Group N streptococci were found to cleave threonine to form acetaldehyde and glycine. Threonine aldolase, the enzyme catalysing this reaction, was found in all strains except Streptococcus cremoris Z8, an organism which had been shown previously to have a nutritional requirement for glycine. The enzyme was strongly inhibited by glycine and cysteine. The inhibition showed characteristics of allosteric inhibition and was pH-dependent. Inhibition by glycine, but not by cysteine, was highly specific. Analogues and derivatives of cysteine which contained a thiol group and a free amino group inhibited the activity of threonine aldolase. The presence of a carboxyl group was not necessary for inhibition. The cleavage of threonine by wholecell suspensions was stimulated by either an energy source to aid transport, or by rendering the cells permeable to substrate with oleate. Threonine did not appear to be degraded by enzymes other than threonine aldolase, as threonine dehydratase activity was low and NAD- and NADP-dependent threonine dehydrogenases were absent.

Twelve spring-calving and twelve winter-calving cows were managed for extended lactation cycles of 18-months duration, with the former group then completing a second extended lactation. Half of the cows were fed according to standard management practice for the herd; the other half received supplementary concentrate from week 9 of lactation onwards. Commencing at the same time, half of the udder of each cow was subjected to increased milking frequency (thrice daily rather than twice daily). Lactation persistency (and hence total milk yield) was significantly increased by frequent milking. Winter calving cows and supplemented cows also exhibited better persistency, but this was only evident up until the point of re-breeding, at around lactation week 33. Milk composition was measured in the spring-calving cows in both their first and second extended lactations. Composition altered during the course of the lactation, protein and fat percentages increasing and lactose percentage decreasing, irrespective of treatment. The quality of the milk for processing into cheese, fermented products, heat-treated products and cream liqueurs was assessed by calculation of casein number (casein protein as a proportion of total protein). Processing quality declined across the course of lactation in those groups that showed poor persistency but not in those that maintained a persistent lactation. Milk hygienic quality (somatic cell counts) showed parallel changes. Body condition score increased during the course of lactation but was not affected by supplementation; none of the cows became excessively fat. All cows remained healthy throughout the extended lactations and the majority (33/36) re-bred successfully. By demonstrating that lactation persistency is plastic and can be improved by simple management interventions, the results lend support to the economic arguments in favour of extended lactation cycles. The likely welfare benefits of extended lactation are also discussed.

Intracellular peptidases of Lactobacillus helveticus may play a major role in the proteolysis of Swiss cheeses, provided that they are released through bacterial lysis. Experimental Swiss cheeses were manufactured on a small scale from thermized and microfiltered milk using as starters (in addition to Streptococcus thermophilus and Propionibacterium freudenreichii) one of two Lb. helveticus strains, ITGLH1 and ITGLH77, which undergo lysis to different extents in vitro. All the cheeses were biochemically identical after pressing. The viability of Lb. helveticus ITGLH1 and ITGLH77 decreased to a similar extent (96–98%) while in the cold room, but the concomitant release of intracellular lactate dehydrogenase in cheeses made with strain ITGLH1 was 5–7-fold that in cheeses made with ITGLH77. Protein profiles and immunoblot detection of the dipeptidase PepD confirmed a greater degree of lysis of the ITGLH1 strain. Free active peptidases were detected in aqueous extracts of cheese for both strains, and proteolysis occurred principally in the warm room. Reversed-phase HPLC revealed a more extensive peptide hydrolysis for ITGLH1, which was confirmed by the greater release of free NH2 groups (+33%) and free amino acids (+75%) compared with ITGLH77. As the intracellular peptidase activities of ITGLH1 and ITGLH77 have previously been shown to be similar, our results indicated that the extent of lysis of Lb. helveticus could have a direct impact on the degree of proteolysis in Swiss cheeses.

Samples of liquid and solid fractions obtained by a commercial process from anhydrous milk fat of softening point (S.P.) 33·5–34°C had S.P. values of 22–23°C and 36–38°C respectively.

