We developed a simple, non-invasive method to measure the intramammary pressure (IMP) response in sows during natural milk ejection and after i.v. injections of oxytocin. The teats of sows were cannulated and the amplitude and duration of IMP changes were measured via a pressure transducer linked to a computer. During natural milk ejections, the amplitude of the response was 5·5±1·8 kPa (mean±SD) and the duration of response at half the peak amplitude was 6·8±1·6 s. In response to oxytocin a threshold response occurred at 60 mIU oxytocin with no further change up to 200 mIU. The amplitude of the response to an i.v. dose of 60–200 mIU oxytocin (5·5±2·2 kPa) and duration (6·4±1·7 s) were similar to that during a natural milk ejection. The responses of two galactophorous ducts of a teat measured simultaneously were similar in peak amplitude and duration. When the responses of four glands of a sow were measured simultaneously, there was a lag time in the onset of the IMP peak from the anterior to the posterior glands, and an increase in the peak amplitude from anterior to posterior. Both during natural milk ejections and in response to oxytocin injections, there were significant differences in peak amplitude between sows. We also aimed to determine whether the pressure or duration of milk flow during natural milk ejections was limiting piglet growth rates. IMP of nine sows and the growth rates of their piglets were measured. Both the peak amplitude of IMP response and the growth rate of piglets increased over the course of lactation, but the relationship was not causal. We found that the growth rate of piglets did not depend on IMP amplitude or duration, but we suggest that it may depend on the frequency of suckling and/or the ability of piglets to withdraw milk while it is available.

Samples of foremilk were collected from 261 clinically normal glands of 150 ewes, and tested using the California mastitis test (CMT). Further samples were collected from 195 of these glands for determination of automated somatic cell counts (SCC), and from 60 of these glands for bacteriological assessment. The sensitivity and specificity of CMT for detecting samples with SCC above different threshold levels and for CMT and SCC in determining bacteriological status were evaluated using two-graph receiver operating characteristics (TG-ROC). Milk samples were obtained subsequently from ten CMT positive, and five CMT negative first- and second-lactation ewes. Samples were cultured using a variety of media, incubation temperatures and atmospheric conditions, immediately after collection, and 1 week after storage at 4°C and −21°C. Results suggested that CMT is best used as a diagnostic test for ovine subclinical mastitis (SCM) with a cut-off of 3 (distinct gel formation), and that automated SCC thresholds of >1200×103 cells/ml are appropriate, especially where low prevalences are expected (e.g. <5%). Additionally, this study showed that routine bacteriological methods were appropriate for isolation of most species of pathogen responsible for ovine SCM, but storage of samples prior to culture, either at 4°C or −21°C, was detrimental to the isolation of several of these organisms.

Effective diagnostic tools for screening herds for mastitis pathogens are important in development and monitoring of mastitis control programmes. A multiplex polymerase chain reaction (PCR) assay for simultaneous detection of Staphylococcus aureus, Streptococcus agalactiae, Streptococcus dysgalactiae and Streptococcus uberis was used in preliminary studies to assess its applicability as an alternative method for monitoring mastitis caused by these organisms at the herd level. PCR was used to detect the presence of these organisms in bulk milk samples. Correlations with bulk milk somatic cell counts (BMCC), total bacteria counts and thermoduric bacteria counts were evaluated. A total of 176 bulk milk samples were collected from 42 herds on five consecutive occasions at approx. 10-d intervals. Str. uberis was the most common organism in these bulk milk samples. There was no relationship between presence of either Staph. aureus, Str. dysgalactiae or Str. uberis and BMCC, total bacteria counts or thermoduric bacteria counts. However, presence of Str. agalactiae was associated with high BMCC and total bacteria counts. The results of this study show that regular analysis of bulk milk using this multiplex PCR assay may be a useful tool for monitoring herd status with respect to Str. agalactiae, but is of less value for monitoring occurrence of Staph. aureus, Str. dysgalactiae and Str. uberis. Further investigations are needed to clarify the relationship between positive PCR results and the prevalence of infected cows in the herd.

