Marker-assisted selection may provide the opportunity to make significant genetic gains in the improvement of economically important traits in livestock (Soller & Beckmann, 1983; Smith & Simpson, 1986). Implementation of this approach will first require identification of candidate genes or anonymous gene markers associated with the traits of interest. Candidate genes are those with a known relationship between physiological or biochemical processes and an economically important trait. In dairy cattle, genes associated with mammary gland growth, development and function are excellent candidate genes for milk production traits.

The polyamines are low molecular mass polycations that influence cell proliferation and growth (Tabor & Tabor, 1984; Pegg, 1986). Ornithine decarboxylase (ODC, EC 4.1.1.17) catalyses the conversion of ornithine to putrescine, the rate-limiting step in polyamine biosynthesis (Pegg, 1986). The level of ODC is induced in quiescent cells exposed to a wide variety of stimuli such as growth hormone, corticosteroids, testosterone and growth factors (Tabor & Tabor, 1984). There is ample evidence that growth factors influence morphogenesis and differentiation of the mammary gland (Imagawa et al. 1994). Trophic hormones that are associated with lactation, such as prolactin and growth hormone, are also required to induce differentiation of bovine mammary epithelial cells (Huynh et al. 1991; Flint & Gardner, 1994), and both mitogenic and lactogenic effects of prolactin, insulin and hydrocortisone appear to be mediated through the polyamine pathway (Rillema et al. 1977; Bedford & Zadworny, 1990; Golden & Rillema, 1993). Strange et al. (1992) have shown that ODC is involved in synthesis of a milk component; ODC has elevated expression in the lactating mammary gland, which declines sharply after weaning. It is therefore possible that particular variants of ODC could be associated with increased mammary gland function and thus influence milk related traits.

The objectives of the present study were to estimate allelic frequencies of ODC polymorphisms in Holstein bulls and to evaluate further the genotypic effects of ODC variants on milk production traits.

Kefir is a traditional fermented milk originating many centuries ago in the Caucasian mountains. It is produced by fermentative activity of ‘kefir grains’ consisting mainly of lactococci, lactobacilli and yeasts in a protein–polysaccharide matrix. The grains contain a relatively stable and specific balance of microorganisms which exist in a complex symbiotic relationship. The grains grow in the process of kefir making only from pre-existing grains (Saloff-Coste, 1996). When kefir grains are allowed to grow in milk, microorganisms are shed from the grains into milk where they continue to multiply with the production of acid, flavour and physicochemical changes.

The traditional method of kefir making is currently by adding kefir grains directly as starter to milk that has been pasteurized and cooled to 20–25°C. After a period of fermentation lasting ∼24 h, the grains are removed by filtration and the beverage is ready for consumption (Saloff-Coste, 1996). Kefir from which the grains have been removed may be used as starter. However, this fermented milk cannot be used for subsequent inoculations to make an acceptable product, because the original balance of microorganisms has been disrupted (Kroger, 1993).

The complex microbiological composition of kefir grains explains why it is difficult to obtain starter with the optimal and constant composition necessary for a regular production of kefir of standard quality (Koroleva, 1988a). Studies have been undertaken to establish cultivation conditions[ratio ]grain[ratio ]milk ratio, cultivation temperature, period of time and conditions prior to separation of grains from the fermented milk, shaking conditions for agitation of milk with the grains in the course of fermentation, washing of kefir grains and so on. All these factors influence the microflora of the kefir starter and fermented milk. There are no rules about household manufacture of kefir. Different reports indicate a wide range of grain[ratio ]milk ratios for kefir making. Bottazzi & Bianchi (1980), Marshall & Cole (1985), Merin & Rosenthal (1986), Mann (1989), Hosono et al. (1990) and Kroger (1993) employed 20–50 g/l while Koroleva (1988a) employed 20–100 g kefir grain/l and Marshall et al. (1984) and Neve (1992) 50–100 g/l. Rea et al. (1996) used 1 g kefir grain/l as starter and 200 g starter in the form of kefir grains is recommended by Hansen for the fermentation of 1 l milk (Marshall & Cole, 1985). A critical control point in kefir manufacture to obtain a product with constant quality is the standardization of the kefir grain[ratio ]milk ratio. Koroleva (1988b) claimed that it is better to use kefir grains as starter for kefir production and, at the same time, to decrease the amount of inoculum.

The purpose of this study was to evaluate the effects of changes in the kefir grain[ratio ]milk ratio (quantity of kefir grains inoculated into the milk) on microflora composition, acidity, apparent viscosity and carbon dioxide content of fermented milk.

The principles of today's machine milking techniques have developed since the early 1900s. The original intention to imitate the sucking action of the calf had to be abandoned owing to technical difficulties. Further developments were made on a largely empirical basis and milking technique became a specific complex of tasks, problems and solutions. Consequently, the sucking of the calf was rejected as a model for machine milking. The sucking behaviour and the application of vacuum and/or pressure by the sucking mammalian offspring have been adapted through evolution. Any eventual changes due to ‘recent’ breeding and development of the milking machine remain marginal compared with evolutionary changes.

