between somatic cell counting, taking > 500000 cells/ml as being indicative of mastitis, and the assay was 0·94, yielding an assay sensitivity of 95·2% and a specificity of 97·3%. In further trials the ELISA could detect elevated PMN antigens in milk with SCC as low as 100000 cells/ml. The results indicate that the monoclonal antibody based direct ELISA has excellent potential in the detection and determination of bovine mastitis.
B-mode ultrasonography of mammary glands of cows, goats and sheep during α- and β-adrenergic agonist and oxytocin administration
Authors: Bruckmaier Rupert M. , Blum Jürg W.
Published: June 2009

Teat and gland cistern of the mammary glands of five dairy cows, five goats and five sheep were scanned in a water bath during α- andβ-adrenergic agonist and oxytocin administration. A 5 MHz linear array scanner was used to create vertical cut pictures with the scan plane longitudinally through the teat channel. The i.v. injection of the α-adrenergic agonist phenylephrine (30 μg/kg) induced diminution of the section area through teat and gland cistern by 38±17% on average within 1 min in all three species. In contrast, the i.v. injection of theβ-adrenergic agonist isoproterenol (1 μg/kg) did not change the cisternal areas. The i.v. injection of oxytocin (2·0 × 10−3i.u./kg) elicited an enlargement of teat and gland cistern area by 48±12% on average. Ultrasonography proved to be a valuable system for visualizing changes of the cisternal volume. Smooth muscle contractions in response to phenylephrine administration are thought to cause area reduction, whereas an expected smooth muscle relaxation after injection of isoproterenol could not be observed by ultrasonography. Milk ejection as induced by oxytocin administration caused dramatic enlargement of the cistern area in all three species.
Milking of cows in late pregnancy: milk production during this period and during the succeeding lactation
Authors: Rémond Bernard , Ollier Alain , Miranda Guy
Published: June 2009

Fifteen lactating cows were milked throughout pregnancy, and the effects on milk performance were studied during this period and during the succeeding lactation, relative to 11 conventionally managed cows (2 months dry before calving) as controls. During the last 2 months of pregnancy, only nine cows did not dry off spontaneously. Protein and fat concentrations in milk increased rapidly, but the concentration of lactose, corrected for milk yield, did not change. The ratios of individual caseins to total protein decreased with the quantity of milk produced, but only for yields below ˜ 6 kg/d. The relative proportion of κ-casein tended to decrease in the last milkings. During the succeeding lactation (first 15 weeks after calving and first 6 weeks of grazing) continuously milked cows yielded 4 kg milk/d less than the cows of the other group. The protein content of their milk was higher (2–3 g/kg depending on the period) than that of the control group, and the lactose content tended (p < 0·10) to be lower. Changes in the relative proportions of nitrogenous fractions with time were not different in the two groups. Differences between the two groups in the concentration of protein in milk, and in the concentration of glucose and non-esterified fatty acids in the plasma, suggest a better energy balance for the continuously milked cows during the succeeding lactation.
Factors in ruminant colostrum that influence cell growth and murine IgE antibody responses
Authors: Watson Dennis L. , Francis Geoffrey L. , Ballard F. John
Published: June 2009

Bovine colostrum was investigated as a source of biologically active molecules capable of stimulating the growth of mammalian cells in culture and modifying the immune response in a murine model. An extract prepared from bovine colostral whey by cation exchange and re versed-phase chromatography stimulated the growth of L6 rat myoblasts, Balb/c-3T3 mouse fibroblasts and BHK-21 baby hamster kidney cells with equal or greater potency than fetal bovine serum. Fractionation of the bovine colostral extract by gel-permeation chromatography in M-acetic acid identified a number of cell-growth factors for each cell type. Bovine colostral extract was compared with an ovine colostral whey preparation for its ability to modulate IgE antibody responses in mice. Doses of 8 and 4 mg/d of ovine colostral whey or bovine colostral extract specifically suppressed IgE antibody responses, whereas at lower doses suppression did not occur. We conclude that bovine colostrum contains cell-growth factors as well as immunomodulatory factors that are able to regulate the IgE response in a heterologous species.
Continuously elevated concentrations of oxytocin during milking are necessary for complete milk removal in dairy cows
Authors: Bruckmaier Rupert M. , Schams Dieter , Blum Jürg W.
Published: June 2009