Determinations of fatty acid composition by GLC showed that unsaturated and short chain fatty acids were present in increased concentration in the liquid fraction (average 37·8 and 12·4% as compared with 35·1 and 10·8% in the original milk fat) and long chain saturated acids in the solid fraction (average 57·8 as compared with 53·8%). There was some concentration of carotene and vitamin A, and to a lesser extent of cholesterol, in the liquid fraction.

Crystallization and melting curves determined by a differential scanning calorimeter (DSC) showed that while the liquid fraction was completely melted at 25°C, the solid fraction contained an increased proportion of fat melting between 30 and 40°C. In addition to these high melting triglycerides the solid fraction also contained some 65% of fat in the liquid phase at the original crystallization temperature of 25°C.

In a series of short-term experiments cows were subjected to exaggerated bacterial challenge and accentuated milking machine conditions known to predispose to new mammary infection. The incidence of new intramammary infection was significantly greater in quarters with peak flow rates > 1·6 kg/min whether they were exposed to impacts (P < 0·05) or milking without pulsation (P <0·001). The infection rates were much lower (P < 0·001) in quarters milked with ‘pulsation and shields’ to protect against these two machine factors. Despite this, quarters with peak flow > 1·6 kg/min still showed a 12-fold increase in mastitis incidence compared with quarters with peak flow < 0·8 kg/min. Rates of milk flow have increased dramatically in the last 40 years through selection and breeding: whole udder peak flow rates in heifers have doubled from 1·9 to 3·8 kg/min. Increased emphasis, therefore, should be placed on hygiene, husbandry and milking techniques to minimize bacterial numbers at teat ends to control mastitis as the drive for higher flow rate and yield make cows increasingly more susceptible to infection. These results suggest that the benefits of reduced infection rate from mastitis control are significantly underestimated since animals are now considerably more susceptible than 40 years ago.

The equipment tested (Fossomatic) is a fully automatic electronic instrument for counting the somatic cells in milk samples. The results are printed on a paper strip and displayed on a digital counter. The capacity is 180 samples/h.

Counting results obtained with the instrument have been compared with direct microscopic cell counts and particle counts by Coulter counter. The correlation coefficient with microscopic cell counts was 0·922–0·988, and with counts by the Coulter counter it was 0·916–0·992.

Reproducibility showed a coefficient of variation of 2–5 %. The carry-over between the samples was 2·0 %. Preserving milk samples with 0·05 % potassium dichromate did not influence cell counts, whereas freezing the samples reduced the counts 30 %.

Milk contains the alkaline proteinase plasmin and lysosomal proteinases; the significance of the latter is ill-defined. The objective of this study was to investigate composition and activities of several different proteolytic enzymes in milk samples of varying somatic cell count (SCC). Increasing milk SCC was correlated with increased plasmin, cathepsin D and cysteine protease activities, with concomitant increases in proteolysis in milk. Addition of plasmin inhibitors confirmed the heterogeneity of proteinase activities in milk, as urea-PAGE analysis of milk samples showed casein hydrolysis in milk after 7 d storage even in samples with inhibitors added; extent and heterogeneity of proteolysis was correlated with milk SCC. Rennet coagulation properties were not significantly correlated with SCC, or activities of measured enzymes. Milk of increasing SCC also exhibited decreased physical stability during incubation of milk at 37 °C. Pasteurized milk was more stable than raw milk, suggesting that the enzyme(s) or mechanisms leading to such instability are impaired by pasteurization. Overall, milk has a very heterogeneous proteolytic enzyme population, with a higher significance of non-plasmin enzymes, such as cathepsin D and cysteine proteinases, than perhaps previously recognised.