Since the cineradiographic techniques used by Ardran et al. (1957, 1958) it was believed that sucking calves use mainly pressure to transfer a squirt of milk from the proximally closed teat cistern and through the teat canal. The calf creates the pressure by compressing the teat between the tongue and the hard palate from the base of the teat towards the teat end. Cowie (1977) summarized these results as follows: ‘Sucking, that is the production of vacuum within the mouth cavity, is not an essential feature in suckling, although it aids the process. … The act of suckling is thus analogous to hand milking …’. These conclusions were based on visually analysed recordings. McDonald & Witzel (1966) measured pressure in teat cisterns and vacuum at the teat end simultaneously during suckling. Average maximum pressure in the teat cistern was 36·6 kPa and vacuum at the end of the teat averaged −34·6 kPa. These authors concluded that the average differential pressure across the teat canal was 71 kPa. However, this method of calculation gives only the maximal pressure difference and not the average pressure applied by the calf. They noted that vacuum in the teat cistern occurred during the resting phase of a suckling cycle. The calf's sucking technique for single teats during a meal has been analysed (Mayntz, 1996). Further details of pressure and/or vacuum application during suckling at a specific teat remain unknown.

The objective of the present study was to extend the current knowledge about milk extraction by sucking calves through continuous and simultaneous measurements of positive and negative pressure in the teat cistern and in the mouth cavity.

In Italy buffalo milk is an important animal product utilized solely for the manufacture of Mozzarella cheese. Of an estimated population of 200000 buffalo there are ∼25000 controlled animals. The average milk production, expressed over 270 d lactation, is 2000 kg/head with average fat and protein contents of 82·6 and 46·4 g/l respectively (Associazione Italiana Allevatori, 1996). In recent years there has been a steady increase in the number of dairy buffalo replacing dairy cows as a consequence of the European Union quota system.

The cheesemaking qualities of milk depend on many factors, the most important of which are the concentrations of intact casein and fat. Milk in which casein has been broken down by proteolytic enzymes is of less value to cheese manufacturers (Lucey & Kelly, 1994). Plasmin (EC 3.4.21.7), the most important endogenous milk proteinase, occurs in milk together with its inactive proenzyme, plasminogen (Schaar & Funke, 1986). Plasmin hydrolyses αs-casein and β-casein, although κ-casein has been reported to be resistant (Fox, 1981). However, Andrews & Alichanidis (1983) found κ-casein to be hydrolysed quite rapidly by plasmin. Plasmin activity is higher in mastitic than normal milk (Bastian & Brown, 1996). Stage of lactation affects plasmin activity: late lactation is associated with higher concentrations of plasmin (Gilmore et al. 1995; Baldi et al. 1996). Thus, plasmin could be a major problem in herds with seasonal breeding such as buffalo, which progress through lactation in synchrony and are therefore at a similar stage of lactation at a given time.

The cascade of reactions leading to plasminogen activation is regulated by a complex network of molecular interactions between plasminogen activators (PA; EC 3.4.21.31) and at least three types of specific PA inhibitors (PAI-1; PAI-2; PAI-3; Saksela, 1985; Politis, 1996). There are two types of highly specific PA: tissue-PA (t-PA) and urokinase-PA (u-PA) (Saksela & Rifkin, 1988). PA activity is higher in mastitic than normal milk (Heegaard et al. 1994a). However, previous studies on PA in milk were carried out on bovine and caprine milk. No information is available on the presence and type of PA in buffalo milk. Thus the objective of the present study was to determine the level and type of PA in different fractions of buffalo milk: casein, serum, and somatic cells.

Economic losses caused by enzootic bovine leukosis (EBL) have been of interest since World War II, when the neoplastic form of EBL increased dramatically in Europe. Olson (1974) and House et al. (1975) showed that animals with lymphosarcoma caused by the bovine leukosis virus (BLV) had reduced milk yields, a less efficient reproductive performance and high veterinary costs and mortality rates, while many carcasses were rejected at slaughter. However, the actual impact of BLV infection in cattle without lymphosarcoma is not clear. The purpose of the study reported here was to compare some productive and reproductive responses of cattle that were antibody-positive (BLV+) or negative (BLV−) for BLV.

Holstein dairy cows in commercial dairy farms were used in this study. Blood samples were collected and subjected to BLV serological examination by the agar gel immunodiffusion test of Miller & van der Maaten (1976). Animals were then grouped as BLV+ or BLV− according to their serological response to the BLV antigen. Productive and reproductive histories were obtained from individual animal records and the following factors were considered: milk production, calving interval and birth rate. For milk production, we had the daily milk yields of 547 animals, and for calving interval the time between two successive parturitions for 444 cows. These values were examined by ANOVA and when this was significant a Student's t test was carried out for each age group. Birth rates, the percentage of animals that calved in 1 year, were available for 557 animals and were examined with the Z-two proportion test. For all analyses, P<0·05 was considered significant.