The importance of elevated concentrations of oxytocin (OT) during the entire milking period was investigated in seven primiparous dairy cows with inherent disturbed milk ejection and in sixteen healthy cows with disturbed milk ejection induced by placing them in an operating theatre. Disturbance of milk removal in both groups has previously been demonstrated to be exclusively due to central blockage of the expected OT release in response to teat stimulation and milking. However, milk ejection can be induced by exogenous OT. OT (0·2 i.u.) was injected i.v. before milking and 49 ± 6% of the total milk was removed. When plasma OT decreased, milk flow stopped. In response to a second and third injection of 0·2 i.u. OT, 30 ± 4 and 7 ± 2% of the milk were removed respectively. The remaining milk was removed with 10 i.u. OT. The lag time from injection of OT to the start of milk flow was inversely correlated with the amount of milk actually removed in response to the OT injection. If 0·2 i.u. OT was injected during intramammary pressure (IMP) recording, IMP immediately increased to its maximum value. After 2·5 ± 0·3 min, IMP decreased to an intermediate IMP (between preinjection and maximum IMP). After two additional injections of 0·2 i.u. OT and after injection of 0·5 i.u. OT, IMP increased to a similar maximum. However, after injection of 0·5 i.u. OT, maximum IMP lasted longer (2·9 ± 0·3 min; P < 0·05) than after injections of 0·2 i.u. If OT was continuously infused (0·15 i.u./min) during milking, milk flow lasted until the udder was completely emptied. IMP increased during OT infusion to a maximum which remained stable until infusion was stopped after 10 min. The same IMP maximum was reached after the first individual OT injection (0·2 i.u.), but when plasma OT decreased towards basal concentrations, milk flow ceased and IMP decreased to an intermediate level. Thus continuously elevated OT concentrations such as those during infusion or during normal milking are necessary for complete milk removal.
Effect of suckling during early lactation and changeover to machine milking on plasma oxytocin and cortisol levels and milking characteristics in Holstein cows
Authors: Tančin Vladimír , Harcek L'Ubomír , Brouček Ján , Uhrinčať Michal , Mihina Štefan
Published: June 2009

Plasma concentrations of oxytocin and cortisol, and milk yield and flow rates, were compared in three primiparous cows and two cows in their second lactation during suckling and subsequent machine milking. After calving, cows suckled their calves for 3–4 weeks and then the experiment was carried out over 4 d. Blood samples were taken prior to, during and after suckling or evening machine milking (EMM) on day 1 of the experiment (the last day of suckling), day 2 (first EMM) and day 4 (third EMM). After weaning and rehousing, cows were machine milked twice daily. During the first EMM, average milk yield and flow rate in the second minute of milking were significantly lower (P < 0·05) than corresponding values for the other 2 d. Plasma oxytocin concentrations were lower during the first than during the third EMM (P < 0·001) and suckling (P < 0·001), and more oxytocin was released during the third EMM than during suckling (P < 0·01). Cortisol concentrations were higher during and after the first EMM than during the third EMM. Thus in cows sucked for several weeks after calving and then separated from their calves and rehoused we found a transient decrease in oxytocin release, milk yield and flow rates during the first machine milking.
Primary structure of ovine αsl-caseins: localization of phosphorylation sites and characterization of genetic variants A, C and D
Authors: Ferranti Pasquale , Malorni Antonio , Nitti Gianpaolo , Laezza Pasquale , Pizzano Rosa , Chianese Lina , Addeo Francesco
Published: June 2009