The aim of this study was to compare the effects of supplementing dairy cows with 1000 IU/day of all-rac-α-tocopheryl acetate (SynAc), RRR-α-tocopheryl acetate (NatAc), or RRR-α-tocopherol (NatAlc), from approximately 3 weeks before estimated calving until 2 weeks after calving, on the concentration of α-tocopherol and its stereoisomers (RRR-, RSS-, RRS-, RSR- and the four 2S-forms of α-tocopherol) in blood and milk. An unsupplemented group was included as control. Blood samples were collected at 3, 2 and 1 weeks before estimated calving, at calving, and 3, 7 and 14 days after calving, while milk samples were taken twice within 24 h after calving and at 7 and 14 days in milk. Overall, time and treatment had significant effects on plasma α-tocopherol with higher concentrations in NatAc than in the other groups. In addition, SynAc had higher concentrations than Control, and NatAlc tended to be higher than Control. The lowest plasma concentrations were observed at calving and 3 days after calving. Independent of treatment, the concentration was higher in colostrum than in milk day 7 and 14 after calving. Analyses of the stereoisomer distribution in plasma and milk showed that, irrespective of dietary treatment, RRR-α-tocopherol was the most predominant form, constituting more than 86%, whereas the remaining part of α-tocopherol was made up by the three synthetic 2R isomers, while the 2S isomers only contributed less than 1% of the total α-tocopherol. In control cows and cows supplemented with natural vitamin E, the proportion of RRR-α-tocopherol in plasma and milk constituted more than 98% of the total α-tocopherol. In conclusion, the results indicate that daily oral supplementation of dairy cows with RRR-α-tocopheryl acetate gives the highest blood concentrations of α-tocopherol in the periparturient period. Analyses of the distribution of the individual stereoisomers of α-tocopherol further indicate that the bioavailability of RRR-α-tocopherol relative to synthetic stereoisomers in cattle is considerably higher than officially accepted until now.

Bifidogenic effect and stimulation of short chain fatty acid (SCFA) production by fractions of oligosaccharides with a DP⩾3 and Gal β(1-6) linkages synthesised from lactose or lactulose by Pectinex Ultra SP-L and Lactozym 3000 L HP G were evaluated in human faecal slurries. Results were compared with those obtained for the commercial oligosaccharide mixture Vivinal®-GOS. Quantitative real-time PCR showed that all galacto-oligosaccharide (GOS) fractions stimulated slightly higher bifidobacteria growth than lactose, lactulose and Vivinal®-GOS. GOS fractions promoted the production of total SCFA and acetic acid in a similar way to Vivinal®-GOS and more than glucose, lactose and lactulose. In conclusion, oligosaccharides derived from lactose and lactulose may represent a suitable alternative to lactulose for the design of new functional food ingredients.

The free glucose concentration in the aqueous phase of samples of goat, sheep, cow, rat and rabbit milk was about 0·1–0·3 mn, while that in human milk was about 2 mM. During starvation the glucose concentration of goat milk fell considerably (by about 80 % in 2 d) in parallel with the decreased rate of lactose production. With rats fed ad lib., glucose concentration in the milk was greater at 12.00 h than at 18.00 h, when lactose synthesis has been shown to decrease. 3-O-Methyl-D-glucose injected into the goat mammary gland via the teat canal specifically entered the blood. These findings support the idea that glucose equilibrates across the apical membrane of mammary secretory cells, so that milk glucose concentrations reflect intracellular glucose concentrations.

In milk samples from 549 cows of the breeds Danish Jersey, Red Danish Dairy Cattle (RDM), and Black and White Danish Dairy Cattle (SDM) the genetic polymorphisms of the αs1, β and K-cascin and β-lactoglobulin (β-Lg) loci were determined by isoelectric focusing in agarose gels. The results of the screening were comparcd with results obtained by Larsen & Thymann (1966). In addition, the genetic linkage of the three casein loci was studied, and the association between milk protein genotypes and yields in first and second lactations of milk, fat and protein were investigated.

The distribution of genotypes of all four milk protein Systems was different from breed to breed.