The responses of five lactating East Friesian milk ewes to experimental mammary infection with Staphylococcus epidermidis and of five control ewes were examined over a period of 10 weeks. Infection caused an influx of neutrophils into milk, the numbers of which started to rise 4 h post infection and peaked 24 h after infection. The initial response was accompanied by mild fever and mild leucopaenia in blood (8 h after infection). No other signs of systemic infection were observed. Milk appeared normal at all times, although the milk yield of infected ewes tended to decline. Staphylococci were absent in milk from four ewes at 2 d and at 3 d after infection, but re-emerged intermittently in four of five ewes at subsequent samplings. Cytokines in milk were measured by ELISA. IL-8 was elevated in infected glands at 2 h and peaked at 8 h. In the four ewes intermittently shedding bacteria, IL-8 remained elevated until the final sampling at 10 weeks. IL-1β was transiently elevated at 1 d and 2 d and showed a pronounced peak in one sheep. Milk samples from this ewe were bacteriologically negative, somatic cell count (SCC) was within the normal range and the concentrations of IL-1β, as well as IL-8, were similiar to the control group (n=5) from 1 week after infection until the final sampling. Histological examination revealed leucocytic infiltrates in the four glands remaining infected at the end of the experiment, and a high level of CD5+ lymphocytes in three ewes. The results suggest that the relationship between the initial neutrophil influx and the proinflammatory cytokines may be responsible for determining the course of infection. Subclinical mastitis due to coagulase-negative staphylococci leads to minor changes in milk yield and milk constituents.

Probiotic microorganisms, especially lactic acid bacteria, are effective in the treatment of infectious diarrhoeal diseases and experimental colitis. Although the mechanisms by which these organisms exert their anti-inflammatory effects are largely unknown, immunomodulating effects are suggested. The objective of this study was to examine the effect of a 5-week oral administration of Lactobacillus rhamnosus subspecies GG (Lb. GG) on the cellular immune response to intestinal microorganisms in ten healthy volunteers. Peripheral blood cells (PB) were stimulated with either ‘self’ or ‘non-self’ preparations of faecal samples and isolated Bacteroides fragilis group-organisms (Bfg) or Escherichia coli (Esch. coli), and pro- and anti-inflammatory cytokines (IL-10, IL-4, IL-6, IFN-γ, TNF-α) were measured in the culture supernatant. CD4+ T-lymphocyte activation was determined by measurement of intracellular ATP following lysis of the cells. The activational response of CD4+ T-lymphocytes towards isolated and heat-inactivated intestinal organisms was increased after the probiotic treatment. Additionally, TNF-α, IL-6 and in part IFN-γ cytokine secretion by PB cells following stimulation with whole stool preparations and single members of the flora was significantly decreased, whereas the IL-10 and in part IL-4 cytokine secretion was increased at the end of the study. In contrast, the activational response of CD4+ T-lymphocytes following stimulation with whole ‘non-self’ intestinal flora was higher than by ‘self’ intestinal flora, but both responses showed a trend towards a reduction at the end of the study. This study documents a direct effect by Lb. GG on the cellular immune system of healthy volunteers and offers a promising tool to investigate systemic immunomodulation due to oral administration of probiotic microorganisms.

It has been hypothesized that certain consumers tolerate untreated cow's milk, but react to processed (i.e. homogenized and pasteurized) cow's milk although they do not suffer from IgE-mediated cow's milk allergy or lactose intolerance. The aim of the study was to compare the tolerance of unhomogenized and homogenized cow's milk in lactose tolerant adults who had repeatedly experienced better tolerance of unhomogenized than homogenized milk. Forty-four subjects were challenged with homogenized and unhomogenized cow's milk for five days in a randomized, double-blind, cross-over study. No differences in the symptoms during the challenges were found. Roughly half the subjects tolerated the homogenized milk better and the other half tolerated the unhomogenized milk better. The results of this study show no difference in the tolerance of homogenized and unhomogenized milk in adults with self-reported symptoms suggestive of hypersensitivity to homogenized milk.