The primary structures of ovine α>s1-casein variants A, C and D (formerly called Welsh variant) were determined. Separation of variants from whole casein was achieved using a fast and reliable reversed-phase HPLC method. Extended structural characterization of the purified proteins using electrospray mass spectrometry, automated Edman degradation and peptide mapping by means of HPLC-fast atom bombardment-mass spectrometry demonstrated that the mature protein was a mixture of two molecular species that differed in the deletion of residues 141–148 and were therefore 199 and 191 residues long respectively. The 199 residue peptide chain, which accounted for ∼ 80% of the entire translated αsl-casein, was as long as its caprine and bovine counterparts, and had a 98 and 89 % degree of identity with those two proteins respectively. Nine serine residues (positions 12, 44, 46, 64 to 68 and 75) were fully phosphorylated in αsl-casein A, whereas Ser115and Ser41were phosphorylated by ∼ 50 and ∼ 20% respectively. The differences between the three genetic variants A, C and D were simple silent substitutions, which however involved the degree to which the protein was phosphorylated. Variant C differed from variant A in the substitution Ser13→> Pro13which determined the loss of the phosphate group on site 12 of the protein chain, SerP12→>Ser12. A further substitution, SerP68→> Asn68caused the disappearance of both phosphate groups in the phosphorylated residues Ser64and Ser66in variant D; in this last casein variant there was no evidence of phosphorylation at Ser41.
Two dimensional electrophoresis and immunoblotting for the study of ovine whey protein polymorphism
Authors: López-Gálvez Gloria , Juárez Manuela , Ramos Mercedes
Published: June 2009

Different electrophoretic techniques have been applied to the study of ovine whey protein polymorphism. Isoelectric focusing (IEF) over the pH gradient 3·5–9·5 was a suitable method for resolving both genetic variants of ovine β-lactoglobulin (β-lg); using this type of IEF, ovine β-lg A seemed to be defined by a major and a minor ‘satellite’ band. Approximate isoelectric point, relative molecular mass, amino acid composition and N-terminal sequence of this minor band were determined. A good separation of ovine β-lg was achieved using ultrathin layer isoelectric focusing (UTLIEF) over the pH gradient 2·5–7·0. However, addition of urea in UTLIEF gels led to some differences in the patterns of the ovine whey proteins when compared with those obtained by IEF on gels not containing urea. Two dimensional electrophoretic techniques provided a considerable separation of ovine whey proteins. Immunoblotting applied to a two dimensional separation permitted the identification of the bands belonging to β-lg and α-lactalbumin fractions.
Mathematical analysis of the relationship between blood flow and uptake of nutrients in the mammary glands of a lactating cow
Authors: Cant John P. , McBride Brian W.
Published: June 2009

A dynamic mathematical model of blood flow regulation in the mammary glands of a lactating cow was constructed from a principle of local vasodilator release in response to changes in intracellular adenylate charge. An equation was derived to predict uptake of the milk precursors acetate, glucose, amino acids and fatty acids, as affected by mammary blood flow rate. Metabolism of the precursors to milk components and CO2 was simulated with a set of empirically derived equations. Relative rates of ATP production and utilization regulated both the number of perfused capillaries and the conductance of arteriolar segments in the mammary glands. The model simulated local control phenomena of functional and reactive hyperaemia, and simulation of autoregulation under changing arterial pressure suggested a predominance of precapillary sphincter regulation. It was predicted that an increase in blood flow without the mammary capacity to utilize blood metabolites efficiently would be detrimental to milk synthesis. Conversely, increased blood flow through changes in mammary activity resulted in predictions of higher milk production. It was proposed that the equation for uptake,

be used in analysis of mammary arteriovenous differences.
Relationship between numbers of α2- and β2-adrenoceptors on blood cells of bulls and milkability of their daughters
Authors: Roets Eddy , Burvenich Christian , Hamann Jorn
Published: June 2009

Milk yield and milking time were measured on one occasion for several daughters (n= 6–44) from 16 bulls at morning milkings. Blood from the bulls was collected, and platelets and mononuclear leucocytes were isolated. The α2-adrenoceptors on platelet membranes were identified by binding of [3H]rauwolscine, whereas for the determination of β2-adrenoceptors on intact mononuclear leucocytes, [3H]CGP–12177 was used. It was found that mean milk flow rate was highly correlated (P< 0·001) with the α2-adrenoceptor densities on blood platelets. No correlation was found with the β2-adrenoceptors on mononuclear leucocytes. It is concluded that estimation of the α2-adrenoceptors on blood platelets from bulls could eventually be used to investigate milking characteristics of cows, and might be useful in the future as a marker in genetic studies.
Teat thickness changes may provide biological test for effective pulsation
Authors: Hamann Jörn , Mein Graeme A.
Published: June 2009