For Jersey cows, significant differences in the gene frequencies from the results of the 1966 investigation were found for αs1 and K-casein and β-Lg. For SDM cows a change in the K-casein frequency had occurred whereas for RDM cows no changes were found.

Linkage between some of the casein loci was found within ail three breeds. For the RDM breed the possible linkage between αs1-casein and the other caseins could not be tested bccause nearly ail thc cows were homozygous for the αs1-cascin-B genotypes.

β-Casein genotypes were associated with yield parameters in ail breeds. The A2A2 genotype of this protein gave higher yields of milk, fat, and protein in the second lactation than thc A1A1 genotype.

Naturally occurring tumours in domestic animals have been recognized as an interesting opportunity for comparative oncology (MacEwen, 1990; Vail & MacEwen, 2000). Cancer is the second most frequent cause of death in humans and the first one in dogs and cats (Jemal et al. 2003). The age-adjusted overall cancer incidence per 100000 individuals per year is comparable in humans and domestic animals, being approximately 300 in humans, 381 in dogs and 264 in cats (Vail & MacEwen, 2000). When analysing the incidence by site, breast cancer is the most frequent (32%) in women, the first of all neoplasia (52%) occurring in bitches and the third (17%) in queens after lymphohaemopoietic and skin tumours (Hayes et al. 1981; Hayes & Mooney, 1985; MacEwen, 1990; MacEwen & Withrow, 1996; Jemal et al. 2003). Several other aspects contribute to the value of domestic animals as models for human cancers (MacEwen, 1990; Vail & MacEwen, 2000). Tumours occur spontaneously in companion animals that share a similar environment with humans and therefore might be exposed to similar risk factors. The high incidence of some tumour types offers large population samples. The shorter overall lifespan of domestic animals associated with a more rapid progression of cancer allows adequate comparison of response time with humans. Biological, anatomical, histopathological, genetic, and molecular similarities between some animal and human tumours are also well established (Hansen & Khanna, 2004). Finally, testing novel therapies is more ethically acceptable when treating spontaneous diseases in companion animals rather than experimentally induced pathologies in animal models. At the same time, there is an increasing interest of owners towards the use of the most advanced therapeutic tools for companion animals despite the higher economic costs associated with these therapies.

Prevalence of mastitis pathogens in milk samples from dairy cows and heifers was studied over a period of 1 year (Aug 2005–Aug 2006) in ten dairy herds in Germany. Milk samples (n=8240) were collected from heifers without clinical mastitis at parturition (n=6915), from primiparous cows with clinical mastitis (n=751) and from older cows with clinical mastitis (n=574). Coagulase negative staphylococci (CNS) were the predominant group of bacteria isolated (46·8% of samples) from clinically healthy quarters of primiparous cows around parturition, followed by streptococci (12·6%), coliforms (4·7%) and Staphylococcus aureus (4·0%). Thirty-three percent of samples were negative on culture (range on farm level, 12·0–46·4%). In cases of clinical mastitis in primiparous and older cows, streptococci were the predominant finding (32·1 and 39·2%) followed by CNS (27·4 and 16·4%), coliforms (10·3 and 13·1%) and Staph. aureus (10·0 and 11·7%). Negative results were obtained from 21·3% (range, 0·0–30·6%) and 19·5% (range, 0·0–32·6%) of these samples. Results indicated substantial differences in the prevalence of pathogens among herds. There was a positive within-herd correlation between the monthly prevalences for Streptococcus dysgalactiae between the three groups of samples. This correlation was also found between clinical samples of primiparous and older cows for Staph. aureus. These correlations were not found for the other pathogens. Besides herd, prevalence of pathogens was influenced by parity, type of sample and season.