The effects of dietary inclusion of freeze-dried goat and cow milk on the utilization of zinc and selenium, and on the metabolic fate of zinc, were studied in transected (control) and resected rats (resection of 50% of the distal small intestine). Intestinal resection reduced the apparent digestibility coefficient and zinc retention in the cow milk diet, whereas these biological indices were similar in transected (control) and resected rats with standard (without milk) and goat milk diets. The apparent digestibility coefficient and retention of selenium were not affected by intestinal resection in the animals fed with the three diets studied. In transected (control) and resected rats, the apparent digestibility coefficient and retention of zinc and selenium were higher for the goat milk diet than for the other two diets. Zinc deposits in the organs, expressed as μg/g dry weight were in order of decreasing concentrations: femur, testes, sternum, liver, kidney, heart, spleen, longissimus dorsi muscle and brain. Deposits were greatest with the goat milk diet, followed by the standard diet and were lowest for the rats given the cow milk diet, both for transected (control) and resected animals.

We conclude that consumption of the goat milk diet produces a greater bioavailability of zinc and selenium and a greater deposit of zinc in key organs, for both the transected (control) and the resected rats, with respect to the standard diet and the cow milk diet.

Acid-precipitated rabbit ‘whole casein’ was digested by trypsin, chymotrypsin, pepsin, and clostripain to screen for possible peptides with antibacterial properties. The peptide fragments were separated by reversed-phase chromatography. The collected fractions were pooled and their antibacterial properties tested against Escherichia coli, Bacillus subtilis and Staphylococcus lentus. Three antibacterial peptide fragments derived from tryptic digestion of rabbit casein were isolated and identified. Their sequences were found as follows: HVEQLLR (residues 50–56 of β-casein), ILPFIQSLFPFAER (residues 64–77 of β-casein), and FHLGHLK (residues 19–25 of αs1-casein). The three peptides were synthesized and found to exert antibacterial effect against Gram positive bacteria only. Proteolytic digestion of rabbit casein by chymotrypsin, pepsin and clostripain yielded several peptide fragments with antibacterial activity. Since antibiotic peptides can be released from casein during the digestion of milk proteins, our results suggest a possible antibacterial function of rabbit caseins. It is conceivable that antibacterial peptides can be generated by endopeptidases of the mammalian gastrointestinal tract possibly providing protection for new-born rabbits against aggression of micro-organisms.

The production of similar or even identical bacteriocins by different lactic acid bacteria is not a rare event. To take advantage of this finding, genetic determinants of the Lactobacillus K7 bacteriocins were tested for putative homologies with previously described bacteriocins of the Lactobacillus acidophilus group through polymerase chain reaction (PCR). Among specific primer pairs of seven known bacteriocins, derived from their respective sequences, only acidocin LF221 A and B primers amplified fragments in chromosomal DNA of K7 strain that revealed strong similarity over small regions of LF221 bacteriocins. Treatment of Lactobacillus K7 with ethidium bromide and mitomycin C was ineffective in generating non-bacteriocinogenic derivatives and had no impact on plasmid loss either. Classification studies elucidated Lactobacillus K7 as a member of the Lactobacillus gasseri species.

An objective method for determining the coagulum cutting time is needed to improve consistency and processing efficiency of goats' cheese. A fibre optic sensor was used to measure the backscatter of near-infrared radiation at 880 nm during the coagulation of skimmed goats' milk for the purpose of predicting coagulum cutting time. A randomised block design, replicated three times, was used to test the effect of three protein concentrations (3, 5 and 7% (w/w)) on diffuse reflectance parameters for cutting time prediction of milk coagulated at five different temperatures (20, 25, 30, 35 and 40°C) to assure a wide range of coagulation rates. The inclusion of a protein term in the existing algorithms was essential to reduce the standard error of prediction to under 6·2 min. An algorithm including a time-based parameter and a protein term, Tcut=β0T2 min (1+γ% Protein) was found to predict cutting time with a SEP of 2·42 min and an R2 of 0·98. γ was considered constant (γ=−0·0674, goats' milk) representing the protein effect on β0. Algorithms using response-based parameters (such as change in reflectance ratio) and the composition parameter protein required additional regression parameters such as temperature and an intercept term to predict the cutting time with the same precision as algorithms using only time-based parameters. Time-based parameters were found to decrease proportionally with increasing temperature and decreasing protein concentration. Response-based and mixed-based parameters were found to decrease with decreasing temperature. Reflectance ratio at cutting time did not significantly change with protein concentration for skimmed goats' milk. The activation energy of κ-casein hydrolysis was calculated based on changes in reflectance profile parameters and was found to be in the range 63–72 kJ mol−1.