Pulsation rates of 40, 60 and 80 cycles/min were combined with pulsator ratios of 50, 60, 70 or 80% in two experiments with different liners. Machine-induced, short-term changes in teat thickness of 14 cows were compared with milk flow rate characteristics and machine strip yields to evaluate the effectiveness of pulsation in relation to liner type. Post-milking teat thickness increased progressively as the b phase of the pulsation cycle was lengthened, and as the d phase was shortened, at all pulsator rates and with either liner. Teat thickness values increased significantly (P< 0·05) when the d phase was <15%. For ratios of 50 and 60%, teat thickness decreased progressively as pulsation rate was increased. At the higher ratios, thickness values were lowest at 60 cycles/min. Pulsation settings that tended to increase teat thickness values also increased both peak milk flow rates and machine strip yields. The influence of liner type on teat thickness changes appeared to be at least as important as the influence of pulsator ratios and greater than the effect of pulsation rate. If so, then international standards for acceptable pulsation characteristics cannot be defined solely in terms of pulsator settings. A limit of ±5% for machine-induced changes in thickness of the teat apex would be an additional guideline for effectiveness of pulsation in relation to both liner type and vacuum level. This could provide a basis for a dynamic test applied to milking cows under field conditions.
Milk yield, oxytocin and β-endorphin gradually normalize during repeated milking in unfamiliar surroundings
Authors: Bruckmaier Rupert M. , Pfeilsticker Hans-Ulrich , Blum Jürg W.
Published: June 2009

For six successive milkings, six dairy cows were relocated immediately before milking to an unfamiliar operating theatre, a procedure previously shown to inhibit oxytocin release and milk ejection. Two control milkings were performed in familiar surroundings. After milk flow had ceased, two i.v. injections of 1 i.u. oxytocin were given to remove the remaining milk. Milk flow was recorded continuously and blood samples were taken every minute during milking and 10 min after milking. During the first milking in unfamiliar surroundings, no oxytocin was released. Thereby, only 13% of the total milk yield, the cisternal milk, was available and the alveolar milk fraction could only be removed after injection of oxytocin. During subsequent relocations oxytocin release steadily increased toward the control level, although the timing of oxytocin release remained delayed as compared with controls. However, the milk fraction available before oxytocin injection increased with increasing number of removals, following an asymptotic approach to control levels. The concentrations of β-endorphin, cortisol (and perhaps also of prolactin) gradually declined with the number of times the animal was moved to unfamiliar surroundings, i.e. hormone concentrations gradually adjusted to control level. During milking, concentrations of prolactin and cortisol increased, while β-endorphin concentrations decreased (except for the first relocation). We conclude that milking-related oxytocin release and therefore milk ejection adapted gradually to repeated relocations to unfamiliar surroundings. This adaptation was inversely related to β-endorphin concentrations, so it is possible that oxytocin release was suppressed by high circulating β-endorphin concentrations.
Leucine and protein metabolism in the lactating dairy cow mammary gland: responses to supplemental dietary crude protein intake
Authors: Bequette Brian J. , Metcalf John A. , Wray-Cahen Diane , Backwell F. R. Colette , Sutton John D. , Lomax Michael A. , Macrae John C. , Lobley Gerald E.
Published: June 2009