The intestinal ecosystem contains a normal microbiota, non-immune cells and immune cells associated with the intestinal mucosa. The mechanisms involved in the modulation of the gut immune system by probiotics are not yet completely understood. The present work studies the effect of a fermented milk containing probiotic bacterium Lactobacillus (Lb.) casei DN114001 on different parameters of the gut immune system involved with the nonspecific, innate and adaptive response. BALB/c mice received the probiotic bacterium Lb. casei DN114001 or the probiotic fermented milk (PFM). The interaction of the probiotic bacteria with the intestine was studied by electron and fluorescence microscopy. The immunological parameters were studied in the intestinal tissue and in the supernatant of intestinal cells (IC). Results showed that the probiotic bacterium interact with the IC. The whole bacterium or its fragments make contact with the gut associated immune cells. The PFM stimulated the IC with IL-6 release, as well as cells related to the nonspecific barrier and with the immune cells associated with the gut. This last activity was observed through the increase in the population of different immune cells: T lymphocytes and IgA+ B lymphocytes, and by the expression of cell markers related to both innate and adaptive response (macrophages). PFM was also able to activate the enzyme calcineurine responsible for the activation of the transcriptional factor NFAT. PFM induced mucosal immune stimulation reinforcing the non-specific barrier and modulating the innate immune response in the gut, maintaining the intestinal homeostasis.

A total of 480 samples of milk from 10 organically and 10 conventionally producing dairy farms in Denmark and covering 8 sampling periods over 1 year (triplicate samplings) were analysed for 45 trace elements and 6 major elements by high-resolution inductively coupled plasma mass spectrometry and inductively coupled plasma atomic emission spectrometry. Sampling, sample preparation, and analysis of the samples were performed under carefully controlled contamination-free conditions. The dairy cattle breeds were Danish-Holstein or Jersey. Sources of variance were quantified, and differences between production systems and breeds were tested. The major source of variation for most elements was week of sampling. Concentrations of Al, Cu, Fe, Mo, Rb, Se, and Zn were within published ranges. Concentrations of As, Cd, Cr, Mn and Pb were lower, and concentrations of Co and Sr were higher than published ranges. Compared with Holsteins, Jerseys produced milk with higher concentrations of Ba, Ca, Cu, Fe, Mg, Mn, Mo, P, Rh, and Zn and with a lower concentration of Bi. The organically produced milk, compared with conventionally produced milk, contained a significantly higher concentration of Mo (48 v. 37 ng/g) and a lower concentration of Ba (43 v. 62 ng/g), Eu (4 v. 7 ng/g), Mn (16 v. 20 ng/g) and Zn (4400 v. 5150 ng/g respectively). The investigation yielded typical concentrations for the following trace elements in milk, for which no or very few data are available: Ba, Bi, Ce, Cs, Eu, Ga, Gd, In, La, Nb, Nd, Pd, Pr, Rh, Sb, Sm, Tb, Te, Th, Ti, Tl, U, V, Y, and Zr.

Whole milks concentrated 1·5–4-fold and acidified and citrated milks concentrated 2·8-fold by ultrafiltration at 50 °C were analysed for chemical changes relevant to further processing, storage or nutrition. Fat and protein were entirely retained in the concentrate. The retention of water-soluble vitamins, Ca, Mg, phosphate and trace minerals depended on the proportion bound to the protein. Ascorbic acid was rapidly destroyed during concentration. Because of the differential retention of nitrogenous components, protein comprised a progressively higher proportion of the total N as the milk became more concentrated. No denaturation of whey protein or disruption of casein micelles was detected during concentration of whole milk, but some solubilization of the casein occurred after citration. Reduction of fat globule size occurred early in the concentration process, damage to the fat globule membrane was indicated and the milk became more susceptible to lipolysis. Apart from a tendency for preacidified or precitrated concentrates to gel, no change in the susceptibility of the milks to heat damage was detected.