In the context of identifying intrinsic time temperature integrators (TTIs) for evaluating thermal processing of milk, the possible influence of seasonal variation in milk composition on the applicability of alkaline phosphatase (ALP), hydroxymethylfurfural (HMF), lactulose and furosine for process impact assessment was studied. Hereto inactivation and formation kinetics of these indicators were analysed in milk samples collected over a one year period. Based on previous research (isothermal and non-isothermal heating conditions) it was assumed that thermal inactivation of ALP followed first order kinetics, and formation of HMF, lactulose and furosine could be described by pseudo-zero order kinetics. This allowed the use of a simplified experimental design to obtain the kinetic parameters. Based on 90% joint confidence regions, kinetic parameter values differed significantly between different batches of milk for all four indicators. Additionally, variable ALP activities and HMF concentrations were observed in the raw milk samples. Nevertheless, the observed differences did not seem to restrict the applicability of the selected TTIs as illustrated by the construction of temperature time tolerance (TTT-) diagrams. Through these diagrams, use of the TTIs implied an uncertainty of approximately 2°C between equivalent processes.

The effects of activation of the lactoperoxidase (LPO) system by H2O2–NaSCN and hydrogen peroxide (H2O2) on the accessibility of sulphydryl groups (SH) in skimmed milk, and on the dynamic rheological properties of the resulting yoghurt were investigated. Four different concentrations of each reagent (20–80 mg H2O2–NaSCN/kg milk and 100–400 mg H2O2/kg milk) were compared. Clear negative correlations were noted between the accessibility of SH groups and both LPO activation rate and H2O2 concentration. Also the native PAGE pattern of the heat-treated samples showed that with increase in the H2O2–NaSCN and H2O2 concentrations, the level of interaction between β-lactoglobulin (β-lg) and κ-casein (κ-CN) decreased. The complex modulus (G*) of skimmed milk yoghurts declined gradually with the decrease in the concentration of accessible SH groups accordingly. Tan δ values of yoghurt samples were found to be different from the control, but close to each other, indicating that protein interaction forces taking place in the formation of gel networks of treated yoghurts were different from the control.

In dry cow therapy (DCT), antibiotics are usually injected direct into the mammary gland after the last milking of the lactation. Most products are designed to eliminate existing intramammary infections (IMI) caused by Staphylococcus aureus and Streptococcus agalactiae and to prevent the establishment of new IMI caused by major pathogens during the early non-lactating period (Nickerson et al. 1999). Systemic therapy has been attempted as a way of improving the cure rates of intramammary treatments of clinical and subclinical mastitis during lactation (Ziv, 1980a; Calvinho et al. 1988; Owens et al. 1988). For systemic therapy to be useful, effective passage of the drug from blood to the foci of infection must be achieved. Maintaining an effective antibacterial concentration in the udder depends on the physicochemical properties of the drug, the dose, the bioavailability of the injected formulation, and the sensitivity of the pathogen (Ziv et al. 1980a, b). The theoretical pharmacokinetic and pharmacodynamic basis for systemic DCT has been reported (Soback, 1988). When used in an attempt to improve cure rates of IMI, subcutaneous injection of tilmicosin at drying-off was ineffective against Staph. aureus IMI (Nickerson et al. 1999) and intramuscular (i.m.) oxytetracycline, in combination with intramammary cephapirin dry-cow treatment did not improve the cure rate for Staph. aureus mastitis (Soback et al. 1990a; Erskine et al. 1994). Nevertheless, i.m. tylosin treatment 2 weeks before the expected day of calving decreased IMI after calving (Zecconi et al. 1999) and subcutaneous norfloxacin nicotinate applied at the cessation of milking proved useful in controlling Staph. aureus infections (Soback et al. 1990a). Moreover, the incidence of this pathogen after systemic DCT may be significantly lower compared with untreated control groups (Soback et al. 1990a, b).