Mammary gland protein metabolism, determined by an arteriovenous difference technique, was monitored in four Holstein-Friesian dairy cows in response to supplemental dietary protein (provided as rumen-protected soyabean meal) during late lactation (weeks 24–30). Each cow was offered two isoenergetic diets composed of grass silage (170 g crude protein/kg dry matter) plus either a low (108 g/kg) or medium (151 g/kg) crude protein concentrate in a single crossover design involving two 21 d periods. On day 21, arteriovenous measurements across the mammary gland were made during a 13 h continuous i.v. infusion of [1-13C]leucine and with frequent (2 hourly) milk sampling during the final 6 h. Although total milk yield was slightly increased (+1 kg/d) by protein supplementation, milk protein yield was not significantly affected. Whole body protein flux (protein synthesis plus oxidation) was not significantly affected by supplementation. Total mammary gland protein synthesis (milk plus non-milk protein) was also not affected by supplementation but on both diets gland synthesis was always greater (by 20–59%) than milk protein output. The fractional oxidation rate of leucine by the mammary gland was significantly increased by protein supplementation (0·047 v. 0·136). Although the enrichment of leucine in secreted milk protein continued to increase, the final value (at 13 h) was 0·94 of the arterial plasma free leucine plateau value (not significantly different), suggesting almost exclusive use of plasma free leucine for milk protein synthesis. Based on current feeding schemes for dairy cattle, a fixed proportion (0·65–0·75) of the additional protein intake (+490 g/d) should have been partitioned into milk protein. Instead, leucine oxidation by the mammary gland was increased. Whether oxidation of other amino acids was also enhanced is unknown but if amino acid oxidation and the ‘additional’ non-milk protein synthesis occurring in the gland are not crucial to milk synthesis, then by reducing such activities improvements in the efficiency of converting absorbed amino acid into milk protein can be achieved.
Detection and localization of peptidases inLactococcus lactiswith monoclonal antibodies
Authors: Laan Harry , Haverkort Roel E. , Leij Loe De , Konings Wil N.
Published: June 2009

Monoclonal antibodies against peptidases ofLactococcus lactiswere isolated and characterized: PEPN1–4 against a lysyl aminopeptidase PepN, PEPT1–5 against a tripeptidase PepT and PEPD1–3 against a dipeptidase PepD. These monoclonal antibodies reacted specifically with their respective antigens in crude cell extracts ofLc. lactissubspp.cremorisandlactis. A number of monoclonal antibodies cross reacted with proteins of other (lactic acid) bacteria. PEPT1, 2, 4 and 5 cross reacted weakly with a 35 kDa protein inLactobacillus delbrueckii, while PEPT1 and PEPT2 reacted with proteins in the cell-free extract ofStreptococcus thermophilusandClostridium fervidus. Of the four isolated monoclonal antibodies against PepN, only PEPN3 cross reacted weakly with a 90 kDa protein inEscherichia colicell-free extract, and the other three antibody species against PepN cross reacted with 80 kDa proteins ofLb. casei, Lb. delbrueckii, andStr. bovis, but not ofEsch. coli. Of the three monoclonal antibodies against PepD, only PEPD1 and PEPD2 cross reacted with 40 kDa proteins ofLb. casei, Lb. delbrueckiiandStr. bovis. All PEPN, PEPD and PEPT antibodies reacted with components in cell-free extracts of eleven differentLc. lactisstrains, indicating that the peptidases of these strains were very similar to those ofLc. lactissubsp.cremorisWG2. However,Lc. lactissubsp.hordniaeappeared to differ from the otherLc. lactissubspecies since only PEPT1, 2 and 5 reacted with a protein in the cell-free extract. Immunogold labelling ofLc. lactisWG2 with the isolated monoclonal antibodies revealed that PepN, PepD and PepT were located intracellularly. The intracellular location of these peptidases is discussed in relation to the supply of essential amino acids and peptides.
Phosphopeptides interacting with colloidal calcium phosphate isolated by tryptic hydrolysis of bovine casein micelles
Authors: Gagnaire Valerie , Pierre Alice , Molle Daniel , Leonil Joelle
Published: June 2009

After extended tryptic hydrolysis of large bovine casein micelles, a mineral-rich peptide fraction was recovered by ultracentrifugation. Its mineral part contained 72% of the colloidal Ca and 49% of the colloidal Pioriginally present in the native micelle. Colloidal nitrogenous components were also present, amounting to 27% of the original N content. They contained most of the phosphopeptides and 82% of the micellar phosphoseryl residues. These tryptic peptides were characterized by reversed-phase HPLC on-line electrospray ion source–mass spectrometry analysis. Among the peptides produced 14 phosphopeptides were identified: αs2-CN(l–24), αs2-CN(1–21), αs1-CN(43–79), αs1-CN(35–79)7P, αs1-CN(35–79)8P, αs1-CN(37–79), αs1-CN(104–119), αs1-CN(104–124), β-CN(1–25), β-CN(1–28), β-CN(1–29), β-CN(30–97), β-CN(33–97) and β-CN(29–97). The proportion of the phosphopeptides interacting with colloidal calcium phosphate was correlated with their relative content of phosphoserine residues, since phosphopeptides containing more than four phos-phoserine residues were consistently present within this fraction. It also appeared that other types of peptides, some of them hydrophobic in nature, were also partly or completely present within the colloidal fraction, including αs1-CN(91–100), αs1-CN(152–193), αs1-CN(23–34), αs1-CN(125–193), αs1-CN(125–199), β-CN(177–209), β-CN( 184–209), β-CN(114–169) and β-CN(108–169). Their possible involvement in the micellar backbone is discussed.
Hyperkeratosis of the teat duct orifice in the dairy cow
Authors: Shearn Martin F. H. , Hillerton J. Eric
Published: June 2009