During the last two decades the traditional subjective appraisal of the body fat stores in farm animals, made by eye and touch, has been rationalized by the introduction of numerical systems of rating specific anatomical points.

Palpation of the lumbar vertebrae, the pin and hook bones (tail head) (Lowman et al. 1973), and, occasionally, width behind the shoulders (Treacher et al. 1986) provides an assessment of the fatness of the animal. This is calibrated from standard photographic charts, in the use of which the operator is trained. Body condition scoring (CS) by this method has been widely developed for dairy cows (Earle, 1976; Mulvany, 1977; MacCarthy, 1978; Scott & Smeaton, 1980; Wildman et al. 1982; Aalseth et al. 1983; Anon. 1986; Garcia Paloma, 1990).

The actual numerical scales have varied among systems. Thus over the range from very thin to very fat cows the scale is 1, 2…7, 8 (Earle, 1976) and 0½, 1,…4½, 5 (Mulvany, 1977), for example. CS is thus a discrete variate with a limited number of readings. Several authors have sought to decrease the division size, for example 1, 1 1/3; 1 2/3;…4 2/3, 5 (Ducker et al. 1985a, see also Dewhurst et al. 1996) and by using the means of the separate values obtained by two operators judging CS independently (W. H. Broster, V. J. Broster, A. J. Clements, J. W. Siviter and T. Smith, unpublished results). In this review CS will be stated in the units given by Mulvany (1977), to which other ranges have been scaled. Inspection of CS data has usually been conducted by analysis of variance, but also by the Mann–Whitney signed rank test (Moorby et al. 1996).

Evans (1978) and Nicoll (1981) studied the variation in recording CS. Both investigators concluded that a system of observation by two independent assessors per recording occasion is advantageous and that revision training is necessary to maintain operator standardization. Of total variance in CS, some 60–70% was found attributable to ‘between-animal’ differences, <5% to assessor variation and <10% to animal×assessor variation. Croxton & Stollard (1976) found good repeatability of CS measurements ‘between’ and ‘within’ operators. It is regrettable that few reports of experimentation give full details of method of and number of operators engaged in body score recording.

Previously we have examined the inactivation of unpurified extracellular proteinase from Pseudomonas fluorescens 22F diluted in demineralized water (Schokker & van Boekel, 1998) in the range 40–70°C. It appeared that the inactivation was most probably caused by intermolecular autoproteolysis, which is the hydrolysis of unfolded proteinase molecules by native (not yet unfolded) molecules. It has been reported that purification of proteinases from Pseudomonas spp. enhances the susceptibility of the proteinase to autoproteolysis (Barach et al. 1976; Griffiths et al. 1981; Leinmüller & Christophersen, 1982; Kroll, 1989; Kumura et al. 1991). On the other hand, when the proteinase is heated in milk or when proteins are added to the enzyme solution, the rate of inactivation by autoproteolysis diminishes (Barach et al. 1978; Kroll & Klostermeyer, 1984; Stepaniak et al. 1991). Apparently, proteins stabilize the proteinase against inactivation by autoproteolysis.

Substrate or other ligands stabilize many enzymes against limited proteolysis. Binding of these substances to the enzyme molecule, either to the catalytic centre or to amino acid residues on the enzyme molecule surface, may impose steric difficulties so that the susceptible peptide bonds are protected against proteolysis (Mihalyi, 1978). Such binding may also cause a conformational change of the enzyme molecule, such that susceptible peptide bonds cannot be attacked or that the conformation is stabilized against unfolding (Mihalyi, 1978). In the latter case an increase in the denaturation temperature (Td) would be expected.

In the case of proteinases, addition of substrate to the enzyme solution may protect the enzyme by a third mechanism. Besides autoproteolysis of the proteinase, the added proteins can be digested. An enzyme molecule digesting a protein is not available at the same time for autoproteolysis, so that the substrate may act as a competitive inhibitor against autoproteolysis.

The aim of this study was to determine the mechanism of protection of the proteinase from Ps. fluorescens 22F by sodium caseinate.

Bovine milk contains a variety of proteins and peptides that are biologically active (Ogra & Ogra, 1978; Duncan & McArthur, 1981; Newby et al. 1982; Juto, 1985; Stoeck et al. 1989; Mincheva-Nilsson et al. 1990; Watson, 1990; Barta et al. 1991; Politis et al. 1991; Fiat et al. 1993). Our laboratory has a long-term interest in some purified milk proteins, particularly lactoferrin (LF), lactoperoxidase (LP) and β-casein (β-CN), which have been shown to be immunologically significant. Some of our recent studies on these bovine milk proteins, particularly β-CN, indicated that their in vitro immunological effects did not always parallel their in vivo activities (Wong et al. 1996a, b; 1997a, b). This study was designed to investigate and compare the capacity of these purified bovine milk proteins to modulate a range of components that are vital to in vivo immune responses in sheep, with a view to providing further information on their potential in biomedical applications. To achieve this objective, a sensitive lymphatic cannulation model was employed that allows in vivo immune components and their functions to be measured in lymph collected under physiological conditions.