While the effect of long-chain fatty acids on adipose tissue (AT) lipogenic activities has been described in non-lactating ruminants (Vernon, 1977), little is known about their effects on the mammary gland and the AT in lactating animals. However, in cows in mid lactation, duodenal rapeseed oil infusion decreased the rate of fatty acid (FA) synthesis in AT and increased milk yield of long-chain FA (18[ratio ]1, 18[ratio ]2 and 18[ratio ]3) and decreased medium-chain FA (14[ratio ]0 and 16[ratio ]0), suggesting a depressive effect of fat feeding on mammary lipid synthesis de novo (Chilliard et al. 1991). On the other hand, in goat species, the addition of vegetable lipids to the diet led to an increase in the milk fat content and yield (Chilliard et al. 2003) suggesting that the possible negative effect of long-chain FA on FA synthesis in the lactating mammary gland could be more than compensated by increasing the supply of FA brought to the mammary gland for milk synthesis. Elsewhere, AT from various anatomical sites are characterized by different FA composition in goat (Bas et al. 1987) together with different patterns of lipogenic gene expression in sheep (Barber et al. 2000). These results suggest that each AT site is characterized by a specific metabolism. However, in lactating ruminants, few data are available on the extent of expression and regulation of genes coding for lipogenic enzymes in AT. Therefore, the current study was performed in three lipogenic tissues of lactating goats, namely the mammary gland, an internal AT site (perirenal AT) and an external AT site (subcutaneous AT).

The maximum and minimum heat stability exhibited by most milks over a relatively narrow range of pH values is shown also by synthetic colloidal calcium caseinate-calcium phosphate systems and even by simple caseinate systems, provided all possess adequate contents of β-lactoglobulin, soluble calcium and phosphate. The phenomenon is not, however, dependent on the presence of the characteristic micellar structure of the casein of milk. The minimum stability observed, usually around pH 6·9, is the most characteristic feature of the phenomenon and arises from heat induced deposition of calcium phosphate on a caseinate/β-lactoglobulin complex. This reaction, which tends to occur to a marked degree at relatively high pH values and calcium ion concentrations, sensitizes the complex to precipitation by calcium ions. The precise pH values at which the maximum and minimum stabilities occur can vary depending on the salt composition of the serum, since the latter can influence the solubility of calcium phosphate.

Bovine mastitis is the most important infectious disease on dairy farms. Conventional antibiotic therapy is often unsatisfactory and alternative treatments are continually under investigation. Lactobacillus (Lb.) perolens CRL 1724 and Lactobacillus plantarum CRL 1716 were previously isolated from milk of dairy cows and selected according to their potential probiotic properties. In the present work the in-vitro capacity of Lactobacillus strains to adhere to bovine teat canal epithelial cells (BTCEC) and to inhibit and co-aggregate 14 mastitis-causing pathogens (MCPs) was investigated. The effect of Lb. perolens CRL 1724 after intramammary inoculation in lactating cows was evaluated through determination of clinical signs of mastitis, milk appearance, somatic cell counts and Lb. perolens CRL 1724 recovery from milk. Lb. perolens CRL 1724 was able to inhibit 12 of 14 MCPs (85·7%) in vitro, especially those considered to be major pathogens. In addition, Lb. perolens CRL 1724 co-aggregated with all of them. Lb. plantarum CRL 1716 was able to inhibit 7 of 14 MCPs (50%) in vitro and showed co-aggregation ability similar to Lb. perolens CRL 1724. Lb. perolens CRL 1724 showed a higher efficacy of adhesion to BTCEC (values of percentage of adhesion and adhesion index of 75% and 14·4, respectively) than Lb. plantarum CRL 1716 (37% and 7·4, respectively). Lb. perolens CRL 1724 was recovered from all mammary quarters and no clinical signs or teat damage were observed after the inoculation of 106 cfu/ml. The udders presented a normal aspect and there were no changes in the appearance of the milk. The results obtained will serve as the basis for further trials to evaluate the potential of Lb. perolens CRL 1724 to be included in a non-antibiotic formulation for the prevention of bovine mastitis.