Teat orifice hyperkeratosis, a commonly observed condition in dairy cows, has been considered a consequence of machine milking and the degree of hyperkeratosis may be increased by a poor milking system. A fully illustrated technique is described which uses a scoring system from 0 for a perfect orifice to 5 for an orifice significantly enlarged with extensively protruding fronds of teat duct keratin. A range of scores found in 25 commercial dairy herds is presented. The scores have been averaged for each cow and the markedly skewed distribution corrected by a square root transformation. This scoring procedure allows comparative measures of hyperkeratosis within and between herds. An 8-fold difference (0·17–1·31) in herd average score was found. Within all herds the score increased with lactational age of the animals and peaked, for any lactation, some 3–4 months post partum, declining as the animals dried off. There was no significant relationship between mean somatic cell count and degree of hyperkeratosis at the herd level. This implies that such chronic pathological changes are unlikely to be related to the level of intramammary infection. It appeared that some hyperkeratosis is an obvious and probably natural response to milking and occurs in a significant proportion of animals in all herds although often only to a slight degree. Much more hyperkeratosis may be a measure of the performance and management of the herd. The genetic influence is unknown. Higher yielding cows will score higher as they milk for longer, but generally high scores may reflect consistent and possibly considerable overmilking. Hyperkeratosis may be an indicator of the quality of management and show the level of attention being paid to the welfare of the herd.
Distribution of the anthelmintic drug albendazole and its major metabolites in ovine milk and milk products after a single oral dose
Authors: Liguoro Marco De , Longo Francesca , Brambilla Gianfranco , Cinquina Annalucia , Bocca Adriana , Lucisano Antonia
Published: June 2009

The distribution of albendazole (ABZ) and its main metabolites albendazole sulphoxide (ABZSO), albendazole sulphone (ABZS02) and albendazole 2-aminosulphone (NH2ABZSO2) were investigated in bulk milk and milk products after administration of a single oral dose of the drug (12·5 mg/kg) to 80 Laticauda sheep. An analytical method was developed for this investigation from an existing procedure used for the determination of these compounds in plasma and digesta samples. No traces of the parent compound or NH2ABZSO2were found in milk or milk products, with the exception of the milk collected 36 h after treatment in which 89 μg NH2ABZSO2/kg was detected. Results indicated that ABZ was rapidly oxidized to ABZSO and then to ABZSO2. These metabolites were found at high levels (1–4 mg/kg) in milk collected within 24 h after treatment. Products derived from such milk also contained high concentrations of the two oxidized metabolites, including up to 5 mg ABZSO/kg in Pecorino cheese. Only small quantities of these two metabolites were found in milk collected during the second day after treatment (range 50–500 μg/kg). They were no longer detectable in milk, collected during the third day after dosing, nor were they found in products made from such milk. These findings confirm that the two polar metabolites ABZSO and ABZSO2were efficiently excreted from the body. Considering that the established maximum residue limit for ovine milk is 100 μg/kg for ABZ plus its metabolites, our results confirmed the appropriateness of the currently prescribed withdrawal time (3 d) after the use of ABZ in lactating sheep. However, considerable levels of ABZSO were detected in milk collected within 24 h after treatment as well as in products and by-products derived from such milk. Owing to the known toxicity of the ABZSO, we stress the need for careful control to ensure adherence to the prescribed withdrawal time.