Caseins (αs1, β, αs2, e κ) represent about 80% of the whole protein content of ruminant milk. Each of these proteins is encoded by single copy genes (CSN1S1, CSN2, CSN1S2 and CSN3, respectively) clustered on a ∼200-kb segment of chromosome 6 (Ferretti et al. 1990; Gallagher et al. 1994) in the order: CSN1S1, CSN2, CSN1S2 and CSN3 (Mercier & Vilotte, 1993). Furthermore, in cattle and goat CSN1S1 and CSN2 are convergently transcribed (Leroux & Martin, 1996; Rijnkles et al. 1997) and are only 20 and 12 kb apart, respectively.

Milk is a heterogeneous fluid in which the colloidal phase is homogeneously dispersed in the aqueous phase. Calcium is partitioned between the colloidal and aqueous phases and is in complex electrochemical equilibrium with several major milk components. In human and bovine milk, calcium is mainly distributed between the aqueous and casein micelle in the colloidal phases (Holt & Jenness, 1984; Neville et al. 1994). Caseins form a complex micelle structure that contains approximately 25000 phosphorylated monomers that react with calcium phosphate complexes in the milk to bind 20–40 mole calcium per mole casein (Holt & Jenness, 1984; Neville et al. 1994). Thus, the distribution of calcium between the colloidal and aqueous phases appears to be governed by the level of casein in the milk (Holt & Jenness, 1984; Neville et al. 1994). In human milk, the casein level is low; ∼25% of calcium is associated with casein, whereas in cows and goats the corresponding figure is higher at ∼65%. In rats, casein levels are among the highest in mammals and 95% of calcium is associated with casein (Neville et al. 1994). In the aqueous phase, calcium is divided among ionic calcium (Ca2+), calcium citrate and calcium phosphate. Calcium citrate and calcium phosphate constitute most of the aqueous calcium in bovine milk in contrast to less than 50% in human milk (Holt & Jenness, 1984; Neville et al. 1994). As the concentration of Ca2+ in milk appears to be essential in preserving the integrity of the mammary tight junctions during lactation (Neville & Peaker, 1981), it is important to follow its concentration precisely in mammary secretions in different stages of the reproduction cycle.

Brucellae are Gram-negative bacteria with a predilection for the reticuloendothelial system and the reproductive tract of cattle, sheep, goats and pigs. In these species brucellosis is of great economic significance because of the resulting abortion and infertility (Samartino et al. 1993; Corbel, 1997; Altecruse et al. 1998). Human brucellosis, caused by contamination from infected animals (percutaneous, conjunctival or nasal infection), is very common and remains a serious health hazard in Mediterranean Europe, western Asia, and parts of Africa and South America (Boschiroli et al. 2001). Raw milk, butter, ice-cream and unpasteurized cheese are also potential vehicles for the transmission of Brucella.

Bacterial decarboxylases and aminotransferases may be involved in the production of flavour or off-flavour compounds from aromatic amino acids during cheese ripening. Transamination is one of the first steps in amino acid catabolism for both lactococci and lactobacilli (Gao et al. 1997; Klein et al. 2001). Biologically active amines, produced by decarboxylation, such as tyramine, phenylethylamine, tryptamine, histamine, cadaverine and putrescine, known as biogenic amines, have been found in cheese and can cause migraine and hypertension in susceptible consumers (McSweeney & Sousa, 2000).

Gelation of acidified milk at pH[ges ]5, after heat treatments is a well known phenomenon, due to the precipitation of whey proteins, and especially β-lactoglobulin onto κ-casein (Sawyer, 1969). High heat treatments cause denaturation of whey proteins which associate with κ-casein through disulphide interchange reactions (Hill, 1989). Since their charge is reduced, the denatured proteins associated with casein micelles become susceptible to aggregation when milk is then acidified, which promotes enhanced protein–protein interactions (Lucey et al. 1997). The gelation phenomenon involves disulphide bonds (Hashizume & Sato, 1988; Goddard, 1996) which are responsible for the gel firmness (Goddard, 1996). However, other interactions between proteins can occur, such as hydrogen and hydrophobic bonds, especially at the initial stage of interactions (Haque et al. 1987; Haque & Kinsella, 1988; Jang & Swaisgood, 1990). It is therefore relevant to investigate a possible contribution of weak linkages to the gel structure and firmness.